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M Stubbs

Publications and source records attributed to M Stubbs.

At least 73 records · Page 4Linked to original sources
PubMed1988

31P-nuclear magnetic resonance spectroscopy studies of the response of rat mammary tumors to endocrine therapy.

We have used 31P-nuclear magnetic resonance spectroscopy to detect the metabolic changes that occur in estrogen-sensitive, N-methyl-N-nitrosourea-induced rat mammary tumors as they regress following ovariectomy. In untreated animals the spectra of the tumors showed a steady loss of high energy phosphates (phosphocreatine and nucleoside triphosphates) and an increase in inorganic phosphate. This was reversed after ovariectomy. Spectral changes occurred before detectable regression of the tumor. Estrogen-insensitive tumors, grown from implanted Rama 600 and 622 cells, did not regress in response to ovariectomy, and their high energy phosphates continued to fall; estrogen-sensitive tumors also failed to respond to sham ovariectomy. These effects are probably due to the reduction in cellular energy requirements that occurs when the hormonal stimulus to growth is removed. Because the nuclear magnetic resonance method is noninvasive, this technique should be applicable clinically as a means of predicting the response of a tumor to endocrine therapy.

Adenosine Triphosphate↗
PubMed1988

Phosphate metabolites in rat skin.

A model for studying the 31P NMR spectrum of rat skin without contribution from other tissue signals has been developed by creating a skin pedicle. 31P NMR spectra were obtained with a solenoidal coil, which was separated from the flank of the rat by a Faraday shield. Phosphomonoesters, inorganic phosphate (Pi) (1.63 +/- 0.12 mumols per g wet wt), phosphodiesters, phosphocreatine (PCr) (1.4 +/- 0.12 mumols per g wet wt) and ATP (1.35 +/- 0.22 mumols per g wet wt) were observed, superimposed on broader signals, probably due to phospholipids. Extracts of freeze-clamped pedicles contained concentrations of phosphorus metabolites similar to those seen by NMR. The exception was Pi which was twofold higher in the extract. The presence of the broader phospholipid contribution suggests that the signals did not arise solely from the panniculus carnosus muscle of rat skin, although this muscle was evident on histological examination of the pedicles. In extracts of normal rat skin levels of creatine, ATP, ADP and Pi were similar to those of pedicles, whereas PCr was about twofold higher. Signals from rat skin are likely to contribute to spectra of subcutaneous organs and tumours. Two kinds of rat hepatoma that contained no PCr frequently gave PCr signals from the overlying skin, whereas in three other subcutaneous tumours the contribution from skin was negligible.

Adenosine Diphosphate↗
PubMed1987

The glucose sensor in HIT cells is the glucose transporter.

The nature of the rate-limiting step for glucose utilization by the clonal insulin-producing cell line HIT-T15 has been investigated. In contrast to the situation in islets of Langerhans, we find that the HIT cell glucose metabolism is limited by the rate of entry of glucose into the cell. This is evidenced by the low rate of sugar transport and by the marked reduction in the rate of glucose utilization elicited by inhibitors of the glucose transporter. As judged by competition with glucose, the HIT cell glucose transporter also transports mannose, 2-deoxyglucose and 3-O-methylglucose but not L-glucose or N-acetylglucosamine. The Km for glucose of the glucose transporter, measured as the concentration of glucose required for a half-maximal rate of glucose utilization, is 4.3 mM, similar to the concentration reported to give half-maximal insulin release. Glucose-stimulated insulin release from HIT cells is inhibited by phloretin or cytochalasin B but not by mannoheptulose. We conclude that the secretory responses of HIT cells are consistent with the substrate-site hypothesis, but that, in contrast to normal B-cells, the glucose sensor which confers concentration-dependence and specificity to sugar-stimulated insulin release, is the glucose transporter.

3-O-Methylglucose↗
PubMed1984

Formation of n.m.r.-invisible ADP during renal ischaemia in rats.

Measurement of the adenine nucleotide and inorganic phosphate content of normoxic and ischaemic kidney in vivo has been made, comparing enzymic assay (after freeze-clamping and acid extraction) with quantification by 31P-n.m.r. Both methods give similar results for ATP, and n.m.r. quantification of Pi gives a value 25-50% of that obtained by enzymic assay. ADP, which is largely invisible to n.m.r. in the normoxic kidney, remains invisible during ischaemia despite a 2-3 fold rise in enzymically assayed ADP. N.m.r. and enzymic assay of the acid extracts give similar values for all metabolites measured. The question of ADP binding in the kidney is discussed, as are the implications for the metabolic regulation of ADP-dependent reactions.

