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Biomedical subjects

M Tepel

Publications and source records attributed to M Tepel.

At least 37 records · Page 2Linked to original sources
PubMed2001

Homocyst(e)ine metabolism in hemodialysis patients treated with vitamins B6, B12 and folate.

Hyperhomocyst(e)inemia is commonly accepted as an independent atherosclerotic risk factor. In most hemodialysis patients, serum homocyst(e)ine is markedly elevated and may contribute to premature atherosclerosis in these patients. Whereas the beneficial effect of folate supplementation on serum homocyst(e)ine has been extensively studied, there are less detailed studies on the effects of cobalamin and pyridoxal phosphate alone, or in combination with folate. We examined the effect of a four-week course of intravenous treatment with folate (1.1 mg), cobalamin (1.0 mg), and pyridoxal phosphate (5.0 mg), administered once (group 1), twice (group 2) or thrice (group 3) weekly in 33 hemodialysis patients divided in three groups of 11 patients. All patients were followed for a further four weeks after treatment was stopped. Serum homocyst(e)ine, cobalamin, folate and pyridoxal phosphate, as well as the metabolites of homocyst(e)ine, methylmalonate, 2-methylcitrate and cystathionine, were determined before, during and after treatment. Baseline serum homocyst(e)ine correlated significantly with serum folate (P=0.0149), cobalamin (P=0.0047) and pyridoxal phosphate (P=0.0408). Correlations independent from the other metabolites or vitamins were found for methylmalonate (P=0.003) and folate (P=0.029). All regimens increased serum cobalamin significantly (in group 1 from 444 +/- 215 to 17,303 +/- 11,989 pg/ml, P<0.01; in group 2 from 542 +/- 633 to 44,896 +/- 15,772 pg/ml, P<0.001; in group 3 from 548 +/- 394 to 77,961 +/- 31,546 pg/ml, P<0.001), but did not increase any of the other vitamin levels. Serum homocyst(e)ine was lowered significantly by 39.8% +/- 31.9% (P<0.05) with two and by 30.1% +/- 26.9% (P<0.05) with three vitamin dosages weekly, but not with one dosage weekly. Since methylmalonate is known to be a sensitive marker of cobalamin deficiency, the data support an important influence of cobalamin levels on baseline homocyst(e)ine levels. Increasing cobalamin levels and additional treatment with folate and pyridoxal phosphate 156 may decrease serum homocyst(e)ine in the same way as high doses of folate alone.

Female↗
PubMed2000

Dinucleotides as growth-promoting extracellular mediators. Presence of dinucleoside diphosphates Ap2A, Ap2G, and Gp2G in releasable granules of platelets.

Dinucleoside diphosphates, Ap(2)A, Ap(2)G, and Gp(2)G represent a new class of growth-promoting extracellular mediators, which are released from granules after activation of platelets. The presence of theses substances was shown after purification from a platelet concentrate. The substances were identified by UV spectrometry, retention time comparison with authentic substances, matrix-assisted laser desorption/ionization mass spectrometry, post-source-decay matrix-assisted laser desorption/ionization mass spectrometry, and enzymatic analysis. Ap(2)A, Ap(2)G, and Gp(2)G have growth-stimulating effects on vascular smooth muscle cells in nanomolar concentrations as shown by [(3)H]thymidine incorporation measurements. The calculated EC(50) (log m; mean +/- S.E.) values were -6.07 +/- 0.14 for Ap(2)A, -6.27 +/- 0.25 for Ap(2)G, and -6.91 +/- 0.44 for Gp(2)G. At least 61.5 +/- 4.3% of the dinucleoside polyphosphates are released by platelet activation. The intraplatelet concentrations suggest that, in the close environment of a platelet thrombus, similar dinucleoside polyphosphate concentrations can be found as in platelets. Intraplatelet concentration can be estimated in the range of 1/20 to 1/100 of the concentration of ATP. In conclusion, Ap(2)A, Ap(2)G, and Gp(2)G derived from releasable granules of human platelets may play a regulatory role in vascular smooth muscle growth as growth-promoting mediators.

