Thermodynamic studies of the interaction of alpha-chymotrypsin with water. I. Determination of the isosteric enthalpies and entropies of water binding to the native enzyme

RNA polymerase enzymes isolated from soybean hypocotyl tissue during successive developmental stages (2-8 days old) have been fractionated by Sephadex column isoelectric focusing. Both the enzymes bound to chromatin and those enzymes free in the soluble phase were investigated during development with respect to their distribution within these two pools. All observed activites were classified according to their alpha-amanitin sensitivity and isoelectric points. Two Class I subspecies (Ia, Ib) and two Class III subspecies (IIIa, IIIb) were continually present bound to chromatin throughout the developmental sequences except the IIb form which was absent at the latest stage. However, a great multiplicity (9 total) of Class II activities (totally inhibited by alpha-amanitin) were observed to be bound to chromatin at the 2nd day stage. These forms were first released from the chromatin complex and recovered in a soluble pool (4th day stage). Subsequent hypocotyl development was accompanied by the gradual disappearance of these Class II subspecies from this pool (6th day) until only two soluble species and one chromatin-bound Class II activity remained (8th day). These observations indicate that the early development of this tissue is accompanied by a dramatic alteration in the conplexity of chromatin-bound RNA polymerase subspecies. Such events may in part determine the domain of RNA secies synthesized at successive developmental stages.     

Response of human blood platelet membrane to sodium selenite.

It was demonstrated that incubation of blood platelets with sodium selenite (1-100 microM) resulted in a dose- and time-dependent loss of platelet thiols (both glutathione and protein -SH groups). The effects of sodium selenite on platelet membrane lipid fluidity by the EPR spin-labelling method was also investigated. We showed there were no alterations in membrane fluidity at the deeper regions (12-DOXYL-Ste) in lipid bilayer, a slight increase (approx. 7%, p < 0.03) of h +1/h0 for spin probe 5-DOXYL-Ste was monitored. The amount of Triton-insoluble protein fraction isolated from platelets after incubation (60 min) with selenite was significantly elevated (p < 0.006). It has been suggested that limited increase in lipid fluidity at the surface regions in the lipid bilayer of the platelet membrane in selenite-treated platelets may be the result of alteration in lipid-protein interactions caused by protein conformational changes.     

In vivo modulation of platelet deposition on human atherosclerotic lesions by various antiaggregatory prostaglandins

The influence of intravenous infusions of various prostaglandins on in vivo platelet function was studied after labelling of autologous platelets with 100 mu ci 111 indium-oxinesulfate in patients with peripheral vascular disease stage II according to FONTAINE. PGI2 (5 ng/kg/min) provoked a significant decrease of platelet deposition and a prolongation in platelet half-life time (74 +/- 6 vs 68 +/- 5 hours). PGE1 (25 ng/kg/min) failed to influence platelet deposition, but prolonged significantly platelet half-life time (82 +/- 6 vs 76 +/- 8 hours). CG 4203 (25 ng/kg/min) decreased significantly platelet deposition and prolonged significantly platelet half-life time (73 +/- 10 vs 67 +/- 11 hours). Iloprost (1 and 2 ng/kg/min) reduced significantly platelet deposition without dose relation. Half-life time was increased significantly after therapy compared to placebo (1 ng: 76 +/- 7 vs 69 +/- 7; 2 ng: 73 +/- 9 vs 67 +/- 9 hours).     

大菜粉蝶颗粒体病毒的形态发生及细胞病理的超微结构观察

本文叙述感染大菜粉蝶颗粒体病毒后,病虫脂肪体细胞超微结构的改变,大菜粉蝶感染后24小时,病虫脂肪体细胞开始出现明显的病变,整个病程是,在开始时细胞核内出现清晰区并出现病毒发生基质,核膜多点成套增生,其后核膜断裂,大量膜样结构聚集在病毒发生基质的周围,核衣壳大量产生,有一部分核衣壳从这些病毒发生基质四周的膜样结构碎片上获得套膜,荚膜蛋白沉积形成成熟的病毒荚膜,或称包含体;另一部分则排列在胞浆内的空泡边缘上;其余的核衣壳则从细胞边缘“芽突”而获得套膜,另外还描述环孔片层及线粒体改变。     

