1228 ENHANCEMENT OF HUMAN KIDNEY ALLOGRAFTS BY COLD B-LYMPHOCYTE CYTOTOXINS YUICHI IWAKI MIN SIK PARK

PAUL I. TERASAKI RONALD BILLING

Department of Surgery, U.C.L.A. School of Medicine, University of California, Los Angeles, California 90024, U.S.A.

The sera of 233 kidney transplant patients before transplantation were tested by cytotoxicity against a panel of B and T lymphocytes at 5°C and 37°C. The results divided the patients into four groups: those whose sera reacted with B lymphocytes at 5°C only; those reacting with B lymphocytes at 5°C and 37°C; those reacting with T lymphocytes at 37°C; and those with no antibodies. The patients with pre-transplant antibodies reactive with B lymphocytes at 5°C had a significantly higher kidney-transplant survival rate at 6 months (70%) and 1 year (65%) than patients who had no antibodies (47% and 46%, respectively). Patients with antibodies reactive at 37°C had a 6-month survival-rate of 38% when reactive against B cells and 43% when reactive against T lymphocytes. The cold cytotoxins were IgM.

Summary

Introduction ALTHOUGH allograft enhancement in animals is well established,1 its role in human transplantation is unclear. Leucocytes from the kidney donor were injected into 4 graft recipients in 1973 by Newton and Anderson2 but none showed evidence of immunisation. If we use "enhancement" in its broadest sense-i.e., induction of lower responsiveness by pretreatment with allogeneic cells-blood-transfusions may be thought to induce enhancement. Without blood-transfusions and with immunosuppressive treatment, approximately 30% of kidney allografts survive a year after transplantation,3 but this can be improved to 50-70% by pre-transplant transfusion of allogeneic blood. If this is considered to be enhancement, it has been achieved since human kidney transplants began. In the early days of the operation most patients were given numerous transfusions before transplantation,4 and the survival rates of recent years may be lower because transfusions have been less fre-

grafts

at

twenty-seven

transplant

centres

between

ranged from 3 months to 2 years. Methods

Lymphocytotoxicity B and T lymphocytes were isolated and tested by the lymphocyte microcytotoxicity test as previously described.6 A serum was tested against B and T lymphocytes from 17-30 normal individuals. Cells from

one

person

were

as B-warm-reactive. Sera classified as T-warm-reactive were those which killed T lymphocytes from >15% of the panel at 37°C whether or not they had cold or warm reactivity or killed T lymphocytes at 5°C. Fig. 1 shows examples of these three patterns of reaction. It should be noted that there are fifteen different patterns of reaction possible by testing sera against B and T cells at 5°C and 37°C. However, the B-cold, B-warm, and T-warm reactive patterns and the absence of lymphocyte reactivity were the most common.

Serum Fractionation 2 ml of B-cold-reactive serum was fractionated by gel filtration on ’Biogel 5M’. 2 ml fractions were eluted in 0-15 mol/1 sodium chloride, 10 mmol/l phosphate buffer (pH 7.2). The fractions under peaks with optical density 280 nm were pooled, concentrated to 2 ml, and tested for IgM or IgG by Ouchterlony immunodiffusion using goat antibodies against human IgG and IgM. There was a clear separation of the IgG and IgM fractions. Each fraction was tested for cold anti-Bcell activity against at least three different B-lymphocyte preparations at 5 IC and 37°C.

Results Of the 233 pretransplant sera tested, 139 (60%) did not react against T or B lymphocytes at 3°C or 37°C. 94 sera had anti-lymphocyte activity, 40 with the

If the state of enhancement could be detected by invitro tests, patients who were properly "prepared" by transfusions and pregnancies could be identified. We describe here a test which may detect enhancing antibodies.

Patients

patients

were

233

recipients of cadaver kidney allo-

Harland, P. S. E. G. in The Child in the African Environment (edited by R. Owor, V. L. Ongom, and B. G. Kirya); p. 349. Kampala, 1975. 22. Kirchner, H., Rühl, H. Exp. Cell. Res. 1970, 61, 229. 23. Rühl, H., Kirchner, H., Bochert, G. Proc. Soc. exp. Biol. Med. 1971, 137, 21.

1089. 24. Williams, R. O., Loeb, L. A. J. Cell Biol. 1973, 58, 594. 25. Chesters, J. K. Biochem. J. 1975, 150, 211 26. Phillips, J. L., Azari, P. Cell. Immun. 1974, 10, 31. 27. Chandra, R. K. J. Pediat. 1972, 81, 1194. 28. Harland, P. S. E. G. Lancet, 1965, ii, 719.

tested separ-

ately against: B lymphocytes at 5°C; B lymphocytes at 37°C; T lymphocytes at 5°C; and T lymphocytes at 37°C. Incubation with complement in all cultures was at 25 °C. Incubationtimes were 1 h with serum and 2 h with complement. Classification of Sera When a serum killed B lymphocytes from > 15 % of the panel at 5°C and all other reactions were negative, it was classified as B-cold-reactive. If it killed B lymphocytes from > 15 % of the panel at 37°C (whether or not it had B-cold reactivity) and killed T lymphocytes from

Enhancement of human kidney allografts by cold B-lymphocyte cytotoxins.

1228 ENHANCEMENT OF HUMAN KIDNEY ALLOGRAFTS BY COLD B-LYMPHOCYTE CYTOTOXINS YUICHI IWAKI MIN SIK PARK PAUL I. TERASAKI RONALD BILLING Department of...
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