Animals↗
PubMed2000

Activation of phosphatidylinositol-specific phospholipase C by HDL-associated lysosphingolipid. Involvement in mitogenesis but not in cholesterol efflux.

Our earlier studies demonstrated that high-density lipoproteins (HDLs) stimulate multiple signaling pathways, including activation of phosphatidylcholine-specific phospholipases C and D (PC-PLs) and phosphatidylinositol-specific phospholipase C (PI-PLC). However, only activation of PC-PLs was linked to the HDL-induced cholesterol efflux. In the study presented here, the role of HDL-induced PI-PLC activation was studied. In human skin fibroblasts, HDL potently induced PI-PLC as inferred from enhanced phosphatidylinositol bisphosphate (PtdInsP(2)) turnover and Ca(2+) mobilization. The major protein component of HDL, apo A-I, did not induce PtdInsP(2) turnover or Ca(2+) mobilization in these cells. Both HDL and apo A-I promoted cellular cholesterol efflux, whereas only HDL induced fibroblast proliferation. Inhibition of PI-PLC with U73122 or blocking intracellular Ca(2+) elevation with Ni(2+) or EGTA markedly reduced the extent of HDL-induced cell proliferation but had no effect on cholesterol efflux. In fibroblasts from patients with Tangier disease which are characterized by defective cholesterol efflux, neither HDL-induced PtdInsP(2) breakdown and Ca(2+) mobilization nor cell proliferation was impaired. HDL-induced fibroblast proliferation, PtdInsP(2) turnover, and Ca(2+) mobilization were fully mimicked by the lipid fraction isolated from HDL. Analysis of this fraction with high-performance liquid chromatography (HPLC) and time-of-flight secondary ion mass spectroscopy (TOF-SIMS) revealed that the PI-PLC-inducing activity is identical with two bioactive lysosphingolipids, namely, lysosulfatide (LSF) and sphingosylphosphorylcholine (SPC). Like native HDL, LSF and SPC induced PtdInsP(2) turnover, Ca(2+) mobilization, and fibroblast proliferation. However, both compounds did not promote cholesterol efflux. In conclusion, two agonist activities are carried by HDL. Apo A-I stimulates phosphatidylcholine breakdown and thereby facilitates cholesterol efflux, whereas LSF and SPC trigger PI-PLC activation and thereby stimulate cell proliferation.

Apolipoprotein A-I↗
PubMed2000

Isolation and characterization of coenzyme A glutathione disulfide as a parathyroid-derived vasoconstrictive factor.

BACKGROUND: Coenzyme A glutathione disulfide (CoA-SSG) was recently isolated from bovine adrenal glands and was shown to be a renal vasoconstrictor. The identification of CoA-SSG in human parathyroid glands and its action on cultured vascular smooth muscle cells (VSMCs) are described here. METHODS AND RESULTS: After purification to homogeneity by several chromatographic steps, CoA-SSG was identified by matrix-assisted laser desorption/ionization mass spectrometry and enzymatic analysis. The dose-dependent growth-stimulating effect of CoA-SSG on VSMCs, measured by the [(3)H]thymidine method, is characterized by a threshold of 10(-)(8) mol/L and a maximum effect of 10 micromol/L, increasing VSMC proliferation 254+/-21% above control. A dose of 10 micromol/L methylmalonyl-CoA and 10 micromol/L CoA increased the rate of proliferation of VSMCs only by 178+/-43% and 50+/-42% above control, respectively. Glutathione has no proliferative effect on VSMCs. The growth-stimulating effect of CoA-SSG (1 micromol/L) was decreased by the antagonists 3,7-dimethyl-1-propargylxanthine (DMPX; 11 micromol/L) (38% compared with CoA-SSG without antagonist) and pyridoxal-phosphate-6-azophenyl-2,4-disulfonic acid (PPADS; 10 micromol/L) (48% compared with CoA-SSG without antagonist; each P:<0. 05 versus control), indicating that the effect is mediated partly via A(2) and partly via P(2)Y(1) and/or P(2)Y(4) receptor. CONCLUSIONS: CoA-SSG may play a regulatory role in VSMC growth as a progression factor and thereby could play an important role in development of hypertension.

