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The GVHRIL foHowing transplantation of small intestine are different from those found after bone marrow transplantation or spleen cell injections in that they show a remarka ble, significant prevalence of lesions within the intestinal mucosa. These findings are consistent with the observation that jntestinal lymphocytes newly formed in mesenteric lymph nodes predominantly home in on the intestine again.& The degree of histologic alteration within different tissues indicates that the graft and the host may survive the lesions of the lymphatic tissues, whereas the severe intestinal lesions following GVHR may easily cause death of the recipient. With regard to clinical sman bowel transplantation two statements can be made: (l) GVHRIL play a significant role in small bowel trans~ plantation. (2) To minimize their biologic importance, a selective elimination of the graft's Jymph nodes by irradiation or surgical resection should be considered in view of the remarkable difference between GVHRIL in lymph nodes and in the graft's intestinal wall itself.
Since systematic hematological studies on blood and bone marrow changes after treatment with 15-Deoxyspergualin (DOS) are lacking, a quantitative assessment was performed fourteen or twenty eight days after intraperitoneal application of DOS to rats. Further observations done 7 and 14 days after discontinuation of DOS administration allowed analysis of banc marrow regeneration. DOS induced lymphocytopenia, granUlocytopenia and anemia with a decrease of bone marrow cellularity due to suppression of cell maturation. The effect was dose-dependent and bone marrow as well as blood changes were observed in animals treated with doses from 0.5 to 10.0 mg/kg DOS. Within 14 days after termination of the treatment, rapid recovery with normalization of all hematological parameters was observed. In the light of our data, these hematological side effects may not be a major disadvantage, if DOS is used in doses below 2.5 mg/kg, and for a course of therapy which is limited to 7 to 14 days.
During the past few years, interest in xenotransplantation of porcine islets of Langerhans for the future therapy of type I diabetes has Increased markedly. Therefore, we established a semiautomated digestion method for isolating islets from the porcine pancreas. However, although the isolation technique was standardized and collagenase of controlled quality was used, we were unable to attain high islet yields with a satisfactory degree of reproducibility. One hypothesis was that varying degrees of interference by donor pancreatic enzymes were responsible for this failure. The aim of this stUdy was to examine the kinetics of four types of enzymatic activity during the isolation procedure, as well as their effects on islet yield: collagenase, trypsin, neutral protease, and clostripaln. Our results indicate that while exogenous collagenase activity decreases slightly during the isolation procedure, the activity of the pancreas enzymes neutral protease and trypsin increases. In some cases, trypsin activity increases very strongly. A strong increase in trypsin activity correlates with poor islet yield, whereas low trypsin activity always correlates with high islet yield. Addition of the protease inhibitor Pefabloc to the isolation medium results in low trypsin activity and reproducible high islet yields.
It is the aim of this study to characterize and quantify the cells within isolated rat islets that express MHC class 11 antigens. A set of five monoelonal antibodies and two polyclonal antisera of defined specificlty were used in combination with a newly devised procedure for three·dimensional immunofluorescence evaluation of intact islets. It is shown that in addition to passen· ger cells, such as Iymphocytes, macro· phages, and dendrlticlike cells, vascular endothelial and endocrine cells are also capable of expressing class 11 antigens. This expression Is strongly influenced by in vitro culture. pregnancy, streptozotocin- induced diabetes, transplantation trauma, and alloantigenic stimuli. The pos· sible role of the above cells in antigen presentation related to islet transplantation is discussed.
Abstract: For isolating islets from the porcine pancreas, we established a semiautomated digestion method. Although the isolation technique was standardized and collagenase of controlled quality was used, until now the reproducibility of high islet yields was unsatisfactory. Our hypothesis was that pancreatic trypsin was responsible for this failure. The aim of this study was to investigate the effect of endogenous trypsin on islet yield. Our results demonstrate that a high trypsin level correlates with poor islet yield, whereas low trypsin activity always correlates with high islet yield. Specific inhibition of trypsin results in low trypsin activity and reproducible, high islet yields.
