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Biomedical subjects

N Zavazava

Publications and source records attributed to N Zavazava.

At least 37 records · Page 2Linked to original sources
PubMed1996

Circumvention of natural killer cell and T-cell mediated allogeneic target killing with tacrolimus (FK506) in small bowel transplantation related graft-vs-host disease.

The role of simultaneous donor-specific transfusion of unprocessed cellular bone marrow (BM) together with solid organ transplantation, a postulated concept to achieve long-term graft acceptance, was investigated in an experimental setting of semiallogeneic transplantation of parental small bowel (SBTx) to F1 hybrids. The established graft-vs-host (GvH) model revealed that simultaneous transfer of SB/BM substantially enhanced GvH-mediated immune responses in recipient target organs, e.g. skin, gut, and liver. In comparison to isolated SBTx, animal survival decreased from 16.1 (+/- 0.9) to 10.1 (+/- 0.8) days after additional BM transfusion, P < 0.001. Severe tissue injury of GvH-susceptible target organs in the setting of simultaneous SB and BMTx was associated with significant changes in recruitment and tissue distribution of NKR-P1+ cells during the GvH-related proliferative immune response. Tacrolimus effectively suppressed these initial events and prevented recipient animals from clinically and histologically observed damage caused by GvH disease.

Animals↗
PubMed1996

In vivo depletion of NKR-P1 positive cells in the recipient prior to small bowel transplantation enhances graft-versus-host disease (GvHD) in the rat.

Recent evidence for major histocompatibility complex (MHC) class I antigen-directed recognition mechanisms of natural killer cells (NKs) have revived interests in investigating non-adaptive immune responses in the framework of solid organ transplantation. A semi-allogeneic rat model of heterotopic small bowel transplantation (HSBTx) from male DA parental to male F1 hybrid rats (DA x LEW) was established to investigate the role of host NKs to attenuate graft-versus-host (GvH)-mediated immunosuppression and tissue injury. By use of anti-NKR-P1 monoclonal antibody (mAb) 3.2.3, host NKs were depleted effectively in vivo after triple intraperitoneal injection prior to HSBTx. In contrast to non-depleted animals, an initial lack of NK activity in F1 hosts significantly decreased the mean survival (P < 0.01) and substantially enhanced graft-versus-host disease (GvHD)-related damage to lymphoid and non-lymphoid target organs. These findings emphasize the important immunoregulatory role of host NKs during the early onset of GvHD.

Animals↗
PubMed1996

Rat MHC class I peptides are immunogenic.

We postulated that indirect recognition of MHC-derived peptides might modulate the alloresponse to donor antigen. In this study, we looked at the potential of two class I peptides derived from the alpha 1 and alpha 2 regions of the DA RT1Aa molecule. Lew responder rats were immunized by varying concentrations of two 25mer peptides covering residues 56-81 and 96-120. The injections were under the footpad and were repeated on day 14. The thickness of the footpads was measured to control delayed-type hypersensitivity (DTH). The animals were sacrificed on day 16 and the splenocytes were tested in mixed lymphocyte culture as responders against DA stimulator cells and CAP third-party splenocytes. In addition, the phenotype of the cells was measured using flow cytometry with antibodies against CD4, CD8, CD5, MHC class II, CD25, CD14 and CD19. Peptide 96-120 induced strong sensitization of the Lew recipient animals at concentrations of 200-500 micrograms (n = 4). The stimulation index was 2-3 times higher than that of untreated animals. Peptide 56-81 failed to induce sensitization at the concentrations used, but surprisingly induced a concentration-dependent immunosuppression that was highest at 400 micrograms (n = 4). In proliferation experiments responder Lew rats proliferated only to peptide 56-81 in vitro.

Amino Acid Sequence↗
PubMed1996

Determination of HLA-B7 T-cell epitopes.

A series of peptides derived from the alpha 1 and alpha 2 regions of the HLA-B7 and HLA-A2 molecules was synthesized with an automatic peptide synthesizer using the FMOC (9-fluorenylmethoxycarbonyl) technique. Peptides were analyzed and purified by reversed-phase high pressure liquid chromatography (HPLC). Peptide purity was > 96%. The effect of B7 peptides on mixed lymphocyte cultures was tested in vivo. The alloresponse in the cultures with B7 peptides was strongly enhanced. The peptides that were most effective inhibited cytotoxic T-lymphocyte (CTL) cytolysis. The data show that the peptides are immunogenic and that they are recognised by both the direct and indirect pathways. Further, the mechanism of peptide recognition was studied. We coupled peptide B7 (residues 62-70 in HLA-B7) and peptide A2 (residues 62-70 in HLA-B2) covalently to fluorescein isothiocyanate (FITC). B7-specific CTLs were incubated with these peptides at 37 degrees C for 90 min and cell fluorescence measured by flow cytometry. The B7 peptide was bound by 60% of the CTLs whereas the A2 peptide, as a negative control, bound only 10%. The molecular size of the ligands to which the peptides bind are being characterized by immunoprecipitation.

