Date: Mon, 30 Nov 1998 20:46:41 -0700

Silicone Breast Implants in Relation to Connective Tissue Diseases and Immunologic Dysfunction A Report by a National Science Panel to the Honorable Sam C. Pointer Jr., Coordinating Judge for the Federal Breast Implant Multi-District Litigation

Betty A. Diamond Barbara S. Hulka

Nancy I. Kerkvliet Peter Tugwell

Chapter I

Review of Animal Studies Relevant to Silicone Toxicity

I. What Is Silicone?

Silicone is the name given to a family of synthetic polymers composed of a repeating Si-O backbone and carbon-linked side-groups. Si-C bonds do not existn in nature but can be formed under appropriate manufacturing conditions. Then most common example of a silicone is poly(dimethylsiloxane)(PDMS), shown in Figure 1. The dimethylsiloxane units are the basic building blocks of silicones (Lane et al., 1996). Depending on the number of dimethylsiloxane units linked together as a linear polymer and degree of cross-linking between polymer chains, products of various textures and strengths are produced, including forms that mimic human body tissues. In general, straight chain polymers are liquids that increase in viscosity as the chain lengthens (liquid gel). Increased cross-linking of the chains leads to increasingly rigid silicone materials (gel elastomer). Substitution of methyl (-CH3) groups with other side chains produces silicone derivatives with varied physical characteristics and chemical reactivities.

Based on a 1950 review article, initial laboratory studies of silicone oils had shown that silicones were "remarkably stable in comparison with other fluids of similar viscosity . . . and more resistant to oxidation and more water repellent than other fluids" (Barondes et al., 1950). They were also shown to have "good resistance to chlorine, nitric acid and hydrochloric acid, sodium chloride, sulfur dioxide and sulfuric acid up to 30% concentration." This early review also cautioned that silicone polymers "are not to be confused with the silicon (Si) compounds as sodium silicate, silica gel, and siliceous earth. Silica gel, for example, is a colloidal silica that absorbs water."

II. Utility and Significance of Animal Studies for Human Toxicity Assessment Experimental animal studies are used for safety assessment purposes prior to the introduction of a chemical or device for use in humans. These studies primarily use laboratory rats and mice, dogs, and rabbits, with additional animal species tested to address specific toxicology questions.

The data obtained from animal studies provide three main types of information. The first tests conducted in animals are generally referred to as "hazard identification." These studies are carried out to determine the possible biological/toxicological effects the chemical is capable of causing, and often incorporate very high exposure levels or unnatural routes of exposure. These studies are not intended to address the likelihood of effects in humans, but allow scientists to understand the basic ways in which the particular chemical interacts with the cells and tissues in a living mammalian organism.

The second major purpose of animal studies is to establish the relationship between exposure and effects and to characterize the dose-response for those effects. Animal studies are carried out using nearly identical groups of animals that differ only in their exposure to the test substance of interest. By controlling for as many other variables as possible (for example, age, sex, genetic background, environment, diet, etc.), any differences in responses between the controls and treated groups can be causally linked to exposure to the test substance.

Furthermore, by testing different levels of exposure (doses), it is possible to see the relationship between severity of effect and dose; that is, how much chemical is necessary to cause specific effects. The results of this phase of testing are useful in predicting human effects to the extent that appropriate animal models are used and good scientific methods are employed. Applicability of results to humans is also enhanced when similar effects are reported by different laboratories and when consistent effects of exposure are seen in more than one animal species. The results of such studies often determine the fate of products prior to marketing. Once a product is marketed, if problems appear to arise from human exposure, the animal data are valuable to support or refute limited or conflicting evidence in humans.

The third main value of animal toxicity studies is to determine the mechanisms by which a chemical interacts with living cells to produce its toxicity, an important factor in understanding how the chemical might induce or aggravate disease. Such mechanistic studies are particularly important if the benefits of the chemical (e.g., drug) outweigh the toxicity (e.g., side-effects) and the product will be marketed in spite of its recognized toxicity. By understanding the mechanisms for the toxicity, measures can be instituted to prevent or reduce the risk of toxicity. In this phase of testing, the approaches used are not dictated by government regulations and are limited only by the ingenuity of investigators and the amount of funding available for such studies. The relevance of such mechanistic studies in animals will depend on how well-defined the toxicity is in humans and the ability to reproduce the same toxic effects in animals.

III. Rationale for Analysis of Specific Animal Studies Relative to Silicone Toxicity When considering the whole data base of animal studies relating to silicone toxicity, several decision points were used during this analysis to determine the relevance of specific papers to SBI toxicity. The rationale for these decisions was as follows:

1. The term silicone has been used to represent many different types of materials that may have very different chemical characteristics when compared to PDMS. Therefore, toxicity studies of silicones that differ significantly from PDMS, are not present in SBIs, and are known to have different chemical reactivities than PDMS, were not included in this analysis. However, in many papers given to the Panel to review, the specific silicone materials tested were not described other than by code. In this case, it was assumed that a relevant form of PDMS was tested, and the data were reviewed and incorporated into this analysis.

2. Studies that examined the toxicity of silicone species that are found only as minor contaminants of SBIs were evaluated, but more critically in terms of their dose-response relationships. In general, minimal effects by minor species that were seen in animals only at high levels of exposure were considered not applicable to the SBI issue.

3. Studies in which relatively large doses of silicone were directly injected into tissues that would not be accessible by the silicone from SBIs in any significant concentration (eg., silicone injected into the brain) have been judged not relevant to the SBI issue.

4. Studies that are based on the oral route of exposure have been judged not applicable. Most silicones are poorly absorbed and do not result in appreciable systemic exposure from this route (Chenoweth et al., 1956). Thus, a lack of toxicity following dietary exposure cannot be used to infer lack of toxicity from SBIs. On the other hand, the possible hydrolysis of some small silicone molecules (e.g., D4) in the acid environment of the stomach also introduces a variable that would not be applicable to the SBI issue .

5. Based on the lack of any definitive evidence that silicone can be degraded to silicon or silica in the body, the toxicology of silicon or silica has not been reviewed for this report. Although very recent studies have provided evidence that D4 can be metabolized via oxidative demethylation (Varaprath et al., 1997), probably through the action of hepatic mixed function oxidase activity (McKim et al., 1988), there is no evidence that PDMS fluid, gel, or elastomer induce hepatic enzymes or are metabolized.

6. This review does not specifically address the issues of silicone leakage or metabolism since there is sufficient animal toxicity data available in which silicone fluids and gels were injected directly into tissue, modeling a worst-case scenario in which all of the silicone in the SBI, including minor species, had leaked through the elastomer and was free in the tissue. Furthermore, the results observed in long-term exposure studies of free fluid and gel would have reflected any migration or metabolism that might have occurred.

7. All literature forms provided to the Panel were reviewed, including peer-reviewed journal articles and non-peer- reviewed book chapters, abstracts, theses and reports. When only the abstract of a report was available, it was not used to provide the sole basis for any conclusions drawn. In judging the quality of individual studies, scientific credibility was strengthened by clearly written reports based on experiments that were hypothesis-driven, had adequate control groups (positive and negative), used well-documented and validated assays, evaluated the dose-response relationship, and were analyzed by accepted statistical tests. Credibility was also increased when conclusions drawn were biologically plausible.

IV. Animal Models for Atypical Connective Tissue Diseases When considering the question of silicones and "atypical connective tissue diseases" (ACTD), the relevance of animal models to human disease becomes an issue. Because most of the symptoms of ACTD are subjective, the disease constellation cannot be modeled in animals unless a surrogate marker for the disease can be identified. However, since the biological basis for the subjective symptoms is not known, only hypothetical causes of ACTD can be examined in animal studies. These hypothetical causes have been articulated by the plaintiffs to the Science Panel and are also found in various publications provided to the Panel. The evidence from animal studies to support or refute these hypotheses has been critically evaluated.