Acetates↗
PubMed2000

[Homocysteine and its metabolites in chronic renal insufficiency and the effect of a vitamin replacement].

BACKGROUND: Hyperhomocysteinemia has been increasingly recognized as an important risk factor for elevated atherosclerotic vascular disease in chronic renal failure. We measured in patients with chronic renal failure homocysteine and metabolites of its 2 metabolic pathways, transulfuration (cystathionine, cysteine) and remethylation (methionine, methylmalonic acid, 2-methylcitric acid). PATIENTS AND METHODS: Eleven patients on conservative treatment (creatinine clearance 10 to 30 ml/min) and 50 chronic uremic subjects on regular hemodialysis were included in the study. Twenty-two of the dialysis patients received daily oral multivitamin supplementation containing 10 mg vitamin B6, 6 micrograms vitamin B12, and 1 mg folic acid during the last year before the study started. RESULTS: In the hemodialysis group homocysteine levels were higher compared with the patients on conservative treatment. Hemodialysis patients with additional vitamin supplementation showed significantly lower homocysteine levels than those without. The pattern of metabolites was different to these results: all metabolites were higher in hemodialysis patients, too (significant for cysteine and methionine), but vitamin supplementation failed to lower all metabolites. CONCLUSION: Analysis of metabolites additional to homocysteine levels may help to understand different results in evaluation of atherosclerotic risk of hyperhomocysteinemia in chronic renal failure.

Adult↗
PubMed2000

Involvement of phospholipase D in store-operated calcium influx in vascular smooth muscle cells.

In non-excitable cells, sustained intracellular Ca2+ increase critically depends on influx of extracellular Ca2+. Such Ca2+ influx is thought to occur by a 'store-operated' mechanism, i.e. the signal for Ca2+ entry is believed to result from the initial release of Ca2+ from inositol 1,4,5-trisphosphate-sensitive intracellular stores. Here we show that the depletion of cellular Ca2+ stores by thapsigargin or bradykinin is functionally linked to a phosphoinositide-specific phospholipase D (PLD) activity in cultured vascular smooth muscle cells (VSMC), and that phosphatidic acid formed via PLD enhances sustained calcium entry in this cell type. These results suggest a regulatory role for PLD in store-operated Ca2+ entry in VSMC.

Animals↗
PubMed2000

Prevention of radiographic-contrast-agent-induced reductions in renal function by acetylcysteine.

BACKGROUND: Radiographic contrast agents can cause a reduction in renal function that may be due to reactive oxygen species. Whether the reduction can be prevented by the administration of antioxidants is unknown. METHODS: We prospectively studied 83 patients with chronic renal insufficiency (mean [+/-SD] serum creatinine concentration, 2.4+/-1.3 mg per deciliter [216+/-116 micromol per liter]) who were undergoing computed tomography with a nonionic, low-osmolality contrast agent. Patients were randomly assigned either to receive the antioxidant acetylcysteine (600 mg orally twice daily) and 0.45 percent saline intravenously, before and after administration of the contrast agent, or to receive placebo and saline. RESULTS: Ten of the 83 patients (12 percent) had an increase of at least 0.5 mg per deciliter (44 micromol per liter) in the serum creatinine concentration 48 hours after administration of the contrast agent: 1 of the 41 patients in the acetylcysteine group (2 percent) and 9 of the 42 patients in the control group (21 percent; P=0.01; relative risk, 0.1; 95 percent confidence interval, 0.02 to 0.9). In the acetylcysteine group, the mean serum creatinine concentration decreased significantly (P<0.001), from 2.5+/-1.3 to 2.1+/-1.3 mg per deciliter (220+/-118 to 186+/-112 micromol per liter) 48 hours after the administration of the contrast medium, whereas in the control group, the mean serum creatinine concentration increased nonsignificantly (P=0.18), from 2.4+/-1.3 to 2.6+/-1.5 mg per deciliter (212+/-114 to 226+/-133 micromol per liter) (P<0.001 for the comparison between groups). CONCLUSIONS: Prophylactic oral administration of the antioxidant acetylcysteine, along with hydration, prevents the reduction in renal function induced by contrast agents in patients with chronic renal insufficiency.

