Biomedical subjects

B Formby

Publications and source records attributed to B Formby.

At least 19 recordsLinked to original sources

PubMed2001

Hyaluronidase can modulate expression of CD44.

CD44 is a family of transmembrane glycoproteins with multiple isoforms generated by alternative exon splicing of a single gene. CD44 and its variants are expressed on a wide variety of cells including cancer cells. The mechanisms by which splice variant exons are selected are unknown. The presence of hyaluronan in the environment of the cell appears to influence that selection process. The expression of particular splice variants of CD44 as well as the simultaneous presence of hyaluronan is important for motility, invasion, and the metastatic spread of some tumors. The influence of hyaluronidase digestion on the expression of CD44 in human cancer cell lines was examined. CD44 isoforms containing alternatively spliced exons were sensitive to hyaluronidase digestion in all lines examined, but differences between cell lines were observed. Expression of CD44s, the standard form, was resistant to digestion in two of three cell lines. A tentative model was formulated proposing that CD44 isoforms containing splice variants are unstable, requiring the continuous presence of ligand for expression. CD44s is relatively more stable, not requiring the continuous presence of hyaluronan. Additionally, a number of new CD44 variant isoforms, not previously observed, were identified.

Alternative Splicing↗

PubMed1999

Bcl-2, survivin and variant CD44 v7-v10 are downregulated and p53 is upregulated in breast cancer cells by progesterone: inhibition of cell growth and induction of apoptosis.

Progesterone inhibits the proliferation of normal breast epithelial cells in vivo, as well as breast cancer cells in vitro. But the biologic mechanism of this inhibition remains to be determined. We explored the possibility that an antiproliferative activity of progesterone in breast cancer cell lines is due to its ability to induce apoptosis. Since p53, bcl-2 and survivin genetically control the apoptotic process, we investigated whether or not these genes could be involved in the progesterone-induced apoptosis. We found a maximal 90% inhibition of cell proliferation with T47-D breast cancer cells after exposure to 10 microM progesterone for 72 h. Control progesterone receptor negative MDA-231 cancer cells were unresponsive to 10 microM progesterone. The earliest sign of apoptosis is translocation of phosphatidylserine from the inner to the outer leaflet of the plasma membrane and can be monitored by the calcium-dependent binding of annexin V in conjunction with flow cytometry. After 24 h of exposure to 10 microM progesterone, cytofluorometric analysis of T47-D breast cancer cells indicated 43% were annexin V-positive and had undergone apoptosis and no cells showed signs of cellular necrosis (propidium iodide negative). After 72 h of exposure to 10 microM progesterone, 48% of the cells had undergone apoptosis and 40% were annexin V positive/propidium iodide positive indicating signs of necrosis. Control untreated cancer cells did not undergo apoptosis. Evidence proving apoptosis was also demonstrated by fragmentation of nuclear DNA into multiples of oligonucleosomal fragments. After 24 h of exposure of T47-D cells to either 1 or 10 microM progesterone, we observed a marked down-regulation of protooncogene bcl-2 protein and mRNA levels. mRNA levels of survivin and the metastatic variant CD44 v7-v10 were also downregulated. Progesterone increased p53 mRNA levels. These results demonstrate that progesterone at relative high physiological concentrations, but comparable to those seen in plasma during the third trimester of human pregnancy, exhibited a strong antiproliferative effect on breast cancer cells and induced apoptosis.

Annexin A5↗

PubMed1998

Contribution of T cells to the development of autoimmune diabetes in the NOD mouse model.

The nonobese diabetic (NOD) mouse spontaneously develops an autoimmune diabetes that shares many immunogenetic features with human insulin-dependent diabetes mellitus (IDDM), type 1 diabetes. The mononuclear cell infiltrates in the islet, which results in the development of insulitis (a prerequisite step for the development of diabetes) are primarily composed of T cells. It is now well accepted that these T cells play important roles in initiating and propagating an autoimmune process, which in turn destroys insulin-producing islet beta cells in the pancreas. T cells are subdivided into CD4+ helper T cells and CD8+ cytotoxic T cells. CD4+ T cells are further subdivided into Th1 and Th2 cells based on profiles of cytokine production, and these two T-cell populations counterregulate each other. Because many autoimmune diseases are Th1 T-cell mediated, current studies have focused on manipulating the Th1/Th2 imbalance to suppress the autoimmune process in the NOD model. Furthermore, the incidence of disease is much higher in females than that in males, suggesting an involvement of sex-steroid hormones in the development of diabetes. Understanding insights of the mechanism of immune-mediated islet cell destruction and the interaction between the immune and the neuroendocrine system may, therefore, provide new therapeutic means of preventing this chronic debilitating disease.