In the present study, an attempt was made to characterize the immunomodulating abilities of the cytostatic drugs cydophosphamide, ifosfamide, vinblastine, vincristine, procarbazine, dacarbazine, 6-mercaptopurine, methotrexate, 5-f/uor-uracil and adriamycine in a defined experimental model. Varying combinations of drug plus transplantation alloantigen, (C3H-lymphocytes) were injected into Balb/c mice at different time intervals in vivo. The resulting T-effector cell reactivity was determined in vitro with the microcytotoxicity assay on day + 5 for primary (r) and day + 7 for secondary (2°) sensitized mice. According to the type of drug (alkylating agent vs. vinca alkaloid vs. antimetabolite vs. cytostatic antibiotic), the dosage (20% LD50 vs. 60% LD50), the state of sensitization (r vs. 2° sensitized recipients), and the time of drug application in relation to the antigen treatment on day 0 (in varying steps from day -6 to day +4), so-called "pharmaconantigen- variation-effects" (PA VE) were established for each of the investigated drugs in form of reaction profiles. The results were as folIows: (1) For almost alt substances, characteristic reaction profiles involving immunostimulation and/or immunosuppression could be established. Similarities in the profiles of different substances made it possible to classify the drugs according to different reaction types. The reaction type however is not definitely correlated to the biochemical mechanism of drug action. (2) The PA VE are decisively inf/uenced by so me of the biological parameters, such as the time of drug application in relation to the antigen treatment and the state of sensitization but relatively !ittle by the dosage of the drug. (3) Considering the different processes occurring du ring primary and secondary immune responses, the PAVE may give hints for a distinct manipulation of the immunoregulation and thus information on the immunobiological mechanism of drug action.
Because successful human islet transplantation requires large quantities of viable islets that must be separated from the highly immunogenic exocrine tissue and because handpicking is too time-consuming and laborious to be clinically relevant, a new approach for solving this problem has been established in rat models. It is based on the principle that magnetic microspheres (MMSs) coupled to lectins with binding specificity for the exocrine tissue portion are trapped in an electromagnetic field, thus providing effluent islets of a high degree of purity. In this study our aim was to adapt this princip'le to human islet preparations. In this context our prime interest was focused on a lectin suitable for human pancreatic tissue. Of 19 different lectins tested, only 1, Wisteria floribunda agglutinin (WFA), is suitable, as shown by immunofluorescence, MMS-Iectin binding, and magnetic separation
An immunogold-silver enhancement technique, which combines effective labeling of viable isolated islets with the ultrastructural resolution of cytological details, was applied in electron microscopy to identify major histocompatibility complex (MHC) structures on islet cells. Incubation of freshly isolated islets from CAP (RT1C) and LEW (RT1') rats with OX18, an MHC class I antibody, showed strong positive reactivity in macrophages and/or dendritic-like cells (M0-DCs) and vascular endothelial cells (VEs) and a comparatively weaker reactivity in endocrine a-, p-, and 8-ce"s. With MHC class" antibody OX6 (anti-I-A), M0-DCs were strongly labeled in both rat strains on the surface and on internal structures. Three of five particularly high titered batches of OX6 revealed MHC class" expression on VE and p-ce"s. Four days of in vitro culture in combination with a high concentration of glucose and interferon-'Y induced strong enhancement of MHC class I structures and, to a lesser extent, class " structures on p-ce"s.
Total immunoreactive insulin (IRI) is conventionally determined by radioimmunoassays. IR! measurement in rats can be made more sensitive, accurate, and practical, as demonstrated by a new modified enzyme-linked immunosorbent assay (ELlSA). It is characterized by indirect binding of an anti-insulin antibody by an antiglobulin antibody and uses the principle of competitive saturation. In this ELlSA, IRI can be determined in a wide range of concentrations, corresponding to the standards. The standard curve ranges from 100 to 0.049 ng/mllRI (1 ng/ml - 23.4 JLU/ml - 172 pM rat insulin). The statistical analysis shows between- and within-assay coefficients of variation of :515%. Diabetes 37:321-26,1988