V. Historical Perspectives on Silicone Toxicity

The first review of silicone toxicology was published in 1950 wherein the results of standard testing of various PDMS fluids (DC200 series) in rats, rabbits, and mice were summarized (Barondes et al., 1950). Routes of exposure to silicone included oral intubation or intraperitoneal (ip) injection in rats; intradermal (id) or subcutaneous (sc) injection in mice and rabbits; intravenous (iv) injection in mice; and eye instillation or skin application in rabbits. The overall conclusions drawn from these studies was that the silicone fluids tested "are practically inert physiologically . . . . and nontoxic to the body tissues. When fed to laboratory animals in doses as high as 2%, no discernable ill effects were noted. There is little if any reaction when administered intradermally, subcutaneously or intramuscularly."

Based on the low toxicity of silicone fluids, and the development of a medical grade silicone rubber, the medical uses of silicone greatly expanded during the 1950s and early 1960s (Agnew et al., 1962; Andrews, 1966 ; Ballantyne et al., 1965; Braley, 1972; ). When certain adverse reactions to injected silicone fluid were reported, they tended to be attributed to the use of nonmedical grade or otherwise adulterated products. This position appears to have evolved from the fact that most clinical experience with silicone was very good, and most animal studies showed little reaction to pure silicone fluid (e.g., Dow Corning IND 2702, Informational Materials, 1968).

In the United States, the first SBI made of a silicone elastomer (rubber) envelope containing silicone gel was developed in 1960 and marketed in1963 (Braley, 1972). The silicone gel matrix was composed of high molecular weight linear PDMS polymers cross-linked via the presence of intermittent methyl vinyl groups within the linear chain (Lane and Burns, 1996). Based on the belief that the envelope protected the patient from exposure to the gel, clinical trials on SBIs were not conducted, and their use in humans was apparently allowed based on already- established successful clinical use of silicone rubber and other silicone prostheses. The major concern regarding silicone toxicity at this time appeared to be possible tumor development at the implant site, predicated on a mechanical theory of tumor induction. However, animal studies in the early 1960s described the tissue response to silicone rubber in rats as a fibrotic capsule formation that was accompanied by a mild chronic inflammatory response in some animals. Histiocytes and giant cells were observed (Agnew et al., 1962). These findings were not considered serious detriments to the clinical use of silicone because of the focus on carcinogenesis and the fact that few tumors were observed (Agnew et al., 1962).

In 1966, the cellular response to silicone fluid was described by Andrews in a preliminary report. In this study, silicone fluid was injected directly into the subcutaneous tissue of mice. Tissue responses were compared to mice injected with saline. Tissue sections were reported to show macrophages that had phagocytosed silicone. Similarly, Rees et al.(1966) and Ben-Hur et al. (1967) reported that silicone fluid injected ip or sc appeared to be phagocytosed and distributed systemically, likely via the lymphatics. Other studies by Ballantyne et al. (1965) showed that massive injections of silicone fluid in guinea pigs, while accompanied by phagocytosis, were well-tolerated by the animals. Sparchu and Clashman (1970) also reported evidence of systemic distribution of ip or sc injected silicone fluid in rats, but noted that much of the silicone appeared to be in extracellular vacuoles and not associated with inflammatory cells. As reviewed by Braley (1973), the use of silicone devices continued to expand in the 1960s, and by 1973, thousands of patients had received various forms of silicones in medical applications, including their growing use as mammary prostheses. Although complications from the clinical use of silicone fluid were recognized, few papers addressed complications from silicone gel implants and those were primarily related to local contracture. It is presumed that there was little concern over the safety of SBIs during this time. This viewpoint was supported by a report by Lilla and Vistnes (1975) who found little reaction to the long-term implantation of various types of SBIs in rabbits. Similarly, two-year dog and rat studies showed little reaction and no toxicity to multiple im, sc or id injections of silicone fluid (DC-360) (West and Jolly, 1976). Similar innocuous effects were seen in dogs that received various implant materials (presumably silicones) over a six-year period (Mastalski et al., 1977). In 1982, a conference at the National Institutes of Health on the safety of clinical applications of biomaterials noted the success of soft tissue augmentation of the breast in its Consensus Statement.

Additional toxicology studies continued to be carried out in the 1980s and into the 90s. The results of two independent two-year chronic toxicity studies of different silicone gels in rats indicated that tissue changes were observed locally at the site of the implant but no systemic toxicity was seen, based on the absence of changes in body weight and food consumption data, or clinical, gross or microscopic pathology results, including data from interim sacrifices (Goodman et al., 1988; Lemen et al., 1992). Tumors were observed at the site of implantation but tumor development was related to the process known as solid-state tumorigenesis. This effect appears to be a process unique to rats injected with free gel since rats injected with liquid silicone (Agnew et al., 1966) or implanted with elastomer (King et al., 1989) did not develop tumors .

Tumors were also not found in rabbits implanted with elastomer-covered gel for up to18 months (Lilla and Vistnes, 1976) or in mice implanted with silicone fluid, gel or elastomer for 180 days (Bradley et al., 1994). Long-term implantation of various synthetic (presumably silicone-based) materials in dogs for as long as six years did not result in tumor development (Mastalski et al., 1977).

VI. Silicone and "Adjuvant Disease"

In the late 1980's, a number of articles began to appear suggesting a possible link between SBIs and autoimmune disorders in women. The concern seems to have been initiated by reports of delayed adverse reactions in some Japanese women that had been injected with silicone fluid admixed with other substances such as paraffin in the breast tissue many years previously (see citations in Picha and Goldstein, 1996). The connective tissue disease that was observed in these women was termed "human adjuvant disease" based on the theory that silicone could act like the experimental adjuvant known as "Complete Freund's Adjuvant" (CFA). CFA is an emulsified preparation of heat-killed mycobacteria in mineral oil. When an antigen is incorporated into CFA, the immune response to that antigen is increased and prolonged, a desirable situation for vaccine delivery. However, CFA itself is not used clinically because of severe local inflammatory reactions as well as possible sensitization to the mycobacterium. The mineral oil component of CFA functions to provide a depot effect for the water-in-oil emulsified antigen; the mycobacterial component induces an inflammatory response that facilitates the immune response to the antigen (Broderson, 1989).

"Adjuvant arthritis" is an experimental inflammatory joint disease in rats that is induced by a single injection of CFA (Glenn and Gray, 1965; Pearson, 1956). The clinical manifestations of the disease resemble some inflammatory rheumatic diseases in humans such as rheumatoid arthritis, ankylosing spondylitis, and Reiter's disease (Muir and Dumonde, 1982). However, "adjuvant arthritis" appears to be a disease unique to rats. Among experimental animals tested, including mice, guinea pigs, rabbits, sheep and monkeys, CFA induces the disease only in the rat, and only in certain strains of rat, indicating that a specific genetic predisposition is required. In some rats (e.g., Dark Agouti [DA]), arthritis can even be induced by the injection of mineral oil alone in the absence of mycobacteria (i.e., Incomplete Freund's Adjuvant [IFA]).

The mineral oil component of CFA functions nonspecifically, and many different types of oils were shown to be effective in inducing "adjuvant disease" in rats when emulsified with killed mycobacteria (Whitehouse et. al., 1974). In these studies, a commercial silicone oil was also shown to be a "potent arthritogen."

However, the silicone oil was described as "a lubricating oil of unknown composition" sold as a lock lubricant, which caused severe weight loss in the rats. Thus, this oil is not representative of the medical grade of silicone found in SBIs.

As summarized in Table 1, more recent studies have shown that neither silicone gel nor silicone oil (PDMS) was capable of eliciting arthritis in Lewis rats when injected alone or emulsified with mycobacteria (Chang, 1993; Picha and Goldstein, 1997). Similarly, in the DA rat, a mixture of silicone gel and oil was not effective in inducing arthritis (Naim et al.,1995) unless injected directly into the joint (Yoshino, 1994). Thus, there are no experimental data to support the labeling of silicone-associated disorders as "human adjuvant disease."

VII. Adjuvant Activity of Silicone

The term adjuvant is a label more widely applied in recent years to describe "any substance that enhances the immune response to an antigen with which it is mixed" (Janeway and Travers, 1994). Effective vaccines for human diseases often depend on the incorporation of an adjuvant in order to generate and enhance the development of protective immunity. Under this broad definition, many diverse substances, acting by diverse mechanisms, have been shown to function as adjuvants. Oily substances that prolong antigen half-life in tissues and may enhance cellular uptake are especially effective adjuvants.

