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U.S. Department of Health and Human Services

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FY 2000 Biomaterial Effects On Immune System

A biomaterial in a medical device in contact with mucous membranes or blood may leach into tissue and initiate a local response such as inflammation. This normal inflammatory process can become pathologic under chronic conditions and result in unresolved granuloma formation. Chronic perturbation of the immune system characterized by episodes of inflammatory bursts can break biological tolerance. Immunological defense mechanisms such as those involving cellular elements of the inflammatory process, the complement cascade, and cell proliferation can (under chronic conditions) damage tissues. This occurs when the normal inflammatory process becomes pathologic and results in granuloma formation and arthritic-like symptoms.

From a regulatory perspective, immunotoxicity is defined as any effect on the structure or function of the immune system or on other systems as a result of immune system dysfunction. An effect is considered adverse or immunotoxic if it impairs humoral or cellular immunity needed by the host to defend itself against infections or neoplastic disease or causes unnecessary tissue damage. It is essential to clearly recognize that change in an immune function or level of immunological mediator may not necessarily appear as an adverse effect, but rather as immunostimulation. Decisions on whether a material/device is immunotoxic must rely on the available evidence from preclinical test results and clinical evaluation, as well as prior history of use. Because the available data will often be less than conclusive, good judgment will play an important part in evaluating immunotoxic risk.

A goal of the OST research program is to provide laboratory assessment of the types of testing available for evaluating potential adverse effect of biomaterials on the immune system and to provide a process for selecting appropriate test methods. OST aims to make it possible for reviewers to obtain adequate information to make confident regulatory decisions; to ask the appropriate questions when negotiating with manufacturers; and to generate independent data concerning immune responses to biomaterials in devices. The data should provide some level of assurance that immunotoxic reactions are unlikely.

OST scientists have served as members of ODE review teams for McGhan Gel-filled breast Implants, for Mentor Silicone Breast Implants; for gene kit components, PDP devices and humanitarian devices exemption submissions (including a request for a toxicological consult regarding the carcinogenicity of gluteraldehyde). Scientists contributed reviews of master files and guidance documents.

Staff developed and coordinated IMMUNOTOX, an online service available on FDA intranet/FIRST to enhance communication among FDA immunologists and immunotoxicologists and to provide online services. The goal of the service is to provide assistance on product-related regulatory review issues.

The regulatory impact of immunotoxicity was addressed in the Immunotoxicity Guidance for Reviewers and Industry, prepared by staff scientists. The guidance was derived, in part, from laboratory studies and literature review and is designed to provide assurance that immunotoxic reactions are unlikely.

Finalization of a Standard Practice for Testing for Alternative Pathway Complement Activation in Serum by Solid Materials
Key words: complement activation, alternative complement pathway, medical materials, standard

This Standard Practice was developed by OST scientists and has been accepted by ASTM as "Standard Practice for Testing for Alternative Pathway Complement Activation in Serum by Solid Materials." Complement activation by the alternative pathway is a potential hazard when a patient’s blood contacts medical device materials. Inappropriate complement activation by blood-contacting medical devices may have serious acute or chronic health effects. This practice provides a protocol for simple, inexpensive, rapid, in vitro screening for alternative pathway complement activating properties of solid materials used in the fabrication of medical devices that will contact blood. The practice is designed for use with other standards that assess the biocompatibility of materials, particularly a previous standard also developed by OST scientists: ASTM F1984-99 "Standard Practice for Testing for Whole Complement Activation in Serum by Solid Materials." The practice is composed of two parts. In the first part, serum from guinea pigs genetically deficient in the complement factor C4 (necessary for classical pathway complement activation) is exposed to a solid material, during which alternative pathway complement activation and depletion of key complement proteins may occur. In the second part, the remaining complement is assayed for its ability to lyse rabbit red blood cells via the alternative pathway. The unsuitability of using human serum depleted of C4 by column immunoabsorption is discussed. Data demonstrating the equivalence of guinea pig serum to human serum for detection of complement activation by biomaterials, such as beaded agarose (Sepharose), is presented. Whereas some assays identify the amount of individual complement proteins in the blood, this assay determines alternative pathway complement functional activity. Therefore, the standard practice examines the ability of complement to exert one of its primary functions: the lysis of target cells. Assessing in vitro alternative complement activation, as described here, provides one method for predicting potential complement activation by materials intended for clinical application in humans when the material contacts the blood. This in vitro test method is suitable for adoption in the specifications and standards used for screening solid materials used in constructing implantable medical devices or devices that come in contact with human blood outside of the body.