Acetylcysteine↗
PubMed2000

[Progressive renal failure. Why blood pressure must now be monitored].

Hypertension alone can lead to chronic nephrosclerosis and in addition promote the progressive worsening of renal function in various forms of renal disease, such as diabetic nephropathy, glomerular nephritis or interstitial nephritis. Thus, the treatment of chronic renal disease associated with reduced excretory function, requires not only general measures and a low-protein diet, but also intensified anti-hypertension treatment with diuretics or beta blockers. In a number of studies, progression or renal insufficiency has been shown to be slowed in particular through the use of angiotensin-converting enzyme inhibitors in patients with chronic renal failure.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2000

Phospholipase A(2) is involved in thapsigargin-induced sodium influx in human lymphocytes.

Previously, we reported that emptying of intracellular Ca(2+) pools with endoplasmatic Ca(2+)-ATP-ase inhibitor thapsigargin leads to the Na(+) influx in human lymphocytes (M. Tepel et al., 1994, J. Biol. Chem. 269, 26239-26242). In the present study we examined the mechanism underlying the thapsigargin-induced Na(+) entry. We found that the thapsigargin-induced increase in Na(+) concentration was effectively inhibited by three structurally unrelated phospholipase A(2) (PLA(2)) inhibitors, p-bromophenacyl bromide, 3-(4-octadecyl)-benzoylacrylic acid (OBAA), and bromoenol lactone (BEL). The thapsigargin-induced Na(+) influx could be mimicked by PLA(2) exogenously added to the lymphocyte suspension. In addition, thapsigargin stimulated formation of arachidonic acid (AA), the physiological PLA(2) product. AA induced Na(+) entry in a time- and concentration-dependent fashion. Both, thapsigargin-induced Na(+) influx and AA liberation were completely inhibited in the presence of tyrosine kinase inhibitor genistein but not in the absence of extracellular Ca(2+). Collectively, these data show that thapsigargin-induced Na(+) entry is associated with tyrosine kinase-dependent stimulation of PLA(2).

Acetophenones↗
PubMed2000

D609-phosphatidylcholine-specific phospholipase C inhibitor attenuates thapsigargin-induced sodium influx in human lymphocytes.

Previously, we reported that the phosphatidylcholine-specific phospholipase C (PC-PLC) inhibitor tricyclodecan-9-yl xanthogenate (D609) potentiates thapsigargin-induced Ca(2+) influx in human lymphocytes. In the present study we examined the effect of D609 on the thapsigargin-induced Na(+) entry. We found that the early phase of the thapsigargin-induced increase in the intracellular Na(+) concentration (approx. 1-2 min after stimulation) was attenuated after preincubation of lymphocytes with D609. By contrast, thapsigargin-induced Na(+) influx was not affected in the presence butan-1-ol, which inhibits phosphatidylcholine-specific phospholipase D (PC-PLD). The thapsigargin-induced Na(+) influx could be mimicked by PC-PLC exogenously added to the lymphocyte suspension, whereas addition of PC-PLD had no effect. In addition, thapsigargin stimulated formation of the physiological PC-PLC products, diacylglycerol. Cell-permeable diacylglycerol analogue, dioctanoyl-glycerol (DOG), produced time- and concentration-dependent increase in the intracellular Na(+) concentration. Both thapsigargin- and DOG-induced Na(+) increases were not affected in the presence of Na(+)/H(+) antiport inhibitor, HOE609, or Na(+)/Ca(2+) antiport inhibitor, dimethylthiourea, as well as in the presence of Co(2+) and Ni(2+), which block store-operated Ca(2+) entry. By contrast, markedly reduced thapsigargin- and DOG-induced Na(+) influx were noted in the presence of flufenamic acid, which blocks the non-selective cation current (I(CRANC)). In conclusion, our results suggest that diacylglycerol released due to the PC-PLC activation contributes to the thapsigargin-induced Na(+) entry.