Amino Acid Sequence↗
PubMed1996

Cross-matches on donor cadaver retinal pigment epithelial cells in corneal risk patients.

BACKGROUND: Allografts can be rejected either through the antibody-mediated or cellular pathways. The objective of this study was to look at the extent of antibody formation in patients awaiting re-keratoplasty using cross-matches on cadaver retinal pigment epithelial (RPE) cells. METHODS: Cadaver RPE cells were derived by trypsin digestion from donor eyes (n = 1200). After 3 days of cell cultivation, the cells were adherent and began to lose their pigment. By day 7 most cells were clear and grew as a polygonal monolayer. MHC class I expression by RPE cells was studied by the W6/32 (anti-HLA-A, B, C) monoclonal antibody (MoAb) and that of class II (HLA-DR) by the 136 MoAb. Normal RPE cells express few class I and no detectable class II antigens. For the induction of MHC expression, cells were subsequently stimulated with 250 U/ml of recombinant gamma-interferon for 5 days. Cells were used for tissue typing and also for cross-matches with recipient serum. Cross-matches were subsequently performed and measured by flow cytometry. RESULTS: Both class I and class II antigens were strongly enhanced, as could be shown by immunohistochemical staining. Some 20% of those patients awaiting rekeratoplasty (n = 60) were positive for anti-HLA antibodies. In one case anti-DR3 antibodies were detected in a recipient who had had several rejection episodes after keratoplasty. CONCLUSIONS: RPE cells are not only useful for cadaver post-mortem HLA typing but also for donor-specific cross-matches. The degree of antibody formation after keratoplasty in rejecting patients was, however, low. This may imply that anti-HLA antibodies are not the major cause of corneal graft loss after keratoplasty.

Antibodies, Monoclonal↗
PubMed1996

Correlation between graft-versus-host induced immunosuppression and host natural killer cell activity in small bowel transplantation.

The occurrence of graft-versus-host disease (GvHD) following small bowel transplantation (SBTx) can be tuned by the recipient's initial natural killer (NK) cell activity, which modifies the immunogeneic balance between donor and host immunocompetent cells. This study was aimed to investigate the role of host NK cells on the incidence and severity of GvHD following SBTx. Intraperitoneal administration of 50 microl ascites fluid of the highly specific anti-NKR-P1 monoclonal antibody (mAb) 3.2.3 into F1 recipient animals on three consecutive days prior to SBTx was performed to suppress NK activity in F1 hybrids. In vivo treatment with 3.2.3 mAb effectively depleted recipient NK activity for at least 10 days in spleens and mesenteric lymph nodes of F1 hosts. In contrast to nontreated F1 recipients, all 3.2.3 mAb-pretreated F1 animals suffered from severe signs of GvHD, and the mean survival time was decreased significantly from 16.0 +/- 0.9 days to 11.0 +/- 0.8 days (p < 0.01) in nontreated and NKR-P1-depleted F1 animals, respectively. Other sequelae included earlier onset of GvH manifestations, pronounced damage of primary and secondary lymphatic organs, substantial increase in spleen index, and lower CD4(+)/CD8(+ )ratios over the course of progressing GvHD. Our results underline the important immunoregulatory role of NK cells as a first defensive line acting on the alloreactivity of donor-derived immunocompetent cells in this model of solid organ transplantation.

Animals↗
PubMed1996

Soluble HLA class I molecules induce apoptosis in alloreactive cytotoxic T lymphocytes.

Soluble HLA class I molecules (sHLAs) have been identified in the serum of patients with inflammatory diseases, allografts and autoimmune diseases and in serum of healthy individuals. The biological significance of these molecules, particularly after allogeneic organ transplantation, has been enigmatic. Here we show that primary alloreactive CD8+ T cells interact with sHLA and undergo apoptosis in the absence of a second signal. Ligation of CD28 rescued T cells from death, implying that sHLAs induce apoptosis through selective stimulation of the T-cell receptor. CD95-L was upregulated after cytotoxic T lymphocytes were incubated with sHLAs, and cell death was blocked by a neutralizing anti-CD95-L antibody, suggesting that sHLAs induce endogenous mutual killing of activated T cells. These results provide a molecular basis for the capacity of sHLAs to downregulate T-cell responses, which may be especially relevant to organ transplantation.

