IMMUNOPATHOLOGY OF CHRONIC INFLAMMATION

[Guest guest]

This is very long and technical . . . but worth taking

the time to read carefully -

It would be a good one to give to doctors who don't

believe implants cause problems! - Rogene

> > From: " Toxic Discovery "

> <toxicdiscovery@...>

> <Undisclosed-Recipient:;>

> Subject: Emailing: IMMOPATH

> Date: Thu, 1 Sep 2005 18:50:29 -0500

>

> Section IV - Immunopathology of

> ChronicInflammation(Plaintiffs' Science Submission

> to National SciencePanel)

> IV. IMMUNOPATHOLOGY OF CHRONIC INFLAMMATION

>

>

>

> A. INTRODUCTION

> The Center for Devices and Radiological Health

> recently redrafted their " Immunotoxicity Testing

> Framework " to provide guidance on the types of

> toxicity testing that should be considered for a

> medical device or constituent materials.(260) Their

> flow chart includes within it consideration of

> immunological effects such as inflammation,

> hypersensitivity, and immunostimulation.

> Specifically, the CDRH was concerned with chronic

> inflammation in prolonged or permanent implants

> because this " could develop into other more serious

> immunological effects. " If testing indicates chronic

> inflammation or any other adverse immunological

> effects from a medical device or constituent

> material, manufacturers are directed to conduct

> testing examining cellular responses:

> histopathology, humoral response, T-cells, natural

> killer cells, macrophages with an emphasis on

> cytokines IL-1, TNF-, IL-6, and TNF-; granulocytes

> (basophils, eosinophils and/or neutrophils), host

> resistance and clinical symptoms (allergy, skin

> rash, urticaria, edema, and lymphadenopathy).

>

> The CDRH's model for analyzing immunological effects

> relating to pathologic findings is a logical model

> for analyzing the findings on silicone gel breast

> implants. This section will track that model in

> reviewing the scientific evidence on the

> significance of chronic inflammation seen in women

> implanted with silicone gel breast implants. The

> Immunology section immediately follows and will

> address related issues of autoimmunity,

> immunostimulation, immunosuppression, and delayed

> hypersensitivity.

>

> There are several sources of information to explain

> how chronic inflammation induces an immune response.

> One source is the literature on chronic

> inflammation, including numerous articles and

> internal manufacturer studies, showing that the

> silicone gel is broken down into thousands of

> microdroplets; the gel bleed itself can consist of

> microdroplets of silicone which are continuously

> released into the body; and that minute particles or

> shards of silicone induce a severe, chronic

> inflammatory reaction and that this reaction is not

> limited to silicone breast implants. Another source

> is the articles documenting that cytokines are

> secreted by activated macrophages found in breast

> implant capsules and silicone-exposed tissue and

> that the inflammatory process is immune mediated.

> This section will also address how the chronic

> inflammatory process manifests itself in women with

> silicone gel breast implants by signs and symptoms

> of fatigue, myalgias, arthralgias, fever, cognitive

> dysfunction, and other neuropathies.

>

>

> B. INFLAMMATION IS MEDIATED BY PARTICLE SIZE

>

>

>

> 1. Silicone Gel Microdroplets From Implant Bleed And

> Gel Subdivision by Exposed Tissue

> As silicone gel slowly and continuously bleeds from

> implants or is released into the body following a

> rupture, thousands of microdroplets are exposed to

> the cellular constituents of the implant capsule and

> ultimately, to the breast tissue. These silicone

> microdroplets are coated with native proteins, some

> of which become denatured and are phagocytized by

> macrophages or surrounded by giant cells if the

> particle is immediately too large for ingestion by

> one cell. Eventually, a whole range of inflammatory

> and immune-mediated cells move to the capsule and

> silicone-exposed tissue. These are usually most

> densely seen around the new blood vessels feeding

> the tissue at the outermost growing edge of the

> implant capsule, where the newest collagen fibers

> are being assembled and laid down. As noted above,

> the cells involved are not only monocytes and

> macrophages, but polymorphonuclear cells,

> eosinophils, plasma cells, lymphocytes, and mast

> cells.

>

> The earliest reports of microdroplets of silicone

> found in capsular tissue dates to a 1974 article by

> Wilflingseder who examined 53 capsular specimens

> from 28 patients who had experienced severe

> contracture.(261) The contracted capsules had

> " droplets " of material which were shown to be

> silicone. The control capsules taken from explanted

> women who had not experienced contracture did not

> contain any silicone. On further examination,

> Wilflingseder found that the silicone microdroplets

> were 2 to 25 in diameter and appeared to be engulfed

> by macrophage cells. He concluded that the

> contractive fibrosis was a result of the " grazing "

> of silicone particles away from the breast implant

> shell causing a phagocytic response that led to

> contracture. Barker also found microdroplets in

> capsular tissue in 1978, but attributed the

> particles to microparticles of silicone released

> through the gel bleed.(262) Gayou's data in 1979

> measured the silicone gel droplets or particles at

> 10 to 75 in diameter.(263) Sanger also observed

> small microdroplets of silicone gel and noted that

> as new small droplets of gel break loose, a more

> mature cellular infiltrate with fibrosis surrounded

> the larger droplets.(264) This contrasted with the

> intense cellular inflammation without fibrosis found

> around the smaller silicone gel droplets.

>

> Dow Corning's internal research produced similar

> results.(265) In 1978, a study by Lentz showed that

> silicone gel implanted subcutaneously in rabbits was

> subdivided by bands of fibrous connective

> tissue.(266) The report states that:

>

> [in] most of the six month subcutaneous implant

> sites, there was obvious subdivision of the gel mass

> by septa of tissue similar in appearance to the

> surrounding capsule. The gross architecture of these

> subdivided sites ranged from apparent discrete

> columns of tissue penetrating the gel to discrete

> spheroidal masses of gel, all separated by the

> intervening tissue. In some cases gross subdivision

> was not noted; however, this fragmentation was

> evident microscopically in many ways as noted below.

> Subdivision of the gel was also noted at earlier

> times, although this reaction was evidently

> progressive as indicated by the increasing frequency

> of subdivision with increasing implant residence

> time.

> Based on these findings and the presence of

> lymphocytes, multinucleated giant cells, macrophages

> and eosinophils which indicated an inflammatory

> process, Dow Corning wrote that use of implantable

> gel was not supported. In deciding not to proceed

> with commercialization, they concluded that " the

> long term localization of the gel is uncertain as is

> the endpoint of the tissue reaction. To permit use

> of this gel by instillation, future studies should

> address the questions of whether the reaction stops

> short of complete dispersal of the gel, whether gel

> fragments are carried to remote sites and if so the

> fate of this material. . . . "

>

> Heyer-Schulte noted a similar fragmentation of the

> gel in a 1978 gel bolus study.(267) Labs,

> which performed the study, reported that tissue

> samples from four rabbits injected subcutaneously

> with silicone gel showed fibroblasts, scattered

> mononuclear cells, scattered particles of gel within

> the capsule, and an inflammatory reaction seen

> around the multiple gel particles. They concluded

> that, " it is the breaking-up of the gel into small

> particles that seems to provoke the greatest

> inflammatory response with a foreign body reaction

> and fibrous encystment [sic] of the gel particles. "

> The reaction varied from minimal subdivision at the

> periphery of the gel mass to extensive fragmentation

> of the mass into several discrete pockets. The

> initial indications of subdivision were discernable,

> however, as delicate fingers of connective tissue

> ingrowth or as small isolated gel masses as early as

> two weeks in some animals and at four weeks in

> others.

>

> Similar reports of gel subdivision by fibrous tissue

> were made in internal manufacturer reports including

> a 1985 Dow Corning report in which gel partitioning

> or subdivision was noted to " greatly increase the

> surface area of gel exposed to tissues. This

> increased surface area may result in more rapid

> removal of the gel from the implant site by

> phagocytic cells. " (268) Further, Dow Corning wrote,

> " [t]he work presented demonstrates that in the rat,

> silicone gel is quickly encapsulated with

> fibrous-tissue. With time, this fibrous tissue

> penetrates the gel mass and partitions it into

> numerous smaller masses. A similar response has also

> been reported to occur in the rabbit. Since this

> growth of fibrous tissue into silicone gel does not

> appear to be species specific, it is reasonable to

> postulate a similar phenomenon may occur in humans. "

>

> In 1987, when plastic surgeon Dr. Vinnik

> inquired whether the body could degrade silicone in

> vivo, Dow responded that:

>

> [w]e do know that silicone gel can be physically

> broken down into smaller particles either

> mechanically or by manipulation in the presence of

> water or body fluids. This phenomenon is similar to

> the effect of mixing oil and water where the oil

> breaks into droplets. Neither the oil nor the water

> has changed in chemical composition but the oil

> behaves differently physically.

> In the body, if (1) both the implant shell and the

> scar capsule surrounding it, tear, allowing the

> silicone gel to mix with tissue fluids, and (2) if

> the gel is then manipulated, such as would occur

> with massage exercise, a change in physical behavior

> can occur. As was discussed during your 1984 tour of

> our medical plant in Michigan, gel, once broken down

> into particles, can migrate ....

> It is the combination of gel in contact with body

> fluids and manipulation which causes the physical

> breakdown....(269)

> On the subject of cohesivity, numerous samples of

> gel returned by you have been compared to retainer

> samples as you requested. There is, as expected, a

> physical change in the gel you returned based upon

> its contact with body fluids. Gel in its particle

> form is less cohesive than uncontaminated gel just

> as oil combined with water has a different physical

> state than oil alone.

>

> Dow Corning again confirmed these findings in

> 1996.(270)

>

> 2. Implant Failure with Migration of Particulate

> Silicone Fragments Is a Complication That

> Demonstrates a Systemic, Rather than Local,

> Complication

> From the 1970s to the present, findings on silicone

> particle debris in exposed tissue have been

> strikingly consistent, i.e., the breakdown of

> silicone into microscopic particles which can

> transported cellularly occurs in virtually all types

> of silicone implants and can result in an intense,

> chronic inflammation in the exposed tissue, joints,

> and lymph nodes. Numerous reports of silicone

> particle reactions are contained in Table 2 in the

> Pathology section,(271) but a 1992 review of the

> literature by Sammarco contains a representative

> summary.(272) Sammarco concluded that:

>

> ilicone, with a high coefficient of friction

> and poor wear characteristics, is unsuitable for

> specific joints, i.e., temporomandibular, radial

> head, trapezium, and metatarsophalangeal joints, in

> which there is excessive loading of the joint

> surfaces and great risk of abrasion of the

> articulating prothesis....

> Implant failure with migration of particulate

> silicone fragments is a complication that

> demonstrates a systemic, rather than local,

> complication.

> (Emphasis added).

> Barrett, at the Department of Urology at the Mayo

> Clinic, also reported on silicone particalization

> from silicone genitourinary devices with subsequent

> migration of the particles to the surrounding tissue

> and the inguinal lymph nodes.(273) Naidu and

> colleagues at the Departments of Orthopedic Surgery

> at Pennsylvania State University, the Department of

> Materials Science and Engineering at the University

> of Pennsylvania, and the Department of Rheumatology

> at the Veterans Administration Hospital in

> Philadelphia concluded in 1996 that, " reakdown of

> silicone elastomer particles evokes a severe

> inflammatory response. " (274) Naidu observed that:

>

> [t]he size of the particulate material plays an

> important role in the type of tissue-cellular

> response. Bulk implantation of metallic or plastic

> objects into bone or muscle results in a fibrous

> membrane. In sharp contrast, implantation of the

> same materials in the form of a powder results in a

> marked cellular inflammatory reaction. . . .

> The exact type of cells that are involved and are

> thought to be most intimately involved in the

> inflammatory response are not well elucidated.