Complement Activation by Implanted or External Blood-Contacting Medical Devices
Key words: alternative pathway, complement activation, standards.

Complement is a series of serum proteins involved in mediating immune reactions. Complement activation is a tightly regulated process that, in addition to direct cell cytolysis, can have profound affects on the immune, vascular, and coagulation systems. Though complement activation is an important defense mechanism against microbial infections, inappropriate activation by implanted or external medical devices may result in serious acute or chronic reactions.

Complement activation can occur by two main pathways. The classical pathway is triggered by antibodies bound to a cell surface. The alternative pathway is triggered by free hydroxyl or amino groups, such as are present on microbial organisms. Medical device materials generally activate complement by the alternative pathway. Complement activation by a candidate material via the alternative pathway in vitro indicates the material’s potential to trigger inappropriate complement activation when implanted in a patient or placed in contact with the patient’s blood outside the body.

OST scientists finalized a Standard Practice, accepted by ASTM, which screens specifically for alternative pathway complement activation by solid materials used in manufacturing medical devices. This supplements ASTM Standard Practice F1984-99, also developed by OST scientists, which screens for complement activation but does not specify which pathway is activated.

Examples of devices whose materials might activate complement by the alternative pathway include perfusion devices (such as dialysis membranes, cardiopulmonary assist systems, biosensor membranes, liver-assist perfusion devices, and columns for removing antibodies and other factors from patient blood), indwelling artificial vascular grafts, encapsulated drugs or cells, and vascular shunts/stents/catheters. OST is conducting research to acquire baseline information related to assessing risk from complement activation, particularly via the alternative pathway, by these devices.

In particular, complement activation by cardiovascular devices is being studied in a pig model. OST scientists are developing standardized methods to study complement activation by cardiovascular shunts, catheters, guide wires, and stents. The serum taken from pigs has been assayed before and after balloon angioplasty, and the scientists have documented a small but significant decrease in systemic whole complement levels, suggesting that complement activation does indeed occur during the procedure. The possible association of device-mediated complement activation in reblockage (restenosis) of unblocked cardiac arteries is also a concern.

Since damage by cardiovascular devices to the endothelial cells lining the inside of the blood vessels is known to be a critical step in subsequent restenosis, methods were developed for determining the effect of complement on pig and human endothelial cells via flow cytometry and fluorescent dyes. OST data demonstrated that complement activation by the classical and alternative methods both produce significant lysis of endothelial cells. OST researchers are now developing methodologies to extend the data to intact monolayers of pig and human endothelial cells, as a model of the interior of device-disturbed blood vessels.

Analysis of Autoantibody Responses to Silicone Gel-Filled Breast Implants
Key words: autoimmune disease, autoantibodies, silicone breast implants, silicone gel

Autoimmune diseases have been reported in women with silicone breast implants. The presence of autoantibodies in some of the women, as well as studies in experimental animals, suggest that silicone may play a role in these adverse effects on the immune system. Earlier OST studies have shown that silicone gel/oil can promote autoantibody production against the connective tissue proteins, collagen, and can migrate from the implant site to other parts of the body. The current project is a confirmation study. There were 240 rats from 2 different strains used in this study: Dark Agouti and Sprague Dawley. Rats were divided into six groups: immunized with Dow Corning oil, silicone oil with/without rat collagen type I/II, Dow Corning gel, silicone gel with/without collagen type I/II, incomplete adjuvant with collagen type I/II, and rat collagen I/II. Rats were pre-bled before the immunization and every 4 weeks after the initial immunization. Sera were tested for antibodies against rat collagen type I, and rat collagen type II using ELISA technique.