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Biomedical subjects

F Manca

Publications and source records attributed to F Manca.

At least 91 records · Page 5Linked to original sources
PubMed1987

Allospecific rat-mouse T hybridoma clones generated from kidney graft-infiltrating cells.

In order to dissect the cellular mechanisms that lead to allograft rejection, infiltrating cells can be isolated and expanded in vitro for functional assays. Since the maintenance of these lines and clones is time-consuming, and large numbers of specific cells cannot be easily obtained, we fused the graft infiltrating cells, after in vitro specific expansion, with the murine T lymphoma BW5147. The rat-mouse TT hybridomas thus generated were screened for antigen-dependent interleukin 2 production, for antigen-dependent polyclonal helper activity, and for surface phenotype. The high frequency of specific clones obtained indicates that this is a convenient approach to generate alloreactive hybridoma clones with specific functions from the inflammatory infiltrates of rejected grafts.

Animals↗
PubMed1987

Frequency and functional characterization of specific T-helper cells infiltrating rat kidney allografts during acute rejection.

T-helper cells (ThC) play an important role in the induction of both cytotoxic T-cell responses and B-cell responses against the grafted organ. Furthermore, ThC alone are capable of causing graft rejection in T cell-deprived mice and rats. In view of these observations we found it important to analyse the frequency and functions of donor-specific ThC in the allograft and in the recipient lymphoid system during the course of acute renal allograft rejection. A limiting dilution assay was developed which, due to the absence of exogenous interleukin 2 (IL-2) and the low numbers of stimulator cells used, appears to be highly selective for the proliferation of specific ThC. Kidney transplants were performed from LBN (RT1n) to congenic Lewis (RT1l) strain differing in major histocompatibility complex (MHC) only. The inflammatory (white) cells were recovered from the graft, and blood and recipient spleen and the frequency of RT1n-responding ThC were determined at different times after transplantation. In the kidney graft itself, the frequency of ThC responding to RT1n MHC antigens was 1:3000 on day 2 and increased to 1:670-1320 at the peak of inflammation. In the spleen, the frequency increased from 1:1000 on day 0 to 1:200 on day 8, and remained high even after the graft was rejected. In the blood, the frequency stayed at the 1:400-1:800 level, and increased to 1:200 only after the graft had been completely destroyed. Individual ThC clones deriving from limited dilution assays of kidney and spleen cells were recovered and expanded with irradiated donor cells without IL-2 and finally with exogenous IL-2 only. All clones showed the T-helper (W3/25) phenotype, seven out of eight tested clones showed a specific anamnestic response to RT1n alloantigens and no response to RT1l or RT1a in a secondary MLC, 12 out of 12 clones produced IL-2 and 11 out of 11 clones produced gamma interferon upon re-stimulation with relevant allogeneic cells, and eight out of ten clones collaborated with syngeneic B cells for Ig synthesis, indicating that they were indeed derived from specific ThC and/or from their precursors. Taken together, the results demonstrate that specific ThC and/or their precursors represent only a very small minority in the graft-infiltrating inflammatory population. This makes it most unlikely that the ThC themselves are responsible for graft destruction; the results indicate rather that a major role of ThC in situ may be instruction of immunologically specific and nonspecific components of inflammation.

Animals↗
PubMed1987

Tuberculous pleural effusions. Evidence for selective presence of PPD-specific T-lymphocytes at site of inflammation in the early phase of the infection.

Patients with tuberculous pleural effusions may show cutaneous anergy to tuberculin purified-protein derivative (PPD). This phenomenon has been attributed either to preferential sequestration of antigen-specific T-lymphocytes to the pleural space or to the presence of suppressor monocytes in the blood. In 2 patients with primary tuberculous infection involving the pleura and with skin anergy to PPD, the in vitro proliferation of pleural and peripheral blood T-cells to PPD was evaluated. Although peripheral blood T-cells were not reactive, pleural T-cells showed a marked proliferative response to PPD. At least in the first patient, the lack of proliferation of circulating T-cells could not be related to the presence of suppressor monocytes. Interestingly, after 4 to 8 wk of specific chemotherapy, PPD skin test became positive and the blood T-lymphocytes responded in vitro to the same antigen. Pleural T-lymphocytes were used to generate long-term, PPD-specific T-cell lines and could be maintained in vitro for more than 3 months with repeated cycles of stimulation. The pleural and the blood T-lymphocytes and the T-cell lines were also characterized phenotypically: although the majority of the T-lymphocytes present in the pleural space after Mycobacterium tuberculosis infection were Leu-3-positive (helper T-cells), T-cell lines proliferating in response to PPD included high numbers of Leu-2-positive cells (suppressor/cytotoxic T-cells). These data suggest that early skin anergy in tuberculous pleurisy may be associated with sequestration of PPD-reactive T-lymphocytes in the pleural spaces involving both Leu-2-and Leu-3-positive T-cells.

Adolescent↗
PubMed1986

Difference in sensitivity to cyclosporine in vitro of human alloreactive lines and clones.

The effectiveness of cyclosporine (CsA) as immunosuppressive agent in human kidney graft rejection is well established. However, in spite of efforts to maintain optimal plasma levels, a fraction of transplanted patients undergo rejection episodes and/or irreversible chronic rejection. This suggests that immunosuppression by CsA cannot control the alloreactive response if there is a high degree of histoincompatibility for HLA or non-HLA antigens, or it has little effect on the "high responder" patient. Both possibilities are difficult to test in the human system. A third hypothesis, the existence of individual CsA resistance, was tested by evaluating the in vitro inhibitory activity of CsA on alloreactive T cell lines from several individuals. A different degree of in vitro sensitivity to the drug was observed among alloreactive lines generated from different individuals and among clones obtained from the same bulk line. The variability at the individual level and at the clonal level may account for the onset of CsA-resistant rejection assuming that in vivo a positive selection in the presence of the drug occurs and allows for the resistant clones, if present, to dominate the sensitive ones.