> Phagocytic cells, including macrophages,

> histiocytes, multinucleated giant cells, and

> osteoclasts probably all have a role in the loss of

> integrity at the bone-elastomer implant interface in

> response to elastomer particulate debris. Monocytes

> and macrophages have the capacity to produce

> cytokines (TNF); cytokines stimulate bone

> resorption, activate osteoclasts, and attract

> polymorphonuclear leukocytes. [footnotes omitted]

> Goldring et al [footnote omitted] described the

> interface membrane surrounding aseptically loosened

> cemented total hip components as synovial-like and

> capable of producing prostaglandin E2 (PGE2) and

> collagenase. Many other pathologic studies have

> reported similar histologic findings in the membrane

> between bone and acrylic cement with implants and

> have also suggested its involvement in osteolysis

> and joint failure. [footnotes omitted]

> Using Dow Corning Silastic silicone finger joints,

> small silicone elastomer pieces were generated were

> in the laboratory ranging in size of under 10 m. The

> particles were filtered using a 0.22 m filter and

> then resuspended in a sterile saline solution.

> Scanning electron microscopy (SEM) characterized the

> size and morphology. The researchers found:

>

> [p]articles in saline were counted on a

> hemocytometer, and an initial pilot dose response

> showed that silicone in the concentration of

> 106particles/mL caused a measurable white blood cell

> (WBC) count increase in the pouch exudate. To elicit

> a measurable inflammatory response, 107particles/mL

> of MSU were needed. After preparing samples with the

> above concentration of particles, the particles were

> sterilized with gamma irradiation (2.5 mrad). Five

> milliliters of each of the various suspensions of

> PMMA, silicone, and MSU were drawn into 10-mL

> syringes using large-bore (16-gauge) needles in

> preparation for injection into the rat subcutaneous

> air pouch.

> Twenty rats were injected with 5mL Silastic silicone

> elastomer particles (106 particles/mL), twenty rats

> were injected with 5 mL MSU particles

> (107particles/mL), and twenty were injected with 5

> mL PMMA particles (107 particles/mL). Exudate was

> retrieved from five animals in each group at the 6,

> 24, 48 and 72 hour time points and analyzed for TNF

> and PGE2. White blood count was the highest in the

> silicone group at 6 and 24 hours after injection.

> TNF levels were highest in the silicone group at 6

> and 24 hours, and PGE2 was the highest in the

> silicone group at 24 hours. Naidu also found silica

> particles in the tissue and suggested that:

>

> during the mechanical grinding process [to prepare

> the silicone particles for the study], a significant

> amount of silica particles is dissociated from the

> elastomer matrix, creating a more concentrated

> solution of particles than what was measured with

> light microscopy. It may simply be a situation of

> particle overload leading to exuberant inflammation.

>

> Conversely, it may be that the silica filler is

> actually more inflammatory. By creating small

> Silastic silicone particles, more silica surface is

> exposed to the in vivo environment. This increase in

> surface area of silica in contact with the in vivo

> environment may also enhance inflammation. However,

> it is still not clear which component of silicone is

> inflammatory; it may be that the cross-linked

> polymer matrix and the silica filler are truly

> different in inflammatory potential....

> In another paper, this one by Peimer in the Journal

> of Hand Surgery, a group of orthopaedic surgeons and

> pathologists from the University at Buffalo, State

> University of New York, reported in a long-term

> follow-up study of patients implanted with hard

> silicone elastomer finger joints that significant

> complications resulted years after

> implantation.(275)

>

> Peimer wrote:

>

> [t]he host tolerance and prosthetic wear of

> silicone implants may be more of a problem than

> originally thought, mainly because, frequently,

> significant complications do not become apparent

> until years after the operation. . . .

> We became aware over several years that a number

> of patients on whom we had performed silicone

> implants (primarily carpal) returned because of

> secondary symptoms.

> Peimer observed that the average size of the

> particulate matter was 15 in size.(276) Further:

>

> [t]he general histologic process was reminiscent

> of invasive pigmented villonodular synovitis and

> presumably involved similar pathophysiologic

> mechanisms of bone invasion via vascular foramina.

> In contrast, the pathologic conditions found in

> these cases were initiated by the tissue response to

> microparticulate silicone. The synovitis and soft

> tissue inflammation were clearly not caused by a

> connective tissue disease in two patients with that

> diagnosis. . . .

> The generation of silicone microparticles leading

> to secondary synovitis and joint changes has

> recently received attention [citing 11 prior

> studies]. This phenomenon has not been widely

> appreciated as a predictable direct consequence of

> normal use of these implants. We did not fully

> understand the underlying microparticulate

> pathophysiology in our early patients.

> The synovitis is dose related, and the progressive

> destruction was arrested by implant removal and

> synovectomy. . . .

> The time lapse between implant surgery and the

> appearance of clinical symptoms reflects the time

> period for wear and destruction of the implant to

> occur. Severity of clinical symptoms was related to

> the extent of the deterioration and the

> proliferation of microparticles. The only means of

> arresting the destructive synovitis was removal of

> the prosthesis and curettage of the lesions. Once

> this procedure was carried out, all of the patients

> recovered without further incident.

> Some of Peimer's conclusions included the following:

>

>

> a.. Microparticulate silicone is poorly tolerated

> and incites an inflammatory foreign-body tissue

> reaction.

> a.. The severity of the clinical, radiographic,

> and surgical pathologic changes is closely related

> to the interval from the time the prosthesis was

> implanted to the second evaluation -- an average of

> almost 3 years.

> a.. Pathologic changes are dose related, secondary

> to the invasive reactive synovitis. Surgery is

> required to arrest the process.

> Hirakawa and colleagues at the Cleveland Clinic in

> 1996 did similar work examining silicone elastomer

> shards that degrade or are abraded from silicone

> implants. Hirakawa found that there are literally

> billions of microparticles of silicone available for

> macrophage phagocytization and potential

> presentation to the immune system by antigen

> presenting cells.(277) His group examined tissue

> from ten cases of failed silicone wrist, finger or

> elbow implants in which silicone wear debris had

> been identified. All ten patients were found to have

> decreased range of motion consistent with a

> fragmented implant. Light microscopy showed

> particles of foreign material in the connective

> tissue and synovial tissue of all cases. He stated:

>

> [T]hese particles were morphologically similar

> [in] all patients, consisting of somewhat

> translucent granules that were refractile which

> transmitted light, but not apparent using polarized

> light. . . . The particles were similar to those

> previously described as representing 'silicone

> synovitis'. In all specimens, there were

> foreign-body giant cells as well as numerous

> mononuclear histiocytes. The reaction was considered

> to be marked in two patients. Lymphocytes and plasma

> cells were common in all samples, including the

> biopsies obtained from patients with avascular

> necrosis of the lunate as well as those with

> rheumatoid arthritis.

> Hirakawa noted the " uniform infiltrate of

> lymphocytes and plasma cells: "

>

> [a]lso of interest in our cases was the uniform

> infiltrate of lymphocytes and plasma cells. Wear

> debris from most metal and polyethylene total-joint

> prostheses is usually associated with

> granular-appearing histiocyte and foreign-body giant

> cells, with relatively few lymphocytes and plasma

> cells (unless the patient has an underlying

> inflammatory arthropathy). This observation suggests

> that in most patients, the traditional immune

> reaction does not play a major role in the reaction

> to wear debris, but rather that the hystiocytic

> [sic] reaction is a nonspecific response, the

> intensity of which is dependent on the chemistry,

> size, number, and possibly the shape of the

> particles. In the present study, however, we were

> surprised to identify a prominent lymphocytic and

> plasmacytic infiltrate in the two patients who

> received implants for post-traumatic ischemic

> necrosis of the lunate. This inflammation was

> histologically indistinguishable from that commonly

> seen in rheumatoid arthritis. An inflammatory

> reaction, including lymphocytes and plasma cells,

> has also been reported and associated with some of

> the retrieved silicone elastomer breast implants.

> Although no conclusions can be made based on this

> study, the prominence of lymphoplasmacytic

> inflammation associated with silicone debris in our

> cases suggests the possibility of an immune-mediated

> response to silicone elastomer debris.

> (Emphasis supplied). Hirakawa attempted to quantify

> the particles and particle size observed in the

> tissue. Given the stated limitations that there was

> probably an underestimate of the amount of debris

> present and that very large particles were excluded

> from the quantitation, the article concluded that,

> " our results suggest that fragmented silicone

> implants release billions of very small particles to

> adjacent tissues. " (278)

>

> There is credible scientific, published evidence

> from which one could conclude that silicone gel

> microdroplets are released into the body via gel

> bleed, that silicone is subdivided into numerous

> microdroplets and particles by tissue and bodily

> fluids, that the small microparticles produce an

> intense, chronic inflammatory reaction, and that the

> reaction is a specific immune-mediated response.

>

> C. CHEMICAL MEDIATORS OF INFLAMMATION

> Chemical mediators of inflammation can originate

> from the plasma (e.g. complement and clotting

> factors) or from cells. Those mediators originating

> from plasma and preformed cellular mediators are

> most important in the early phases of the

> inflammatory process. Since this discussion focuses

> on the progression of acute to chronic inflammation

> caused by silicone gel breast implants and, since

> this represents a long term, persistent chronic

> process, this discussion will be limited to newly

> synthesized mediators released from cells. NEWLY

> SYNTHESIZED MEDIATORS MAJOR CELLULAR SOURCES

>

> Prostaglandins All leukocytes, Platelets,

> Endothelium

> Leukotrienes All Leukocytes

> Platelet Activating Factors All Leukocytes,

> Endothelium

> Nitric Oxide Macrophages, Endothelium

> Cytokines Macrophages, Endothelium

>

>

> 1. Prostaglandins and Leukotrienes

>

> These chemicals affect a variety of biologic

> processes, including inflammation. They are local,

> short-range hormones which are formed rapidly, exert

> their affects locally, and either decay

> spontaneously or are destroyed enzymatically.

> Prostaglandins and leukotrienes act to augment the

> inflammatory process and are also involved in the

> systemic symptoms of inflammation including fever,

> myalgias and night sweats.

>

> 2. Platelet Activating Factors

>

> Platelet activating factors work locally on the

> endothelial cells and inflammatory cells to augment

> the inflammatory process. These factors are

> indirectly associated with systemic symptomatology

> in that they enhance the production of

> prostaglandins and leukotrienes.

>

> 3. Nitric Oxide

>

> Nitric Oxide (NO) acts as a free radical which can

> be cytotoxic to certain bacteria and tumor cells. NO

> is likely to be responsible for the initial pain

> associated with the acute inflammatory process.

>

> 4. Cytokines

>

> Cytokines are generally small polypeptides with

> short half lives. (279) They exert many influences

> over the immune system including controlling the

> development and differentiation of leukocytes,

> promoting cell activation and proliferation, and

> regulation and suppression of immune responses

> including activated helper T-cells. One of the key

> features of activated helper T-cells is their

> ability to activate macrophages. Macrophages serve a

> number of functions including the secretion of

> cytokines. Specifically, the macrophage-derived

> cytokines which mediate inflammation include

> Interleukin-1 (IL-1), Tumor Necrosis Factor (TNF),

> and Interleukin-6 (IL-6).