The human fibroblast class II extracellular matrix receptor mediates platelet adhesion to collagen and is identical to the platelet glycoprotein Ia-IIa complex

A monoclonal antibody, P1H5, to the human fibroblast class II extracellular matrix receptor (ECMR II) specifically inhibits human fibroblast adhesion to collagen and immunoprecipitates a cell surface receptor containing an alpha and beta subunit of approximately 140 kilodaltons each (Wayner, E. A., and Carter, W. G. (1987) J. Cell Biol. 105, 1873-1884). We report here that P1H5 also specifically inhibits adhesion of unactivated human platelets to type I and III collagens, but not to fibronectin. Immunoprecipitation of the class II ECMR from Triton X-100 detergent lysates of platelets, after cell surface iodination, identified the platelet collagen receptor. Peptide mapping confirmed that the II alpha and II beta subunits immunoprecipitated from platelets are structurally homologous with those derived from fibroblasts. The platelet ECMR II alpha and -beta subunits comigrate with platelet membrane glycoproteins Ia and IIa, respectively, on two-dimensional nonreduced-reduced sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels. These results indicate that platelet and fibroblast adhesion to collagen are both mediated by a similar receptor and that the alpha and beta subunits of this receptor are identical to platelet membrane glycoproteins Ia and IIa, respectively. Although glycoprotein Ia has been previously implicated as a collagen binding protein, our results are the first direct evidence that platelet glycoprotein Ia is associated with glycoprotein IIa in a heterodimer complex and that this complex, by mediating platelet attachment, is an actual receptor for platelet adhesion to collagen.     

Inactivation of SLIT2-ROBO1/2 pathway in premalignant lesions of uterine cervix: clinical and prognostic significances

The SLIT2-ROBO1/2 pathways control diverse biological processes, including growth regulation. To understand the role of SLIT2 and ROBO1/2 in cervical carcinogenesis, firstly their RNA expression profiles were screened in 21 primary uterine cervical carcinoma (CACX) samples and two CACX cell lines. Highly reduced expressions of these genes were evident. Concomitant alterations [deletion/methylation] of the genes were then analyzed in 23 cervical intraepithelial neoplasia (CIN) and 110 CACX samples. In CIN, SLIT2 was deleted in 22% samples compared to 9% for ROBO1 and none for ROBO2, whereas comparable methylation was observed for both SLIT2 (30%) and ROBO1 (22%) followed by ROBO2 (9%). In CACX, alteration of the genes were in the following order: Deletion:ROBO1 (48%) > SLIT2 (35%) > ROBO2 (33%), Methylation:SLIT2 (34%) > ROBO1 (29%) > ROBO2 (26%). Overall alterations of SLIT2 and/or ROBO1 (44%) and SLIT2 and/or ROBO2 (39%) were high in CIN followed by significant increase in stage I/II tumors, suggesting deregulation of these interactions in premalignant lesions and early invasive tumors. Immunohistochemical analysis of SLIT2 and ROBO1/2 in CACX also showed reduced expression concordant with molecular alterations. Alteration of all these genes predicted poor patient outcome. Multiparous (≥ 5) women with altered SLIT2 and ROBO1 along with advanced tumor stage (III/IV) and early sexual debut (<19 years) had worst prognosis. Our data suggests the importance of abrogation of SLIT2-ROBO1 and SLIT2-ROBO2 interactions in the initiation and progression of CACX and also for early diagnosis and prognosis of the disease.     