Biological Transport↗
PubMed2000

Platelet function in patients with obstructive sleep apnoea syndrome.

Patients with obstructive sleep apnoea syndrome (OSAS) are subject to an increased cardiovascular morbidity including myocardial infarction and stroke. Platelets play an important role in the pathogenesis and triggering of acute cardiovascular syndromes. So far, the influence of OSAS on platelet function is not fully understood. Platelet aggregability to epinephrine, collagen, arachidonic acid, and adenosine diphosphate in vitro was measured in 17 consecutive male patients (53.0+/-2.1 yrs) with polysomnographically verified OSAS and compared with that of 15 male controls (50.1+/-3.6 yrs) at 20:00 h, 24:00 h, and 06:00 h. In addition, the long-term effects of continuous positive airway pressure (CPAP) therapy on platelet aggregability was assessed after 6 months. Platelet aggregation in vitro induced by epinephrine showed a slight increase overnight in the untreated OSAS patients (NS) whereas it decreased slightly (NS) in the controls and in the treated OSAS patients. Pretherapeutic platelet aggregability was significantly lowered by CPAP therapy both at 24:00 h (64.0+/-6.5 versus 55.3+/-6.7%, p<0.05) and at 06:00 h (64.1+/-6.5 versus 45.8+/-7.6%; p=0.01). Platelet aggregability during sleep in the controls resembled that found in patients with OSAS during CPAP therapy. The results suggest that obstructive sleep apnoea syndrome contributes, at least in part, to platelet dysfunction and that long-term continuous positive airway pressure treatment may reduce platelet aggregability.

Adult↗
PubMed2000

Increased sodium-proton antiporter activity in patients with obstructive sleep apnoea.

Cytosolic pH (pH(i)) and the activity of the sodium-proton antiporter (Na(+)/H(+) antiporter) were measured in lymphocytes from 22 patients with obstructive sleep apnoea and from 24 age-matched healthy subjects (Controls). The cellular Na(+)/H(+) antiporter was measured spectrophotometrically using a pH-sensitive fluorescent dye after intracellular acidification using sodium propionate. Resting pHi was similar in lymphocytes from patients with obstructive sleep apnoea and from controls (7.36 +/- 0.20, n=22; vs. 7.35 +/- 0.19, n=24; mean +/- SD). The Na(+)/H(+) antiporter activity was significantly higher in patients with obstructive sleep apnoea than in controls (11.87 +/- 3.26 x 10(-3) pH(i)/s vs. 4.38 +/- 1.40 x 10(-3) pH(i)/s; P < 0. 0001). The apparent affinity of the Na+/H+ antiporter was not significantly different between the groups (6.90 +/- 0.23 vs. 6.87 +/- 0.20). In patients with obstructive sleep apnoea the activity of the Na(+)/H(+) antiporter remained stable during the night. The activity of the Na(+)/H(+) antiporter was 13.49 +/- 4.80 x 10(-3) pH(i)/s at 20.00 and 13.26 +/- 6.13 x 10(-3) pH(i)/s at 02.00. From the present results it is concluded that an increased cellular Na(+)/H(+) antiporter activity may be a genetic marker for patients who are predisposed to obstructive sleep apnoea.

Aged↗
PubMed2000

Increased intracellular reactive oxygen species in patients with end-stage renal failure: effect of hemodialysis.