As summarized in Table 2, several studies have examined the ability of silicones to function as adjuvants to increase antibody production or cell-mediated immune responses when injected as an emulsified preparation with the antigen. In general, it appears that adjuvant activity is seen more often with silicone gel than with silicone oil. The low molecular weight cyclosiloxane D4 has also been shown to possess adjuvant activity in terms of enhancing antibody production to some antigens.

Adjuvants have also been used experimentally to induce autoimmune disease in

animals following immunization with autoantigens or cross-reacting foreign antigens. For example, an arthritic disease can be induced in animals following immunization with either homologous or heterologous type II collagen (Ellis et al., 1992). In this collagen-induced arthritis model, collagen protein is emulsified in CFA or IFA and injected into rats or mice. After one or more immunizations, the onset of the disease is identified by severe swelling and erythema in the paws, which is associated with a massive inflammatory infiltrate into the synovium (Ellis et al., 1992). The histological changes in the joints of these animals resemble those observed in rheumatoid arthritis patients.

The ability of silicone to substitute for mineral oil in the induction of collagen-induced arthritis has been examined in both rats and mice (see Table 2). Following the injection of bovine collagen emulsified in a silicone oil:gel mixture, Naim et al. (1995) reported that arthritis was induced in 4/10 rats compared to 8/9 rats injected with collagen in IFA. When silicone oil was tested independently from silicone gel, the incidence of arthritis was higher with the gel(7/10) than the oil (3/10). In contrast, D4 was not an effective adjuvant for induction of arthritis (Naim et al., 1995). Using the DBA/1 mouse model, Schaefer (1997) reported that silicone oil was not an effective adjuvant for the induction of collagen-induced arthritis, even if the inoculum included killed mycobacteria, whereas 80% of the mice injected with collagen in CFA developed arthritis.

It is important to recognize that the successful induction of arthritis in rats with collagen emulsified in silicone does not reflect silicone-induced "adjuvant arthritis", which develops in the absence of active immunization. Rather, these results point to the successful immunization of the rat to the foreign collagen protein when silicone gel or oil was used as the adjuvant. On the other hand, in an animal model of experimental autoimmune thyroiditis, the injection of rat thyroglobulin (Tg) emulsfied in a silicone oil:gel mixture was unable to induce thyroiditis, while 100% of rats injected with Tg in CFA developed thyroid disease (Naim et al., 1993).

VIII. Effects of Silicone in Animal Models of Autoimmune Disease

The question of whether or not silicone is capable of causing or aggravating autoimmune disease can be addressed most directly by laboratory animal studies using different experimental models of autoimmune disease. These established models of autoimmune disease have been developed to study the biological processes responsible for the symptoms associated with the disease, the predisposing genetic and environmental factors that influence the disease process, and the effectiveness of potential therapies. While no single animal model perfectly matches human disease, there are usually many parallels, and data obtained from different animal models can provide insight into the disease process in humans.

Animal models in which autoimmune disease develops spontaneously are most relevant for the evaluation of the ability of silicone to exacerbate (promote) autoimmune disease. Promotion could be associated with the early appearance, increased severity, and/or increased incidence of autoimmune disease in animals that were destined to develop autoimmune disease due to genetic predisposition. Animal models in which autoimmune disease is induced by specific antigen injection are also useful to evaluate promotion when the severity of the induced disease in controls can be minimized. In contrast, it is much more difficult to evaluate whether or not silicone causes an autoimmune disease because of the multifactorial nature of disease induction. Causation could perhaps be deduced if a novel disease developed in an autoimmune- prone strain or if autoimmune disease was induced in a non-susceptible strain. As summarized in Table 3, the effects of silicone have been assessed in several experimental paradigms of autoimmunity.

Arthritis-prone DBA/1 Mice

Using arthritis-prone DBA/1 mice, Schaefer et al (1997) examined the ability of silicones to induce arthritis. The results of these studies showed that mice implanted with silicone oil, gel or elastomer for as long at 12 months did not develop arthritis. However, it should be noted that DBA/1 mice injected with CFA also failed to develop arthritis in this study; thus the positive control group failed to document the sensitivity of the model.

In a different animal model, genetically susceptible BALB/cAnPt mice injected in the peritoneal cavity with various silicone gels did not develop the arthritis that is frequently found in this strain when they are treated with pristane oil (Potter et al., 1994). Similarly, single or multiple sc injections of silicone gel failed to induce arthritis in BALB/cAnPt mice even if the implant site was co-injected with Staphylococcus bacteria (MacDonald et al., 1998).

Taken together with the previously described ineffectiveness of silicone in rat adjuvant arthritis models, these findings indicate that silicones do not directly induce arthritis in arthritis-prone mice or rats.

MRL lpr/lpr Model of Lupus

MRL/lpr mice carry a spontaneous lymphoproliferative mutation (lpr/lpr) that results in the development of an autoimmune syndrome at approximately eight weeks of age. The disease progresses over 16–24 weeks and is characterized by high levels of circulating autoantibodies leading to an immune-complex mediated glomerulonephritis, diffuse vasculitis and arthritis (Hang et al., 1982). Approximately 50% of the mice die by 24 weeks of age due to renal failure. The clinical symptoms in MRL lpr/lpr mice closely resemble systemic lupus erythematosus (SLE) in humans. The arthritis that develops in MRL lpr/lpr mice is similar to RA in humans. The mice also develop a Sjögren's- like inflammation of the conjunctiva. MRL+/+ mice, which lack the lpr gene mutation, develop a milder autoimmune disease later in life as compared to MRL lpr/lpr mice. Schaefer (1997) investigated the ability of silicones to modify disease in the MRL strain. At five weeks of age, prior to the onset of autoimmune symptoms, MRL lpr/lpr and MRL+/+ mice received sc implants of silicone gel, silicone oil or a sham implant. During the next 12 weeks, clinical parameters of disease were measured by palpation of lymph nodes, urinary protein, and serum titers of collagen and DNA antibodies. Serum levels of several cytokines were also monitored. At sacrifice, kidneys were fixed and stained for immune complex deposition.

All MRL lpr/lpr mice had severe glomerulonephritis by the time they were sacrificed, and silicone exposure did not influence the severity of the disease. MRL+/+ mice showed minimal renal changes, and this too was unaffected by the silicone implants. Lymph node enlargement was also not influenced by silicone. Anti-DNA antibody titers were significantly higher in MRL lpr/lpr mice that received silicone gel and in MRL+/+ mice that received gel or oil implants as compared to sham controls. Some differences in the levels of certain cytokines were noted at various times during the experimental time period, but no pattern of change was revealed that would suggest that silicone altered disease by modifying cytokine production.

In these and other experiments, Schaefer (1997) presents data that purport to demonstrate the presence of autoantibodies to silicone-bound proteins. However, the unorthodox procedures that were used to quantify the proteins, the lack of positive controls, and the manner in which the data were presented, do not allow such conclusions to be made. The data are not convincing of anything more than nonspecific binding of protein to the implant.

In similar studies using a different strain of mouse with the lpr mutation, Osborn et al. (1995) reported that silicone oil containing 5% D4 did not alter the incidence of mortality at 48 weeks of age when compared to saline-injected mice. The frequency and latency of other disease symptoms also did not differ between the groups.

New Zealand Black (NZB) x New Zealand White (NZW) F1 Murine Model of SLE NZB/W mice spontaneously develop severe systemic autoimmune disease characterized by elevated titers of anti-nuclear antibodies (ANA), increased levels of serum IgG, polyclonal activation of B cells and the subsequent development of a fatal immune-complex mediated glomerulonephritis (Rose and Bhatia, 1995). The disease symptoms resemble human SLE.

White et al. (1998) evaluated the ability of silicone gel implanted in the mammary region of female NZB/W mice to alter the course of the disease over a 78-day period. The effects of silicone were compared to two known inducers of autoimmune disease, mercuric chloride and D-penicillamine. These positive control groups are helpful in demonstrating the sensitivity of the model to exogenous autoimmune-inducing substances. The results of these studies showed that silicone gel-implanted mice did not differ from their sham controls in terms of total IgG levels or antibody titers to dsDNA, laminin, DNP-HSA or SRBC. Spleen weight was also not affected by silicone exposure. In contrast, all of these parameters were significantly elevated in the positive control groups when compared to their own controls. Although actual disease was not measured in this study, silicone gel exposure did not appear to be promoting the clinical signs that have been associated with development of the disease in this model.