Microtubule inhibitors alter the secretion of beta-glucuronidase by human blood platelets: involvement of microtubules in release reaction II

The effects of the antimicrotubular drugs colchicine and vinblastine on the blood platelet release reaction were studied by measuring release of ¹⁴C-5-hydroxytryptamine (¹⁴C-5-HT, release I) and β-glucuronidase (release II) from gel-filtered human platelets. β-glucuronidase release induced by thrombin was significantly inhibited by colchicine (0.01-1 mM) or vinblastine (0.05–0.1 mM). Release of ¹⁴C-5-HT, however, was unaffected at low concentrations of colchicine and only slightly inhibited at higher concentrations. Inhibition of β-glucuronidase release depended on colchicine or vinblastine concentrations and decreased with longer time intervals (1′, 5′, 20′) after thrombin stimulation. Levels of the cytoplasmic enzyme, lactic acid dehydrogenase, in supernatants of colchicine treated platelets were not significantly different from controls. Colchicine also inhibited β-glucuronidse release, but not ¹⁴C-5-HT release, induced by trypsin and sodium arachidonate. Binding of ¹⁴C-colchicine by platelets was measured and it was found that platelet aggregation and release of 5-HT induced by adenosine diphosphate, epinephrine and collagen proceeded without any alteration in colchicine binding. However, significant increases in the rate and degree of colchicine binding were observed when platelets were stimulated by thrombin, trypsin and arachidonic acid which induced aggregation, release of both 5-HT and β-glucuronidase. The results suggest that an alteration in platelet microtubules is correlated with the physiologic response resulting in release II and that the cellular mechanisms effecting release I and II by platelets differ qualitatively in that the microtubules may facilitate release II.     

Contribution of Zinc Deficiency to Insulin Resistance in Patients with Primary Biliary Cirrhosis

The relationship between metabolic abnormalities of trace elements and insulin resistance has been established. Recent studies have revealed that insulin resistance is associated with autoimmune responses. The purpose of this study was to examine the correlation between zinc or copper metabolism and insulin resistance in patients with primary biliary cirrhosis (PBC). Sixteen patients with PBC were divided into two groups: early and advanced stage disease. The overall value of the homeostasis model assessment of insulin resistance (HOMA-IR) in patients with advanced stage PBC was significantly higher than that in patients with early stage PBC, although the mean value in advanced stage PBC was significantly lower than that in hepatitis C virus (HCV)-related liver cirrhosis. There was an inverse correlation between serum zinc concentrations and HOMA-IR values in patients with PBC, while we found no correlation between serum copper levels and HOMA-IR values. HOMA-IR values were inversely associated with peripheral platelet counts, indicating the relationship between insulin resistance and hepatic fibrosis. These results suggest that zinc deficiency plays important roles of insulin resistance and subsequent hepatic fibrosis in patients with PBC, although insulin resistance in advanced stage PBC was significantly milder than that in HCV-related liver cirrhosis.     

Reduction of the chloroplast membrane system caused by disorders in early stages of chlorophyll biosynthesis

A chlorophyll-deficient xantha mutant of cotton (Gossypium hirsutum L.) was examined with respect to development and structural organization of the chloroplast membrane system as affected by disruption of early stages of chlorophyll biosynthesis in the light. The analysis of early chlorophyll precursors showed that the mutant is unable to synthesize 5-aminolevulinic acid (5-ALA) in the light. The disorders in early stages of chlorophyll biosynthesis arrested the development of chloroplast membrane system at the stage of vesicles and single thylakoids. The accumulation of 2–5% chlorophyll in the mutant was related to the formation of light-harvesting chlorophyll-a/b-protein complexes I and II, whereas pigment-protein complexes composing reaction centers of photosystem I and photosystem II were lacking. It is concluded that the chloroplast membrane system in the mutant with impaired 5-ALA synthesis is incapable of development and is even reduced upon long-term growing under light.     

The thrombin high-affinity binding site on platelets is a negative regulator of thrombin-induced platelet activation. Structure-function studies using two mutant thrombins, Quick I and Quick II.