BACKGROUND: Reactive oxygen species (ROS) have been implicated in various forms of cellular injury. ROS may cause cell damage and are involved in the pathophysiology of several diseases, including atherosclerosis and chronic inflammation. METHODS: Disturbances of intracellular ROS levels were investigated in 28 patients with end-stage renal failure. The intracellular ROS levels were measured in lymphocytes before and after hemodialysis using biocompatible membranes and were compared with those from 11 patients with end-stage renal failure, not yet on renal replacement therapy, and 27 healthy control subjects. ROS levels were measured spectrophotometrically using the intracellular dye dichlorofluorescin diacetate. RESULTS: The spontaneous production of ROS was significantly higher in lymphocytes from patients with end-stage renal failure compared with healthy control subjects (P < 0.01). The addition of 100 nmol/L phorbol-myristate-acetate (PMA) produced a significant increase of ROS, both in lymphocytes from patients with end-stage renal failure and healthy control subjects. The PMA-induced ROS increase was significantly higher in lymphocytes from patients with end-stage renal failure compared with healthy control subjects (P < 0.01). In patients with end-stage renal failure, not yet on renal replacement therapy, the PMA-induced ROS was also significantly higher compared with healthy control subjects. The PMA-induced ROS increases were significantly inhibited by catalase, but not by superoxide dismutase or the superoxide dismutase mimetic, tempol. PMA-induced ROS was significantly reduced by tyrphostin A51 in lymphocytes from patients with end-stage renal failure and from healthy control subjects (each P < 0.01), indicating the involvement of a tyrosine kinase-dependent pathway. In patients with end-stage renal failure, the spontaneous and the PMA-induced production of ROS was not significantly different before and after hemodialysis. CONCLUSIONS: Regular hemodialysis sessions using biocompatible membranes have no effect on the elevated intracellular ROS in patients with end-stage renal failure.

Aged↗
PubMed2000

Increased intracellular generation of reactive oxygen species in mononuclear leukocytes from patients with diabetes mellitus type 2.

Since increased cellular production of reactive oxygen species is a source of oxidative stress and thus may contribute to the development of diabetic complications, the baseline and stimulated concentrations of intracellular reactive oxygen species were measured in 16 patients with diabetes mellitus type 2 and 19 healthy control subjects. Reactive oxygen species and cytosolic calcium were monitored spectrophotometrically using dihydrorhodamine-123 and fura-2, respectively, in a suspension of mononuclear leukocytes. Measurements were made in the presence or absence of superoxide dismutase, sodium azide, genistein, or bisindolylmaleimide 1. Baseline reactive oxygen species concentrations were significantly higher in diabetic patients compared with control (p<0.001). Activation of mononuclear leukocytes by formyl-Met-Leu-Phenylalanine and phytohemagglutinin significantly increased reactive oxygen species in diabetic patients compared with control (p<0.05). The formyl-Met-Leu-Phenylalanine-induced increases were unchanged in the presence of superoxide dismutase, but dropped significantly in the presence of sodium azide by 80% and 73% in diabetic patients and control, respectively (each p<0.01). The formyl-Met-Leu-Phenylalanine-induced responses were significantly inhibited by genistein and bisindolylmaleimide 1 (p<0.01), suggesting the involvement of tyrosine kinase and protein kinase C. Resting calcium (p<0.05) and stimulated calcium were significantly greater in diabetic patients than in control. The results show that patients with diabetes mellitus type 2 generate increased reactive oxygen species under stimulated conditions, suggesting increased risk for oxidative stress and associated complications.

Calcium↗
PubMed1999

Identification and characterization of P(1), P(7)-Di(adenosine-5')-heptaphosphate from human platelets.