Tight Skin Mouse Model of Scleroderma

Mice bearing the TSK mutation (TSK/+) spontaneously develop skin fibrosis and characteristic autoantibodies which resemble human scleroderma. Frondoza et al. (1995) evaluated the influence of silicone on the pathogenesis of the disease in this mouse model. TSK/+ mice as well as their phenotypically normal TSK/- litter mates were injected with low molecular weight silicone fluid, high molecular weight silicone gel, IFA as a positive control, or saline as a negative control. One month later, skin was examined histologically for development of hyperplasia and thickening. Other tissues (kidney, liver, spleen) were examined for pathological changes. Circulating autoantibodies to RNA polymerase I , topoisomerase and bovine serum albumin (BSA) were also measured.

The results indicated that the normal progression of the disease seen in saline-treated control TSK/+ mice as revealed by histological examination was not altered by silicone exposure. It was also not altered by IFA. No evidence of hyperplasia or pathology was seen in the TSK/- mice with any of the treatments. None of the mice showed pathological changes in the other organs. Circulating antibodies to RNA polymerase I, topoisomerase or BSA were not altered in TSK/+ mice treated with silicone or IFA. as compared to saline-treated controls. The lack of effect in the IFA-treated positive control group limits the ability to interpret the lack of effects with silicone.

Type II Collagen-induced Arthritis

Because collagen provides the basic framework of cartilage, the experimental induction of an immune response to collagen can produce the symptoms of arthritis. As previously described in Section VII, animal models of collagen-induced arthritis have been characterized in which the intradermal injection of bovine type II collagen emulsified with CFA or IFA induces an arthritis in genetically susceptible rats or mice (Holmdahl et al., 1989). The lesions found in the affected joints are quite similar to those found in humans with rheumatoid arthritis. Anti-collagen antibodies develop in immunized rats and mice and are also found in RA patients, but the specific role they play in the pathogenesis of the disease is controversial. Circulating levels of inflammatory cytokines such as IL-1 , TNF- , and IL-6 are elevated during the disease process, and experimental manipulation of cytokine activity (production or receptor blockade) modifies the disease. Thus, proinflammatory substances might be expected to promote arthritic disease.

Schaefer et al. (1997) used the mouse model of collagen-induced arthritis to examine the ability of various forms of silicone implants to promote the disease process. Mice were injected with silicone oil, gel or elastomer for three days or nine months prior to immunization with collagen in CFA. The results showed that silicone in all forms had no influence on the incidence or severity of arthritis as compared to sham- treated mice. However, because the incidence and severity of disease was high in the controls, significant promotional effects would have been difficult to demonstrate. Time-to-onset of disease was not reported.

In a second study, Schaefer (1997) used collagen immunization with IFA instead of CFA to induce a lower incidence of disease in the controls. In this study, 9/10 mice implanted with silicone elastomer nine months prior to immunization exhibited disease compared to 3/10 control mice. The severity of the disease was also increased in the silicone elastomer-treated mice. In mice treated with silicone gel or oil, 6/9 mice in each group developed disease and their arthritic scores also tended to be higher than the controls, although these changes were not statistically different from controls. Taken together, the results suggest that exposure to different forms of silicone may promote the development of arthritis in this model of autoimmune disease. However, these results must be considered preliminary and interpreted cautiously in light of the small number of animals tested and the fact that only 3 control mice developed disease. Furthermore, the findings are less than compelling based on the fact that anti-collagen antibody titers were not altered by silicone, and cytokine levels were not consistently altered in a manner that might explain the increased incidence of disease.

NZB Mouse Model of Autoimmune Hemolytic Anemia

NZB mice develop a form of autoimmune hemolytic anemia that closely resembles the human disease. The disease begins at about three months of age and by nine months almost all mice show reduced hematocrits as evidence of the disease process (Howie and Helyer, 1968). MacDonald et al.(1998) used the NZB model to study the influence of silicone gel implants on this autoimmune disease process. Groups of mice were injected with saline as a negative control, pristane as a positive control, or silicone gel. Injections given one time were compared to injections given three times to examine the effect of multiple exposures. Some mice were given an injection of silicone followed three months later by a capsulotomy to evaluate the effect of "traumatic rupture." Other mice were given an injection of silicone followed three months later by a low dose of Staphylococcus epidermidis, intended to mimic infection with "a common contaminant found on the surface of breast implants and hypothesized to be involved in capsular contracture." Appropriate controls were included for all of the procedures except the capsulotomy. Mice were examined daily for ten months after which time all surviving mice were sacrificed. Blood was used to measure hematocrits and serum was analyzed for autoantibody production. Urinary protein levels were monitored bi weekly during the course of the study for the possible development of glomerulonephritis.

By ten months of age, some mortality had occurred in all groups, including the controls (20-30%). Only multiple treatments with pristane, the positive control, significantly increased the mortality of the mice to 80%. Increased mortality (60%, P < 0.085) was also noted after multiple treatments of silicone. Hematocrits were much lower in all NZB mice compared to normal BALB/c mice. Hematocrits were further reduced in mice given pristane three times, and in all mice injected with silicone, although it was not clear what pairwise comparisons were made to determine statistical significance. In contrast, hemagglutination titers were similar in all groups compared to controls. ANA titers were elevated in silicone-implanted mice that had undergone a capsulotomy at three months. Anti-collagen IgM titers were elevated in mice that were injected with silicone and infected with S. epidermidis whereas anti-collagen IgG titers were similar in all groups.

Multiple injections of pristane or silicone increased urinary protein levels.

Based primarily on the mortality and hematocrits, the results of this study provide limited evidence for a promotion of autoimmune hemolytic anemia by silicone in NZB mice. However, the results would have to be repeated before such a conclusion could be drawn. The data would also be more compelling if additional endpoints relevant to the disease process had been measured, as it is not clear what caused the death of the animals. In addition, the clinical data might have been more insightful if it had been collected prior to the onset of mortality. On the other hand, the relevance of this disease model to women with SBIs is unclear.

Normal BALB/c mice were also tested in parallel with the NZB mice and given identical pristane and silicone treatments to determine if a non-genetically disposed mouse could be induced to develop disease by exposure to silicone. However there was no significant mortality in any treatment group, and all had normal hematocrits. These data indicate silicone was not able to induce autoimmune disease in genetically resistant mice.

IX. Immunotoxicity of Silicone in Animals

Although the majority of results from the animals models of autoimmune disease do not support an enhancement of the disease process by silicone, which was tested in several different forms and for various periods of time, arguments can be put forth as to why the animal models are different from the human situation and therefore not reflective of the human response to silicone.

Thus, it is appropriate to review the animal studies that have examined the ability of silicone to alter processes that are believed to contribute to the development of autoimmunity. Based on a general conceptual understanding of autoimmune disease pathogenesis, several hypothetical mechanisms have been proposed by which silicone could induce or exacerbate the process. These include:

1. Silicone causes immune system "dysregulation" resulting in abnormal T cell and/or B cell activity leading to the generation of the autoimmune response. For example, polyclonal B cell activation or loss of suppressor T cell functions have been associated with some autoimmune diseases.

2. Silicone induces specific T cell activation by modification of self-proteins resulting in a novel autoimmune disease.

3. Silicone causes inflammation and the resulting inflammatory cytokine production initiates or exacerbates autoimmune disease development. The evidence available to support or refute these hypotheses will be summarized below.

Evidence that Silicone Alters Immune Responsiveness of Animals Several studies have been conducted in laboratory animals in an effort to determine the influence of silicone exposure on immune function. Comprehensive immunotoxicology studies were carried out in the early1990s by the Medical College of Virginia under contract to the National Toxicology Program. A standard immunotoxicology screen was utilized in mouse studies to examine the immunomodulatory effects of various doses of silicone oil, silicone gel, or silicone elastomer disks implanted subcutaneously in the breast area of female B6C3F1 mice.