Animals↗

PubMed1998

Phosphorylation stabilizes alternatively spliced CD44 mRNA transcripts in breast cancer cells: inhibition by antisense complementary to casein kinase II mRNA.

CD44, the predominant vertebrate cell surface receptor for hyaluronan, exists in a variety of isoforms resulting from alternative splicing of a single gene. Particular spliced variants of CD44 correlate with increased cell motility, and with poor clinical prognosis in several kinds of carcinomas. Combinations of 9 variant exons that confer this enhanced motility on tumor cells are inserted into a single site in the middle of the extracellular domain of CD44. Evidence suggests that phosphorylation of 2 serine residues in the intracellular domain of CD44 are involved in controlling these events. However, evidence is lacking as to the nature of such kinases. Acidic amino acids in close proximity to these 2 serine residues suggests casein kinase II (CKII) is involved. We now show an antisense phosphorothioate oligonucleotide designed to hybridize to the AUG translation initiation codon of subunit CKII alpha' mRNA blocks in vivo phosphorylation of CD44 in MDA231 breast tumor cells, and at the protein level decreases ectopic expression of total CD44 as well as the metastatic v-7 CD44 isoform. Furthermore subplateau RT-PCR analysis demonstrated antisense transfected MDA231 tumor cells had significant down-regulated or eliminated mRNA transcripts of metastatic CD44 isoforms. CKII as a CD44-associated serine kinase therefore may serve as an important molecule in a signaling cascade that produces a variety of cellular responses in MDA231 breast cancer cells. Since the 3'-untranslated region of CD44 mRNA contain 4 dispersed AUUUA sequences which serve as signals targeting mRNA for rapid turnover, a mechanism is proposed by which CD44 phosphorylation mediates labile message stabilization, hence providing insights into the processes involved in cancer cell growth, invasion and metastasis.

Alternative Splicing↗

PubMed1998

Expression of CD44 isoforms in cultured human trabecular meshwork cells.

PURPOSE: To determine the expression of CD44 isoforms in cultured human trabecular meshwork (HTM) and to discuss their possible relationship with outflow facility. METHODS: CD44 isoform expression in cultured HTM was qualitatively examined using immunohistochemistry and RT-PCR analysis. RESULTS: Immunohistochemistry of cultured HTM showed intense staining with a CD44s antibody, and with antibodies against CD44 exon 7, exon 11-12 and exon 14. By RT-PCR, at least three isoforms of CD44 were expressed in HTM: CD44s, CD44v-III and CD44v-I. CONCLUSIONS: At least three isoforms of CD44 are expressed in the HTM. CD44 may play a role in binding and turnover of hyaluronic acid in the trabecular meshwork, thereby regulating outflow facility.

Cells, Cultured↗

PubMed1997

Hyaluronan is a prerequisite for ductal branching morphogenesis.

Hyaluronan, a macromolecular carbohydrate polymer of the extracellular matrix is prominent early in embryogenesis, coinciding with rapid tissue growth. CD44, the predominant receptor for hyaluronan on vertebrate cells, is a variably expressed transmembrane glycoprotein. Mouse anterior prostate glands obtained at various postnatal time points were examined for the expression of hyaluronan and CD44. Reverse transcriptase polymerase chain reaction analysis was used to map the temporal regulation of specific CD44 variant isoforms. In each age group, hyaluronan was localized exclusively in the stromal matrix. Hyaluronan was greatly reduced in the later ages and was entirely absent around the developmentally quiescent proximal regions of the ducts. Early in prostate development, CD44 was prominent in the mesenchyme. However, in the later phases, CD44 expression became associated with membranes of epithelial cells. The role of hyaluronan-CD44 interactions in ductal branching morphogenesis was studied by serum-free organ culture of mouse anterior prostate. In the presence of optimal levels of testosterone, the organs underwent ductal branching morphogenesis. Treatment with either neutralizing anti-CD44 antibodies, hyaluronan hexasaccharides or the enzyme hyaluronidase inhibited androgen-stimulated ductal branching morphogenesis. These results are suggestive of the significant role played by hyaluronan-CD44 interactions in mediating androgen-induced prostatic growth and morphogenesis.

