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Biomedical subjects

G Soslau

Publications and source records attributed to G Soslau.

At least 37 records · Page 2Linked to original sources
PubMed1991

Mitochondrial DNA topoisomerase I from human platelets.

An anucleated cell system has been used for the first time to study mitochondrial topoisomerase activity. Mitochondrial extracts from human blood platelets contained type I topoisomerase. The type I classification was based on ATP-independent activity, inhibition by ATP or camptothecin, and the lack of inhibition by novobiocin. Platelet mitochondrial topoisomerase I relaxation activity was inhibited linearly by increasing concentrations of EGTA. Topoisomerase activity greater than 90% inhibited by 175 microM EGTA was partially restored to 16 and 50% of the initial level of activity by the subsequent addition of 50 and 100 microM Ca2+, respectively. Additionally, results from studies of partially purified platelet mitochondrial topoisomerase I were consistent with the crude extract data. This work supports the hypothesis that platelet mitochondria contain a type I topoisomerase that is biochemically distinct from that previously isolated and characterized from cell nuclei.

Blood Platelets↗
PubMed1991

Effect of tranexamic acid on platelet ADP during extracorporeal circulation.

Seventeen adults received the antifibrinolytic drug tranexamic acid during cardiac surgery utilizing extracorporeal circulation (ECC). In 8 patients, drug administration began prior to skin incision (pre-ECC); infusions commenced after ECC and protamine administration in another 9 patients (post-ECC). Compared with the post-ECC group, the pre-ECC group exhibited less bleeding via mediastinal drains (420 vs. 655 mL/12 h median, P = 0.024), decreased frequency of the presence (greater than or equal to 10 micrograms/mL) of fibrin split products (P less than 0.05), and greater platelet dense granule content of adenosine diphosphate after surgery (15.47 vs. 4.05 nmoles/mg protein median, P = 0.021). Follow-up in vitro study of tranexamic acid inhibition of plasmin-induced platelet activation utilizing normal human platelet rich plasma and porcine plasmin revealed a 13-fold lower concentration of tranexamic acid for 50% inhibition when plasmin was preincubated with the drug (1.2 micrograms/mL, 95% CI = 1.13-1.60 micrograms/mL) compared to when platelet rich plasma was preincubated with the drug (16 micrograms/mL, 95% CI = 7.3-99. micrograms/mL). Plasmin inactivated with tranexamic acid retained its ability to inhibit thrombin-induced platelet activation, thus suggesting that tranexamic acid inhibits plasmin's catalytic activity and not its binding to platelets. Both clot lysis and platelet dysfunction may contribute to bleeding after ECC. Tranexamic acid blocks plasmin-induced partial platelet activation during ECC, thus preserving platelet function and promoting hemostasis after ECC.

Adenosine Diphosphate↗
PubMed1990

Selective reduction of serotonin storage and ATP release in chronic renal failure patients platelets.

Hemorrhagic complications, as monitored by skin bleeding times, occur in a significant number of chronic renal failure (CRF) patients. The etiology of hemostatic defects in these patients is complex and ill defined. Our studies demonstrate, for the first time, that activated platelets, derived from CRF patients, release significantly (P less than .001) less ATP than controls while the percent of releasable serotonin (5HT), assumed to be co-stored with ATP, is unaltered. Analysis of the CRF-derived platelets reflects a selective acquired storage pool defect with significantly (P less than .001) reduced 5HT levels while their dense granule contents of ATP and ADP are normal. The comparison of ATP release from platelets derived from CRF patients whose bleeding times were less than 9 min to those with bleeding times of 9 min or greater was significantly different (P less than .02). This report demonstrates for the first time that there is a statistically significant correlation of ATP release and 5HT content to bleeding times (P less than .001). The perturbation of platelet 5HT uptake, 5HT dense granule content, and ATP release appears to result from newly described altered plasma factors, detected by our in vitro mixing studies. It is proposed that the reduced level of releasable platelet 5HT and ATP contributes to bleeding disorders commonly encountered in CRF patients.

Adenosine Diphosphate↗
PubMed1990

Desmopressin-induced improvement in bleeding times in chronic renal failure patients correlates with platelet serotonin uptake and ATP release.

