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PubMed1977

The derivation and characterization of neuronal cell lines from rat and mouse brain.

This study shows that permanent cell lines can be established from rat and mouse brain by direct tissue culture methodology without the aid of exogenous chemical or viral transforming agents. These cells were derived from specific areas of the brain, such as the cerebellum and hippocampus, at chosen times during fetal and neonatal development. Success in establishing neuronal cell lines was dependent upon the use of selection pressures designed to keep the background of glial cells and fibroblasts at a minimum. These manipulations included care in the choice and processing of the original tissue, utilization of cytotoxic anti-glial sera, and continuous manual isolation of cells with neuronal morphology. Slow-growing nerve cells were thus allowed to adapt spontaneously to culture with a minimum of competition from faster-adapting cell types. Many of these cell lines are judged to be neuronal on the basis of their electrical excitability and their characteristic surface antigens. The cells respond positively in a sodium flux assay which has been shown to correlate well with the ability to generate an action potential, and also express one or more of three antigens previously found to be specific for nerve cells.

Animals↗
PubMed1979

Mixed agglutination in microtiter U-trays.

The principle of mixed agglutination with cell cultures was applied to those cells which could not be obtained as monolayer cell cultures. Artificial monolayers of freshly isolated cells from various tissues were prepared in poly-L-lysine-coated wells of a microtiter U-tray. By means of this technique, murine alloantigens, H-2 private specificities and Thy-1 antigens were demonstrated on cells from thymus, brain, spleen, kidney and liver. Ia.2 antigen was detected on splenocytes but not on thymocytes or fibroblast L cells. Evidence was also presented that HLA A and B locus antigens could be demonstrated on human thymocytes. Sensitivity of this test was shown to be at least 100 times higher than that of the standard lymphocytotoxicity test for detection of HLA antigens.

Absorption↗
PubMed1977

Recirculating, suppressor T cells in transplantation tolerance.

An adoptive transfer system was used to examine the capacity of cellular inocula from rats fully tolerant of Ag-B antigens to transfer tolerance to irradiated recipients. Permanent tolerance in these irradiated recipients involved specific suppression of the regenerating immune response. Cells obtained from tissues rich in recirculating lymphocytes were the most effective suppressors. Highly purified inocula of T cells from tolerant donors were potent suppressors in irradiated hosts, but were not capable of direct suppression of peripheral antigen-sensitive T cells.. The role of the thymus in maintaining the complement of recirculating suppressor T cells in tolerant animals was examined after adult thymectomy. Thymectomized tolerant rats did not reject their tolerated grafts, and the longevity of the suppression in tolerant rats was confirmed by showing that adoptive transfer of cells from thymectomized tolerant donors was effective in suppressing irradiated recipients up to 180 days after thymectomy. Cellular inocula from these donors appeared to lose their suppressor function marginally faster than they lost effector function (as measured by their capacity to mediate rejection of third party control grafts). Thymectomy made tolerant rats more vulnerable to the termination of tolerance by challenge with normal cells. Transplantation tolerance is maintained in adult rats by long-lived rapidly recirculating suppressor T cells. The target for the suppressor action of these cells is probably the precursor of alloantigen-sensitive lymphocytes, and the effect of suppression may be deletion or inactivation of the relevant clone of these cells.

Animals↗
PubMed1979

Binding of purified, soluble major histocompatibility complex polypeptide chains onto isolated T-cell receptors. I. Reactivity against allo- and self-determinants.

In this study, we tried to get information about the fine antigen-binding ability of purified, soluble, idiotype-positive T-cell receptor molecules. Lewis anti-DA T-cell receptors were purified from normal Lewis serum by the use of anti-idiotypic immunosorbent and sodium dodecyl sulfate-polyacrylamide gel, and were coupled to cyanogen bromide-activated Sepharose 4B. In parallel, Lewis anti-DA, Lewis anti-BN, and DA anti-Lewis alloantibody immunosorbents were prepared. The major Ag-B chain (44,000 daltons) and the two polypeptide chains (34,000 and 27,000 daltons) of Ia were purified from Lewis, DA, and BN lymphocytes and absorbent on the above-mentioned immunosorbents. We found that the major Ag-B chain as well as the two Ia chains were bound to the alloantibody columns if they were derived from the corresponding allogeneic strain. No retaining ability for self-major histocompatibility complex (MHC) or third-party MHC chains was noted with the alloantibody immunosorbents. When using immunosorbents made up of idiotypic T-cell receptors, only two MHC polypeptides of the relevant allo-MHC type were retained, namely, the Ag-B and the heavy Ia chains. No detectable activity was observed when testing the same column for reactivity against third-party MHC polypeptide chains. However, the Lewis anti-DA T-cell receptors could be shown to display weak, but significant, reactivity toward one Lewis MHC polypeptide chain, that is, the heavy chain of Ia type.

Animals↗
PubMed1977

In vivo and in vitro effects of acute graft-versus-host serum in the rat.

