Distinction of anti-K562 and anti-allocytotoxicity in in vitro-stimulated populations of human lymphocytes.
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Biomedical subjects
Publications and source records attributed to E Klein.
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A characteristic alkaline phosphatase (orthophosphoric monoester hydrolase, alkaline pH optimum, EC 3.1.3.1) was detected in the sera of most patients with infectious mononucleosis, acute and chronic lymphatic leukaemia, non-Hodgkin's lymphoma, Burkitt's lymphoma and nasopharyngeal carcinoma. The enzyme was also present in the sera of nine out of 26 patients with cancer of the cervix. N-APase in these cases counted 30-100% of the total alkaline phosphatase activity. N-APase was absent from the sera of healthy individuals and of patients with acute and chronic granulocytic leukaemia, breast cancer, colon cancer, rheumatoid arthritis, ulcerative colitis, systemic lupus erythematosis, hepatitis and obstructive jaundice. Only three of 22 patients with Hodgkin's disease showed n-apase activity in the serum. In infectious mononucleosis the presence of N-APase activity was well correlated with the clinical course. In 13 cases studied, the clinical improvement was associated with the decrease or disappearance of N-APase activity. N-APase activity could not be detected in white cells of acute myeloid leukaemic patients, nor in the cells of myeloid blastic crisis of chronic granulocytic leukaemia. It was present in the cells of lymphoid blastic crisis of chronic granulocytic leukaemia.
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Lymphocytes were separated from the synovial fluid of 34 arthritic patients. The majority of lymphocytes rosetted with SRBC and were thus of T origin. A proportion of these cells were activated as indicated by their high 3H-thymidine incorporation in vitro, the formation of "stable" E rosettes, and their ability to attach to autologous monocytes and to human cells. Such attached activated T cells enhanced the cytotoxic activity of monocytes. Human and murine-activated T cells generated in mixed lymphocyte cultures also attached to cell lines of the same species, and after their attachment they enhanced the cytotoxic activity of monocytes and natural killer cells. It is suggested that one of the possible roles of activated T cells in immunologically damaged tissues is the attraction of circulating nonspecific killer cells to the site of the response.
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A case report of a parathyroid cyst with high calcium levels which dropped to normal after excision of the cyst, and a short discussion of the literature is herewith presented. It is believed that this case presents the rare association of a parathyroid cyst and hyperparathyroidism.
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Three cytotoxic systems, NK, ADCC and the seemingly indiscriminative cytotoxicity of in vitro activated lymphocytes (cultivated with K 562) were found to require Ca++ and did not function in Mg++. The optimum concentration of Ca++ was identical for the three systems and for various lymphocyte subsets. 15 minutes after initiation of the interaction cell damage had already occurred in all three systems.
Permeability and rejection properties of new, high flux cellulose membranes and fibres have been compared with Cuprophan PM 150. The greater solute and water flux is explained in terms of larger "pores", which permit greater transport of large molecules.
A T-cell-enriched lymphocyte subset of samples from 15 tumor patients was tested for primary cytotoxicity against autologous tumor cell preparations and against 1-3 different allogeneic tumor cell preparations from biopsy material. Allogeneic cytotoxicity occurred in only 1 of 10 patients with autologous reactivity. The lymphocytes of 14 patients were cultured with autologous cells from biopsy material for 6 days. These lymphocytes killed autologous targets, but only 1 patient's lymphocytes were cytotoxic against 1 of the 4 allogeneic tumors tested. Cocultivation with allogeneic cells from biopsy specimens generated cytotoxicity toward the sensitizing allogeneic cells in 3 of 9 test combinations. In 2 of 3 instances the effectors were also active against the autologous tumor cells. Cytotoxicity in primary and secondary tests occurred thus only rarely against allogeneic targets. This indicated either the presence of individual tumor-related antigens on the cells from biopsy material or reflected the histocompatibility restriction of T-cell-mediated cytotoxicity.
Blood lymphocytes from 22 cancer patients were examined for cytotoxicity against autologous tumour cells in a short-term 51Cr release assay. Only three showed reactivity. In an attempt to increase cytotoxic potential in these and induce reactivity in non-reactive cases, the lymphocytes were cultured alone or with autologous tumour cells for 6 days. Upon subsequent testing against frozen, stored targets, nine samples reacted, including two of those with primary reactivity. In seven cases augmented cytotoxicity was evident in mixed cultures compared with lymphocytes cultured alone. Two of 10 cases showed cytotoxicity against the K562 cell line after culture and, in two of 13 tests in which allogeneic tumour biopsy targets were used, weak reactivity was bound. Cytotoxicity for autologous tumour biopsy cells was uniformly accompanied by positive blastogenesis in MLTI assays. In four cases blastogenesis occurred without the induction of cytotoxicity.
