SUPPRESSED NATURAL KILLER CELL ACTIVITY IN PATIENTS

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SUPPRESSED NATURAL KILLER CELL ACTIVITY IN PATIENTS

WITH SILICONE IMPLANTS: REVERSAL UPON EXPLANTATION BY:

ANDREW W, CAMPBELL, M.D., Clinical assistant professor, University

of

Texas

Health Science Center, Medical Director, Center For Immune,

Environmental

and Toxic

Disorders, Houston, Texas.

NACHMAN BRAUTBAR, M.D., clinical Professor of Medicine, University

of

Southern California School of Medicine, Medical Director, Center for

International

Occupational and Toxicological Medicine, Los Angeles, California.

ARISTO VOJDANI, PH.D., Associate Professor of Medicine, Drew

University

School

of Medicine and Science, Department of Medicine and Dermatology,

Director,

Immunosciences Lab, Inc., Los Angeles, California.

PUBLISHED, TOXICOLOGY AND INDUSTRIAL HEALTH 10: 3 MAY - JUNE

1994

Address all correspondence to:

W. , M.D.

14441 Memorial Drive, Suite 6

Houston, Texas 77079

Reprinted with the permission of W. , M.D.

SUPPRESSED NATURAL KILLER CELL ACTIVITY IN PATIENTS WITH

SILICONE

EXPLANTATION - ABSTRACT

We have previously shown that natural killer cell activity is

significantly suppressed in

patients with silicone breast implants. These patients were

symptomatic

and the suppression

of natural killer cell activity was associated with additional

significant

immunological

abnormalities. (1) Our studies have recently been confirmed by

Srnith et

al., (2) who

described natural killer cell activity suppression following

exposure to

silicone gel. and

reversal upon removal of the gel.

This study has been designed to evaluate the natural killer cell

activities in symptomatic

women with silicone breast implants after implantation and after the

explantation of the

implants. Each patient served as its own control. Our findings show

a

marked significant

increase in previously suppressed natural killer cell activity.

These

findings are compatible

with the recent studies in experimental animals, showing that

administration of silicone

reduces natural killer cell activity. and that this is reversible

upon

removal of silicone.

Since NK cells are important in the control of tumor cell growth we

propose here that

patients with reduced NK cell activity are at a higher risk of

developing

cancer. a notion

recently described in experimental animals. (3 & 4) *Nachman

Brautbar,

M.D. to whom all

communications should be addressed at: 2222 Ocean View Avenue, Los

Angles.

California

90057

INTRODUCTION

NK cells are sensitive indicators of activation by biologic response

modifiers, and their

monitoring has been used to document alteration in the activity of

circulating immune cells,

Abnormalities in NK cell activities have also been described in

Autoimmune

Disorders.

Most recently, NK cell activities have been shown to be affected in

patients exposed to

chemicals, and in patients with silicone breast implants.

Since the symptomatology of patients with silicone breast implants

has

been associated with

multiple immunological abnormalities, including production of

autoantibodies.(',',',')

stimulation and suppression of T cells, (1) elevation of circulatory

immune complexes, (1)

and since the reversal of symptomology upon removal of the implants

has

been associated

with reversal of the immunological abnormalities, (5 )it is logical

to

suggest that

immunological marker, such as NK cell activity, will follow these

patterns: Suppression as a

result of exposure to silicone and reversal to normalization upon

removal

of the silicone.

Indeed, our studies here support this notion and further support the

concept of an immune

response to silicone breast implants.

PATIENTS AND METHODS

Forty women who underwent silicone breast implants and were

evaluated for

symptoms

ranging from joint pain, muscle pain central nervous system

symptomatology, skin rashes

and myalgias (3) have been studied. Natural Killer cell activities

were

studied prior to the

explantation and 3 to 15 months after explantation with an average

of 8

months: +/- 1.2 S.

E.

ANALYSIS OF NATURAL KILLER CELL ACTIVITIES

Separation of Human PBL Mononuclear cells from patients and controls

were

separated

from the fresh whole blood by Ficoll-hypaque density gradient

centrifugation (Litton

Bionetics, Rockville, MD). The lymphocyte band at the interface was

collected and cells

pelleted by centrifugation, washed twice in RPMI- 1640 and suspended

in

complete medium,

consisting of RPMI-1640 supplemented with 10)% human AB serum, 2mM

glutamine,

25mM Hepes (pH 7.2), 50 units penicillin, and 50 units streptomycin

per mi.

NK Cell Cytotoxicity Assay A modified (51)CR-release assay, as

described

previously, was

employed. Briefly, 1X10(4) (51)Cr-labeled K562 target cells (New

England

Nuclear

Corporation, Boston, MA) in 0. I mil CM were added per well in

microliter

plates. Effector

cells were pipetted into quadruplicate wells to give effector:target

cell

ratios of 100: 1, 50: 1

and 25: 1. These cells were allowed to interact at 3 7degC for 4 hr

in an

atmosphere of 5%

C02/95% air.