>

> IL-1 and TNF are separate cytokines that bind to

> different cellular receptors but which have very

> similar biologic effects. Much of their importance

> stems from their ability to enhance activation of

> helper T lymphocytes by antigen-presenting cells

> (APCs). IL-1 and TNF- are each secreted by APCs on

> contact with antigen and MCH-specific helper T cell,

> and they can provide a co-stimulatory signal that

> promotes T cell activation.(280) In addition, IL-1

> and TNF- act in a paracrine (hormone like) fashion

> on the T cell, augmenting IL-2 secretion, expression

> of surface receptors for IL-2 and Interferon (INF)

> and all subsequent events leading to clonal

> proliferation.(281) Through their ability to

> potentiate helper cell activation, IL-1 and TNF- can

> promote nearly all types of humoral and cellular

> immune responses. In this regard both cytokines

> often act together with IL-6, thereby producing

> synergistic effects.(282)

>

> As will be discussed at length below, one of the

> side effects of chronic stimulation with cytokines

> is the exacerbation of underlying diseases or immune

> dysfunctions.(283) Many of these proinflammatory

> cytokines, including IL-1 and TNF-(284) induce the

> expression of autoreactive receptors on immune

> cells. In addition, alteration of the cytokine

> production patterns has been shown to shift the

> predominant immune response from a T-1 response to a

> T-2 response thereby enhancing an autoimmune

> reactivity.(285) Systemically, these cytokines cause

> fever, sleep disorders, cognitive dysfunctions,

> appetite disorders, myalgias and arthralgias.(286),

> (287), (288),(289),(290),(291)

>

> D. MANUFACTURERS' EXPERTS URGED THEM TO CONDUCT

> RESEARCH ON CHRONIC INFLAMMATION AND CYTOKINES IN

> WOMEN WITH SILICONE GEL BREAST IMPLANTS BUT THEY

> DECLINED

>

> As numerous reports were published in the literature

> documenting severe, prolonged chronic inflammation

> in silicone implanted individuals, the manufacturers

> - primarily Dow Corning(292) - were urged to conduct

> or fund studies on this issue with regard to women

> implanted with silicone gel breast implants. One of

> Dow's experts, Dr. Noel Rose, wrote letters to Dow

> Corning urging that they fund a study on the

> relationship of chronic inflammation, cytokine

> production, and atypical manifestations of disease

> symptoms. Notes of a meeting attended by Dr. Rose

> and Dow Corning in early 1993 reflect the following:

>

>

> Dr. Rose believes that silicone produces chronic

> inflammation. Cytokines are certain to result from

> this chronic inflammation and that is an area that

> needs further investigation. He also mentioned that

> a chronic inflammatory focus can serve as an

> adjuvant.(293)

> Dr. Rose prepared a funding proposal on this issue

> and submitted it to Dow Corning in 1993.(294) Dow

> Corning, in line with its " litigation strategy " and

> apparently after consultation with its General

> Counsel and outside legal counsel (who are shown as

> receiving blind carbon copies of the letter)

> declined to fund this particular study.(295) Dr.

> Rose responded that waiting for epidemiology was a

> mistake. " While epidemiology can show association,

> only experimental investigations can demonstrate a

> cause-and-effect relationship. " (296) He urged them

> to reconsider funding based on a pilot experiment he

> had conducted which showed that " injected mice have

> a small but statistically significant increase in

> antibody to topoisomerase, one of the autoantibodies

> characteristic of human scleroderma. " Further, he

> wrote:

>

> [t]here is now reasonably strong evidence that

> silicone gel (but not silicone oil) may serve as an

> adjuvant. While I know of no reason why injection of

> an adjuvant by itself necessarily triggers an

> autoimmune process, these recent findings do raise a

> number of questions about the longer-term effects of

> silicone implants.

> My special concern is that silicone implants as

> sources of chronic inflammation or as adjuvants may

> initiate or exacerbate autoimmune responses in

> predisposed individuals.

> Additionally, he noted, in separate correspondence,

> that his research interest had turned to the

> long-term effects of chronic inflammation and

> especially of cytokine production and release in

> genetically predisposed individuals.(297) The

> release of cytokines, Dr. Rose wrote, could explain

> " many of the 'atypical' conditions " reported by

> women with silicone gel breast implants.

>

> Despite Dow Corning's refusal to fund this study on

> chronic inflammation and cytokine release from

> activated macrophages, others - not funded by Dow

> Corning - were able to study this issue.

>

> E. RESEARCH ON WOMEN WITH SILICONE BREAST IMPLANTS

> SHOWS THE ELEVATION OF CYTOKINES IN CHRONICALLY

> INFLAMED SILICONE-EXPOSED TISSUE

>

> Several groups of researchers have found that human

> cells produce increased amounts of pro-inflammatory

> cytokines IL-1, IL-6, and TNF- when in contact with

> silicone or silicones that have been pre-absorbed

> with either albumin, fibrinogen, or IgG. (298),

> (299), (300), (301) In a study by Naim and van Oss,

> they noted that silicone placed in the body becomes

> immediately coated with plasma proteins.(302)

> Subsequent events involve migration of leukocytes to

> the implant site. Interaction among the

> monocytes/macrophages, lymphocytes and neutrophils,

> mainly through cytokine release and cell to cell

> interactions, determines whether the body will

> tolerate the implant over time.(303) Naim found that

> monocytes secreted nearly twice the amount of

> cytokines IL-1, IL-6 and TNF- when in contact with

> silicones adsorbed with proteins (albumin,

> fibrinogen and Ig).(304) Their results, listed

> below, show that concentrations of each of these

> cytokines were released when in contact with the

> silicone-gel, silicone-oil and silicone gel-oil

> combinations.

>

> TABLE 1 FROM NAIM'S ARTICLE

>

> Polymer Adsorbed Protein Cytokines* ± SEM

> (pg/ml)

> IL-1 IL-6 TNF-

> Tissue culture polystyrene No protein

>

> Albumin

>

> Fibrinogen

>

> IgG

> 5.0 ± 1.4

>

> 10 ± 0.9

>

> 8.0 ± 0.6

>

> 9.0 ± 1.0

> 3.0 ± 0.3

>

> 5.0 ± 2.0

>

> 4.0 ± 1.0

>

> 6.0 ± 4.0

> 18 ± 4.0

>

> 36 ± 8.0

>

> 36 ± 5.0

>

> 158 ± 34

>

> Silicone elastomer No protein

>

> Albumin

>

> Fibrinogen

>

> IgG

> 2.0 ± 1.0

>

> 106 ± 39

>

> 89 ± 6.7

>

> 101 ± 8.0

> 3.0 ± 0.4

>

> 89 ± 6.0

>

> 90 ± 17

>

> 90 ± 4.0

> 16 ± 6.0

>

> 357 ± 16

>

> 306 ± 33

>

> 388 ± 16

>

> Silicone gel No protein

>

> Albumin

>

> Fibrinogen

>

> IgG

> 5.0 ± 3

>

> 138 ± 20

>

> 164 ± 7.0

>

> 168 ± 4.0

> 5.0 ± 2.0

>

> 159 ± 31

>

> 160 ± 33

>

> 143 ± 16

> 24 ± 11

>

> 673 ± 43

>

> 731 ± 84

>

> 601 ± 18

>

> Silicone oil No protein

>

> Albumin

>

> Fibrinogen

>

> IgG

> 12 ± 1.0

>

> 202 ± 0.5

>

> 137 ± 5.0

>

> 153 ± 3.0

> 4.0 ± 0.6

>

> 189 ± 23

>

> 150 ± 12

>

> 139 ± 6.0

> 12 ± 2.0

>

> 699 ± 63

>

> 585 ± 24

>

> 685 ± 28

>

> Silicone gel + oil No protein

>

> Albumin

>

> Fibrinogen

>

> IgG

> 9.0 ± 3.0

>

> 180 ± 41

>

> 166 ± 16

>

> 200 ± 6.0

> 3.0 ± 0.5

>

> 149 ± 7.0

>

> 152 ± 14

>

> 147 ± 11

> 73 ± 21

>

> 702 ± 32

>

> 511 ± 31

>

> 720 ± 48

>

>

> *Cytokines values were derived from the pooled

> contents of six wells per treatment. The mean

> cytokine values were calculated from duplicate

> assays from two separate experiments.

>

> Similarly, Naidu, discussed above with regard to

> silicone particles inciting chronic inflammatory

> reactions, found that the particles produced a

> pseudosynovial-like lining, and that cytokine TNF-

> and Prostaglandin E2 were produced, with the highest

> levels noted at six and 24 hours following injection

> of the silicone particles into rats.(305) He

> reported:

>

> [t]he overall time course of the inflammatory

> response suggests that there is an initial release

> of TNF by the pseudosynovial lining cells. The WBC

> influx into the pouch is concomitant with the

> increase in TNF. Further rise in TNF at 24 hours is

> probably because of additional TNF release by the

> WBCs. There is a lag in PGE2 response. Peak

> PGE2levels occur at 24 hours; this appears to be

> temporally related to the influx of WBCs into the

> pouch tissue.

> Krause, in a study published in 1990, also reported

> finding IL-1 on the surface of Dow Corning silicone

> elastomer which was placed inside an artificially

> created synovial lining (composed of macrophages and

> fibroblasts) in male Sprague Dawley rats.(306) At 2

> or 7 days, the rats were killed and the implants

> removed. The cells were harvested and, after 48

> hours, radioactivity was measured in counts per

> minute using a liquid scintillation counter. Krause

> found that there was IL-1 produced on the surface of

> the silicone elastomer (SE), and that:

>

> [t]he control surface of the SE is characterized

> morphologically by a smooth, nonporous surface. At

> the 7-day harvest, we observed cells with epithelial

> and fibroblastlike features. These cells formed

> sheets over the surfaces of the SE.

> Lossing reported that inflammatory cells in silicone

> breast implant capsules expressed immunoreactivity

> for TGF-, IGF-II, IGF-I, and, to a lesser extent,

> PDGFB, NGF and TNF-.(307) He reviewed 12 biopsy

> specimens from 11 patients implanted with silicone

> gel breast implants who had experienced varying

> degrees of capsular contracture including nine who

> had Grade III contracture. Controls were biopsy

> specimens from patients with subpectoral implants

> after breast augmentation from an expander

> prosthesis, a three year old contracted scar after

> full-thickness burn injury and from surgical skin

> incisions for breast reconstruction. He found

> chronic inflammatory reaction (characterized by

> fibroblast-like cells, macrophages, lymphocytes,

> scattered polymorphonuclear leukocytes, plasma cells

> and mast cells) in all of the implant capsules and

> myofibroblasts in some macrophages that could be

> identified by electron microscopy.

> Immunohistochemical analyses showed the following:

>

> [t]he myofibroblasts and some of the macrophages

> in the capsules expressed TGF- and IGF-II

> immunoreactivities, most prominently in the

> contracted capsules. Cells of implant capsules

> showed IGF-I immunoreactivity, most prominently the

> abundant myofibroblasts in the contracted capsules.

> Macrophages, common fibroblasts, and vascular cells

> usually expressed IGF-1 immunoreactivity as well.

> Scattered inflammatory cells in the interface region

> showed IGF-1 immunoreactivity at variable intensity.

> Extracellular IGF-1 immunoreactivity was recognized

> restricted to the interfacial amorphous coating

> covering the implants. Scattered macrophages showed

> as well NGF and TNF- immunoreactivities.

> Smooth-muscle cells in arteries and scattered

> fibroblasts and macrophages expressed PDGF

> immunoreactivity to variable extents. Cells in the

> tissue adjacent to the capsules sometimes expressed,

> at low intensities and in variable frequencies,

> peptide growth factor immunoreactivities, as did

> pericapsular fibroblasts.

> Lossing noted similar experiments in animals which

> showed peptide growth factor immunoreactivities in

> capsule cells, followed by removal of the implant

> which made the peptide growth factors

> immunohistochemically subside and eventually vanish.

> Subcutaneous infusion of cytokines accelerated the

> differentiation of fibroblasts into myofibroblasts

> in vivo. TNF- also acts as an immunomodulator of

> chronic inflammatory reactions in addition to

> promoting growth and influencing the expression of

> growth factor receptors. It is also of crucial

> importance in the early phases of scar tissue

> formation (collagen content and wound tensile

> strength). Lossing concluded that:

>

> t is likely that activated macrophages and

> other cells in an inflamed tissue influence the

> formation and release of factors that exert strong

> effects on neighboring cells. The inflammatory

> reaction going on in the capsule must therefore be

> considered to be of primary importance for its

> cellular composition and the tendency of a capsule

> to become contracted. . . .

> Several factors in addition to the implant per se

> are likely to modulate the prolonged foreign-body

> reaction in the interface zone. Seepage of silicone

> from the breast implant increases the inflammatory

> response. [footnotes omitted] The physicomechanical

> stimulation exerted by movements between the tissue

> and the implant is another factor of importance. . .

> .

> We conclude that the chronic low-grade

> inflammatory foreign-body reaction around

> gel-filled, low-bleed silicone breast implants is a

> major factor responsible for the persistence of the

> high local levels of peptide growth factors and

> various cytokines in the capsules. We presume that

> such highly potent factors stimulate, e.g.,

> granulation tissue fibroblasts to turn into

> myofibroblasts. This model of response is beneficial

> in ordinary wound healing but may in patients with

> soft implants, intended to stay for long time

> periods, constitute a negative factor.