To elucidate the thrombin domains required for high-affinity binding and platelet activation, the platelet binding properties of thrombin and two mutant thrombins, thrombin Quick I and Quick II, were compared to their agonist effects in elevating intraplatelet [Ca2+]. In Quick I, a mutation within the fibrinogen binding groove results in decreased clotting and platelet aggregating activities, whereas in Quick II, a mutation in the primary substrate binding pocket abolishes both activities. Dysthrombin binding was decreased compared to thrombin. The fibrinogen binding groove appeared more important than the primary substrate pocket for high-affinity binding since Quick I showed drastically reduced, and Quick II only slightly reduced, binding affinity (Kd approximately 200 and approximately 10 nM, respectively). The deduced interaction of thrombin with its high-affinity binding site indicated that the thrombin catalytic site is directed toward the platelet surface and therefore, when bound, is proteolytically inactive. Quick I (0.5-5 nM) elicited intraplatelet [Ca2+] fluxes at concentrations where high-affinity binding was undetectable. Saturation of high-affinity binding sites with active-site-modified thrombin did not affect thrombin-induced (0.5 nM) or Quick I-induced (5 nM) responses. In contrast, addition of D-Phe-Pro-Arg chloromethyl ketone (FPRCK) subsequent to thrombin or Quick I stimulation of platelets abolished agonist-induced responses. Since Quick I was only 10-17% as effective as thrombin in increasing intraplatelet [Ca2+], our data support a model in which thrombin acts enzymatically on a platelet membrane "substrate", through an interaction mediated in part by the fibrinogen binding groove of thrombin. This conclusion is consistent with the inhibition observed with high concentrations (greater than 100 nM) of Quick II and FPRCK-modified thrombin (FPR-thrombin) in platelets stimulated with low concentrations of thrombin (less than 0.5 nM) or Quick I (less than 2 nM), consistent with inhibition by substrate depletion. In contrast, concentrations of FPR-thrombin or Quick II (less than 100 nM), which saturated predominantly the high-affinity binding sites, enhanced the platelet responses induced by thrombin (less than 0.5 nM). Thus, occupation of the high-affinity sites with inactive thrombin increased the concentration of active thrombin available for substrate interaction. Quick I-induced responses were not enhanced, consistent with its inability to interact with the high-affinity site. Since thrombin bound to the high-affinity site is proteolytically inactive, we hypothesize that the thrombin high-affinity binding site on platelets functions to alter thrombin activity and platelet activation.     

Regulation of angiotensin II receptors and extracellular matrix turnover in human retinal pigment epithelium: role of angiotensin II

The early stage of age-related macular degeneration (AMD) is characterized by the formation of subretinal pigment epithelium (RPE) deposits as a result of the dysregulation in the turnover of extracellular matrix (ECM) molecules. However, the mechanism involved remains unclear. Hypertension (HTN) is an important risk factor for AMD, and angiotensin II (ANG II) is the most important hormone associated with HTN. However, the relevance of ANG II receptors and ANG II effects on RPE have not been investigated yet. Therefore, the expression and regulation of ANG II receptors as well as the ECM turnover were studied in human RPE. ANG II receptors were expressed and upregulated by ANG II in human RPE. This regulation resulted in functional receptor expression, since an increase in intracellular concentration of calcium was observed upon ANG II stimulation. ANG II also increased matrix metalloproteinase (MMP)-2 activity and MMP-14 at the mRNA and protein levels as well as type IV collagen degradation. These ANG II effects were abolished in the presence of the ANG II receptor subtype 1 (AT1) receptor antagonist candesartan. In contrast, ANG II decreased type IV collagen via both AT1 and AT2 receptors, suggesting a synergistic effect of the two receptor subtypes. In conclusion, we have confirmed the presence of ANG II receptors in human RPE and their regulation by ANG II as well as the regulation of ECM molecules via ANG II receptors. Our data support the hypothesis that ANG II may exert biological function in RPE through ANG II receptors and that ANG II may cause dysregulation of molecules that play a major role in the turnover of ECM in RPE basement membrane and Bruch's membrane, suggesting a pathogenic mechanism to explain the link between HTN and AMD.     