Hemostatic defects resulting in life-threatening hemorrhagic episodes are a common occurrence in the chronic renal failure patient. Hemorrhagic tendencies correlate best with laboratory tests of bleeding times. The identification of a specific hemostatic defect and its role in bleeding dyscrasias has yet to be elucidated. Our studies demonstrate that factor VIII coagulant activity and factor VIII related antigen (vWF:Ag) are normal or greatly elevated in uremic renal failure patients with greatly prolonged bleeding times. The multimeric state of the von Willebrand factor is also normal in these patients. The bleeding times were normalized in all 15 patients, 90 minutes post-infusion with desmopressin (DDAVP). No significant changes in factor VIII/vWF associated properties, blood cell counts, or coagulation factors were observed post-DDAVP treatment. However, a significant increase in platelet serotonin uptake (p less than .025) and ATP release (p less than .025) was detected after DDAVP treatment. These results indicate that DDAVP acts on the platelet membrane. This is further substantiated by the ability of DDAVP to block vasopressin-induced platelet aggregation in a dose- and time-dependent fashion. Perturbations in the movement and storage of serotonin and the release of adenosine 5'-triphosphate (ATP) in the platelets of uremic individuals are proposed to play a critical role in regulating bleeding times.

Adenosine Diphosphate↗
PubMed1989

Modulation of platelet function by extracellular adenosine triphosphate.

A potential physiologic role of extracellular adenosine triphosphate (ATP) on platelet function is proposed in this report. It is widely accepted that ATP competitively inhibits adenosine diphosphate (ADP)-induced platelet aggregation. Our observations of platelet aggregation with the agonists, collagen, epinephrine, and ADP in the presence of 180 mumol/L ATP could support this competitive nature of ATP. However, the disaggregation of maximally aggregated platelets induced by ATP, theophylline, or ATP plus theophylline indicates that additional mechanisms of ATP action may be present. Extracellular gamma-32P-ATP (7 pmol) labels surface-membrane proteins in intact platelets as demonstrated by several criteria. The reaction is Ca++-dependent. Stimulation by calcium occurs in the physiologic range of 1 to 5 mmol/L. Significant levels of phosphorylation occur within one minute with near maximal levels reached by five minutes. Platelet cyclic AMP (cAMP) levels were elevated in a dose-dependent fashion in cells incubated for four minutes with increasing amounts of extracellular ATP (18 to 540 nmol). The addition of ATP plus theophylline resulted in a synergistic stimulation of cAMP levels. ATP was not being hydrolyzed to adenosine by plasma nucleotidases, as demonstrated by the lack of effect of ten U of adenosine deaminase. The phosphorylation of surface proteins by extracellular ATP released from activated platelets may modulate platelet responsiveness to agonists at distances removed from the site of vascular injury. Phosphorylation may also play a role in signal transduction to regulate the levels of intracellular cAMP, which further inhibits platelet activation.

Adenosine Triphosphate↗
PubMed1989

Hereditary sideroblastic anemia with associated platelet abnormalities.

A 62 year old male (R.H.) presented with a mild anemia (Hb 11-12 gm%) and a history of multiple hemorrhagic episodes. The marrow had 40-50% sideroblasts. Marrow chromosomes were normal. His wife was hematologically normal, while one daughter, age 30 years, had a sideroblastic anemia (Hb 11-12 gm%) with 40-50% sideroblasts in the marrow. Her anemia was first noted at age 15 years. Administration of vitamin B6 did not correct the anemia in either the father or daughter. Platelet abnormalities inherited jointly with this disorder are described for the first time. Both R.H. and his daughter had prolonged bleeding times, with normal PTT, PT times, fVIII:C, fVIII:Ag levels, and vWF multimers, which may rule out a von Willebrand's disease. They have normal platelet numbers but abnormally low platelet adhesiveness and greatly depressed ADP, collagen, and epinephrine responsiveness. Response to ristocetin was in the low normal range, and aggregation with thrombin was normal. While desmopressin completely normalized R.H.'s bleeding time, none of these platelet parameters were improved. No differences in the SDS PAGE protein patterns of RH platelets could be detected in comparison to normal samples. His platelets took up and released serotonin (5HT) normally, and electron micrographs defined no morphological abnormalities. However, no ATP was released from platelets activated with collagen, and when followed by thrombin about fourfold greater ATP was released by control platelets as compared to RH platelets. The dense granule fraction derived from RH platelets contained about 20% the level of ATP, 40% the level of ADP, and 50% the level of 5HT detected in a normal sample. The results indicate that the bleeding disorder is related to a non-classical heritable storage pool defect. The connection between the inherited sideroblastic anemia and platelet defects is obscure.