Pooled serum from 6- to 8-week-old female Lewis x Brown Norwegian F1 hybrid (LBNF1) rats undergoing acute local graft-versus-host (GVH) reactions was found to have the ability, when compared with normal F1 serum, to increase node weight in a popliteal lymph node weight gain assay in syngeneic animals. Suspension of donor cells in normal F1 serum on one side and corresponding concentrations of GVH serum on the contralateral side resulted in an average increase in popliteal node weight of 28.8 per cent on the side receiving GVH serum. Of the 64 animals tested, 50 had larger nodes on the GVH serum side. This effect was not impaired by heating (56 C, 30 min) or by rapid freezing (-75 C) and thawing of the serum. Preinjection of the serum into the footpads 2 hr before donor cells also gave increased node weights as did injection of the serum i.p. In contrast, GVH serum was found to inhibit unidirectional mixed lymphocyte cultures of seven different allogeneic rat strain combinations, the inhibition ranging from 35 to 74 per cent. In addition, the blastogenic cell responses of three different strains to phytohemagglutinin P were inhibited between 45 and 77 per cent. The relationship between these two phenomena, in vivo increase in node weight and in vitro inhibition of blastogenesis, is not clear and is the object of further investigation.

Animals↗
PubMed1977

Mixed leucocyte interaction suppression generated after alloimmunization.

Utilizing a bone marrow cell immunization protocol designed to actively enhance renal or cardiac allografts, the in vitro mixed lymphocyte interaction has been studied. After immunization with allogeneic cells over a wide dose (10(5) to 10(8)) and time range, the mixed lymphocyte interaction using peripheral blood lymphocytes is reduced, and this reduction is shown in mixing experiments to be attributable to a circulating suppressor cell that has adherence characteristics of T lymphocytes. Specificity for in vivo generation of these circulating cells as well as specificity in the in vitro reaction was shown, although a nonspecific suppression background was observed. Allogeneic thymocytes, splenocytes, and platelets were effective in immunizing to generate these cells also. There was no general correlation with the ability to generate these cells and to actively enhance cardiac allografts, since suppression was seen after alloimmunization in strain combination that cannot be actively enhanced.

Animals↗
PubMed1978

Transplantation unresponsiveness induced by allogeneic serum in combination with allogeneic cells and hydrocortisone in adult rats.

Attempts were made to optimize the treatment by using serum as antigen source for the induction of transplantation unresponsiveness in adult rats. With the scheme described it was possible to induce by serum injections the transplantation unresponsiveness not only in the week non-H-1, but also in the strong H-1 antigenic difference; this shows that there is no essential difference in the presence of H-1 and non-H-1 antigens in the serum. Allogeneic serum in conjunction with allogeneic cells was most effective in the non-H-1-different strain combination, whereas the combined treatment consisting of allogeneic serum, allogeneic cells, and hydrocortisone produced the longest skin graft survival in the H-1 antigenic difference. Some paradoxical results are discussed.

Animals↗
PubMed1978

Effect of azathioprine and prednisolone on passive enhancement of rat renal allografts.

Passive enhancement provides only partial suppression of rejection in the (DA X Lewis)F1 to Lewis renal allograft model. Suboptimal (8 mg/kg/day) and supraoptimal (30 mg/kg/day) doses of azathioprine administered with enhancing serum failed to suppress the rejection reaction in enhanced animals. Similarly, suboptimal (4 mg/kg/day) and optimal (16 mg/kg/day) doses of methylprednisolone were ineffective. However, the onset of rejection in enhanced animals was delayed by the use of both azathioprine (30 mg/kg/day) and methylprednisolone (16 mg/kg/day). The survival times of enhanced animals treated with azathioprine were significantly shorter than those of animals treated with enhancing serum alone, suggesting that this agent may prevent the development of autoenhancement. Although suboptimal doses of antilymphocyte serum suppress rejection in this enhancement model, the dose requirements of conventional immunosuppressive agents appear to be maximal rather than minimal.

Animals↗
PubMed1978

Non-major histocompatibility system immunogenetic influences in rat renal allograft survival.

H-1-incompatible rat renal allograft survival rates are strongly influenced by the non-major histocompatibility system (MHS) genes. The same H-1 (MHS) incompatibility tested on four different non-MHS backgrounds gave widely differing results ranging from 11 to 60% fractional survival between the best and the poorest (P less than 0.01) at 105 days. These results can best be explained if non-H-1 and intra-MHS amplifiers and/or suppressors are invoked.

Animals↗
PubMed1978

Prolongation of rat cardiac allograft survival by donor pretreatment. Screening of antineoplastic drugs.

We have made preliminary investigations into the effect of 33 different drugs on the survival of rat cardiac allografts, the drugs being administered to the allograft donor. All drugs were administered at LD50 i.v. to the graft donor 6 hr prior to removal of the organ. Especially effective were alkylating agents and antimetabolites. Pretreatment with cyclophosphamide, busulfan, methotrexate, azauridine, or bromodeoxyuridine prolonged the survival from 7 to more than 20 days. Pretreatment with chlorambusil, mannomustine, mannosulfan, 1-(2-chlorethyl)-3-cyclohexyl-1-nitrosourea, DTC, fluoruracil, or hudroxyurea prolonged the survival to more than 14 days. Several other alkylating agents and antimetabolites prolonged the survival moderately, i.e., to approximately 10 days or more. Purine antagonists, mercaptopurine and azathioprine, were totally ineffective as were also anticancer antibiotics and vinca alkaloids. Pretreatment with procarbazine or methylprednisolone alone increased the survival only moderately, whereas pretreatment with both of these drugs together increased the survival up to 27 days.

Alkylating Agents↗