> Wells also reported finding increased hyaluronan

> (HYA) and IL-2 in women with silicone gel breast

> implants.(308) Wells examined biopsy specimens from

> fourteen women with silicone gel breast implants to

> determine if there was a local increase in

> hyaluronan, a marker of active inflammation, and to

> characterize the inflammatory cells and their

> secreted factors in tissue surrounding the implants.

> He reported in an abstract in 1993 that there was an

> increase in hyaluronan in implanted patients versus

> controls. The abstract states:

>

> [t]he hyaluronan was localized extracellularly in

> areas containing fibrosis and cellular infiltrates.

> Using various monoclonal antibodies, the

> infiltrating cells were determined to be macrophages

> and T cells. The number of infiltrating macrophages

> was greater than that of the T cells based on the

> relative staining intensity. No IL-6 was localized

> in any of the tissue sections. In contrast, large

> amounts of IL-2 were found in regions of

> infiltrating lymphocytes.

> Wells suggested that activated macrophages played a

> central role in the inflammatory process and could

> induce fibroblasts to proliferate synthesizing

> hyaluronan which in turn may lead to local edema and

> fibrosis.

>

> Data currently exists on increased amounts of HYA in

> various inflammatory diseases; one of the more

> prominent diseases is rheumatoid arthritis.(309)

> Wells suggest that HYA may be one of the molecules

> readily produced by actively proliferating cells in

> and around breast implant capsules, which may in

> turn act as a " Chemo attractant " signaling cells to

> the site of inflammation. This chronic activation of

> T cells could be responsible for the cutaneous T

> cell lymphoma reported by Duvic.(310)

>

> Garland found that there was a strong trend for a

> positive correlation between serum IL-6 levels and

> duration of implantation.(311) The correlation was

> highly significant for women less than 45 years of

> age (p < 0.01), but not for older women. Similarly,

> the correlation was significant among women with

> their silicone breast implants still in place (p =

> 0.01) but not for those who had their implants

> removed. Among women age 45 or younger who were

> studied with their silicone breast implants in place

> (n = 31), the correlation between serum IL-6 levels

> and duration of implant was highly significant (p =

> 0.009).

>

> The findings of elevated IL-6 levels are consistent

> with the work of Potter, NCI, who found that

> repeated injections of silicone gel induced

> plasmacytomas in mice. Potter noted that the best

> known factor that produces plasma cell

> differentiation and plasmacytoma growth is

> IL-6.(312) Felix also reported in the Immunology of

> Silicones that IL-6 was secreted by activated

> macrophages after a 20 hour exposure to

> siloxanes.(313) He observed, " The B9 cell bioassay

> of these treated cells showed as much as a 10 fold

> higher production (500 U/ml) of IL-6 than did the

> untreated cells. The degree of increase was

> dependent on the compound and concentration used. "

>

> Also, not surprisingly, other researchers have

> failed to detect elevated levels of inflammatory

> cytokines in circulating serum of silicone

> patients.(314), (315), (316), (317) None of these

> researchers studied cytokine levels locally at the

> site of the chronic inflammatory reaction to the

> silicone. Therefore, their failure to find elevated

> cytokine levels in serum is to be expected since the

> inflammatory cytokines are very short lived, most

> living only several hours.(318) In fact, it is

> generally accepted that assays of local cytokine

> production at the site of inflammation or organ

> injury and repair are likely more biologically

> relevant than those in the peripheral blood.(319)

> For example, researchers bred mice to produce

> excessive amounts of TNF-. All of the mice developed

> histological characteristics compatible with human

> rheumatoid arthritis, yet there were no detectable

> levels of serum TNF- in any of the mice.(320) In

> that case, TNF- was the known agent responsible for

> ongoing disease in the mice and yet it was not

> detectable in sera.

>

>

>

>

> F. CYTOKINES IN WOMEN WITH SILICONE BREAST IMPLANTS

> CAN CAUSE SYSTEMIC SIGNS AND SYMPTOMS OF DISEASE

>

>

>

> 1. TNF-[alpha]

> The two types of TNF-[alpha] receptors elicit

> distinct responses: the Type I receptor generally

> promotes cytotoxic activity and fibroblast

> proliferation, whereas the Type II receptor promotes

> T lymphocyte proliferation.(321) TNF-[alpha] plays

> an important part in silica-induced pulmonary

> fibrosis.(322)

>

> Both IL-1 and TNF-[alpha] can activate endothelial

> cells and thus promote neutrophil migration into an

> inflamed site. They induce the production of

> endothelial growth factors and have angiogenic

> activity. Acting alone or synergistically, they can

> induce a number of effects that are mediated through

> the hypothalamus: they are endogenous pyrogens

> (i.e., they induce fever) and directly induce the

> secretion of corticotropin-releasing factor, which

> stimulates the release of adrenocorticotropic

> hormone from the pituitary and thus induces

> glucocorticoid production by the adrenals. Both IL-1

> and TNF-[alpha] stimulate alkaline phosphatase

> activity in osteoblasts, and proliferation by

> fibroblasts and synovial cells. Increased levels of

> IL-1 and TNF-[alpha] are found in inflammatory joint

> fluids and contribute to the fibrosis and thickening

> of arthritic joints.(323)

>

> Both IL-1 and TNF- stimulate epithelial cell

> proliferation and function, including the production

> of basement membrane collagen. Moreover, IL-1 and

> TNF-[alpha] regulate one another, and their ability

> to synergize enables them to achieve maximal effects

> at sub-optimal concentrations. This is economical,

> results in enormous amplification of host reactions,

> and increases the efficiency of the host defense

> system.(324)

>

> 2. IL-6

> IL-6 is a cytokine with multiple biologic activities

> on a variety of cells. Its major activities include

> synergizing with IL-1 and TNF-[alpha] to

> co-stimulate immune responses, inducing the

> acute-phase response in liver cells, and supporting

> the growth of transformed hepatocyte and myeloma

> cell lines in tissue culture. IL-6 was initially

> called interferon- 2 (IFN-2) because it appeared to

> exhibit antiviral activity and to cross-react with

> some antisera to IFN-. IL-6 can be produced by many

> cell types, including activated T and B lymphocytes,

> monocytes, endothelial cells, epithelial cells, and

> fibroblasts. Its expression is induced by a variety

> of stimuli, including TNF-[alpha], IL-1,

> platelet-derived growth factor, and any factors that

> activate T lymphocytes.

>

> IL-6 has certain biologic activities that are not

> exhibited by other known members of the family.

> Alone among these cytokines, IL-6 acts as a

> co-stimulant that synergistically augments the

> mitogenic effects of IL-1 and TNF-[alpha] on helper

> T-cells. IL-6 is also very effective in enhancing

> TNF-[alpha] or IL-1 induced cachexia and

> glucocorticoid synthesis and is able independently

> to stimulate osteoblast activity and keratinocyte

> growth. This suggests that its main immunologic

> function is to potentiate the effects of other

> cytokines. Since malignant B cells of multiple

> myeloma both produce and respond to IL-6, it may act

> as an autocrine growth factor for these cells.(325)

>

> 3. IL-2

> Interleukin-2 (IL-2) is synthesized and secreted

> primarily by CD4+ T Lymphocytes. T cells activated

> by mitogens or by antigen-specific interactions

> resulting in the activation of CD4+T cells caused

> the production and release of IL-2.(326) IL-2 has

> also been referred to as T cell growth factor. One

> of the important things about IL-2 is that resting T

> lymphocytes do not synthesize or secrete IL-2

> protein but can be induced to do both by appropriate

> combinations of antigen and costimulatory factors or

> by exposure to Polyclonal mitogens.(327)

>

> IL-2 is also very short lived. When normal human

> lymphocytes are exposed to a T cell mitogen, IL-2

> mRNA expression becomes detectable after 4 hours,

> reaches peak concentration at 12 hours, and

> thereafter declines rapidly. The abrupt

> disappearance of the MRNA reflects not only the

> cessation of IL-2 gene transcription, but also the

> instability of IL-2 in mRNA, which has a half-life

> of less than 30 minutes. T cells, once activated by

> antigens or mitogens, express an IL-2 receptor that

> reaches its maximum expression within two to three

> days after the cells become activated.(328), (329)

> The expression then declines to undetectable levels

> by six to ten days after activation. If the T cell

> is reactivated, the IL-2 receptors on the cell's

> surface again activate IL-2 production in the same

> basic pattern. The presence of IL-2 in and around

> breast capsules conclusively shows that T cells have

> been activated within days of this finding. Again,

> as with the presence of plasma cells, the

> demonstration that IL-2 is present in the capsules

> and tissues surrounding breast implants positively

> proves that an antigen activated T cells, regardless

> of what that antigen is.

>

> The combination of IL-2 with activated lymphocytes

> promotes several other cellular activities as well

> as T cell proliferation. For instance, IL-2

> stimulated T cells exhibit enhanced cytotoxicity and

> produce lymphokines such as Interferon gamma,

> TNF-[alpha], and T cell growth factor beta, B cell

> growth factors such as IL-4 and IL-6, and

> hematopoietic growth factors such as IL-3 and

> IL-5.(330)

>

> 4. Hyaluronic acid (HYA)

> Hyaluronic acid is normally present in many tissues

> and is particularly abundant in loose connective

> tissue and in synovial fluid. Increased levels of

> circulating HYA have been found in patients with

> various liver diseases and during the active stages

> of inflammatory joint disease.

>

> 5. Cytokines Communicate With Other Inflammatory

> Cells, B-cells and Macrophages

> Inflammatory mediators such as the

> macrophage-induced cytokines, IL-1, TNF-[alpha] and

> IL-6 are not only important in modulating local

> tissue injury, they also have deleterious systemic

> effects.(331) These events are due to the autocrine,

> paracrine, and endocrine nature of many of these

> mediators generated in the context of an

> inflammatory response.

>

> IL-1 and TNF-[alpha] also act directly on many other

> types of immune and inflammatory cells. For example,

> they can directly promote growth and differentiation

> of B-cells -- particularly during the transitions

> from pre-B-cells into mature B lymphocytes and from

> lymphocytes into plasma cells. They also can

> activate neutrophils and macrophages, stimulate

> hematopoiesis, and induce expression of numerous

> other cytokines and inflammatory mediators.(332)

>

> Most cytokines have a very short half life and the

> majority of cytokines normally act locally on cells

> in their immediate vicinity.(333) Cytokines serve as

> messengers in regulating the amplitude and duration

> of the immune inflammatory responses by

> communicating with cells and in stimulating cell

> growth.(334) For instance, researchers have proven

> that a macrophage population remote from the primary

> site of inflammation was conditioned (primed) for

> increased TNF-[alpha] production even though these

> remote macrophages did not have direct contact with

> the particular antigenic material, in this case

> silica.(335) Thus, systemic immunostimulation is

> known to occur.

>

> 6. Cytokines Communicate Directly with the Brain

> from Local Sites of Inflammation

> Cytokines can signal the brain that inflammation has

> occurred. The cytokine to brain communication can

> result in marked alterations in brain function and

> behavior.(336) Scientists have known for decades

> that the brain responds to inflammation in distal

> parts of the body by causing fever and the familiar

> tired and achy feelings that accompany

> infections.(337) It has recently been demonstrated

> that cytokines such as IL-1 act directly with local

> nerve cells which relay the message to the brain

> that inflammation is occurring, thus triggering the

> brain's response which alters behavior systemically.

