Gastrointestinal carcinoma-associated antigen defined by a murine monoclonal antibody.

A murine monoclonal antibody, CHIP, has been prepared against a human pancreatic carcinoma cell line, SHAW. With the use of the avidin-biotin immunoperoxidase technique, the CHIP antibody detected an antigen found in 11 of 20 fixed tissue sections of tumors obtained from patients with pancreatic carcinoma. The antibody also detected the antigen in 25 of 26 colon carcinoma specimens, 4 of 6 gastric carcinoma specimens, and 1 of 2 esophageal adenocarcinoma specimens. The antigen was also found in normal proximal jejunum and colon and in small amounts in pancreatic islets and parathyroid. There was no reactivity with normal pancreatic ductal or acinar cells or with mesenchymal tissues.

Preferential lysis of pancreatic B-cells by islet cell surface antibodies.

Sera from patients with insulin-dependent diabetes mellitus (IDDM) containing islet cell surface antibodies (ICSA) were studied for their capacity to lyse cultured rat islet cells. The uptake of ethidium bromide was used to identify lysed cells and immunofluorescent staining with antisera to insulin, glucagon, somatostatin, or pancreatic polypeptide was used to identify the different islet cell types (B-, A-, D-, and PP-cells, respectively). Our experiments showed that in the presence of complement, sera containing ICSA lysed 81% of the B-cells, but 10% or less of the A-, D-, and PP-cells. Normal control sera resulted in lysis of less than 4% of each of the islet cell types. The demonstration that ICSA are preferentially lytic for B-cells may be important in defining the role of these autoantibodies in the pathogenesis of IDDM, particularly since the B-cell mass in diabetics is markedly reduced relative to the other islet cell types.

Microculture system for studying monolayers of functional beta-cells.

A method is described for growing monolayers of newborn rat beta-cells in microculture trays. After disruption of the pancreas with collagenase, islets were isolated by Ficoll density gradient centrifugation, trypsinized to obtain individual cells, and plated in 96-well tissue culture trays. The cells were incubated for the first 3 days in growth medium containing 0.1 mM 3-isobutyl-1-methylxanthine to promote monolayer formation. The cultures could be maintained in a functional state, as defined by their responsiveness to known modulators of insulin secretion, for at least 2 weeks. As few as 1 X 10(3) islet cells/well gave results that were reproducible within +/- 10%. It is suggested that the microculture system for islet cells might prove to be a rapid and reproducible screening technique for studying drugs, viruses, or other agents that affect beta-cell function.

Specific identification of lysed target cells in a heterogeneous population by a double-fluorescent label technique.

A double-fluorescent label method for the specific identification of target cells during cytotoxicity testing of a mixed cell population is described. When used for the detection of pancreatic islet cell surface antibodies, damaged cells are identified by the uptake of ethidium bromide (see as cells with orange nuclei when examined under rhodamine filters) and the various islet cell types are simultaneously identified by indirect immunofluorescent staining with the appropriate islet hormone antiserum and FITC-conjugated second antibody (seen as cells with green cytoplasma when examined under fluorescein filters). In this way, we have shown the insulin-containing beta cell to be the target of islet cell surface antibodies. This technique may be particularly useful in the study of autoimmune endocrine diseases where in vitro cytotoxicity testing would involve target cells intermixed with other cell types (e.g., adrenal gland, pituitary of gonadal tissue).

Advanced education for the role of rural nurse generalist.

The Montana State University College of Nursing has developed a master's degree program which prepares nurses as generalists with advanced knowledge for understanding and addressing rural health care needs. The programs is clear about its goals and objectives and does not attempt to be "all things for all people." The emphasis is on rural nursing, and this emphasis is present in recruiting, teaching, research and publication at the College. Classroom and clinical experiences challenge students to develop a broad range of skills, and most importantly to enhance critical thinking and problem-solving abilities. Since delivering high quality health care in rural areas requires the ability to understand health care from the consumer's perspective, both data collection and clinical experience in rural communities are required. The enthusiasm for rural nursing--practice, teaching and research--displayed by faculty members, alumnae and students is both a major factor in, and an indication of, the program's success.