Aggregation of spectrin and PKCθ is an early hallmark of fludarabine/mitoxantrone/dexamethasone-induced apoptosis in Jurkat T and HL60 cells

It has been shown that changes in spectrin distribution in early apoptosis preceded changes in membrane asymmetry and phosphatidylserine (PS) exposure. PKCθ was associated with spectrin during these changes, suggesting a possible role of spectrin/PKCθ aggregation in regulation of early apoptotic events. Here we dissect this hypothesis using Jurkat T and HL60 cell lines as model systems. Immunofluorescent analysis of αIIβII spectrin arrangement in Jurkat T and HL60 cell lines revealed the redistribution of spectrin and PKCθ into a polar aggregate in early apoptosis induced by fludarabine/mitoxantrone/dexamethasone (FND). The appearance of an αIIβII spectrin fraction that was insoluble in a non-ionic detergent (1% Triton X-100) was observed concomitantly with spectrin aggregation. The changes were observed within 2 h after cell exposure to FND, and preceded PS exposure. The changes seem to be restricted to spectrin and not to other cytoskeletal proteins such as actin or vimentin. In studies of the mechanism of these changes, we found that (i) neither changes in apoptosis regulatory genes (e.g., Bcl-2 family proteins) nor changes in cytoskeleton-associated proteins were detected in gene expression profiling of HL60 cells after the first hour of FND treatment, (ii) caspase-3, -7, -8, and -10 had minor involvement in the early apoptotic rearrangement of spectrin/PKCθ, and (iii) spectrin aggregation was shown to be partially dependent on PKCθ activity. Our results indicate that spectrin/PKCθ aggregate formation is related to an early stage in drug-induced apoptosis and possibly may be regulated by PKCθ activity. These findings indicate that spectrin/PKCθ aggregation could be considered as a hallmark of early apoptosis and presents the potential to become a useful diagnostic tool for monitoring efficiency of chemotherapy as early as 24 h after treatment.     

Serum steroid hormones in female Russian sturgeon (Acipenser gueldenstaedtii Br.) with normal oocyte maturation and with gonadal function disorders after hormonal stimulation

Blood serum cortisol, testosterone, and 11-ketotestosterone levels were determined by ELISA methods in female Russian sturgeon after hormonal stimulation for induction of ovulation. All females were divided into 5 groups depending on their initial reproductive status and response to hormonal stimulation; 1) normal ovulation, 2) ovulation followed by release of non-viable eggs, 3) normal state of ovary without ovulation, 4) lack of ovulation, early stage of oocyte resorption, 5) lack of ovulation, advanced stage of oocyte resorption. Cortisol levels in all five groups did not differed significantly. Testosterone levels were low (15-24 ng/ml) in females after ovulation and in females without response to hormonal stimulation because of early stage of oocyte resorption. Non-matured females with normal state of oocytes had significantly higher testosterone levels--82 +/- 16 ng/ml. Non-matured females with advanced stage of resorption were subdivided into 2 subgroups--with low (approximately 7 ng/ml) and high (> 100 ng/ml) levels of serum testosterone. 11-ketotestosterone levels were similar in all investigated groups of sturgeon.     

In vivo inhibition of megakaryocyte and platelet production by platelet factor 4 in mice.

The in vivo effect of human platelet factor 4 (PF4) on murine megakaryocytopoiesis and thrombopoiesis was studied. Administration of PF4 induced a dose-dependent decrease in the numbers of megakaryocytes and their progenitor cells (CFU-MK), continuing for 1 week after the injection. However, the size of megakaryocytes and their colonies was not changed following PF4 injection. Platelet levels were significantly decreased at days 3-4. The number of CFU-GM was decreased at days 1-2. White blood cells and hemoglobin were unaffected by PF4. These data indicate that PF4 inhibits megakaryocyte and platelet production in vivo by acting on the early stage of megakaryocyte development.     

脊髓损伤后脊髓神经细胞膜PAF受体特性的变化

采用３ＨＰＡＦ放射配体结合试验方法测定脊髓神经细胞膜上ＰＡＦ受体的特异性位点,观察脊髓损伤后２、６ｈ、１、３周脊髓神经细胞膜ＰＡＦ受体结合特性变化。结果显示,脊髓神经细胞膜上存在ＰＡＦ高、低亲和力结合位点,脊髓损伤后２、６ｈ、１周组ＰＡＦ受体高、低亲和力位点Ｋｄ值和Ｂｍａｘ均有不同程度下降,与对照组比较,有显著性差异（Ｐ＜０．０５）。表明在伤后早期ＰＡＦ受体亲和力增加,结合位点减少。提示ＰＡＦ受体在脊髓损伤后继发性损害病理生理过程中起一定作用。     