Animals↗

PubMed1996

Effect of 5-alpha dihydrotestosterone on T-cell proliferation of the female nonobese diabetic mouse.

Nonobese diabetic (NOD) mice develop type I diabetes spontaneously and have been utilized as a model for human autoimmune insulin-dependent diabetes. The disease is caused by the destruction of insulin-producing beta cells in the pancreatic islet of Langerhans by infiltrating inflammatory cells, which are primarily T lymphocytes. The incidence of diabetes in NOD mice is increased in females compared with males, suggesting that sex steroid hormones play an important role in the development of the disease. We therefore investigated the effect of a male steroid, 5-alpha-dihydrotestosterone (5DHT), on disease development, T-cell phenotype, T-cell proliferation, and cytokine profiles in this model. None of the mice that received 5DHT for 120 days (n = 7) developed insulitis, whereas all control mice (n = 8) developed the disease. The percentage of CD4+ T cells in peripheral blood mononuclear cells was markedly decreased in the 5DHT-treated females compared with those in controls (37.1 +/- 4.8 vs 51.3 +/- 9.3, P < 0.02), whereas no significant differences in the percentage of CD8+ T cells were observed between treated and control female mice. Results of a syngeneic mixed lymphocyte reaction (SMLR) also suggested that T cells are major target cells of 5DHT administration. An increased expression of IL-4 mRNA, representing T helper 2 (Th2) T cells, was observed in the SMLR. On the basis of these results, a systemic administration of 5DHT appears to have direct effects on the expansion of Th2 cell populations with subsequent restoration of normal immune responses.

Animals↗

PubMed1995

Association of an androgen-responsive T cell phenotype with murine diabetes and Idd2.

T cells are involved in the induction and suppression of autoimmune diabetes in nonobese diabetic (NOD) mice. Because the incidence of diabetes is 13-fold greater in NOD/Smrf females, we searched for T cell phenotypes that showed sexual dimorphism and associated with diabetes in backcross segregants. The percentage of CD4+PBL was higher in NOD/Smrf females than males, was intermediate in [NOD X NON] F1 mice and approximated a 1:1 distribution in F1 mice backcrossed to either NOD or NON parental strains, suggesting primary control of the phenotype by an incompletely dominant gene, but not excluding additional effects by other genes. We term this primary gene Tlf(T lymphocyte frequency) because it also influenced the percentage of CD8+ T cells, although to lesser extent and independently from the MHC previously shown to lower the CD8+ T cell fraction in NON mice. Tlf segregated with diabetes in BC1 females, suggesting linkage with at least one diabetic locus. Genotyping of markers for Idd1, Idd2, and Idd3/10 revealed that Tlf mapped with Idd2 on chromosome 9. Dihydrotestosterone simultaneously lowered CD4+ PBL levels and prevented diabetes in NOD females while, in vitro, it had a differential effect on Con A elicited cytokines, increasing IL-2 22% and decreasing IL-4 39% (p < 0.0001). Thus the Tlf phenotype in NOD females, like diabetes, can be modulated by androgens.

Animals↗

PubMed1995

Immunologic response in pregnancy. Its role in endocrine disorders of pregnancy and influence on the course of maternal autoimmune diseases.

During gestation, the immune system is challenged with establishing tolerance to the partial allograft represented by the paternal contribution to the fetal genome. That fact that cell-mediated immunity is decreased while T-cell-dependent immunoglobin production remains intact or is increased is generally accepted. This latter phenomenon can be placed in teleological perspective in terms of providing passive immunity to the fetus. T-cell lineages that diverge based on function and cytokine production have been identified. Pregnancy is associated with a relative increase in Th2-associated immunity, characterized by increased production of the cytokines IL-4 and IL-10. These changes occur concomitantly with increased immunoglobulin production and decreased Th1 immunity. IL-2 and interferon secretion characterize Th1 immunity, facilitating allograft rejection. These changes in Th1 and Th2 have implications for the clinical course of a several autoimmune diseases that may complicate pregnancy and the postpartum period.

Animals↗

PubMed1994

In situ islet cytokine gene expression during development of type I diabetes in the non-obese diabetic mouse.