Adenosine Triphosphate↗
PubMed1988

Cerastes cerastes (Egyptian sand viper) venom induced platelet aggregation as compared to other agonists.

The Egyptian Sand Viper (Cerastes cerastes) crude venom and subfractions were, for the first time, shown to induce platelet aggregation with agonist activities present in two subfractions. The combined activities of the crude venom components behaved in a unique fashion as compared to the platelet agonists, ADP, collagen and thrombin. The action of the venom was inhibited by conditions that increased cAMP, partially required the formation of thromboxane A2 and was inhibited by the serine protease inhibitor PMSF while being only partially sensitive to leupeptin or soybean trypsin inhibitor. One of the fractionated venom agonists strongly induced serotonin release while the other venom agonist essentially did not. Further characterization of the Cerastes cerastes venom components should broaden our knowledge of the pathology of snake venoms, platelet aggregation and their potential therapeutic value.

Adenosine Diphosphate↗
PubMed1988

Loss of low-affinity serotonin receptors upon storage of human platelets.

Platelets actively accumulate virtually all plasma serotonin within their dense granules. As a readily isolated, homogeneous cell type, platelets have served as a model for serotonin uptake into neurological tissue, in addition to defining the role of serotonin in hemostasis. The number of serotonin receptor types on the platelet membrane and the function of these receptors has not been conclusively demonstrated. The presence of different receptor types that may be altered or lost in disease or upon aging (in vitro storage or in vivo) could have significant physiological effects on platelet function. This report demonstrates that at least two receptor types are present on freshly prepared human platelets. However, after 3 to 4 days of storage in autologous plasma, the low-affinity, high-capacity serotonin receptor appears to be lost. This phenomenon probably accounts for some of the discrepancies reported in the literature. The high-affinity receptor present in both freshly isolated and stored platelets binds about 9 x 10(3) serotonin molecules per platelet. Binding can be completely blocked by imipramine; however, some passive diffusion appears to occur even at the low level of extracellular serotonin concentrations employed in these studies (nanomolar range). The influx of serotonin into platelets appears to be poorly reversible, even in reserpine-treated cells, where the extravesicular cytoplasmic concentration would be high. The loss of the low-affinity serotonin receptor type reported in these studies may be directly or indirectly associated with the reduced responsiveness observed in stored platelets.

Biological Transport↗
PubMed1987

Herpes simplex virus replication and protein synthesis in a human blood-derived cell line.

Herpes simplex virus (HSV) types 1 and 2 were shown to replicate in a newly described human cell line (Meg) derived from the peripheral blood of a healthy volunteer. The cell line has both megakaryocyte-like and B cell-like properties. Upon infection with HSV-1 or -2, at a m.o.i. between 0.5 and 5, unlike B and T cells, the Meg cells were growth-arrested and this was accompanied by cytopathic effects and virus replication. The HSV proteins and glycoproteins B and D (gB and gD) made in the blood-derived Meg cells were compared to the corresponding proteins made in the non-blood-derived cell lines, Vero (African green monkey kidney cell) and HEp-2 (human epidermoid carcinoma cell). The maximum level of HSV protein synthesis occurred earlier in the Meg cells than in the Vero and HEp-2 cells. The electrophoretic pattern of HSV-1 and -2 proteins made in the Meg cell line was similar to the corresponding proteins made in the Vero and HEp-2 cell lines; however, some qualitative and quantitative differences were evident. There were no apparent differences detected in the migration pattern of gB made in all three cell lines while significant differences were observed with the gD species. However, upon hydrolysis with Staphylococcus aureus V-8 protease of the monoclonal antibody-purified gB and gD, distinct differences were observed in the electrophoretic pattern of the generated peptide fragments of both gB and gD made in the three cell lines. The results demonstrate that a human blood cell can support HSV replication and that species-specific post-translational modification of gB and gD occurs in HSV-infected Vero cells as compared to HSV-infected human cells.