> (338), (339), (340) The overall pattern that emerges

> suggests that a great deal of IL-1 signaling to the

> central nervous system (CNS) is accomplished by

> activation of vagal afferents rather than by direct

> access to brain. IL-1 produced hyperalgesia, fever,

> conditioned taste aversions, and now hypothalamic NE

> depletion and corticosterone increases are all

> completely or largely eliminated by vagotomy. The

> fact that different afferent branches of the vagus

> might mediate different aspects of the response to

> IL-1 is not expected if cytokine release is viewed

> as a local event as well as a long range hormonal

> signal.(341), (342)

>

> 7. Cytokines Cause Clinical Symptoms and Autoimmune

> Disease

> Processes occurring within the immune system can

> alter neural function. Cytokines released by cells

> of the immune system during illness are key

> messengers in immune-to-brain communication. IL-1

> for instance is known to stimulate a myriad of

> illness-related outcomes such as fever, sickness

> behavior, aphagia, adipsia,

> hypothalamic-pituitary-adrenal activation, and

> changes in pain reactivity.(343) Thus peripherally

> released IL-1 has potent neural effects and is a

> critical mediator of the impact of immune processes

> on the brain. A primary route of peripheral cytokine

> signaling is through stimulation of peripheral vagal

> afferents rather than or in addition to direct

> cytokine access to brain.(344)

>

> It is well accepted that products of the immune

> system (cytokines) can signal the brain that

> infection has occurred. This cytokine-to-brain

> communication can result in marked alterations in

> brain function and behavior.(345) Many mechanisms

> have been proposed to explain how immune products

> can reach the brain via the blood to cause

> centrally-mediated " illness " responses. One of those

> mechanisms is through local stimulation of

> peripheral nerve cells at local sites of

> inflammation.(346), (347), (348), (349), (350)

> Several series of investigations have convincingly

> demonstrated that the non-specific " sickness

> symptoms " such as lethargy(351) and fever(352) are

> due to the effects of the proinflammatory cytokines

> IL-1, TNF-[alpha], and IL-6 on the central nervous

> system.(353)

>

> In addition to changes in body temperature and other

> metabolic and physiologic responses corresponding to

> immune activation, pyrogens, such as IL-1 and TNF

> can induce profound behavioral changes including

> depressed cognitive functioning.(354) IL-1 is

> specifically implicated as the mouse brain expresses

> both type 1 and type 2 IL-1 receptors which further

> support the idea that type 1 IL-1 receptors are

> synthesized and expressed by neurons.(355)

>

> Autoimmune diseases, such as rheumatoid arthritis

> and inflammatory bowel disease are characterized by

> chronic inflammatory responses resulting in tissue

> damage. These diseases have a number of common

> denominators including: abnormal cytokine

> expression, aberrant antigen-antibody complexes,

> T-cell anomalies, and increased numbers of

> neutrophils and macrophages. The interaction between

> neutrophils and macrophages induces a state of

> chronic inflammation which contributes to the

> disease state.(356) Several spontaneous autoimmune

> diseases in mice or humans have been reported to be

> associated with TNF overproduction in the affected

> organ: lupus nephritis, diabetic pancreatitis,

> rheumatoid arthritis, the demyelinating plaque of

> multiple sclerosis, and the epidermis of

> cirrhosis.(357)

>

>

> G. PHENOTYPES OF LYMPHOCYTES WHICH RESPOND TO

> SILICONE

> As set forth above, recent work by Naim and van Oss

> determined that human monocytes previously unexposed

> to silicones but now cultured on silicones produce

> significantly greater quantities of various

> cytokines than human monocytes cultured on other

> substances.(358) This is particularly significant

> because several studies have determined the

> phenotype of the lymphocytes which have responded to

> the silicone in the capsules of women with breast

> implants, and they have found evidence of a specific

> immune response to something in the capsule. For

> example, Katzin, Mt. Sinai, found eighty-nine (89%)

> percent of the implant-associated lymphocytes were

> T-cells.(359) Twenty-five (25%) percent of the CD3+

> T-cells co-expressed HLA-DR compared with only 7.9%

> of matched peripheral blood lymphocytes. Sixty-eight

> (68%) percent of the implant-associated T-cells

> co-expressed CD4 and CD29, while only three (3%)

> percent of the T-cells co-expressed CD4 and CD45RO.

> These results led them to conclude that, " [t]he

> expression of HLA-DR and predominance of CD29+, CD4+

> T-cells indicate that there is immune activation

> with a potential for stimulating antigen-specific

> antibody production. "

>

> In a study by O'Hanlon, the researchers evaluated

> the cellular phenotypes of 22 silicone breast

> implant capsules.(360) They found that although the

> number and type of inflammatory cells varied among

> the different capsules, when present, they were

> " usually localized around silicone vacuoles,

> suggesting an association between silicone release

> and local immune responses. " The inflammatory

> infiltrates contained multinucleated giant cells,

> activated T-lymphocytes, activated B-lymphocytes,

> macrophages, and plasma cells. The study then looked

> for any pattern in the genetic activation of T-cell

> receptor genes (TCR). Having found a pattern, they

> concluded that " the data suggest that at least in

> certain individuals, antigenic selection may account

> for observed similarities in TCR V gene expression. "

>

>

> Another study by Ladin at the Henry Ford Hospital in

> Detroit confirmed the above results.(361) Ladin also

> attempted to identify the phenotypes of the T-cells

> in the infiltrate around silicone breast implant

> capsules. The results of their study were as

> follows:

>

> [t]he pericapsular cellular infiltrate consisted

> of predominantly CD2+ T cells with a CD4+/CD8+

> ration of 8:1. An activated phenotype was seen

> including human leukocyte antigen (HLA)-DR+cells

> (>75%)and CD25+cells (10-20%). Most T cells express

> the / TCR (>50%+) while / cells were occasionally

> seen (1-2/40 x field). Mononuclear cells of

> dendritic morphology were scattered throughout the

> infiltrate and expressed CD36, HLA-DR, Thy-1, CD1b,

> and CD1c. Southern analysis revealed evidence of a

> non-germline TCR gene rearrangement found in HindIII

> digests from all implant patients. Normal breast and

> scarred skin were also found to have evidence of

> minor TCR rearrangements, while none were seen in

> normal skin or patients' blood. Pericapsular tissue

> immunostaining also demonstrated V TCR subsets in

> four of five patients, confirming that protein was

> expressed at the cell surface.

> They concluded:

>

> These findings suggest that a silicone-related

> substance is inducing an activated, clonotypic

> T-cell response in the pericapsular tissue, which

> develops around silicone gel breast implants. The

> activated profile indicates that these T cells do

> not represent a passive infiltrate but rather are

> engaged in cell-mediated immune processes. The

> clonotypic response means that T cells are

> responding to a single focal antigenic stimulus.

> These T cells may be involved in the pathogenesis of

> a silicone-mediated immune adjuvant disease, which

> could cause local or systemic inflammation.

> (Emphasis added). Stark's work in 1990 further

> provides support that the capsular tissue

> surrounding silicone gel breast implants involves an

> immune-mediated process.(362)

>

> Supporting Ladin's finding of protein expressed at

> the cell surface, Tang noted in 1995 that

> polydimethylsiloxane (Dow Corning Silastic)

> immediately acquires a layer of host proteins after

> implantation.(363) As discussed more fully in the

> Immunology section, Tang noted that at least part of

> the chronic inflammation may arise from interactions

> between the protein-coated surfaces of the

> biomaterial and host tissues and phagocytes, such as

> macrophages and foreign body giant cells. Spread of

> these particles has been associated with

> lymphadenopathy, fever and lymphoma, and is believed

> to mediate seronegative and seropositive synovitis.

> During the chronic inflammatory response to implants

> and implant materials, products generated by

> adherent inflammatory cells may damage the implant

> and/or react with the biomaterial to generate toxic

> catabolites.(364)

>

> The cellular response in animals has also been

> meticulously documented by Picha.(365) In addition,

> a number of other significant animal studies looking

> at the cellular response to silicones are documented

> in the Pathology section of this brief.

>

> These studies demonstrate that there is some form of

> specific immune response involved with silicone

> materials.(366)

>

> H. EXPLANTATION OF THE SILICONE BREAST IMPLANT AND

> THE SILICONE-CONTAINING CAPSULE RESOLVES THE CHRONIC

> INFLAMMATION

> The medical community acknowledges that silicone

> incites and prolongs the chronic inflammatory

> process. Reports from almost 50 articles (some

> including case series of more than one hundred

> women) are set forth in the Clinical Experience

> section and show improvement and resolution of

> systemic disease symptoms and resolution of chronic

> inflammation following removal of the silicone

> device.(367) Lossing recently noted that:

>

> [r]emoval of the implant made the peptide growth

> factors immunohistochemically subside and eventually

> vanish in about a week and, concomitantly, the

> myofibroblasts . . . .

> The myofibroblasts obviously depended on trophic

> compounds for their maintenance and survival, since

> they vanish when the low-grade chronic inflammatory

> reaction subsides after removal of the foreign

> body.(368)

> Likewise, Peimer reported that the synovitis seen in

> persons implanted with other types of silicone

> devices, and the progressive destruction was

> arrested by implant removal and synovectomy.(369)

> The significance of the silicone-containing capsule

> is demonstrated by Ahn's 1995 report on four cases

> in which the local and systemic complaints recurred

> intermittently following earlier explantation of the

> silicone breast implant.(370) Multiple silicone

> granulomas, pain from extensive fibrosis, chest pain

> and tightening, and systemic symptoms continued

> months after explantation. Further surgery revealed

> that the capsule surrounding the silicone breast

> implants had been left in during the original

> explantation surgery. Capsulectomies - or surgical

> excision of the capsules - were performed which

> resulted in resolution of the complaints, thus

> substantiating the numerous studies discussed in the

> Pathology and Immunology sections that the

> silicone-containing capsule is immune-mediated.

>

> CONCLUSION

> From the prolific data, it is now clear that the

> cellular responses to most silicone gel breast

> implants involves chronic inflammation which is

> mediated by particle size and other chemical

> mediators of inflammation. Research on women with

> silicone breast implants documents the presence of a

> variety of cytokines along with activated

> T-lymphocytes representing specific cell-mediated

> immune responses. It is well established that

> cytokines along with chronic inflammation can cause

> systemic signs and symptoms, immune dysfunction and

> autoimmune disease in predisposed individuals.

>

> This section began with the CDRH's analysis for

> immunotoxicity, and the prior Pathology section

> began with a similar discussion on biocompatability.

> Plaintiffs submit that 1) the histopathology of

> chronic inflammation, 2) the release of cytokines by

> activated macrophages, 3) the presence of

> granulocytes (basophils, eosinophils and

> neutrophils), and 4) the resolution of the chronic

> inflammatory and synovitis after removal of the

> silicone implant and silicone-containing capsule

> support that silicone gel breast implants are

> immunotoxic and are notbiocompatible. The clinical

> symptoms in implanted women, e.g., myalgias,

> arthralgias, fatigue, fever, lymphadenopathy, and

> cognitive dysfunction, correspond to the symptoms

> associated with these cellular reactions and

> supports a finding of immunotoxicity as well.

>

> Continue to Section V of Plaintiffs' Submission.

>

> 260. Center for Devices and Radiological Health

> (CDRH), Immunotoxicity Testing Framework (11/21/96)

> [Record No. 7079].

>

> 261. Wilflingseder, T., Propst, A., Mikuz, G.,

> " Constrictive fibrosis following silicone implants

> in mammary augmentation, " Chir. Plastica 2:215

> (1974) [Record No. 1318].

>

> 262. Barker, D.E., Retsky, M.L., Shultz, S.,

> " 'Bleeding' of silicone from bag-gel breast

> implants, and it clinical reaction to fibrous

> capsule reaction, " Plast. Reconstr. Surg. 61:836-841

> (1978) [Record No. 0998].

>

> 263. Gayou, R., " A Histological Comparison of

> Contracted and Non-Contracted Capsules Around

> Silicone Breast Implants, " Plast. Reconst.

> Surg.63(5):700-707 (1979) [Record No. 1016]; see

> also , W., Balogh, K., Abraham, J.L.,

> " Silicone Granulomas: Report of Three Cases and

> Review of the Literature, " Human

> Pathology16(1):19-27 (1985) [Record No. 2474](tiny

> residual microdroplets of silicone observed);

> Ferreira, M.C., Spina, V., Iriya, K., " Changes In

> The Lungs Following Injections Of Silicone Gel, "

> British J. Plast. Surg. 28:173-176 (1975) [Record

> No. 0383] (tendency of the gel to break up into

> smaller droplets); Vistnes, L., Bentley, J.,

> Fogarty, D., " Experimental Study of Tissue Response

> to Ruptured Gel-Filled Mammary Prostheses, " Plast.