Polyurethane disks were also tested. Controls were injected with saline. Immunological testing was carried out ten days or 180 days after initiation of silicone exposure (Bradley et al., 1994a,b). The ten-day period was selected to represent the peak inflammatory response. The 180-day exposure was intended to reflect the chronic condition of SBIs wherein a well-developed fibrous capsule had formed. Endpoints assessed included body weight, histopathology, hematology, serum complement levels, bone marrow colony formation, spleen cell subpopulations (B cells, T cells and T cell subsets); primary antibody response to SRBC (IgM and IgG PFCs); proliferative responses to B and T cell mitogens and to allogeneic lymphocytes, cytotoxic T lymphocyte response, NK cell cytotoxicity, reticuloendothelial system clearance of antigenic particles, peritoneal macrophage phagocytosis and IFN- production. The ability of the animals to resist infection by Streptococcus pneumonia or Listeria monocytogenes and to control B16 melanoma metastases were assessed as holistic measurements of overall immune status.

The results of these immunotoxicology studies showed that silicone in any form tested did not induce systemic toxic effects or alter immune function. The only noteworthy silicone-related treatment effect was a decrease in NK cell activity in the spleen of mice injected with silicone gel or implanted with the elastomer disk for 180 days. However, this effect on NK activity did not translate into an increase in B16 tumor metastasis, which is the host resistance model considered sensitive to changes in NK cell activity.

Follow-up studies were carried out to validate the effect of silicone on NK activity in a dose-response study. The results of the study confirmed the suppression of NK cell activity but only at the highest dose level. Comparative studies using F344 rats also showed a suppressive effect on NK cell activity from silicone treatment. In rats, NK activity could be boosted in silicone-implanted rats by polyI:C but not to levels observed in controls (Wilson and Munson, 1996).

Taken together, these results indicate a modest and somewhat consistent effect of silicone exposure on NK cell activity. The importance of the finding may derive from the fact that any systemic alteration in an immunological endpoint could be induced by the presence of a silicone implant. However, the importance of the finding in terms of autoimmune disease is not known since a role for NK cells in autoimmune disease development is not widely recognized.

The mechanism by which silicone alters NK cell activity was not elucidated.

Although the NTP-sponsored immunotoxicity assessment of silicone

was thorough and of high quality, there were some shortcomings in the experimental design. For example, these studies utilized a standard screening procedure to identify immunotoxic substances. The assays were previously validated to detect immunosuppressive substances and were not specifically oriented to address autoimmune-relevant endpoints. Another potential shortcoming is that the animals used in the studies were not predisposed to develop autoimmune disease and may therefore be more resistant to the effects of silicone than an autoimmune-prone strain.

Evidence for Antigenicity of Silicone

The question of silicone antigenicity has been addressed in a limited number of animal studies that have attempted to demonstrate silicone-specific antibodies or silicone-specific T cell responses, including the possible development of specific immunologic memory. The data from these studies will be reviewed here.

Delayed-type hypersensitivity (DTH) responses are useful measures of T-cell mediated immunity in animals. A true DTH response follows delayed kinetics thatreflect the response of pre-sensitized antigen-specific memory T cells. TheseT cells proliferate in response to their specific antigen and secrete cytokinesthat activate macrophages, causing tissue swelling that peaks 48–72 hours afterantigen injection. The kinetics of the DTH response is crucial fordifferentiating T cell immunity from nonspecific inflammatory responses that occur within a few hours of injection and are not antigen-specific. The DTH response is also distinguished by its kinetics from a rapid contact hypersensitivity response that is mediated by antigen-specific IgE antibodies.

Brantley et al.(1990) used a creative approach to ask if there is a cell-mediated immune response to silicone. They "immunized" rats to silicone by injecting silicone in CFA. Four weeks later, the animals were given silicone implants. Reactions to the implants were measured by capsule formation and histology of the capsule. Based on the similar histology of the capsules of immunized (CFA + silicone) and nonimmunized (CFA only) rats, there was no evidence of an immune response to silicone. One would have expected a characteristic tissue reaction if the rats had been immune to the silicone.

However, these negative results must also be interpreted cautiously given that a positive control was not included to demonstrate the sensitivity of the technique.

In related studies, Brantley et al. (1990) immunized rats with silicone gel sonicated in CFA. Four weeks later, lymphocytes from the spleen were obtainedand tested for their ability to respond to silicone in vitro. There was no difference in the proliferative response from silicone-immunized mice compared to mice injected only with CFA. Thus, there was no measurable memory response to silicone, the most unambiguous measure of antigen-specific immunity.

Smith et al. (1990) immunized rats with fine particles of solid silicone from a bone prostheses emulsified with CFA. After six sensitizing injections, they reported evidence for an immune response to the silicone in a positive skin reaction to challenge and IgG deposition around the silicone implant. However, the study lacked appropriate controls.

LeBeau (1967) reported that silicone gel strips did not induce a hypersensitivity response in the skin of guinea pigs. Naim et al (1993) found no evidence for a DTH response to silicone in rabbits.

Silicone-specific antibody production by B lymphocytes has also been examined to determine if there is a specific immune response to silicone. In order to demonstrate antigen specificity, one must be able to show specific binding of the induced antibody to the antigen in vitro. In regard to silicone antibodies, these in vitro assays have been fraught with problems associated with nonspecific protein binding to the silicone (Butler et al., 1996; Rosenau et al., 1996). The studies are also complicated by the hydrophobic nature of silicone materials and the difficulties of working with them in antibody assays

. Unfortunately, because of the unconventional methods that have been used in past studies in an attempt to circumvent these problems, along with a failure to provide adequate assay validation, the data currently available are not convincing of silicone-specific antibody production.

Evidence that Silicone Induces Inflammation

Several studies have reported that subcutaneous or intramuscular injection of various types of silicone in experimental animals induces an inflammatory response similar to a foreign body reaction leading to the formation of a fibrous capsule around the implant (Lilla and Vistnes, 1975; Goodman et al., 1988; Lemen et al., 1992). The extent of the inflammatory response in any particular study depends on the type of silicone material that is implanted, the size and shape of the implant, its location, as well as other unidentified factors (Picha and Goldstein, 1991). In addition, differences in surgical techniques cannot be minimized since surgical trauma alone can account for a part of the early inflammation noted, and inadvertent bacterial contamination would likely increase the severity of the inflammatory response. However, in most studies, the inflammatory response declines over time, and the implants are usually found surrounded by a mature connective tissue capsule of varying thickness containing minimal numbers of macrophages, neutrophils and lymphocytes (Grasso et al., 1965; Malczewski, 1984; Mudgett et al., 1990; Picha and Goldstein, 1991; Rasmussen, 1988). Recently, Fabre et al. (1998) used a novel technique to demonstrate this process. They measured the cellular response to a cylindrical elastomer implant using flow cytometry. Two days after implantation, a mixture of monocytes and neutrophils predominated in the exudate that formed inside the tube. At day nine, cell subpopulations could still be identified, whereas by day 23, the cellular components had declined to undetectable levels. Fibrinogen levels rose progressively during this time.

These results indicate a resolution of the active inflammatory response to the silicone elastomer within the 23-day time frame.

Injection of silicone fluid directly into the joint of DA rats induced arthritis, suggesting that a local inflammatory reaction was induced by silicone (Yoshino, 1994). This response is not particularly surprising since the DA rat is highly susceptible to arthritis induction. However, there is no evidence that silicone from SBIs is transported to joints at any significant concentration to directly induce arthritis. This is supported by the animal studies discussed previously that show silicone injections outside of the joint do not induce arthritis, even in the DA rat (see Table 1).

While it is generally accepted that silicones elicit varying degrees of local inflammation at the site of the implant, there is little evidence from controlled animal studies that suggest silicone causes systemic inflammatory responses. The most extensive examination of this possibility was carried out by Schaefer (1997), who measured levels of circulating cytokines at various times after silicone implantation in relationship to the development of autoimmune disease. The results of these studies, documented in Table 3, failed to provide evidence that silicones induced a systemic alteration in inflammatory cytokine production.