Animals↗
PubMed1986

Methylation and restriction endonuclease cleavage of linear Z-DNA in the presence of hexamminecobalt (III) ions.

These studies employed the synthetic linear DNA, poly dGdC, in the B and cobalt hexammine chloride (Co)-induced Z form to determine the effect of conformation on protein-DNA interactions. The rate of the reaction of the restriction endonucleases, Hha I and Cfo I, are reduced with Z DNA as compared to B DNA. The ability of both restriction endonucleases to react with an aggregate form of Z DNA (Z* DNA) is found to depend upon how the Z* DNA is formed. When Z* DNA is induced by low concentrations of Co (50 microM), the endonucleases remain active. In the presence of 100 microM Co, which causes increased aggregation, the endonucleases are inactive. The Hha I DNA methyltransferase reacts at equal rates with the B, Z and low cobalt Z* forms and at a greatly reduced rate with the high cobalt Z* form. These results are significantly different than those observed with Z form dGdC tracts inserted into circular DNA molecules.

Chlorides↗
PubMed1984

Phosphoproteins altered by antiproliferative doses of human interferon- beta in a human bladder carcinoma cell line.

Phosphoproteins of control and IFN-beta treated human bladder carcinoma cells (RT4) were labelled in vitro with [32P]-ATP and analyzed by polyacrylamide gel electrophoresis. Cells treated with antiproliferative doses of IFN had reduced levels of phosphorylated 60 Kd and 40 Kd proteins. IFN also induced within 24 hours the modification of a low molecular weight phosphoprotein doublet in the 22-24,000 molecular weight range. The ability to phosphorylate high molecular weight proteins by the in vitro procedures was generally depressed by IFN treatment. There was a dramatic shift in the phosphorylation of alkali stable phosphoamino acids associated with proteins in the 43-50,000 molecular weight range in IFN-treated cells. Preliminary studies indicate that at least some of the IFN-induced modifications of cellular phosphoproteins may result from transcriptional control of specific oncogenes.

Adenosine Triphosphate↗
PubMed1983

De novo synthesis of DNA in human platelets.

Platelets, incubated with radiolabeled thymidine and purified free of contaminating nucleated cells, were analyzed for their ability to synthesize DNA. The only DNA species isolated from these purified platelets was mitochondrial DNA. The CsCl gradient-purified platelet DNA was treated with the restriction endonucleases EcoRI, HindIII and HpaI yielding the expected pattern for human mitochondrial DNA. Nitrocellulose blots of the electrophoresed, restriction endonuclease-treated DNA were fluorographed. All of the DNA fragments generated by the restriction enzymes were labeled, indicating de novo synthesis. This was further substantiated by inhibition of DNA synthesis by ethidium bromide and 2',3'-dideoxythymidine. Platelet DNA appeared to become greatly fragmented after 4 to 7 days storage while all of the thymidine incorporated was observed in intact mitochondrial DNA. These results indicate a continuous degradation of platelet mitochondrial DNA with no apparent repair mechanism. The ability of platelets to synthesize DNA may be associated with the protein synthetic capacity of platelets previously described.

Blood Platelets↗
PubMed1983

Selective inhibition of restriction endonuclease cleavage by DNA intercalators.

The preferred dye binding sites and the microenvironment of known nucleotide sequences within mitochondrial and plasmid pBR322 DNA was probed in a gross fashion with restriction endonucleases. The intercalating dyes, ethidium bromide and propidium iodide, do not inhibit a given restriction endonuclease equally at all of the restriction sites within a DNA molecule. The selective inhibition may be explained, in part, by the potential B to Z conformation transition of DNA flanking the restriction site and by preferred dye binding sites. Propidium iodide was found to be a more potent inhibitor than ethidium bromide and the inhibition is independent of the type of cut made by the enzyme.