Adenosine Diphosphate↗
PubMed1984

Effect of hibernation on liver and kidney metabolism in 13-lined ground squirrels.

Metabolic rates and adenine nucleotide content of liver and kidney from hibernating ground squirrels were measured and compared to rats to study the biochemical adaptation to hibernation. High rates of renal and hepatic gluconeogenesis were observed in squirrels, particularly from propionate and glycerol compared to rat. During hibernation and starvation soluble phosphoenolpyruvate carboxykinase activity was increased in both liver and kidney. Although metabolic rates are decreased during hibernation the results suggest that the enzymic complement is maintained at high activity even during torpor.

Adenine Nucleotides↗
PubMed1983

Studies on the mechanism of the antifungal action of benzoate.

A method is described for the determination of the pH of intracellular water based on the distribution of [14C]benzoate (0.01 mM) between intra- and extra-cellular water. Benzoate at higher concentrations (2-10mM) enters the yeast cell in the undissociated form, and its neutralization within the cell can cause a shift of the pH of the intracellular water by more than 1 pH unit. Benzoate causes an accumulation of the two hexose monophosphates of yeast glucose fermentation and a decrease in intermediates beyond phosphofructokinase, suggesting inhibition at this stage. Benzoate also causes a concomitant fall in [ATP]. Phosphofructokinase is inhibited to a greater extent than hexokinase at acid pH. There is a relationship between intracellular pH, phosphofructokinase inhibition and CO2 production, suggesting that the antifungal action of benzoate is caused by an accumulation of benzoate at low external pH, which lowers the intracellular pH into the range where phosphofructokinase is sensitive. The subsequent inhibition of glycolysis causes a fall in [ATP] and thus restricts growth.

Adenine Nucleotides↗
PubMed1981

Evidence for expression of Ly-6.2 on non-bone marrow-derived cells in kidney, skin and connective tissues.

Mouse alloantigen Ly-6.2 is detectable in various non-lymphoid tissues such as kidney, but it is not clear whether or not this expression is due to bone-marrow derived passenger leukocytes. To determine whether non-marrow derived cells express Ly-6.2, we examined the expression of this antigen in kidney and on isolated connective tissue and epidermal cells. Studies in radiation chimeras demonstrated that the kidney did not become Ly-6.2 positive when negative animals were reconstituted with positive marrow. Thus, passenger leukocytes cannot account for the renal expression of Ly-6.2, indicating that most of this antigen is on non-marrow-derived (parenchymal) cells in kidney. Various isolated cell types--fibroblasts, osteocytes, chondrocytes and skin epidermal cells--were found to be Ly-6.2 positive. Indeed, absorption and cytotoxicity results suggested that the amount of Ly-6.2 on fibroblasts exceeded the amount of an H-2 antigen on these cells. Comparison of fibroblasts to lymphocytes indicated that fibroblasts had 13--60 times more Ly-6.2 than spleen cells and three times more than PHA blasts. The results indicate that the Ly-6.2 detected in non-lymphoid tissues is predominantly on the parenchymal or connective tissue elements of those tissues.

Animals↗
PubMed1980

Expression of Ia in mouse kidney.

Expression of Ia antigens in mouse kidney was studied by absorption of diluted anti-Ia sera with crude suspensions of kidney issue. Specific absorption of anti-Ia activity was seen for all Ia specificities tested: Ia.1,2, Ia.3, Ia.4,5,12, Ia.7, Ia.8, Ia.9, Ia.15, and Ia.16. Certain polyspecific antisera (against the I-Ak products Ia.1,2,3,15) were more difficult to absorb than oligo-specific antisera against other Ia specificities (e.g., Ia.7 and Ia.9). This observation may indicate that polyspecific sera are less absorbable by limited numbers of antigenic sites because of steric hindrance, although differences in the extent of antigen expression in kidney have not been excluded. Ia absorption could be demonstrated either in microcytotoxicity or in 51Cr release assays. Both mechanically disrupted and enzyme-disrupted kidney tissue suspensions absorbed Ia specifically, although the former method was used routinely. As estimated from the efficiency of absorption, the amount of Ia in kidney was small, about 2 to 5% of that in spleen. One kidney was equivalent in absorptive capacity to about 3 X 10(6) splenocytes, and to greater than 3 X 10(6) buffy coat cells. Comparisons of the rates of absorption indicated that the amount of Ia in kidney was less than the amount of an H-2K or D alloantigen. Ia was expressed in kidney in an immunogenic form, since animals immunized repeatedly with I region-incompatible kidney tissue produced anti-Ia antibodies. Thus, Ia antigens are expressed in and are immunogenic in mouse kidney and can be studied by conventional serological techniques.