Evidence that Silicone Activates Macrophages

As previously discussed, studies carried out in the late 1960s had shown that silicone fluid injected ip or sc appeared to be phagocytosed and distributed systemically via the lymphatics (Ben Hur et al., 1967; Rees et al., 1966; Sparchu and Clashman, 1970). In more recent studies, macrophages containing silicone were also found in rats injected with silicone fluid (Hill et al.,1996; Malczewski et al., 1994), but not in rats implanted with silicone gel or elastomer (Malczewski et al., 1994). Likewise, several other studies on silicone gel have found no evidence for the phagocytosis or systemic distribution of silicone gel in rats that had been implanted for as long as two years (Goodman et al.,1988; Raposo do Amaral et al.,1993; Tiziani et al., 1995). Taken together, these results suggest that phagocytosis of silicone by macrophages and systemic distribution of silicone is a phenomenon primarily associated with the injection of free silicone fluid rather than gel.

However, because macrophages have been shown to be capable of engulfing silicone in any form, many of the hypotheses related to silicone-induced disease invoke a role for silicone-induced activation of macrophages. These activated macrophages are then hypothesized to secrete cytokines that lead to the disease symptomology. Unfortunately, there are no definitive data available that have characterized the influence of silicone on such macrophage activation or cytokine production.

Das et al. (1990) examined the ability of silicone sonicated with CFA to cause long-term activation of macrophages as measured by their secretion of IL-1. At eight months after silicone injection, there was no difference in IL-1 production compared to mice injected with CFA or saline. However, the negative results must be tempered by the fact that no relevant positive control was included. The addition of LPS in vitro to activate macrophages was not an appropriate positive control

MacDonald et al. (1996) reported that silicone gel injected into the peritoneal cavity of certain strains of mice induced a population of predominately macrophages that was able to induce a small degree o proliferation in CD4+ T cells in a nonantigen-specific manner. The author suggested that these "silicone-laden macrophages" induce a proliferative response that is "unique to silicone" because macrophages from pristane- or thioglycollate-injected mice did not induce proliferation. However, the authors failed to speculate on what unique factor silicone-laden cells produce that other inflammatory macrophages do not, nor did they demonstrate by any other criteria that the macrophages were indeed activated. Furthermore, they did not demonstrate that the activity was mediated by the macrophage component of the peritoneal exudate cells, nor that the macrophages indeed contained silicone. Thus, the data do not support the conclusions of the authors and provide no insight into the effect of silicone on macrophage activity.

The studies of Bradley et al. (1994a,b) addressed the potential for silicone to alter systemic macrophage activity in mice that received short-term(ten day) or long-term (180 day) implants of silicone oil, gel, or elastomer. Reticuloendothelial clearance of particles by tissue macrophages present in the liver, spleen, lymph nodes and lungs was evaluated by measuring the vascular learance and tissue uptake of radioactively labeled foreign particles (SRBC or Covaspheres). The functional activity of adherent peritoneal cells (primarily macrophages) was evaluated by their ability to phagocytose radiolabeled particles in vitro, and by their ability to be activated by IFN and LPS to kill tumor cells in vitro.

The results of these studies revealed no change in any of these parameters ten days after the implant surgery except for a decrease in macrophage tumoricidal function under various in vitro conditions. However, this decrease in tumoricidal function did not translate into a change in the resistance of the mice to tumor growth. On the other hand, in one experiment, all of the mice in the silicone groups were more resistant to infection by Listeria bacteria, which could possibly reflect enhanced phagocytic activity.

In mice that had been implanted with silicone materials 180 days previously, there was increased uptake of SRBC by the liver of mice exposed to silicone gel, but this effect was not confirmed in a subsequent dose-response study. Peritoneal cells from silicone fluid- implanted mice had significantly increased phagocytic activity for Covaspheres, but this too was not confirmed in a dose-response study. Finally, the growth of iv-injected tumor cells inn the lung was decreased in all silicone-treated mice compared to vehicle controls, which could reflect increased macrophage tumoricidal activity. However, since macrophage tumoricidal activity was not evaluated, additional studies would be required to document this effect.

Recently, Rhie et al. (1998) published a study in which macrophages were cultured on silicone gel that was centrifuged onto the bottom of tissue culture plates, allowing for direct contact between the cells and the gel. Subsequent analysis of the function of these macrophages demonstrated a significantly enhanced responsiveness compared to macrophages cultured directly on the plastic plate. The authors conclude that silicone gel activated the macrophages to augment immune function. However, the authors do not consider an equally plausible explanation for the data; that is, that by preventing the adherence of the macrophages to the plastic plate with the gel coating, the suppressive effect of adherence-induced activation was prevented. This possibility arises from the well-known and widely used technique of coating tissue culture plates and tubes with silicone to prevent macrophage adherence, and the equally well-known fact that an excess of activated macrophages usually suppresses immune function in vitro. Unfortunately, Rhie et al. failed to provide an important control group that would have characterized a normal immune response without any added macrophages. They also failed to provide any direct evidence for the state of activation of the macrophages cultured on gel vs plastic (e.g., adhesion molecule expression or cytokine production). Thus, these studies do not provide convincing evidence that silicone gel induces the activation of macrophages.

X. Potential Contribution of Other Materials in SBIs to Toxicity Low Molecular Weight Cyclosiloxanes D4 and D5 are low molecular weight, cyclic silicones that have been detected in SBIs. D4 has been analyzed at levels of approximately 500 ppm in the gel and in the elastomer at a level of 100-300 ppm (Van Dyke et al., 1993). D5 levels are similar. There has been some speculation that these molecules play a part in the health effects of SBIs.

The toxicities of D4 and D5 have been tested independently of silicone because they were formulated and are marketed for use in a variety of products. Inhalation of relatively large doses of D4 and D5 have been shown to produce few toxic effects other than liver enlargement and induction of hepatic drug metabolizing enzymes (McKim et al., 1988; Mehendale, 1989; Siddiqui, 1989). Such hepatic effects are not seen following the implantation of silicone gel (Bradley et al., 1994b; Selwyn and Danner, 1988). High doses of D4 have also been reported to enhance NK cell activity (Wilson and Munson, 1997), where as exposure to silicone gel suppresses NK function (Bradley et al., 1994a,b). Thus, based on current studies, there is no data to implicate these low molecular weight silicones in any health effect associated with SBIs.

Silanols

The toxicity of silanols is considered not relevant to the SBI issue. Silanols are highly toxic chemicals with a profile of toxicity in rats that includes liver, kidney, and neural damage, and bleeding (Dow Report No. 2964). However, none of these toxicities are seen in rats injected with large amounts of silicone fluid, gel or elastomer.

Platinum

Platinum (Pt) is used as a catalyst in the preparation of silicone gels and elastomers. According to Lykissa et al. (1997), Pt was detectable in silicone gel at a level of approximately 700 µg/kg (parts per billion) using ICP-MS. Furthermore, they indicate that when the gel was incubated in lipid-rich media, Pt diffused into the media at a rate of approximately 20–25 µg/day/250g implant. Since the whole implant used in these studies would only contain 175 µg of Pt, it suggests that all of the Pt would diffuse from the gel into the media within seven days. This is not logical. There are no data available that address the level of Pt in blood or tissues of animals or humans who have SBIs.

However, if a worst-case exposure scenario was calculated based on the value published by Lykissa, wherein all of the Pt in two 300-ml implants was released into the body of a 50-kg woman, the Pt dose would be 8.4 µg/kg or 8.4 ppb. This concentration of Pt is approximately equivalent to the five ppb level found naturally in the environment. Pt toxicity is dependent on its chemical form. Pt salts primarily cause liver and kidney toxicity, which have not been associated with SBI materials. Thus, there is no evidence that Pt plays any role in the health effects associated with SBIs.

XI. Conclusions

This chapter has reviewed the experimental animal data to evaluate the evidence that silicone breast implants have the potential to cause systemic disease in humans. The results of this review indicate that the silicones used in SBIs are of very low toxicity to animals. Although there is documented evidence of local inflammatory reactions to silicone breast implant materials in animals, there is no convincing evidence for a significant systemic inflammatory response. The local reaction to silicone is similar to other "foreign body reactions" described with other implanted materials.

There is some evidence that macrophages can phagocytose small droplets of silicone which may then be transported via the lymphatics to other tissues in the body. This process appears to occur primarily with low molecular weigh silicone fluid rather than the high molecular weight gel. However, even with phagocytosis, there is currently no definitive evidence for systemic effects on the immune system or for processing of silicone as an antigen for T cell activation. There is also no convincing evidence from animal studies that T cells can be activated by silicone.