> Reconstr. Surg.59(1):31-31 (1977) [Record No. 1969]

> (tendency of gel to break up into smaller droplets);

> and Sanger, J., Kolachalam, R., Komorowski, R.A., et

> al., " Short-Term Effect of Silicone Gel on

> Peripheral Nerves: A Histologic Study, " Plast.

> Reconstr. Surg. 89(5):931-940 (1992) [Record No.

> 1604] (gel migration with breakdown of the gel into

> smaller droplets leading to an intense cellular

> inflammation).

>

> 264. Sanger, J., Kolachalam, R., Komorowski, R.A.,

> et al., " Short-Term Effect of Silicone Gel on

> Peripheral Nerves: A Histologic Study, " Plast.

> Reconstr. Surg. 89(5):931-940 (1992) [Record No.

> 1604].

>

> 265. Lentz, A.J., Chandler, M.L., LeVier, R.R.,

> " Biological Evaluation of an Implantable Silicone

> Gel: Summary of Acute and Chronic Studies, " Dow

> Corning Report No. 4856, DCCKMM 174130-174159 (1978)

> [Record No. 7017]; see also Reuter, B. Memo to Dr.

> Vinnik, M 420161 - 420166 (2/24/87) [Record

> No. 7292].

>

> 266. Lentz, A.J., Chandler, M.L., LeVier, R.R.,

> " Biological Evaluation of an Implantable Silicone

> Gel: Summary of Acute and Chronic Studies, " Dow

> Corning Report No. 4856, DCCKMM 174130-174159 (1978)

> [Record No. 7017]. Several weeks later, Dow Corning

> wrote that, " The evidence for progressive

> subdivisions of gel is sufficient to warrant the

> conclusion that a hypothesis favoring efficacy in

> mammary augmentation and/or mammary reconstruction

> cannot be supported without further

> experimentation. " LeVier, R., Memo to Bey and

> re: Proposal for Development of the Implantable Gel,

> F 744 - 746 (3/16/78) [Record No. 7021].

>

> 267. Pudenz, B., Talcott, T., Heyer-Schulte Memo to

> Tom Hyans attaching report on production of gel

> bolus studies, MD 114595 - 114598 (5/23/78) [Record

> No. 7039].

>

> 268. Boley, W.F., Bejarano, M.A., " Fate of Q7-2159A

> Gel Injected Subdermally In Rats: Macro

> Observations, " Dow Corning Report No. 150, DCC

> 80031717 - 80031729 (1985) [Record No. 7042]

>

> 269. Reuter, B., Letter to Dr. Vinnik, M

> 420161 - 420166 (2/24/87) [Record No. 7292].

>

> 270. Malczewski, R.A., Mudgett, S.L., Geil, R.G, et

> al., " A Histological Description Of The Local

> Cellular Response Associated With The Subcutaneous

> Implantation Of Dow Corning Q7-2159A Mammary Gel,

> Dow Corning 7-2317 Fluid, Dow Corning

> Q7-2423/Q7-2551 Elastomer, Polytetrafluoroethylene,

> Ultra High Molecular Weight Polyethylene, And A

> Titanium Alloy (Ti-6AI-4V) In CD-1 Mice, " Dow

> Corning Report No. 1994-I0000-38978, DCC 807220001 -

> 807220049 (1994) [Record No. 6258].

>

> 271. Small silicone particles were reported to cause

> synovitis marked by swelling and discomfort by

> , R.J., Atkinson, R.E., Jupiter, J.B.,

> " Silicone Synovitis of the Wrist " , J. Hand

> Surg.10A:47-60 (1985) [Record No. 7020]; Christie,

> A.J., Weinberger, K.A., Dietrich, M., " Silicone

> Lymphadenopathy and Synovitis: Complications of

> Silicone Elastomer Finger Joint Prostheses, " JAMA

> 237:1463-1464 (1977) [Record No. 0971]. Silicone

> particles from silicone tubing also were reported to

> cause a foreign-body reaction leading to

> pancytopenia. Bommer, J., Ritz, E., Waldherr, R.,

> " Silicone-Induced Splenomegaly: Treatment of

> Pancytopenia by Splenectomy in a Patient on

> Hemodialysis, " NEJM 305 (18): 1077-1079 (1981)

> [Record No. 1053].

>

> 272. Sammacro, G.J., Tabatowski, K., " Silicone

> Lymphadenopathy Associated With Failed Prosthesis of

> the Hallus: A Case Report and Literature Review, "

> Foot & Ankle 13(5):273-276 (1992) [Record No. 3038];

> see also Worshing, R.A., Engber, W.D., Lange, T.A.,

> " Reactive Synovitis from Particulate Silastic, " J.

> Bone and Joint Surgery 64(4):581-585 (1982) [Record

> No. 1068].

>

> 273. Barrett, D.M., O'Sullivan, D.C., Malizia, A.A.,

> et al., " Particle Shedding And Migration From

> Silicone Genitourinary Prosthetic Devices, " J.

> Urology 146:319-322 (1991) [Record No. 1574] (Biopsy

> revealed shedding of particles by most types and

> brands of prostheses, the development of foreign

> body silicone granulomas associated with the abraded

> particles, and lymphadenopathy. The explanted

> devices were examined revealing pitting and flaking

> of the silicone surface.).

>

> 274. Naidu, S.H., Beredjiklian, P., Adler, L., et

> al., " In Vivo Inflammatory Response to Silicone

> Elastomer Particulate Debris, Journal of Hand

> Surgery 21A(3):496-500 (1996) [Record No. 7008].

>

> 275. Peimer, C.A., Medige, J., Eckert, B.S., et al.,

> " Reactive Synovitis After Silicone Arthroplasty, "

> Journal of Hand Surgery 11A(5):624-638 (1986)

> [Record No. 7105].

>

> 276. Light microscopy studies showed silicone

> particles in the synovial and capsular tissue of

> every patient, ranging in size from 60 - 80 .

> Findings from TEM studies on selected synovial

> tissue specimens showed a number of amorphous

> foreign particles, ranging in size from 0.2 - 1

> distributed among the collagen fibers of the tissue.

>

>

> 277. Hirakawa, K., Bauer, T., Culver, J., et al.,

> " Isolation and Quantification of Debris Particles

> Around Failed Silicone Orthopedic Implants, " The

> Journal of Hand Surgery 21A:5 (9/96) [Record No.

> 7076].

>

> 278. Because they could only detect and count

> particles down to approximately a half-micron in

> size, Hirakawa noted that there were probably

> billions of additional particles below that limit

> which were present in the tissue and inside the

> cells.

>

> 279. Detrick, B., Hooks, J.J., " Cytokines in Human

> Immunology, " Handbook of Human Immunology, Chapter

> 7, pp. 233-266 (1997) [Record No. 7126]; see

> discussion infra.

>

> 280. See e.g., Oppenheim, J.J., Ruscetti, F.W. ,

> Faltynek, C., et al, " Cytokines; Complement &

> Kinin, " Basic & Clinical Immunology 8thEd., Chapters

> 9-10, pp. 105-136 at 105 (1994) [Record No. 7127].

>

> 281. Kindler, V., Sappino, A., Gray, B. et al., " The

> Inducing Role of Tumor Necrosis Factor in the

> Development of Bactericidal Granulomas during BCG

> Infection, " Cell 56:731-740 (3/10/89) [Record No.

> 7128].

>

> 282. See e.g., Oppenheim, J.J., Ruscetti, F.W. ,

> Faltynek, C., et al, " Cytokines; Complement &

> Kinin, " Basic & Clinical Immunology 8thEd., Chapters

> 9-10, pp. 105-136 at 105 (1994) [Record No. 7127].

>

> 283. Vial, T., Descotes, J., " Immune-Mediated

> Side-Effects of Cytokines in Humans, " Toxicology

> 105:31-57 (1995) [Record No. 7095].

>

> 284. Wilheim, M., Silacci, P., Gessl, A., et al.,

> " Tumor Necrosis Factor-Alpha Induction of Major

> Histocompatibility Complex Class II Antigen

> Expression is Inhibited by Interferon-Gamma In A

> Monocytic Cell Line, " Eur. J. Immunol. 25:3202-3206

> (1995) [Record No. 7089]; , S.D., McRae, B.L.,

> Vanderlugt, C.L., et al., " Evolution of the T-Cell

> Repertoire During the Course of Experimental

> Immune-Mediated Demyelinating Diseases, " Immunol.

> Rev. 144:225-244 (1995) [Record No. 7090]; ,

> A., Lanir, N., Shapiro, S., et al.,

> " Immunoregulatory Effects of Interferon-Beta and

> Interacting Cytokines on Human Vascular Endothelial

> Cells: Implications for Multiple Sclerosis and Other

> Autoimmune Diseases, " J. Neuroimmunol. 64:151-161

> (1996) [Record No. 7091].

>

> 285. Takacs, K., Douek, D.C., Altmann, D.M.,

> " Exacerbated Autoimmunity Associated With A T

> Helper-1 Cytokine Profile Shift in H-2E-Transgenic

> Mice, " Eur. J. Immunol. 25:3134-3141 (1995) [Record

> No. 7092]; Waisman, A., Ruiz, P.J., Hirschberg,

> D.L., et al., " Suppressive Vaccination with DNA

> Encoding A Variable Region Gene of the T-Cell

> Receptor Prevents Autoimmune Encephalomyelitis and

> Activates The Immunity, " Nature Medicine

> 2(8):899-905 (1996) [Record No. 7093].

>

> 286. Borish, L., Rossenwasser, L., " Update on

> Cytokines, " J. Allergy Clin. Immunol. 97:719-734

> (1996) [Record No. 7146].

>

> 287. Tews, D.S., Goebel, H.D., " Cytokine Expression

> Profile in Idiopathic Inflammatory Myopathies, " J.

> Neuropathol. Exp. Neurol. 55:342-347 (1996) [Record

> No. 7147].

>

> 288. Middleton, G.D., et. al., " Effect of Alpha

> Interferon on Pain Thresholds and Fibromyalgia, "

> Arthritis & Rheumatism 37(9)(Supp):S14 (1994)

> [Record No. 7148].

>

> 289. Iwakura, Y., " Autoimmunity Induction by Human T

> Cell Leukemia Virus Type 1 in Transgenic Mice That

> Develop Chronic Inflammatory Arthropathy Resembling

> Rheumatoid Arthritis in Humans, " J. Immunol.

> 155:1588-598 (1995) [Record No. 7149].

>

> 290. Vial, T., Descotes, J., " Clinical Toxicity of

> Cytokines Used as Hemopoietic Growth Factors, " Drug

> Saf.13(6):371-406 (1995) [Record No. 7150].

>

> 291. Gause, B.L., Sznol, M., Kopp, W.C., et. al.,

> " Phase 1 Study of Subcutaneously Administered

> Interleukin-2 in Combination with Interferon

> Alpha-2A in Patients with Advanced Cancer, " J. Clin.

> Oncol. 14(8):2234-2241 (1996) [Record No. 7151].

>

> 292. Following the adverse publicity on silicone gel

> breast implants in 1992, Dow Corning announced that

> it would fund $10 million to research the effects of

> silicone gel breast implants in the body. As now

> demonstrated by the Affidavit of the General Counsel

> of Dow Corning, Dow sought reimbursement of this

> research as part of its litigation defense costs,

> stating that the studies were funded only after

> considering their impact on Dow's " litigation

> strategy. " J. R. Affidavit, ¶ 4 (7/10/95)

> [Record No. 0486].

>

> 293. on, M., Dow Corning Memo to LeVier and

> Cook re: PSEF Meeting of 3/12/93, DCC 279061322 -

> 279061325 (3/17/93) [Record No. 7111].

>

> 294. Dow Corning had announced in 1992 that they

> were devoting $10 million to research on silicone

> gel breast implants and solicited research proposals

> directly and through the Plastic Surgery Educational

> Foundation.

>

> 295. Cook, R.R., Dow Corning Letter to Noel Rose,

> DCC 279022039 - 279022041 (12/22/93) [Record No.