Animals↗
PubMed1980

Expression of Ia in mouse kidney. II. Evidence for expression on resident marrow-derived and nonmarrow-derived cells.

The origins of Ia antigens in perfused mouse kidney were investigated. Three possible sources were considered: leukocytes in residual blood which was trapped in kidney, bone marrow-derived cells resident in kidney, and nonbone marrow-derived renal parenchymal or vascular cells. Leukocytes in trapped blood seemed to make no significant contribution to renal Ia expression because (1) perfused kidney had approximately as much Ia as nonperfused kidney, even though the perfusion reduced the blood content by 90%; (2) the estimated number of leukocytes in trapped blood was at least three orders of magnitude less than that needed to account for Ia expression by kidney; and (3) perfused kidney, volume for volume, absorbed more anti-Ia than did whole blood, so that no amount of blood contamination could account for all renal Ia expression. Thus most Ia in kidney must be on resident cells, either bone marrow-derived or parenchymal. To demonstrate bone marrow-derived Ia-positive cells, we created radiation chimeras of (B10 X B10.D2)F1 bone marrow into B10 hosts. Ia of (B10 X B10.D2)F1 bone marrow donor origin was easily detectable in kidneys of these chimeras at 4 months. However, we also demonstrated Ia of nonbone marrow donor origin in chimera kidney: long-term B10.A into (BALB/c X A)F1 chimeras and C57BL/6 into (C57BL/6 X DBA/2)F1 chimeras continued to express renal Ia of bone marrow recipient origin. Thus, some renal Ia is produced by bone marrow-derived cells, and some is produced by cells which are nonmarrow derived (or are marrow derived but are resistant to replacement in bone marrow chimeras). The cells expressing Ia in kidney were unlikely to be thymus derived because anti-Thy-1.2 was not absorbable by the same kidney preparations which absorbed anti-Ia.

Animals↗
PubMed1979

Loss of cell constituents from hepatocytes on centrifugation.

In studies of the metabolism of isolated hepatocytes, it is often necessary to measure the concentrations of cell constituents both in cells and medium. When hepatocytes are separated in the special tubes of Hems, Lund & Krebs (1975) (Biochem. J. 150, 47--50), they lose much glucose, urea and Na+, whereas there is no loss of K+, glutamate, aspartate and adenine nucleotides. Cell water is also lost, as measured by the distribution of 3H2O. This loss is mainly due to an exchange of cell water with the aqueous solution in the stems of the tubes through which the cells pass on centrifugation. In general, substances are lost only when the intracellular concentration is equal to, or lower than, the extracellular concentration. Probably solutes are lost because they travel with the water unidirectionally out of the cell. A loss of solute does not occur when the cells are centrifuged in conical tubes with a layer of silicone oil between the cell suspension and the deproteinizing layer. The reasons for the loss occurring in the special separation tubes are discussed.

Animals↗
PubMed1978

Is the adenine nucleotide translocator rate-limiting for oxidative phosphorylation?

1. The effects of atractyloside and carboxyatractyloside (between 5 and 40mum) on O(2) uptake, glucose synthesis, urea synthesis, the adenine nucleotide content and the intracellular K(+) concentration were measured in isolated hepatocytes. 2. Urea synthesis was much less inhibited than glucose synthesis by both atractylosides. Measurements of intermediary metabolites of carbohydrate metabolism in freeze-clamped liver after injection of atractyloside into rats indicate that inhibition of gluconeogenesis is due to interference at the cytosolic reactions requiring ATP (phosphoenolpyruvate carboxykinase and 3-phosphoglycerate kinase). 3. The decrease in [ATP]/[ADP]x[P(i)] after addition of atractyloside or carboxyatractyloside was restricted to the cytosol. 4. Dihydroxyacetone can be converted either into glucose with the consumption of 2mol of ATP (per mol of glucose) or into lactate with the production of 2mol of ATP. In the presence of high concentrations of atractyloside and carboxyatractyloside more ATP was produced than was used for the synthesis of glucose from dihydroxyacetone, probably for the maintenance of intracellular [K(+)]. 5. When the rates of respiration were altered by changing substrates, the degrees of inhibition of respiration and translocation by a given concentration of the atractylosides were the same, whereas at a given concentration of HCN the degree of inhibition was high at higher initial rates, and low at lower initial rates. 6. Inhibition of a complex series of reactions by atractyloside does not necessarily indicate that the translocator is a rate-limiting step in that sequence as Th. P. M. Akerboom, H. Bookelman & J. M. Tager [(1977) FEBS. Lett.74, 50-54] assume. This point is discussed.

Adenine Nucleotides↗