Expression of cytokine genes in the islets of non-obese diabetic (NOD) female mice was examined. RNA samples were prepared from the islets and spleens of NOD mice at different time points at prediabetic stages during the natural disease process. Cytofluorometric analyses showed that the majority of lymphocyte infiltrates in the islets at 14 weeks of age consisted of T cells (68%). Of these, 80% of Thy1.2+ cells were CD4+ T cells. Less than 1% of in situ islet immune cells expressed a cell surface marker specific for the macrophage (Mac-1). Results of polymerase chain reaction using RNA (RT-PCR) prepared from spleens, and isolated and purified islets demonstrated that IFN-gamma message was detectable in the islets at 7 weeks of age (an early stage of insulitis). No message for this gene was detected in the spleen at any stage studied (7, 14 and 16 weeks of age). In contrast, TNF-alpha message was detected in both spleen and the islets at all stages, although the level of expression of TNF-alpha in the islets was much higher than that in the spleen. These results suggest that both cytokines are produced by in situ islet T cells, possibly activated T cells, which may be responsible for initiating or perpetuating autoimmune reactions in the islets.

Animals↗

PubMed1994

Maternal hyperglycemia is not the only cause of macrosomia: lessons learned from the nonobese diabetic mouse.

Maternal hyperglycemia has been implicated as the major cause of neonatal macrosomia, yet clinicians frequently report the birth of large-for-gestational-age infants in normoglycemic pregnancies. We examined the relationship between birthweight, maternal blood glucose (BG), glycosylated hemoglobin (GHb) levels, litter size, maternal age, gestational duration, and parity using the non-obese diabetic (NOD) mouse model. We observed 133 litters and analyzed the birthweight in relation to BG, GHb, litter size, maternal age, gestational duration, and parity. We found that mean litter birthweight was significantly negatively correlated with the total number of pups in each litter (r = -0.39; P < 0.01) and significantly positively correlated with parity (r = 0.19; P < 0.05) and maternal age (r = 0.22; P < 0.05). The total number of pups was significantly negatively correlated with parity (r = -0.33; P < 0.01) and with parent age (r = -0.21; P < 0.05). The relationship between birthweight and GHb was bimodal. No relationship was found with a GHb less than 2.5%, a significant positive correlation was found for GHb between 2.6% and 4.0% (r = 0.67; P < 0.01), and a negative relationship was found when GHb was above 4.0%. Thus, increased parity, maternal age and glucose are associated with increased birthweight. Mild hyperglycemia plays the major role when age, maternal size, gestational duration, and parity are controlled.

Animals↗

PubMed1993

Adhesion and costimulation of proliferative responses of human gamma delta T cells by interaction of VLA-4 and VLA-5 with fibronectin.

The very late antigens, VLA-4 and VLA-5 belong to the beta 1 subfamily of integrins and have been identified as receptors for different binding regions of fibronectin (FN). We have detected VLA-4 and VLA-5, but not VLA-3 and VLA-6 expressed on human CD3+CD4-CD8- gamma delta TCR T cells by flow cytometry. Binding assays, performed on FN-coated plates, showed that activated CD25high (IL-2 receptor) but not resting CD25low gamma delta T cells specifically adhere to FN. The binding capacity is inhibited by the synthetic peptide GRGDSP which inhibits adhesion mediated by VLA-5 and a functional mAb directed against the alpha 4 subunit. Most FN binding is mediated by VLA-4. Additionally, resting gamma delta T cells cultured on coimmobilized anti-TCR delta 1 mAb and FN or the 40 kDa fragment (which contains the adhesion site in the IIICS domain recognized by VLA-4) for 96 h in the absence of exogeneous IL-2 showed significant increase in proliferation when compared to that of resting gamma delta T cells cultured on immobilized anti-TCR delta 1 mAb alone. Also expression of CD25 was significantly enhanced on cells cultured on coimmobilized anti-TCR delta 1 mAb and FN, indicative of T cell activation. Cross-linking of VLA-4 and VLA-5 molecules costimulated expansion of resting gamma delta T cells induced by cross-linked TCR delta 1. These results suggest that the gamma delta T cell beta 1 integrins, VLA-4 and VLA-5, may function in a dual capacity as signalling and adhesion molecules.

Antibodies, Monoclonal↗

PubMed1992

In situ islet T cell receptor variable region gene usage in the nonobese diabetic mouse.