> 7270] (A blind copy of the letter was sent to

> (Dow Corning General Counsel) and

> Bernick (Kirkland & Ellis)).

>

> 296. Rose, N.R., Letter to P. Klykken, DCC 279022042

> - 279022043 (12/8/93) [Record No. 7172].

>

> 297. Rose, N.R. Letter to Ralph Cook, DCC 279022038

> (1/5/94) [Record No. 7109].

>

> 298. Naim, J.O., Zhang, J.W., Van Oss, C.J., " In

> Vitro Activation of Human Monocytes by Various

> Plasma Proteins Adsorbed onto Silicone Elastomers,

> Gels and Oils, " Surfaces in Biomaterials, Abstract,

> pp. 105-106 (1996) [Record No. 7106] (finding high

> levels of inflammatory cytokines (IL-1, TNF alpha,

> and Il-6) in association with plasma absorbed onto

> silicone).

>

> 299. Bommer, J., Weinreich, T., Lovett, O.T.T., et

> al., " Particles from Dialysis Tubing Stimulate

> Interleuken-1 Secretion by Macrophages, " Nephrol

> Dial Transplant 5: 208-213 (1990) [Record No. 7166]

> (finding that macrophages from silicone and

> PVC-loaded animals spontaneously release high levels

> of IL-1).

>

> 300. Cabral, A. R., Alcocer-Varela, J.,

> Orozco-Topete, R., et al., " Clinical,

> Histopathological, Immunological and Fibroblast

> Studies in 30 Patients with Subcutaneous Injections

> of Modelants Including Silicone and Mineral Oils, "

> La Revista de Investigacion Clinics 46(4): 257-266

> (July-August 1994) [Record No. 0469] (finding an

> association between injected silicone oil and the

> development of autoimmune disease and implicating

> IL-1 in the process).

>

> 301. Naidu, S.H., Beredjiklian, P., Adler, L., et

> al., " In Vivo Inflammatory Response to Silicone

> Elastomer Particulate Debris, " Journal of Hand

> Surgery 21A(3):496-500 (1996) [Record No. 7008].

>

> 302. Naim, J.O., Zhang, J.W., van Oss, C.J., " In

> Vitro Activation of Human Monocytes by Various

> Plasma Proteins Adsorbed Onto Silicone Elastomer,

> Gels and Oils, " Surfaces in Biomaterials, 105-106

> (1996) [Record No. 7106].

>

> 303. Id.

>

> 304. Id. Naim placed silicone elastomer from a

> McGhan silicone breast implant, silicone gel from a

> Dow Corning silicone breast implant, and Dow Corning

> 360 medical fluid in 100 microliters of human

> albumin, human fibrinogen, and human IgG, and

> allowed these to adsorb overnight. Using commercial

> immunassay kits, they determined the concentration

> of human IL-1, IL-6 and TNF-[alpha].

>

> 305. Naidu, S.H., Beredjiklian, P., Adler, L., et

> al., " In Vivo Inflammatory Response to Silicone

> Elastomer Particulate Debris, Journal of Hand

> Surgery21A(3):496-500 (1996) [Record No. 7008].

>

> 306. Krause, T.J., on, F.M., Liesch, J.G., et

> al., " Differential Production of Interleukin 1 on

> the Surface of Biomaterials, " Arch. Surg.

> 125:1158-1160 (1990) [Record No. 1219].

>

> 307. Lossing, C., Hansson, H., " Peptide Growth

> Factors and Myofibroblasts in Capsules Around Human

> Breast Implants, " Plastic and Reconstruct Surgery

> 91(7): 1277-1286 (6/93) [Record No. 2929].

>

> 308. Wells, A.F., s, S., Gunasekaran, S., et.

> al., " Local Increase in Hyaluronic Acid and

> Interleukin-2 in the Capsules Surrounding Silicone

> Breast Implants, " Ann. Plast. Surg. 33: 1-5 (1994)

> [Record No. 0436] (finding large amounts of Il-2 in

> the inflamed areas surrounding breast implants). See

> also Wells, A.F., Klareskog, L., Lindblad, S.,

> " Correlation Between Increased Hyaluronan Localized

> in Arthritic Synovium and the Presence of

> Proliferating Cells: a Role for Macrophage-Derived

> Factors, " Arthritis & Rheum35:391-396 (1992) [Record

> No. 7139].

>

> 309. Evered, D., Whelan, J., " The Biology of

> Hyaluronan, " Ciba Foundation Symposium 143,

> Chichester, UK, Wiley and Sons (1989) [Record

> No. 7136]; Engstrom-t, A., Hallgren, R.,

> " Circulating Hyaluronic Acid Levels Vary with

> Physical Activity in Healthy Subjects and in

> Rheumatoid Arthritis Patients - Relationship to

> Synovitis Mass and Morning Stiffness, " Arthritis &

> Rheum 30:1333-1338 (1987) [Record No. 7138]; Wells,

> A.F., Klareskog, L., Lindblad, S., " Correlation

> Between Increased Hyaluronan Localized in Arthritic

> Synovium and the Presence of Proliferating Cells: A

> Role for Macrophage-Derived Factors, " Arthritis &

> Rheum 35:391-396 (1992) [Record No. 7139]; Dahl,

> I.M.S., Husby, G., " Hyaluronic Acid Production in

> Vitro by Synovial Lining Cells from Normal and

> Rheumatic Joints, " Ann Rheum Dis 44:647-657 (1985)

> [Record No. 7140].

>

> 310. Duvic, M., , D., Menter, A., et al.,

> " Cutaneous T-cell lymphoma in association with

> silicone breast implants, " Journal of the American

> Academy of Dermatology 32(6):939-942 (1995) [Record

> No. 0135].

>

> 311. Garland, L.L., Ballester, O.F., Vasey, K.B., et

> al., " Multiple Myeloma in Women with Silicone Breast

> Implants: Serum Immunoglobulin and Interleuken-6

> Studies of Women at Risk, " Immunology of Silicones,

> pp. 361-366 (1996) [Record No. 0448].

>

> 312. Potter, M., on, S., Wiener, F., et al.,

> " Induction of Plasmacytomas With Silicone Gel In

> Genetically Susceptible Strains of Mice, " J. Natl.

> Cancer Inst. 86(14):1058-1065 (7/20/94) [Record No.

> 1772]; Nordan, R.P., Potter, M., " A

> Macrophage-derived Factor Required by Plasmacytomas

> for Survival and Proliferation in Vitro, " Science

> 233:566-569 (1986) [Record No. 7131]; Shacter, E.,

> Avzadon, G.K., , J., " Elevation of

> Interleukin-6 in Response to a Chronic Inflammatory

> Stimulus in Mice: Inhibition by Indomethacin, " Blood

> 80:194-202 (1992) [Record No. 7132]; Suematsu, S.,

> Matsusaka, T., Matsuda, T., et al., " Generation of

> Plasmacytomas with the Chromosomal Translocation

> t(12;15) in Interleukin-6 Transgenic Mice, Proc.

> Nat'l. Acad. Sci. 89:232-235 (1992) [Record No.

> 7133].

>

> 313. Felix, K., Janz, S., Pitha, J., et al.,

> " Cytotoxicity and Membrane Damage in vitro by

> Inclusion Complexes Between -Cyclodextrin and

> Siloxanes, " Immunology of Silicones, pp. 93-99

> (1996) [Record No. 0140].

>

> 314. Blackburn, W.D., Grotting, J.C., Everson, M.P.,

> " Lack of Evidence of Systemic Inflammatory Rheumatic

> Disorders in Symptomatic Women with Breast

> Implants, " Plast. Reconstr. Surg. 99: 1054-1060

> (1997) [Record No. 6051] (failing to find elevated

> levels of circulating inflammatory cytokines).

>

> 315. Das, S.K., , M., Ellsaesser, C., et al.,

> " Macrophage Interleukin-1 Response to Injected

> Silicone in a Rat Model, " Ann. Plast. Surg. 28:

> 535-537 (1992) [Record No. 1611].

>

> 316. Cuellar, M.L., Gutierrez, M., Cabrena, G., et.

> al., " Soluble Intercellular Adhesion Molecule 1

> (Sicam-1) and Other Acute Phase Reactants Such as

> Esr and Crp Are Not Elevated in Silicone Breast

> Implant Patients, " Abstract, Arthritis and

> Rheumatism 36(9):5219, Abstract C90 ( 9/93) [Record

> No. 0673] (failing to find elevated markers of

> inflammation in sera of patients with silicone

> breast implant associated human adjuvant disease).

>

> 317. Garland, L.L., Ballester, O.F., Vasey, K.B., et

> al, " Multiple Myeloma in Women with Silicone Breast

> Implants. Serum Immunoglobulin and Interleukin-6

> Studies in Women at Risk, " Immunology of Silicones,

> pp. 361-366 (1996) [Record No. 0448].

>

> 318. Roitt I., " The Production of Effectors, "

> Essential Immunology 8th Ed, Chapter 10, pp. 173-193

> (1994) [Record No. 7144].

>

> 319. Whiteside, T.L., " Cellular Immunology:

> Monitoring of Immune Therapies, " Handbook of Human

> Immunology , Chapter 10, pp. 343-380 at p. 362

> (1997) [Record No. 7167].

>

> 320. Keffer, J., Probert, L., Cazlaris, H., et al,

> " Transgenic Mice Expressing Human Tumour Necrosis

> Factor: a Predictive Genetic Model of Arthritis, "

> The EMBO Journal 10(13): 4025-4031 (1991) [Record

> No. 0411].

>

> 321. Id.

>

> 322. Mohr, C., " Systemic Macrophage Stimulation in

> Rats with Silicosis: Enhanced Release of Tumor

> Necrosis Factor Alpha from Alveolar and Peritoneal

> Macrophages, " Am. J. Respir. Cell Mol. Biol. 5:

> 395-402 (1991) [Record No. 5243]; Piguet, P.F.,

> Grau, G.E., Vessalli, P., " Tumor Necrosis Factor and

> Immunopathology, " Immunol. Res.10:122-140 (1991)

> [Record No. 7165].

>

> 323. Id. at p. 111.

>

> 324. Id.

>

> 325. Id. at pp. 113-114.

>

> 326. Oppenheim, J.J., Ruscetti, F.W., Faltynek, C.,

> et al., " Cytokines; Complement & Kinin, " Basic &

> Clinical Immunology 8th Ed. Chapters 9-10, pp.

> 105-136 at p. 112 [Record No. 7127].

>

> 327. Id.

>

> 328. , K.A., " Interleukin-2, " Sci Am 262:26-30

> (1990) [Record No. 7134].

>

> 329. Dorshkind K., " Bone Marrow Stromal Cells and

> Their Factors Regulate B Cell Differentiation, "

> Immunol Today8:7-10 (1987) [Record No. 7135].

>

> 330. Oppenheim, J.J., Ruscetti, F.W., Faltynek, C.,

> et al., " Cytokines; Complement & Kinin, " Basic &

> Clinical Immunology 8th Ed., Chapters 9-10, pp.

> 105-136 (1994) [Record No. 7127].

>

> 331. Oppenheim, J.J., Ruscetti, F.W., Faltynek, C.,

> et al., " Cytokines; Complement & Kinin, " Basic &

> Clinical Immunology, Chapters 9-10, pp. 105-136

> (1994) [Record No. 7127]; Detrick B., Hooks, J.J.

> " Cytokines in Human Immunology, " Handbook of Human

> Immunology, pp. 233-266 (1997) [Record No. 7126].

>

> 332. See, e.g., Becker, Reece, Poeni, " Cellular

> Aspects of Immune Response, " The World of the Cell

> 3rd Ed., Chapter 24, pp. 784-800 (1996) [Record No.

> 7143].

>

> 333. Roitt, I., " The Production of Effectors, "

> Essential Immunology, 8th Ed., Ch. 10, 173-193

> (1994) [Record No. 7144].

>

> 334. Oppenheim, J.J., Ruscetti, F.W., Faltynek, C.,

> et al., " Cytokines; Complement & Kinin, " Basic &

> Clinical Immunology, pp.105-136 (1994) [Record No.