Diadenosine pentaphosphate and diadenosine hexaphosphate have been isolated in human platelets and have been postulated to play an important role in the control of vascular tone. Here we describe the isolation and identification of diadenosine heptaphosphate from human platelets. Dinucleoside polyphosphates were concentrated by affinity chromatography from a nucleotide-containing fraction from deproteinated human platelets. Dinucleoside polyphosphates were purified by anion-exchange and reversed phase high performance liquid chromatography to homogeneity. Analysis of one of these fractions with matrix-assisted laser desorption/ionization mass spectrometry revealed a molecular mass of 1076.4 (1077.4 = [M + H](+)) Da. UV spectroscopic analysis of this fraction showed the spectrum of an adenosine derivative. Comparison of the postsource decay matrix-assisted laser desorption/ionization mass spectrum of the fraction minus that of diadenosine heptaphosphate (Ap(7)A) demonstrated that the isolated substance was identical to Ap(7)A. The identity of the retention times of the authentic and the isolated compound confirmed this result. Enzymatic analysis demonstrated an interconnection of the phosphate groups with the adenosines in the 5'-positions of the riboses. With thrombin-induced platelet aggregation, Ap(7)A is released from the platelets into the extracellular space. The vasoconstrictive action of Ap(7)A on the vasculature of the isolated perfused rat kidney Ap(7)A was slightly less than that of Ap(6)A. The threshold of the vasoconstrictive action of Ap(7)A was 10(-5) mol/liter. The vasoconstrictive effect was abolished by suramin and pyridoxal phosphate 6-azophenyl-2', 4'-disulfonic acid, suggesting an activation of P(2x) receptors. Furthermore, Ap(7)A inhibits ADP-induced platelet aggregation. Thus, the potent vasoconstrictor Ap(7)A derived from human platelets, like other diadenosine polyphosphates, may play a role in the regulation of vascular tone and hemostasis.

Adenosine Diphosphate↗
PubMed1999

High-performance liquid chromatographic assay of the diadenosine polyphosphates in human platelets.

Diadenosine pentaphosphate (Ap5A) and diadenosine hexaphosphate (Ap6A) were recently identified in human platelets and were shown to be important modulators of cardiovascular function. Here we describe an HPLC assay for quantitating Ap3A, Ap4A, Ap5A, and Ap6A contents in human platelets simultaneously. Di(1,N6-ethenoadenosine) hexaphosphate was used as internal standard. The extraction procedure consists of (a) deproteinization, (b) selective concentration of the diadenosine polyphosphates with a boronate affinity chromatography, and (c) desalting prior to the HPLC analysis. The assay was validated by PSD-MALDI-mass spectrometry and by addition of authentic diadenosine polyphosphate to platelet samples. The assay was carried out by an ion-pair reversed-phase perfusion chromatography. In platelets from human blood the following amounts of diadenosine polyphosphates were determined: Ap3A, 192.5 +/- 151.0 nM; Ap4A, 223.8 +/- 172.3 nM; Ap5A, 100.2 +/- 81.1 nM; Ap6A, 32.0 +/- 19.6 nM (mean +/- SD, n = 105). The described assay can be used with less than 20 ml blood and allows quantitation of the diadenosine polyphosphates in the picomole range.

Blood Platelets↗
PubMed1999

Electron-probe X-ray Microanalysis of Magnesium and Sodium Ion Content in Vascular Smooth Muscle Cells from Spontaneously Hypertensive and Normotensive Rats.

Whereas in blood cells decreased magnesium concentrations and increased sodium concentrations in essential hypertension have often been described, only sparse data exist on cellular magnesium or sodium content in vascular smooth muscle cells. Therefore, in aortic smooth muscle cells from seven spontaneously hypertensive rats (SHR) of the Münster strain and seven normotensive Wistar-Kyoto rats (WKY), the intracellular magnesium and sodium content were measured by electron-probe X-ray microanalysis. Measurements were performed in aortic cryosections 3 µm thick. The magnesium ion content was 0.93 +/- 0.17 g/kg dry weight in SHR vs 1.14 +/- 0.12 g/kg dry weight in WKY (p < 0.05). Vascular smooth muscle sodium ion content was measured at 6.85 +/- 0.59 g/kg dry weight in WKY and 12.47 +/- 1.62 g/kg dry weight in SHR (p < 0.05). In conclusion, aortic smooth muscle cells from SHR are characterized by a markedly lowered intracellular magnesium ion content and increased sodium ion concentrations compared with normotensive cells. The results may be due to genetically determined disturbances in transmembrane magnesium and sodium ion transport. Cellular magnesium and sodium handling may be disturbed in SHR aortic smooth muscle as it is in hypertensive blood cells.

Journal Article↗