Animals↗
PubMed1982

The loss of sialic acid and its prevention in stored human platelets.

The loss of sialic acid was determined in human platelets stored during a seven day period in their homologous plasma. Approximately 30% of the sialic acid was lost during the first three days of incubation at room temperature and a total of 73% was lost after seven days. The rapid in vitro loss of sialic acid may mimic a slower in vivo loss. It was found that platelets with a greater density had a higher sialic acid content than the less dense platelets. The loss of sialic acid from stored platelets could be completely inhibited by the addition of silyl compounds to the incubation plasma. The trisaccharide, N-acetylneuramin-lactose, gave a greater degree of protection than fetuin at comparable concentrations.

Blood Platelets↗
PubMed1981

Control and virus-transformed baby hamster kidney cells resistant to ethidium bromide. II. Membrane properties.

Aspects of membrane structure and functions were studied in ethidium bromide resistant cells. Submitochondrial particles were solubilized and electrophoresed. The gel patterns, representing mitochondrial membrane proteins, demonstrated qualitative and quantitative alterations in mitochondrial preparations derived from virus-transformed cells and ethidium bromide resistant cells as compared to the control cells. The plasma membrane glycoproteins were labelled by the sodium borohydride method. The glycoproteins were pleased with Triton X-100 and electrophoresed. Fluorograms of the gels demonstrated some marked differences between the ethidium bromide resistant cells and their parental strain. The observed alterations in the membrane glycoproteins did not result in altered glucose transport properties or in the elution patterns of plasma membrane glycopeptides as analyzed by Sephadex G-50 chromatography. Dye uptake and binding studies with intact parental and drug resistant cells and their isolated mitochondria demonstrated no alteration of the membrane permeability or the number of binding sites for ethidium bromide. Similar results were also obtained with a cyanine dye. This latter finding was significant in that it permitted one to exclude dye exclusion as a mechanism for ethidium bromide resistance.

Animals↗
PubMed1980

Control and virus-transformed baby hamster kidney cells resistant to ethidium bromide. I. Characterization and the respiratory enzymes.

Cell lines resistant to ethidium bromide have been developed from cultured mammalian BHK21/C13 cells and these same cells transformed by Rous sarcoma virus (C13/B4). Cells resistant to 2 micrograms ethidium bromide per milliliter have been cloned. One clone of the control and one of the virus-transformed cell lines has been employed for characterization. The resistant cells, in the presence of 2 micrograms ethidium bromide/ml, grow at approximately the same rate as the untreated parental cells. The control cells possess a "normal" karyotype (44 chromosomes), while the corresponding ethidium bromide mutant has a reduced chromosome number of 41 and a number of translocations. The mitochondria displayed morphological alterations compared to the parental lines during the transition phase prior to the isolation of the ethidium bromide-resistant cells. The mitochondria of the ethidium bromide-resistant mutants appear somewhat enlarged with a normal morphology. The effect of ethidium bromide on selected respiratory enzymes in normal and virus-transformed ethidium bromide-resistant baby hamster kidney cells was determined. Ethidium bromide-resistant cells exhibited a depressed level of cytochrome aa3. This depression could not be reversed by growth in ethidium bromide-free media. Ethidium bromide-resistant cells possessed the same cytochrome b, c, and c1 levels per cell as their corresponding parental lines. Purified mitochondria isolated from virus-transformed ethidium bromide-resistant cells exhibited a depression in cytochrome oxidase-specific activity, while the ethidium bromide-resistant control cells did not. All cell lines studied showed a depression in NADH-ferricyanide and NADH-cytochrome c reductase-specific activities relative to their parental BHK21/C13 cells. No increase was observed in virus-transformed ethidium bromide-resistant cells. Ethidium bromide-resistant control cells exhibited a two-fold increase in oligomycin-insensitive adenosine triphosphatase activity relative to their parental cells. All of the cell lines studied possessed equivalent oligomycin-sensitive adenosine triphosphatase-specific activity except for the virus-transformed, dye-resistant mutant, whose activity was increased.

Adenosine Triphosphatases↗