Cell Line↗
PubMed1985

Constraints in T-B cooperation related to epitope topology on E. coli beta-galactosidase. I. The fine specificity of T cells dictates the fine specificity of antibodies directed to conformation-dependent determinants.

Experiments to test the relationship between the epitopes on a protein antigen recognized by T and B cells in their collaboration to produce antibody cannot rely solely on hapten-carrier models. In the present work we used E. coli beta-galactosidase, a molecule whose tertiary and quaternary epitopes have been well characterized, as the model antigen. T helper cells were raised by stimulating mice with the intact or the denatured molecule or with any of several beta-galactosidase cyanogen bromide peptides. In a series of in vitro helper T cell assays we confronted the various T populations with B cells preimmunized with the native antigen, and we tested their capacity to help production of (a) binding antibodies and (b) antibodies directed to single conformational epitopes, characterized by their capacity to protect the enzyme from heat denaturation or to activate defective beta-galactosidase. According to our results, (a) equivalent T cell help can be provided by T helper cells primed with native or denatured antigen, even for the production of "conformational" antibodies; (b) one of the peptides (CB-18) is most efficient in raising help for binding antibodies; and (c) two peptides (CB-20 and CB-21) rank highest in priming T helper cells for the eventual production of protecting and activating antibodies, respectively. Thus, not every beta-galactosidase-specific T helper cell is useful in providing help to B cells specific for any particular epitope on the molecule, but rather preferential pairings exist, possibly governed by a proximity rule.

Animals↗
PubMed1985

Physico-chemical and thermodynamic properties of monomeric concanavalin A.

An alkylated monomer of Concanavalin A was prepared photochemically according to the method of Tanaka et al. (1981). Its affinities for methyl-alpha-D-gluco, methyl-alpha-D-manno and p-nitrophenyl-alpha-D-manno pyranoside were calculated. The enthalpies of these binding reactions were measured calorimetrically and the thermodynamic parameters were calculated. The values obtained suggest that the structure of the monomer differs from that of the dimeric and tetrameric molecules. Calorimetric studies also showed that the monomeric derivative reacts with IgM but not IgG. The enthalpy per binding site in the monomer-IgM reaction is equal to that of the monomer-mannose derivative reaction; mannose is the terminal residue of the saccharide chains of the IgM molecule. The stoichiometry of the reaction is ten ConA-m per IgM molecule.

Amino Acids↗
PubMed1985

Inhibition of the accessory function of murine macrophages in vitro by cyclosporine.

The accessory function of macrophages, which is strictly related to the induction of T cell activation, has been studied to determine whether it is affected by cyclosporine (CsA). Irradiated spleen cells, used as a source of macrophages, were pulsed overnight with beta-galactosidase (GZ) in the presence of CsA. After washing of the pulsed macrophages, cells from a GZ-specific T cell line were added to cultures and 3H-thymidine incorporation was measured 72 hr later. We found that 500 ng/ml CsA present during macrophage pulsing with GZ reduced T cell proliferation to 5%. On the other hand, 100 ng/ml CsA almost completely abrogated the proliferative response when present for the duration of the culture. Similar results were also obtained using antigen-pulsed peritoneal-adherent macrophages to stimulate the T cell line to proliferate, or a T hybridoma clone to produce interleukin-2 (IL-2). The possibility that CsA actually affects interleukin-1 (IL-1) production by macrophages by inhibiting uninvolved T cells could be ruled out. We conclude that CsA-induced inhibition of T cell functions (proliferation and IL-2 production) is partially due to the effect of the drug on the accessory function of macrophages. This immunosuppressive mechanism of action of CsA on macrophages has not been previously described.

Animals↗
PubMed1985

[Value of splenic embolization in patients with hypersplenism].

The authors describe the importance of embolization of the splenic artery by means of Gianturco coils in splenomegalic portal hypertension. They propose their first experience of six cases and the good results on the basis of clinical parameters, of bleeding of esophageal varices, of hematological values. They also show the influence of this method on hemodynamic, metabolic, and respiratory parameters, altered in cirrhotic patients.

Adult↗
PubMed1984

A simple procedure for freezing and storing lymphocyte panels in trays.

Lymphocytes from different donors may be frozen in Terasaki trays by a simple procedure. This method provides the tissue typing laboratory with a readily available panel that can be routinely used to test for lymphocytotoxic antibodies sera from patients waiting for kidney transplant, transplanted patients and polytransfused individuals. The advantage of this procedure is that expensive programmed freezing apparatus is not required.

Blood Preservation↗
PubMed1984

Calorimetric study of rheumatoid factor binding to human IgG.

The binding of human IgM monoclonal rheumatoid factor (RF) to native human IgG was studied in a flow calorimeter. Thermal titration gave a enthalpy of binding of 5.5 X 10(4) J/mole site for the reaction, with 1 mole of RF binding to 2.5 moles of IgG; this corresponds to one RF subunit per IgG heavy chain. Similar experiments performed with heat-aggregated human IgG gave almost identical results. If the IgG molecule undergoes a conformational change upon heating, this is not such as to affect the enthalpic contribution of binding.

Antigen-Antibody Reactions↗