Crude membrane (CM) extracts from three different cultured human melanoma lines that were "virus-augmented" (infected with vesicular stomatitis virus (VSV) and subsequently inactivated by ultraviolet light) produced positive skin tests in 17 of 20 (85%), 11 of 20 (55%), and 13 of 18 (72%) tests, respectively, performed in 20 melanoma patients. Identical CM extracts from the same melanoma lines that had not been infected with VSV gave positive skin tests in 2 of 20 (10%), 4 of 20 (20%), and 2 of 18 (11%) tests, respectively, performed in the 20 melanoma patients, and no positive tests in the control patients. The 3 virus-augmented extracts were positive in only 2 of 18 (11%), 0 of 18 (0%), and 1 of 17 (6%) control subjects, respectively. The controls consisted of six normal volunteers and 12 patients with cancers other than melanoma. The "virus-augmented" CM extracts thus exhibited markedly greater sensitivity without significant loss of specificity as compared to nonvirus augmented extracts when used as tumor-specific melaonma skin test antigens.
This report describes model systems which show low primary in vitro syngeneic cytotoxic responses to a Moloney-induced YAC tumor (syngeneic in A mice) and a Rauscher-induced RBL5 tumor (syngeneic in C57BL/6 mice) and examines different approaches to overcome these defects. Two major findings were obtained: (a) spleen cells from A mice, injected with tumor cells from an in vitro tumor line YAC-1, derived from YACL, could generate a significant syngeneic cytotoxic response. In contrast, spleen cells from A mice injected with tumor cells from the in vivo tumor line failed to generate a syngeneic cytotoxic response. Thus, tumor cells from the in vitro line were more immunogeneic that those from the in vivo line. (b) Spleen cells from A mice which were injected with the cross-reactive allogeneic tumor RBL5, could generate significant cytotoxic responses to the syngeneic tumors YAC and YAC-1. Similarly, spleen cells from C57BL/6 mice injected with the cross-reactive allogeneic tumor YAC-1, could generate a significant cytotoxic response to the syngeneic tumor RBL5. Thus, cross-reactive allogeneic tumors could stimulate syngeneic cytotoxicity. The theoretical and the practical implications of these studies are discussed.
Unprimed spleen cells from A and C57BL/6 mice could not produce cytotoxic responses to their syngeneic tumors: a Moloney virus-induced in vitro subline YAC-1 and a Rauscher virus-induced in vitro subline RBL5, respectively. Spleen cells from A and C57BL/6 mice immunized with YAC-1 OR RBL5 (which cross-react serologically) generated significant syngeneic cytotoxicities after cultivation in vitro. The in vivo carried tumor of A mice, unlike the in vitro sublines, could not stimulate a priming effect. In contrast, YAC stimulated the formation of suppressor cells in both A and C57BL/6 mice. The suppressor cells abrogated the priming effect of the syngeneic tumors, but not the priming effect of the allogeneic tumors. Furthermore, YAC did not suppress normal allogeneic anti-tumor responses. The theoretical and the practical implications of these studies are discussed.
The T-cell nature of the responding cells in autologous tumor stimulation (ATS) was proven by (1) E-rosette formation of lymphocytes attached to tumor cells during the early period of co-cultivation; (2) blast-transformed E-rosetting cells detectable at the end of the test period (day 6) and (3) the reactivity of T-enriched lymphocyte subset and unresponsiveness of T-depleted fractions. Preparation of tumor suspensions devoid of lymphocytes made it possible to carry out cross-tests between patients. Blastogenesis with allogeneic tumors was rare. The restricted autologous nature of the reaction is a strong indication for genuine tumor-relatedness.
Special procedures are: (1) Short-term treatment with steroid hormones; (2) antithyroid substances; (3) radioactive iodine; (4) operation. The first two methods are complementary to the definitive procedures of isotope and surgical therapy. The indications conform to the age of the patient and the size of the goiter. The treatment of choice in 60% of all cases is radioactive iodine; 20-25% are treated with antithyroid substances only for several years. The remaining cases, complicated by goiters of size II and more, should be handled surgically. Discussion based on personal experience with 3580 cases over 20 years).