The ability of silicone to act as an adjuvant has received a lot of attention. Even though some silicone gels and fluids have been shown to possess adjuvant activity when antigen is emulsified with the silicone prior to immunization, this capability has little bearing on the issue of silicone- induced autoimmune disease. It most likely reflects a depot effect of the non-degradable silicone. There are no convincing data that show silicone acts like an adjuvant when it is present at a site distant from the antigen injection, and there is no biologically plausible mechanism for antigen emulsification to take place in the body.

Immunotoxicity testing of silicones revealed only one fairly reproducible effect which was suppression of NK activity in both rats and mice. The degree of suppression was variable between experiments and not of sufficient magnitude to affect a disease model responsive to NK cell activity. Although these results are of interest, the specific role that NK cells may play in autoimmune disease development is not well understood.

The greatest weight of evidence in this review has been given to studies that evaluated the ability of silicone to induce or promote autoimmune disease in whole animal models. Such animal models provide the most holistic approach to identifying biologically relevant effects induced by silicone exposure that might lead to autoimmune disease induction or promotion in humans. The use of animals that are genetically predisposed to develop autoimmune disease provide the advantage of a high and predictable incidence of spontaneous disease. If an alteration in disease induction occurs with silicone treatment and it correlates with changes in relevant clinical endpoints, the evidence for a cause-effect relationship becomes more credible. If clinical correlates of a promotional effect by silicone can be identified in animals, it would provide a focus for human clinical investigations.

Several adequately designed animal studies have been published that address the question of silicone's ability to induce or exacerbate autoimmune disease. The human autoimmune diseases that are simulated in these animal models include rheumatoid arthritis, systemic lupus erythematosus, scleroderma, and autoimmune hemolytic anemia. In the 17 experimental regimens outlined in Tables 1 and 3, 15 indicate that silicone did not induce or promote the development of autoimmune disease and/or alter diagnostic clinical endpoints. The other two experiments must be viewed as providing weak but suggestive preliminary evidence of a promotional effect by silicone exposure. However, these preliminary findings must be confirmed in independent studies. Curiously, one of models that shows some evidence of promotion by silicone is the model of autoimmune hemolytic anemia, which is not of obvious relevance to the SBI issue. The other is a bovine collagen-induced model of arthritis using incomplete Freund's adjuvant to lower the degree of disease in the controls. Limitations of these studies relate to the small number of animals in the treatment groups and the lack of clinical endpoints that verify the exacerbation of disease.

On the other hand, there are also limitations with some of the studies that show no effect of silicone on autoimmune disease. The biggest problem with several of the studies is that disease incidence is so high in the controls, it would be difficult to demonstrate an increase in disease in the silicone-treated mice. Although a promotional effect in such cases might be evidenced by an early appearance of the disease, latency was not an endpoint that was documented in most of the studies. The second limitation of several of these studies is the lack of positive controls that demonstrate the sensitivity of the model to exogenous modulation. Without a positive control, it is difficult to put the failure of silicone to alter disease in a relevant context. Nevertheless, the data from these studies cannot be ignored for their null effects on the disease process.

In conclusion, the preponderance of evidence from animal studies indicates little probability that silicone exposure induces or exacerbates systemic disease in humans.

References

Agnew WF, Todd EM, Richmond H, et al. Biological evaluation of silicone rubber for surgical prosthesis. J Surg Res 2:357–63,1962.

Andrews JM. Cellular behavior to injected silicone fluid: a preliminary report. Plast Reconstr Surg 38:581–83, 1966.

Ballantyne DL, Jr., Rees TD, Seidman I. Silicone fluid: response to massive subcutaneous injections of dimethylpolysiloxane fluid in animals. Plast Reconstr Surg 36:330–38, 1965.

Barondes R. Silicones in medicine: New organic derivatives and some of their unique properties. Military Surgeon 106:379–87, 1950.

Ben-Hur N. Local and systemic effects of dimethylpolysiloxane fluid in mice. Plast Reconst Surg 39:423–26, 1967.

Bradley SG, Munson AE, McCay JA, et al. Subchronic 10 day immunotoxicity of polydimethylsiloxane (silicone) fluid, gel and elastomer and polyurethane disks in female B6C3F1 mice. Drug Chem Tox 17:175–220, 1994a.

Bradley SG, White KL, McCay JA, et al. Immunotoxicity of 180 day exposure to polydimethylsiloxane (silicone) fluid, gel and elastomer and polyurethane disks in female B6C3F1 mice. Drug Chem Tox 17:221–69,1994b.

Braley SA. The use of silicones in plastic surgery. Plast Reconstr Surg 51:280–88, 1973.

Brantley SK, Davidson SF., St. Arnold PA, et al. The effects of prior exposure to silicone on capsule formation, histology, and pressure. Ann Plastic Surg 25:44–47, 1990.

Brantley SK, Davidson SF, St.Arnold PA, et al. Assessment of the lymphocyte response to silicone. Plast Reconstr Surg 86:1131–37, 1990.

Broderson JR. A retrospective review of lesions associated with the use of Freund's Adjuvant. Lab Animal Sci 39:400–5, 1998.

Butler JE, Lu EP, Navarro P, et al. The adsorption of proteins on a polydimethylsiloxane elastomer (PEP) and their antigenic behavior. In: Potter

M, Rose NR, eds., Immunology of Silicones. Springer, pp 75–84, 1996.

Chang Y. Adjuvanticity and arthritogenicity of silicone. Plast Reconstr Surg 92:469–73, 1993.

Chenoweth MB, Holmes B, Stark F. Dow Corning Report No. 1377. The Physiological Assimilation of Dow Corning 200 Fluid. Midland, MI, 1956.

Das SK, Johnson M, Ellsaesser C, et al. Macrophage interleukin 1 response to injected silicone in a rat model. Ann Plast Surg 28:535–37, 1992.

Fabre T, Bertrand-Barat J, Freyburger G, et al. Quantification of the inflammatory response in exudates to three polymers implanted in vivo. J Biomed Mater Res155:637–41, 1998.

Frondoza C, Jones L, Rose NR, et al. Development of scleroderma-like syndrome in Tsk/+ mice is not enhanced by silicone administration. In: Potter M, Rose NR, eds., Immunology of Silicones, Springer, pp 299–306, 1996.

Gish J, Gaspari AA, Klykken P, et al. Adjuvancy of octamethylcyclotetrasiloxane (D4) in the skin immune system of normal mice. J Invest Dermatol 106:924 (Abstract).

Glenn EM, Gray J. Adjuvant-induced polyarthritis in rats: biologic and histologic background. Am J Vet Res 26:1180–94, 1998.

Goodman DG, Gail RG, Kitchen DN. Two-year gel implant study (B7811) in Sprague- Dawley rats. Jamesville, MD: Pathco, Inc., 1988.

Grasso P, Fairweather FA, Golberg L. A short-term study of epithelial and connective tissue reactions to subcutaneous injection of silicone fluid. Food Cosmet Toxicol 3:263–69, 1965.

Hang L, Theofilopoulos AN, Dixon FJ. A spontaneous rheumatoid arthritis-like disease in MRL/1 mice. J Exp Med 155:1690–1701, 1982.

Hill SL, Landavere MG, Rose NR. The adjuvant effect of silicone gel and silicone elastomer particles in rats. In: Potter M, Rose NR, eds., Immunology of Silicones, Springer, pp123–37, 1996.

Holmdahl R, Andersson ME, Goldschmidt TJ, et al. Collagen induced arthritis as an experimental model for rheumatoid arthritis. APMIS 97:575–84, 1989.

Howie JB, Helyer BJ. The immunology and pathology of NZB mice. Adv Immunol 9: 215–66, 1968.

Janeway, CA, Travers P. Immunobiology. The Immune System in Health and Disease. New York: Garland Publishing, 1994.

King DW, Zimmer MA, Rasmussen CR. Two-year tissue implant study with Q7-4750, X7-4780 and Q7-4840 in Fischer 344 rats. Dow Corning Report reference 226, 1989.