Apoptosis↗
PubMed1995

Porcine valves are reendothelialized by human recipient endothelium in vivo.

The degeneration of human allogeneic and porcine xenogeneic heart valves has not been clearly understood. The question is whether the observed loss of function and calcification is primarily an immunologic process or a mechanical process or is influenced by both factors. In the current study, we looked at explanted xenogeneic heart valves for the presence of recipient endothelium. Explanted valves were shock frozen and stored at -80 degrees C before use. They were subsequently examined by immunohistochemical staining with a variety of monoclonal antibodies. Xenogeneic valves showed clearly positive results for the human major histocompatibility complex class I and class II antigens and morphologically showed a thin layer of viable endothelium restricted to the annular region of the valve. Additionally, they were also positive for intercellular adhesion molecule-1 and the H-Y antigen. Although the xenogeneic valves were significantly degenerated, the endothelium was clearly defined and could be identified immunohistochemically as being of recipient origin. The grafts remained negative for endothelial cell-leukocyte adhesion molecule-1 and factor VIII. These data allow speculation on whether reendothelialization of valvular grafts with recipient endothelium is a normal repair mechanism in vivo.

Animals↗
PubMed1994

Soluble MHC class I molecules in human body fluids.

Apart from their well-known function of antigen presentation in the form of peptides, major histocompatibility antigens (MHC) have been found to be unique markers of individual body odors in murine experimental models. In the current study we examined the nature and expression of soluble human MHC class I molecules in body fluids. Biochemical analysis of affinity purified serum class I molecules revealed a variety of molecules within the molecular weight region of 45 to 21 kD. SDS-Western blotting of HLA derived from hepatocytes and spleen cells suggested that much of the small molecular mass fraction of sHLA (< 45 kD) found in serum is derived from the liver. sHLA were detected and quantitated in serum, plasma, urine, cerebrospinal fluid, and sweat. No sHLA were detectable in cerebrospinal fluid (n = 20). In addition, sHLA was measured in serum of women during the menstrual cycle. A significant increase in sHLA was observed during the first half of the cycle, suggesting that sexual hormones may increase sHLA concentration. The observed increase was most prominent in women that were HLA-A24.

Adult↗
PubMed1994

In vitro effects of solubilized HLA-DR--role in immunoregulation?

To test the hypothesis that soluble HLA-DR antigens (sHLA-DR), binding to the T-cell receptor (TCR) and/or CD4 structures, compete with and abrogate functions of their cell-bound counterparts, we studied effects of detergent-solubilized, affinity-purified HLA-DR molecules on the DNA synthesis, IL-2, and IL-1 secretion by human peripheral blood mononuclear cells (PBMC). While resting T cells did not show any response, there was a dose-dependent suppression of T-cell responses induced by mitogen (phytohemagglutinin, PHA), recall antigen (purified protein derivative of tuberculin, PPD), or HLA class II alloantigens (Daudi cells). In the PHA system, sHLA-DR affected DR-identical and DR-disparate PBMC with equal efficiency, suggesting a nonspecific interference with accessory functions of cell-bound HLA class II molecules. A competitive ligation of lymphocyte (rather than of monocyte) CD4 is suggested based on the failure of sHLA-DR (i) to potentiate inhibitory effects of anti-CD4 mAb, (ii) to suppress mitogen responses after depletion of CD4+ lymphocytes and also following separate monocyte (vs lymphocyte) pretreatment, and (iii) to induce a reproducible IL-1 secretion inhibition. In the PPD system, suppressive sHLA-DR effects on autologous or DR-identical PBMC significantly exceeded that on DR-disparate PBMC, and in the MLR, third party allogeneic sHLA-DR was exceeded in its suppressive potency both by sHLA-DA pertinent to responder cells and by sHLA-DR pertinent to stimulator cells. These additional specific effects may result from competition (with cell-bound class II restriction and recognition determinants) at the TCR level rather than from peptide competition at the antigen-presenting cell level. Interference by sHLA-DR with the primary and/or accessory signaling may offer new therapeutic strategies in allotransplantation and autoimmunity.

Dose-Response Relationship, Drug↗