(51)Cr-release was determined by centrifuging the plates at 1000 x g

for 5

min and

harvesting 0.1 ml of the culture supernatant for counting gamma

counter.

Total release was

determined by adding 100 ul of 1.0% Triton X-100 and spontaneous

release

by adding

labeled target cells alone in CM.

The percent " Cr-release was determined by the experimental (R(e)),

spontaneous (R(s)). and

total (R(t)) release by the following formula:

" Cr-release = (R(e)) - (R(s))

------- x 100%

(R(t))- (R(s))

Lytic units (LU) were calculated from effector titration curves, and

(1)LU

was defined as

the number of effector cells required to achieve 20% lysis. LU/ 10

(6) is

the number of LU

in 10(6) effector cells. For assay reproductability, recommended

criteria

were employed,

Statistical analysis of significance utilized the paired + test and

the

difference of the means

of the of the 2 groups studied.

RESULTS

Table I shows the individual values for natural killer cell

activities

prior to explantation and

after explantation.

There was a marked and significant increase in natural killer cell

activity after explantation.

Natural killer cell activity was 27.00Ò3.0 (S.E.) Prior to

explantation

and 36.89+- 3,7 (S.E.)

after explantation. The average age of implants was 9.5 years Ò

0.96. P

value for the

difference of the means between NK cell I and NK cell III was

P<0.032.

DISCUSSION

Previous studies reported from our laboratory have shown a

significant and

marked

reduction of natural killer cell activities in patients with

silicone

breast implants. (1) This

reduction was attributed to direct effect by silicone through

activation

of cellular or

humoral mechanisms secondary to an immune response to adjuvant

effects of

silicone.

The following functions have been ascribed to natural killer cells:

1) control of tumor cell growth

2) involvement in the control of microbial infections,

3) immunoregulatory properties

4) involvement in the development of graft versus host disease,

5) contributes to the development of some forms of diabetes, and

6) involvement in various gastrointestinal diseases. (7) Natural

killer

cells mediate a

non-major histocompatibility complex with restricted killing against

target cells, which are

termed " NK susceptible. " This lytic activity can be observed

against a

variety of neoplastic

and viral-infected cells in non-major histocompatibility complex

restricted function.

Inhibition of natural killer cell activity has been described using

prostaglandin A and E

glucocorticoids and prolactin as well as luteinizing hormone.

Exposure of

murine or human

natural killer cells to agents such as anti-asialo-GNfl or

Prostaglandins

induce dose-response

suppression. Other cellular mechanisms have been described in

association

with suppression

of natural killer cells, including suppressor cells, which play a

physiological role in

regulation of NK cell activity in vivo. Most recently, exposure to

orgarfic solvents has been

shown to cause suppression of NK cell activity and stimulation of

several

autoantibodies. (8)

Our findings here show that the suppressed activity of the natural

killer

cells is reversed and

improved markedly upon removal of the silicone implants, indicating

that

the direct or

indirect effect of the silicone breast implants, which caused

suppression

of natural killer cell

activities, has been eliminated by removal of the silicone, the

causative

agent. This was not

correlated with the length of implantation which ranged anywhere

from 0.6

to 16 years

with an average of 9.5 +- 0.96 years, nor was this correlated with

the

length of time period

from the explantation.

Our studies here are in agreement with the studies reported recently

by

et al., (2)

which reported in rats implanted with silicone gel a marked

reduction, in

natural killer cell

activity, and reversal of this suppression immediately following

removal

of the implants.

Our studies here demonstrate clearly that symptomatic patients with

silicone breast implants

significantly are associated with a marked reduction of natural

killer

cell activity, and that

this reduction is improved upon removal of the silicone breast

implants.

The issue of cancer

development in patients with silicone breast implants has not been

addressed at the

molecular level, nor has it been addressed at the clinical level.

Recent studies (9) have shown carcinogenicity of silicone in

experimental

animals. Indeed,

these studies were followed by an editorial calling for

epidemiological

studies in patients

with silicone breast implants to further evaluate the incidence of

hematological

malignancy.(10) These investigators did not measure natural killer

cell

activities in these

studies, however, based on our clinical studies here, and on the

experimental studies by

et al, (2) it is strongly suggested that one mechanism

contributing to the

carcinogenicity is the reduced activity or inhibition of natural

killer

cells. The actual

mechanisms by which silicone inhibits NK cell activity is not clear.

Additional studies at the

molecular level are required to further understand the mechanism of

suppression of NK cell

activity by silicone.

Our studies described here indicate that silicone does inhibit the

functions of natural killer

cell activities and that this inhibition is reversible, however, may

be

associated with

reduction of ability to control tumor cells, and in those patients

who

have chronic reduction

of natural killer cell activity, may be associated with

carcinogenicy. We

propose clinical

studies to find the incidence of cancers in patients with silicone

breast

implants.

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