Klykken P, White KL. The adjuvancy of silicones: dependency on compartmentalization. In: Potter M, Rose NR, eds., Immunology of Silicones, Springer, pp.113–21,1996.

Lane TH, Burns SA. Silica, silicon and silicones. Unraveling the mystery. In: Potter M, Rose NR, eds., Immunology of Silicones, Springer, pp 3–12, 1996.

LeBeau JE. Evaluation of Dow Corning Gel Strip as to its ability to produce a sensitization phenomena upon guinea pig skin. Report I0065-1306-4, 1967.

Lemen J, Wolfe GW. Combined Chronic Toxicity and Oncogenicity Study in Rats. Hazleton Washington, Inc. Report, 1992.

Lilla JA, Vistnes LM. Long-term study of reactions to various silicone breast implants in rabbits. Plast Reconstr Surg 57:637–49, 1976.

Lykissa ED, Kala SV, Hurley JB, et al. Release of low molecular weight silicones and platinum from silicone breast implants. Anal Chem 69:4912–16, 1997.

Malcrewski R. Ninety-day implant study of Dow Corning Q7-2146/50 gel. Report 2332-6, 1985.

Mastalski K, Jenkins DH, Kinoshita FK, et al. Subcutaneous implant study with 114-189171-22571, 114-189171A-113070, 114-189171B-113070, Prosthesis, 114-P847F-22371 and 114-1102B1/2-22371 in female beagle dogs. Report IBT No. 8580-09424, 1977.

McDonald AH, Weir K, Schneider M, et al. Silicone gel enhances the development of autoimmune disease in New Zealand black mice but fails to induce it in BALB/cAnPt mice. Clin Immunol Immunopath 87:248–55, 1998.

McKim JM, Wilga PC, Kolesar GB, et al. Evaluation of octamethylcyclotetrasiloxane (D4) as an inducer of rat hepatic microsomal cytochrome P450, UDP- glucuronosyltransferase, and epoxide hydrolase: a 28-day inhalation study. Tox Sci 41:29–41, 1998.

Mehendale HM. Evaluation of the liver microsomal enzyme induction potential of D-5. Report 3724-10,1989.

Mudgett SL, Zimmer MA, Ruhr LP. A 90-day implant study of Dow Corning Q7-2412 elastomer. Report 1990-I0000-35219, 1990.

Muir VY, Dumonde DC. Different strains of rats develop different clinical forms of adjuvant disease. Ann Rheum Dis 41:538–43, 1982.

Naim JO, Ippolito KML, Lanzafame RJ, et al. Induction of type II collagen arthritis in the DA rat using silicone gel as adjuvant. In: Potter M, Rose NR, eds, Immunology of Silicones, Springer, pp 103–11, 1996.

Naim JO, Ippolito KML, Lanzafame RJ, van Oss CJ. The effect of molecular weight and gel preparation on humoral adjuvancy of silicone oils and silicone gels. Immunol Invest 24: 537–47, 1995.

Naim JO, Ippolito KML, van Oss CJ. Adjuvant effect of different types of silicone gel. J Biomed Mater Res 37:534–38, 1997.

Naim JO, Lanzafame RJ, van Oss CJ. The adjuvant effect of silicone-gel on antibody formation in rats. Immunol Invest 22:151–61, 1993.

Naim JO, van Oss CJ, Lanzafame RJ. The induction of autoantibodies to thyroglobulin in rats with silicone gel as adjuvant. Surg Forum 44:676–78, 1993.

Nicholson JJI, Wong GE, Frondoza CG, et al. Silicone gel and octamethylcyclotetrasiloxane potentiate antibody production to bovine serum albumin in mice. In: Potter M, Rose NR, eds., Immunology of Silicones, Springer, pp. 140–44, 1996.

Osborn TG, Nesher G, Moore TL. Effect of silicone injection of skin thickness and antinuclear antibody in the tight-skin mouse. Arth Rheum 37(9 suppl.):S271, 1995 (Abstract)

Osborn T, Moore T, McMurtry P. Effect of polydimethylsiloxane on autoimmune parameters in C57Bl/6 lpr/lpr (B6-lpr) mice. Am Coll Rheum 38:Abstract 1033,1995.

Pearson CM. Development of arthritis, periarthritis and periostitis in rats given adjuvants. Proc Soc Exp Biol Med 91:95–101, 1956.

Picha GJ, Goldstein JA. Investigation of silicone oil and fumed silica in an adjuvant animal model. Plast Reconstr Surg 100:643–52, 1997.

———. Analysis of the soft-tissue response to components used in the manufacture of breast implants: rat animal model. Plast Reconstr Surg 87: 490–500, 1991.

Potter M, Morrison S, Wiener F, et al. Induction of plasmacytomas with silicone gel in genetically susceptible strains of mice. J Natl Cancer Inst 86:1058–65, 1994.

Potter M, Wax JS. Genetics of susceptibility to pristane-induced plasmacytomas in BALB/cAn: reduced susceptibility in BALB/cJ with a brief description of pristane-induced arthritis. J Immunol 127:1591–95, 1981.

Raposo do Amaral CM, Tiziani V, Cintra ML, et al. Local reaction and migration of injected silicone gel: experimental study. Aesth Plast Surg 17:335–38, 1993.

Rasmussen CR. A 90-day implant test of Dow Corning Q7-2423 in rabbits. Report 5194-7, 1988.

Rees TD, Ballantyne DL, Jr., Seidman I, et al. Visceral response to subcutaneous and intraperitoneal injections of silicone in mice. Plast Reconstr Surg 39: 402–10, 1967.

Rhie JW, Han SB, Byeon JH, et al. Efficient in vitro model for Immunotoxic assessment of mammary silicone implants. Plast Reconstr Surg 102:73–77, 1998.

Rose NR, Bhatia S. Autoimmunity: Animal models of human autoimmune disease In: Methods in Immunotoxicology. Wiley-Liss, Inc., 427–45, 1995.

Rosenau B, Schneebaun AB, Schur PH. The development of an ELISA method for the detection of "antibodies" to silicone. In: Potter M, Rose NR, eds,. Immunology of Silicones, Springer, pp. 69-74, 1996.

Schaefer CJ. The influence of silicone implantation on experimental models of autoimmunity. PhD dissertation, Wayne State University, 1997.

Schaefer CJ, Whalen JD, Knapp T, et al. The influence of silicone implantation on type II collagen-induced arthritis in mice. Arthr Rheum 41:1064–72, 1997.

Selwyn MR, Danner VM. Statistical analyses for two-year gel-implant study of Q7-2159A and MDF-0193 in Sprague-Dawley rats. Report M8518-0:M8518-0, 1988.

Siddiqui WH. Evaluation of liver microsomal enzyme induction potential of decamethyl-cyclopentasiloxane in the rat. Report I-0005-2586, 1989.

Smith DJ, Jr., Sazy JA, Crissman JD, et al. Immunogenic potential of carpal implants. J Surgical Res 48:13–20, 1990.

Sparschu GL, Clashman A. Pathology Report on the Effects of Dow Corning 360 Fluid-350 Centistokes after Administration to Rats Intraperitoneally or Subcutaneously. Midland, MI: Dow Chemical Co., 1970.

Van Eden W, Holoshitz J, Nevo Z, et al. Arthritis induced by a T-lymphocyte clone that responds to Mycobacterium tuberculosis and to cartilage proteoglycans. Proc Natl Acad Sci USA 82:5117–20, 1985.

Varaprath S, Salyers KL, Plotzke KP. Non-regulated study: identification of major metabolites of octamethylcyclotetrasiloxane (D4) in rat urine. Report 1997-I0000-43454, 1997.

West B, Jolly ER. Two year safety evaluation study in dogs with DOW Corning 360 liquid. 1976.

Whitehouse MW, Orr KJ, Beck FWJ, et al. Freund's adjuvants: relationship of arthritogenicity and adjuvanticity in rats to vehicle composition. Immunology 27:311, 1974.

Wilson SD, Munson AE. Modulation of NK1.1 splenocytes after exposure to octamethylcyclotetrasiloxane (D4). The Toxicologist (Abstract), 1996

Yoshino S. Silicone-induced arthritis in rats and possible role for T cells. Immunobiol 192:40–47, 1994.

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