Several features of the genetics and immunopathology of diabetes in the nonobese diabetic (NOD) mouse, which spontaneously develops type I diabetes, are shared with the human disease. Immunohistochemical studies support the concept that T lymphocytes are the major components of inflammatory cells in the pancreatic islets and these cells may play a critical role in the destruction of the beta cells leading to diabetes. Therefore, we examined whether particular TCR-beta variable region genes were utilized by in situ islet T cells at different stages (4 - 5, 7, 14 - 15 and 16 weeks of age) of the disease process. Dot-blot hybridization was performed using RNA prepared from isolated islets, thymus, spleen, peripheral blood leukocytes and axillary lymph nodes of 10 to 15 mice pooled for each data point. Ten different TCR V-beta probes were used for the analyses. Limited usage of islet V-beta genes was observed only at the early prediabetic stage (4 - 5 weeks old) of the disease. At later stages of the disease (7 - 16 weeks old), no preferential usage of TCR genes was observed in the islets compared to those of peripheral lymphoid organs. These data suggest that only certain types of T cells bearing particular TCR V-beta genes may be responsible for initiating and perpetuating infiltration of immune cells into the islets and these particular T cells are only identified at the very early stages of the autoimmune process.

Animals↗

PubMed1992

Exogenous administration of IL-1 alpha inhibits active and adoptive transfer autoimmune diabetes in NOD mice.

Diabetes susceptibility in non-obese diabetic (NOD) mice may involve immune dysregulation resulting from cytokine deficiencies. The cytokine IL-1 plays a role in various immune as well as endocrine responses and may be hypoexpressed in NOD mice. Treatment with low levels of exogenous IL-1 alpha for 22 weeks prevented the naturally occurring insulitis and diabetogenic process in NOD mice during and at least 33 weeks after cessation of IL-1 alpha treatment. Treatment with IL-1 alpha also inhibited insulitis and hyperglycemia induced by adoptive transfer of pathogenic, polyclonal CD4+8- T cells. Even after islet-cell destruction, IL-1 alpha injections in diabetic NOD mice normalized plasma glucose levels when administered in combination with insulin, whereas equivalent levels of IL-1 alpha alone did not. Our studies support the hypothesis that IL-1 alpha suppresses autoimmune diabetes and hyperglycemia in NOD mice by pleiotropic effects on both immune and metabolic systems. Thus, IL-1 treatment could clinically be an effective immunotherapeutic modality for autoimmune diabetes mellitus by suppressing early disease progression or normalize plasma glucose levels when insulin is present.

Animals↗

PubMed1992

Quantitative and functional analyses of spleen and in situ islet immune cells before and after diabetes onset in the NOD mouse.

Cytofluorometric analysis using specific monoclonal antibodies directed against the T cell antigens Thy-1.2, CD4, CD8, CD4V beta(8.1 + 8.2 + 8.3), and the antigen Mac-1 expressed by mature macrophages and NK cells were used to characterize and quantify the phenotypes of (1) unfractionated and Percoll gradient fractionated in situ islet immune cells isolated from prediabetic and diabetic female NOD mouse spleens. We found in prediabetic female mice that the majority (approximately 70%) of the in situ islet immune cells were Thy-1.2 positive T cells. CD4 positive T cells (approximately 40%) were the most abundant phenotype together with double negative T cells (approximately 20%). The percentage of CD8 positive T cells were approximately 10%, and only approximately 4% of the immune cells were Mac-1 positive. The percentages of CD4V beta (8.1 + 8.2 + 8.3) positive and double negative T cells in diabetic spleens were significantly higher in comparison to prediabetic spleens. In C57B1/6J control nondiabetic mice the percentage of double negative T cells in the spleens was significantly 4-fold lower when compared to diabetic NOD spleens. The specific cytolytic activity mediated by in situ islet immune cells against 51Cr-labeled dispersed syngeneic single-cell islet cells at an effector to target ratio of 20 was twenty- to thirty-fold higher than that mediated by prediabetic splenic lymphoid cells. It is concluded that prediabetic NOD mouse in situ islet immune cells are mostly CD4 positive and double negative T cells, and that CD4 and CD8 positive T cells in the intra-islet infiltrate warrants further evaluation as potential effector T cells in target beta-cell destruction.

Animals↗

PubMed1991

Comparison of cytolytic and proliferative activities of human gamma delta and alpha beta T cells from peripheral blood against various human tumor cell lines.