> 7127]; Detrick, B., and Hooks J.J., " Cytokines in

> Human Immunology, " Handbook of Human Immunology,

> Chapter 7, pp. 233-266 (1997) [Record No. 7126].

>

> 335. Mohr, C., " Systemic Macrophage Stimulation in

> Rats with Silicosis: Enhanced Release of Tumor

> Necrosis Factor-alpha from Alveolar and Peritoneal

> Macrophages, " Am. J. Respir. Cell Mol. Biol.

> 5:395-402 (1991) [Record No. 5243].

>

> 336. Watkins, L. R., Maier, S.F., Goehler, L.E.,

> " Cytokine to Brain Communication: a Review and

> Analysis of Alternative Mechanisms, " Life Sciences

> 57(11):1011-1026 (1995) [Record No. 5375].

>

> 337. Borish, L., Rosenwasser, L.J., " Update on

> Cytokines, " J. Allergy Clin Immunol. 97:719-734

> (1996) [Record No. 7146]; Tews, D.S., Goebel, H.D.,

> " Cytokine Expression Profile in Idiopathic

> Inflammatory Myopathies, " J. Neuropathol. Exp.

> Neurol. 55:342-347 (1996) [Record No. 7147];

> Middleton, G.D., McFarlin, J.E., Lee, W., et al.,

> " Effect of Alpha Interferon on Pain Thresholds and

> Fibromyalgia, " Arthritis & Rheum. 370 (Supp): S214

> (1994) [Record No. 7148]; Iwakura, Y., Saijo, S.,

> Koika, Y., et al., " Autoimmunity Induction by Human

> T Cell Leukemia Virus Type 1 in Transgenic Mice That

> Develop Chronic Inflammatory Arthropathy Resembling

> Rheumatoid Arthritis in Humans, " J. Immunol

> 155:1588-1598 (1995) [Record No. 7149]; Vial, T.,

> Descotes, J. " Clinical Toxicity of Cytokines Used as

> Hemopoietic Growth Factors, " Drug Saf. 6:371-406

> (1995) [Record No. 7150]; Gause, B.L., Sznol, M.,

> Kopp, W.C., et al., " Phase I Study of Subcutaneously

> Administered Interleukin-2 in Combination with

> Interferon Alpha-2A in Patients with Advanced

> Cancer, " J. Clin. Oncol. 8:2234-2241 (1996) [Record

> No. 7151].

>

> 338. Goehler, L.E., Busch, C.R., Tartaglia, N., et

> al., " Blockade of Cytokine Induced Conditioned Taste

> Aversion by Sub-diaphragmatic Vagotomy: Further

> Evidence for Vagal Mediation of Immune Brain

> Communication, " Neuroscience Letters 185:163-166

> (1995) [Record No. 7152].

>

> 339. Watkins, L. R., Maier, S.F., Goehler, L.E.,

> " Cytokine to Brain Communication: a Review and

> Analysis of Alternative Mechanisms, " Life Sciences

> 57(11):1011-1026 (1995) [Record No. 5375].

>

> 340. Id.

>

> 341. Fleshner, M., Goehler, L.E., Hermann, J., et

> al., " Interleukin-1 Beta Induced Corticosterone

> Elevation and Hypothalamic NE Depletion Is Vagally

> Mediated, " Brain Research Bulletin 37(6):605-610

> (1995) [Record No. 7154].

>

> 342. Goujon, E., Parnet, P., Laye, S., et al.,

> " Adrenalectomy Enhances Pro-Inflammatory Cytokines

> Gene Expression, in the Spleen, Pituitary and Brain

> of Mice in Response to Lipopolysaccharide, "

> Molecular Brain Research 36:53-62 (1996) [Record No.

> 7155].

>

> 343. Goujon, E., Parnet, P., Cremona, S., et al,

> " Endogenous Glucocorticoids Down Regulate Central

> Effects of Interleukin-1B on Body Temperature and

> Behaviour in Mice, " Brain Research 702:173-180

> (1995) [Record No. 7156].

>

> 344. Fleshner, M., Goehler, L.D., Hermann, J., et

> al., " Interleukin-1 Beta Induced Corticosterone

> Elevation and Hypothalamic NE Depletion Is Vagally

> Mediated, " Brain Research Bulletin37(6):605-610

> (1995) [Record No. 7154].

>

> 345. Aubert, A., Vega, C., Dautzer, R., et al,

> " Pyrogens Specifically Disrupt the Acquisition of a

> Task Involving Cognitive Processing in the Rat, "

> Brain, Behavior, and Immunity 9:129-148 (1995)

> [Record No. 7157].

>

> 346. Watkins, L.R., Maier, S.F., Goehler, L.E.,

> " Cytokine-to-brain Communication: a Review &

> Analysis of Alternative Mechanisms, " Life

> Sciences57(11):1011-1026 (1995) [Record No.

> 5375](cytokines signal brain by stimulating afferent

> terminals of peripheral nerves at local sites of

> synthesis and release).

>

> 347. Watkins, L.R., Wiertelak, E.P., Goehler, L.E.,

> et al., " Characterization of Cytokine-induced

> Hyperalgesia, " Brain Research 654:15-26 (1994)

> [Record No. 7158].

>

> 348. Watkins, L.R., Eoehler, L.E., Relton, J.K., et

> al., " Blockade of Interleukin-1 Induced Hyperthermia

> by Subdiaphragmatic Vagotomy: Evidence for Vagal

> Mediation of Immune-brain Communication, "

> Neuroscience Letters 183: 27 (1995) [Record No.

> 7159] (IL-1 beta elicits various illness symptoms

> including hyperthermia)

>

> 349. Goehler, L.E., Vbusch, C.R., Rartaglia, N., et

> al., " Blockade of Cytokine Induced Conditioned Taste

> Aversion by Subdiaphragmatic Vagotomy: Further

> Evidence for Vagal Mediation of Immune-brain

> Communication, " Neuroscience Letters 1985: 163-166

> (1995) [Record No. 7152] (IL-1 beta and TNF-alpha

> elicit various illness symptoms including avoidance

> of novel tastes with which they have been paired

> (conditioned taste aversion)).

>

> 350. Parnet, P., Amindari, S., Wu, C., et al.,

> " Expression of Type I and Type II Interleukin-1

> Receptors in Mouse Brain, " Molecular Brain Research

> 27: 63-70 (1994) [Record No. 7160] (mouse brain

> expresses both type I and type II IL-1 receptor mRNA

> and proteins and offer further support to the idea

> that type I IL-1 receptors are synthesized and

> expressed by neurons).

>

> 351. Kent, S., Bluthe, R., Kelley, K., et al.,

> " Sickness Behavior as a New Target for Drug

> Development, " Trends Pharmacal. Sci. 13: 24-28

> (1992) [Record No. 7161].

>

> 352. Kluger, M.J., " Fever: Role of Pyrogens and

> Cryogens, " Physiol. Rev. 71: 93-127 (1991) [Record

> No. 7162].

>

> 353. Laye, S., Parnet, P., Goujon, E., et al.,

> " Peripheral Administration of Lipopolysaccharide

> Induces the Expression of Cytokine Transcripts in

> the Brain and Pituitary of Mice, " Molecular Brain

> Research 27:157-162 (1994) [Record No. 7164].

>

> 354. Aubert, A., Vega, C., Dantzer, R., et al.,

> " Pyrogens Specifically Disrupt the Acquisition of a

> Task Involving Cognitive Processing in the Rat, "

> Brain, Behavior, and Immunity 9:129-149 (1995)

> [Record No. 7157].

>

> 355. Parnet, P., Amindari, S., Wu, C., et al.,

> " Expression of Type I and Type II Interleuken-1

> Receptors in Mouse Brain, " Molecular Brain Research

> 27:63-70 (1994) [Record No. 7160].

>

> 356. Lefkowitz, D.L., Mills, K., Lefkowitz, S.S. et

> al., " Neutrophil--macrophage Interaction: a Paradigm

> for Chronic Inflammation, " Medical Hypotheses 44:

> 58-62 (1995) [Record No. 1539] (proposing that the

> release of myeloperoxidase (MyPo) from neutrophils

> and subsequent binding to macrophages initiates a

> cascade of events which enhance the production of

> reactive oxygen intermediates and Cytokine

> expression resulting in the chronic inflammatory

> state associated with autoimmune diseases).

>

> 357. Piquet, P.F., Grau, G.E., Vessalli, P., " Tumor

> Necrosis Factor and Immunopathology, " Immunol. Res.

> 10:122-140 (1991) [Record No. 7165].

>

> 358. Naim, J.O., Zhang, J.W., van Oss, C.J., " In

> Vitro Activation of Human Monocytes by Various

> Plasma Proteins Adsorbed onto Silicone Elastomer,

> Gels & Oils, " Surfaces in Biomaterials, Abstract,

> pp. 105-106 (1996) [Record No. 7106].

>

> 359.Katzin, W.E., Feng, L., Abbuhl, M., et al,

> " Phenotype of Lymphocytes Associated with the

> Inflammatory Reaction to Silicone Gel Breast

> Implants, " Clinical and Diagnostic Laboratory

> Immunology 3(2):156-161 (3/96) [Record No. 2286].

>

> 360. O'Hanlon, T.P., Okada, S., Love, L.A., et al,

> " Immunohistopathology and T Cell Receptor Gene

> Expression in Capsules Surrounding Silicone Breast

> Implants, " Immunology of Silicones, pp. 237-242

> (1996) [Record No. 0279].

>

> 361. Ladin, D.A., Saed, B.M., Fivenson, D.P.,

> " T-Cell Response in Silicone Gel Breast Implant

> Capsules, " Surgical Forum 45:730-731 (1994) [Record

> No. 7107].

>

> 362. Stark, G.B., Gobel, M., Jaeger, K.,

> " Intraluminal Cyclosporine A Reduces Capsular

> Thickness Around Silicone Implants in Rats, " ls

> of Plastic Surgery 24:156-161 (1990) [Record No.

> 1206].

>

> 363. Tang, L., Eaton, J.W., " Inflammatory Responses

> to Biomaterials, " Am. J. Clin. Pathol.103:466-471

> (1995) [Record No. 5349].

>

> 364. Id.

>

> 365. Picha, G., Goldstein, J., " Analysis of the

> Soft-Tissue Response to Components Used in the

> Manufacture of Breast Implants: Rat Animal Model, "

> Plastic and Reconstructive Surgery 87(3): 490-500

> (3/91) [Record No. 1568].

>

> 366. Katzin, W., Feng, L. Abbuhl, M., et al.,

> " Phenotype of Lymphocytes Associated with the

> Inflammatory Reaction to Silicone Breast Implants, "

> Clinical and Diagnostic Laboratory Immunology

> 3(2):156-161 (1996) [Record No. 2286]; Wells, A.F.,

> s, S., Gunasekaran, S., et al., " Local

> Increase in Hyaluronic Acid and Interleukin-2 in the

> Capsules Surrounding Silicone Breast Implants, "

> ls of Plastic Surgery 33(1):1-5 (1994) [Record

> No. 0436]; Ladin, D.A., Saed, G.M., Fivenson, D.P.,

> " T-Cell Response in Silicone Gel Breast Implant

> Capsules, " Surgical Forum 45:730-731 (1994) [Record

> No. 7107].

>

> 367. See Clinical Experience Section.

>

> 368. Lossing, C., Hansson, H., " Peptide Growth

> Factors and Myofibroblasts in Capsules Around Human

> Breast Implants, " Plastic and Reconstruct Surgery

> 91(7): 1277-1286 (6/93) [Record No. 2929].

>

> 369. Peimer, C.A., Medige, J., Eckert, B.S., et al.,

> " Reactive Synovitis After Silicone Arthroplasty, "

> Journal of Hand Surgery 11A(5):624-638 (1986)

> [Record No. 7105].

>

> 370. Ahn, C.Y., Shaw, W.W., Narayanak, K., et al.,

> " Residual Silicone Detection Using MRI Following

> Previous Breast Implant Removal: Case Reports, "

> Aesthetic Plastic Surgery 19:361-367 (1995) [Record

> No. 7050].

>

>