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C R Stopford

Publications and source records attributed to C R Stopford.

7 recordsLinked to original sources
PubMed1989

3-Deazaadenosine 5'-triphosphate: a novel metabolite of 3-deazaadenosine in mouse leukocytes.

Evidence has been obtained for the metabolic formation of small amounts (1-2% of the ATP pool) of 3-deazaadenosine 5'-triphosphate (c3ATP) from 3-deazaadenosine (c3Ado) in mouse cytolytic lymphocytes and mouse resident peritoneal macrophages. With intact leukocytes, pharmacological evidence was obtained that adenosine kinase was not the enzyme chiefly responsible for the phosphorylation of c3Ado. Moreover, in the presence of MgCl2, NaCl and IMP, purified rat liver 5'-nucleotidase catalyzed the phosphorylation of c3Ado to 3-deazaadenosine 5'-monophosphate (c3AMP). Two lines of evidence suggest that the metabolic formation of c3ATP is not involved in the inhibition of leukocyte function caused by c3Ado. First, the inhibitory action of c3Ado on antibody-dependent phagocytosis and lymphocyte-mediated cytolysis was reversed markedly upon removal of the drug from the medium. However, the intracellular content of c3ATP remained constant in lymphocytes and macrophages after removal of c3Ado. Second, in macrophages and in lymphocytes, similar intracellular amounts of c3ATP were formed from both c3Ado and 3-deazaadenine under conditions in which the former was biologically active and the latter was essentially inactive. Thus, it appears unlikely that the novel c3ATP metabolite is of relevance for the mechanism of action of c3Ado in mouse leukocytes.

5'-Nucleotidase↗
PubMed1985

3-Deazaadenosine-induced disorganization of macrophage microfilaments.

3-Deazaadenosine (c3Ado) has been reported to inhibit a number of cellular functions. These biological effects of c3Ado have generally been attributed to its ability to act as inhibitor and substrate of S-adenosylhomocysteine hydrolase. In this report, it is revealed by fluorescence microscopy that c3Ado caused disorganization of the microfilament system of mouse macrophages at concentrations (greater than or equal to 5 microM) similar to those that inhibited antibody-dependent phagocytosis and zymosan-stimulated H2O2 production by these cells. Inhibition of phagocytosis and perturbation of microfilaments by c3Ado were completely abrogated by washing the macrophages free of this agent and allowing the cells a 30-min recovery period. Furthermore, these effects of c3Ado on phagocytosis and microfilaments appeared to be independent of the increase in S-adenosylhomocysteine and S-3-deazaadenosylhomocysteine that occurred in these macrophages. First, periodate-oxidized adenosine and 3-deaza(+/-)aristeromycin, two other inhibitors of S-adenosylhomocysteine hydrolase that caused greater increases in macrophage S-adenosylhomocysteine than did c3Ado, had no effect on either phagocytosis or microfilaments. Second, pretreatment of macrophages with periodate-oxidized adenosine (to inhibit S-adenosylhomocysteine hydrolase) prevented the subsequent metabolism of c3Ado to S-3-deazaadenosylhomocysteine but did not diminish the effects of c3Ado on phagocytosis or microfilaments. These results demonstrate that c3Ado can perturb the microfilament system of cells and provide an alternative mechanism for the biological effects of c3Ado.

Actins↗
PubMed1984

Antagonism by taxol of effects of microtubule-disrupting agents on lymphocyte cAMP metabolism and cell function.

Several microtubule-disrupting agents (colchicine, demecolcine, vinblastine, vincristine, podophyllotoxin, and nocodazole) have been shown to inhibit lymphocyte-mediated cytolysis. These agents also enhanced the prostaglandin E1-induced rise in cAMP levels in these cytolytic lymphocytes. Taxol, a natural product alkaloid that has been shown to enhance microtubule polymerization and to stabilize microtubules, antagonized both of these effects of the microtubule-disrupting agents in the cytolytic lymphocytes. Taxol also antagonized the enhancement of cAMP increases by colchicine in lymphocytes stimulated by 2-chloroadenosine, isoproterenol, and cholera toxin. The enhancement of the prostaglandin E1-induced cAMP response caused by treatment of the lymphocytes with either cytochalasin B or 3-deazaadenosine in the presence or absence of L-homocysteine was not antagonized by taxol. Taxol, colchicine, or the combination of these two agents did not affect ATP levels in cytolytic lymphocytes. These results support a modulatory role for microtubules in both the cytolytic process and the production of cAMP in these lymphocytes.