Selective killing of T cells by immunotoxins directed at distinct V beta epitopes of the T cell receptor.

The potency and specificity of anti-T cell receptor (TcR)-directed immunotoxins were studied in two T cell leukemia lines, HPB-ALL and Jurkat, and in primary T cells. Immunoconjugates were synthesized using anti-CD3, or distinct anti-V beta antibodies cross-linked to CRM9, a binding site-mutant of diphtheria toxin. All TcR-expressing cells display the CD3 complex on the plasma membrane. HPB-ALL cells express the V beta 5 gene product in the beta subunit of the TcR, while Jurkat cells express V beta 8. V beta expression in primary T cells isolated from buffy coats is heterogeneous. Primary T cell populations expressing specific V beta epitopes in the TcR were generated by plating CD3+ T cells on V beta-specific antibody-coated flasks or by positive immunomagnetic selection. Immunotoxins directed against the invariant CD3 epsilon epitope target and kill all T cells. Immunoconjugates targeted at distinct anti-V beta epitopes are specific for cells that express the corresponding gene product in the TcR. The results demonstrate the ability of anti-TcR-based immunotoxins selectively to kill T cells with defined V beta epitopes. These reagents may be clinically useful in disorders mediated by autoreactive T cell populations exhibiting V beta restriction and in the treatment of clonal TcR-expressing lymphomas.

Cytotoxic autoantibodies to beta cells in the serum of patients with insulin-dependent diabetes mellitus.

We studied serum from 36 patients with insulin-dependent diabetes mellitus (IDDM) for the capacity to lyse beta cells. Immunofluorescence revealed an islet-cell cytoplasmic antibody (ICA) in 20 patients with IDDM and an islet-cell-surface antibody (ICSA) in 23. Neither ICA nor ICSA was found in any of 21 normal controls or 15 patients with non-insulin-dependent diabetes. In the presence of complement. ICSA-positive serum caused significant lysis as measured by release of 51Cr (50.1 +/- 8.8 per cent) from cultured rat islet cells, but ICSA-negative serum did not (17.7 +/- 7.3 per cent) (P < 0.001). Proof that ICSA-positive serum was lytic for beta cells was obtained by a double-fluorescence technique that identified lysed cells by their capacity to take up ethidium bromide and beta cells by their staining with fluorescein-conjugated antibody to insulin. These findings suggest that cytotoxic ICSA contributes to the pathogenesis of IDDM, but the mere presence of ICSA does not appear to be sufficient to produce diabetes; family studies showed that one fourth of the serum samples from nondiabetic first-degree relatives of diabetic probands were ICSA-positive and cytotoxic for beta cells.

Epstein-Barr virus-transformed lymphocytes produce monoclonal autoantibodies that react with antigens in multiple organs.

Peripheral blood lymphocytes from normal individuals and patients with autoimmune abnormalities such as insulin-dependent diabetes mellitus and thyroiditis were infected with Epstein-Barr virus, and the culture supernatants were tested for autoantibodies that reacted with normal tissues. Between 58 and 86% of Epstein-Barr virus-transformed cultures produced immunoglobulin M antibodies, and between 9 and 24% of the transformed cultures produced immunoglobulin G antibodies that reacted with normal tissues. Ten Epstein-Barr virus-transformed clones secreting human immunoglobulin M monoclonal autoantibodies were isolated. Four of these monoclonal autoantibodies were studied in depth and found to react with antigens in multiple organs, including thyroid, pancreas, stomach, smooth muscle, and nerves. It is concluded that Epstein-Barr virus can trigger the production of autoantibodies without infecting the target cells to which the autoantibodies are directed.