Peripheral blood lymphocytes (PBLs) from healthy donors were more than 95% enriched for gamma delta T cells. V gamma 9/V delta 2 was the predominant T-cell subset and represented more than two thirds of the isolated gamma delta T cells. When activated by surface-immobilized anti-TCR delta 1 monoclonal antibody (MoAb) during 5 days of culture, gamma delta T cells demonstrated a significant fourfold to 15-fold stronger cytolytic activity against various tumor cell lines than did PBL polyclonal alpha beta T cells analogously activated by surface-immobilized anti-CD3 MoAbs. Blocking experiments with specific MoAbs demonstrated that the cytolytic activity of gamma delta T cells was non-major histocompatibility complex (MHC) restricted and did not involve the gamma delta T-cell antigen receptor (TCR) but that cytolytic activity was dependent on LFA-1 beta/ICAM1 interaction. The proliferative responses of gamma delta and alpha beta T cells to Ovcar-3 tumor cells and irradiated allogeneic PBLs were inhibited by anti-HLA-A,B,C MoAb. Our findings suggest that gamma delta T cells activated via the TCR have a significant advantage in non-MHC-restricted lysis of various tumor cell lines over PBL alpha beta T cells stimulated analogously with anti-CD3, which may be important in terms of applicability of activated gamma delta cells for adoptive immunotherapy of metastatic cancers.

Antibodies, Monoclonal↗

PubMed1991

Studies of concanavalin A in nonobese diabetic mice. II. Lymphocyte tracking and phenotype responses.

We studied the tracking of 51chromium-labeled (resting) and Con A-induced (activated) tritiated thymidine-labeled syngeneic lymphocytes to adrenal, blood, brain, heart, liver, lymph nodes, pancreas, spleen, testis/ovary, thymus and thyroid in prediabetic, nonobese diabetic (NOD) mice and in age- and sex-matched C57BL/6 mice. 51Chromium-labeled cells showed no significant difference between strains in tracking to any tissue except lymph nodes (decreased in NOD vs. C57, P less than .05). Con A incubation resulted in no difference between strains in lymphocyte tracking to lymph nodes, but NOD mice had increased pancreatic tracking with Con A-incubated cells compared to C57 mice (P less than .05). Female NOD mice had reduced thymic tracking (P = .001) compared to C57 controls. Positive selection experiments showed the Con A-responsive cell to be a T cell. Both Lyt2+ (CD8+) and L3T4+ (CD4+) enriched T cell populations showed a labeling response to Con A. After 48 h of Con A incubation, the L3T4+/Lyt2+ ratio increased in splenocytes from NOD mice (P less than .05), whereas it decreased in C57 controls (P less than .01). Over the course of 6 days in culture, NOD splenocytes exposed to Con A characteristically exhibited a delayed expansion of the Lyt2+ population. We conclude that Con A incubation of NOD splenocytes results in T cells that, when reinfused, avoid the thymus and track preferentially to the pancreas. Con A immunomodulation as potential treatment for autoimmune disease warrants further study in this murine model.

Animals↗

PubMed1990

Autologous CD4 T-cell responses to ectopic class II major histocompatibility complex antigen-expressing single-cell islet cells: an in vitro insight into the pathogenesis of lymphocytic insulitis in nonobese diabetic mice.

We investigated by flow cytometric analysis the expression of class II major histocompatibility complex (MHC) molecules by viable single-cell islet cells (SCICs) prepared from male and female 4- and 10-week-old nonobese diabetic (NOD) mouse islets. With anti-I-Ak monoclonal antibody (specific for I-Ak,f,r,s beta and produced by clone 11-5-2), and fluorescein isothiocyanate-conjugated goat anti-mouse IgG as second-step antibody, we found that SCICs from both sexes aberrantly expressed class II MHC molecules, which was not altered after SCICs were cultured for 24 hr or 120 hr in the presence of 10 ng of recombinant murine interferon gamma per ml. Double-indirect immunofluorescence of male SCICs indicated that the expression of class II MHC molecules was a property of beta cells. Control experiments documented that macrophages and mononuclear cells did not contaminate the SCIC preparations. Coculture experiments with responder splenic CD4 T cells isolated from diabetic NOD mice and stimulator male SCICs indicated a recognition event evidenced by a 12-fold increase in proliferative response. Monoclonal antibodies to class II MHC and CD4 antigens blocked the proliferative response. Results from control autologous and allogeneic mixed lymphocyte reactions suggest that the responder CD4 T cells are autoreactive self-class II MHC restricted. We tentatively conclude that the ability of SCICs from both sexes of NOD mice to express class II MHC molecules as early as 4 weeks of age may represent a mechanism for targeting immune reactions to beta cells and initiate lymphocytic insulitis.

Animals↗