Alkaloids↗
PubMed1983

Inhibition of lymphocyte function by 9-deazaadenosine.

9-Deazaadenosine (c9Ado), a novel C-nucleoside, has been found to inhibit lymphocyte-mediated cytolysis (LMC) in a time-dependent manner. c9Ado inhibited LMC by 50% at concentrations of 10 and 0.07 microM after drug-pretreatment periods of 3 and 22 hr, respectively, although a 1-hr pretreatment of cytolytic lymphocytes with 100 microM c9Ado had no effect upon this lymphocyte function. c9Ado was metabolized rapidly and extensively to 9-deazaadenosine 5'-triphosphate (c9ATP) both by mouse cytolytic lymphocytes and by human erythrocytes. Adenosine kinase purified from rabbit liver phosphorylated c9Ado with a Km of 200 microM and a Vmax of 8% that for adenosine. The metabolic buildup of c9ATP in lymphocytes was accompanied by a large, time-dependent decrease in cellular ATP and by smaller percentage decreases in CTP, UTP and GTP. Among other biochemical effects examined, c9Ado was found to cause a decrease in lymphocyte cAMP content and appeared to be neither an inhibitor nor a substrate for S-adenosylhomocysteine hydrolase. Consistent with this latter result, L-homocysteine thiolactone had no effect on the inhibition of LMC by c9Ado. Neither the inhibition of LMC by c9Ado nor the metabolic formation of c9ATP in lymphocytes was affected by erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), indicating that c9Ado is not a substrate for adenosine deaminase. 5-Iodotubercidin, a non-competitive inhibitor (Kis = 9 nM, Ku = 20 nM) of adenosine kinase, prevented the above effects of c9Ado on lymphocyte function, c9ATP formation, and ATP levels. Either complete preservation (with coformycin) or partial replenishment (with adenosine plus EHNA) of ATP levels in c9Ado-treated lymphocytes resulted in partial restoration of cytolytic function to cells containing large amounts of c9ATP. These results suggest that c9Ado is inhibitory to LMC both because it causes a decrease in the absolute concentration of ATP within the cytolytic lymphocytes and because it permits the establishment within these cells of an unfavorable c9ATP:ATP ratio which impedes the utilization of ATP in a reaction essential to the execution of this lymphocyte function.

Adenosine Triphosphate↗
PubMed1983

In vivo and in vitro antitumor activity expressed by cells of concomitantly immune mice.

Growth of a primary tumor is often accompanied by the development of resistance to subsequent challenge implants of the same tumor, i.e., concomitant immunity. Using the P815 mastocytoma tumor, the kinetics of concomitant immunity was found to be governed by duration of exposure to the tumor and tumor mass. By implanting small "challenges" prior to the immunizing tumor, resistance to the growth of existing tumor foci was demonstrated. Winn-type assays revealed that antitumor activity was present in cell populations from the peritoneal exudate and lymph node draining the tumor. Peritoneal exudate cells, when infused systemically, were also able to confer protection against P815 mastocytoma challenge, suggesting their role as mediators of concomitant immunity. The 51Cr release technique indicated that cytolytic activity in lymph node cells, peritoneal exudate cells, and the spleen was present over a time course parallel to the kinetics of in vivo challenge. The peritoneal resident cell population was only slightly active; thus, effectors accumulated in the inflammatory exudate. Removal of specific subsets of cells from effector populations with antibody to surface markers and complement produced similar effects on both Winn and cytolytic assays. Anti-Thy 1.2 ablated measurable activity. It was substantially but not completely reduced by anti-Lyt 1.1 and only to a small degree by anti-Lyt 2.1.

Animals↗
PubMed1981

In vitro cytotoxicity expressed by cells active against established tumors in vivo.

Although antitumor activity by host cells has been documented in vivo and in vitro, the cellular relationships between these two classes of studies are not clear. Cells capable of causing the regression of solid tumors are generated in lymph nodes draining sites of immunization with Corynebacterium parvum:irradiated P815 mastocytoma admixtures. These cells are active in a 51Cr release assay at a low effector:target ratio producing a characteristic low level of specific 51Cr release which required 24 hr for optimal development. The activity is immunologically specific for the immunizing tumor and is mediated by nonadherent, rapidly dividing (vinblastine-sensitive) cells. They are absent in thymectomized animals and susceptible to alpha-Thy 1.2 antibody and complement. They are present in peritoneal exudates, consistent with the systemic resistance demonstrable in the animal model. The properties and development kinetics of effector cells measured by 51Cr release correlate closely with those of cells showing in vivo activity, supporting the identity of the two populations.

Animals↗