不同CRRT治疗时机对脓毒症合并急性肾功能不全患者疗效及预后的影响

目的:探讨不同持续性肾脏替代治疗(CRRT)治疗时机对脓毒症合并急性肾功能不全患者的临床疗效及预后的影响。方法:将我院ICU收治的60例脓毒症合并急性肾功能不全患者,按照CRRT治疗时机分为早期组(1-2期,n=30)和晚期组(3期,n=30)。比较两组患者治疗前后不同时点平均动脉压(MAP)、白细胞(WBC)计数、血红蛋白(HB)、血小板(PLT)计数、急性生理学与慢性健康状况(APACHE)Ⅱ评分等临床资料的变化,机械通气时间,肾功能恢复率及28 d病死率等。结果:与早期组比较,晚期组治疗后WBC计数明显升高(P0.05)。治疗后12 h、24 h、72 h,早期组ACHEⅡ评分较晚期组显著降低(P0.05)。与晚期组比较,早期组机械通气时间显著缩短,肾功能恢复明显升高,28d内病死率也明显降低(P0.05)。结论:脓毒症合并急性肾功能不全患者应早期启动CRRT治疗,最佳介入时间是KDIGO-AKI 3期之前,有助于改善患者预后。     

Comparison between in vitro lipid peroxidation in fresh sheep platelets and peroxidative processes during sheep platelet ageing under storage at 4 degrees C.

Incubation of sheep platelet crude membranes with xanthine oxidase (XO)/hypoxanthine/Fe(2+)-ADP revealed: (i) a fast peroxidative response - with a maximal linear rate of 14 nmol malondialdehyde (MDA) equivalents/mg protein, as evidenced by the thiobarbituric acid test - and a decrease in the polyunsaturated fatty acid (PUFA) content of the platelet crude membranes; (ii) a decrease in the lipid fluidity in the deep lipid core of the membranes but not at the membrane surface; (iii) a dramatic inhibitory effect on glucose 6-phosphatase (Glc-6-Pase) but not on acetylcholinesterase activity. Platelets were also aged by storage at 4 degrees C in their own plasma or in Seto additive solution. In these media, platelet aggregates were visible and the effects on platelet phospholipids, PUFA, lipid extract fluorescence, crude membrane fluidity and membrane-bound enzyme activities were assessed for comparison with those observed in in vitro lipid peroxidation. The sensitivity of membranes from stored platelets to lipid peroxidation was also assessed. Storage of platelets in plasma for 5 days was associated with different changes in their crude membranes such as decreases in arachidonic acid contents, the decrease not being avoided by the presence of phospholipase A(2) inhibitors, increases in MDA equivalents, conjugated dienes and lipid extract fluorescence, decreases in the amounts of MDA equivalents formed by platelet crude membranes treated with the oxidizing agents, changes in membrane fluidity and inhibition of Glc-6-Pase. All these alterations were less pronounced or even abolished after platelet storage in Seto. These findings suggest that platelet lipid peroxidation due to XO/hypoxanthine/Fe(2+)-ADP and platelet membrane alterations observed after platelet ageing under storage at 4 degrees C share common features. Also, as regards the prevention of peroxidative processes, Seto solution permits better storage of sheep platelets than plasma.     

Global methylation analysis identifies PITX2 as an upstream regulator of the androgen receptor and IGF-I receptor genes in prostate cancer

The insulin-like growth factor-I (IGF-IR) and androgen (AR) receptors are important players in prostate cancer. Functional interactions between the IGF-I and androgen signaling pathways have crucial roles in the progression of prostate cancer from early to advanced stages. DNA methylation is a major epigenetic alteration affecting gene expression. Hypermethylation of tumor suppressor promoters is a frequent event in human cancer, leading to inactivation and repression of specific genes. The aim of the present study was to identify the entire set of methylated genes ("methylome") in a cellular model that replicates prostate cancer progression. The methylation profiles of the P69 (early stage, benign) and M12 (advanced stage, metastatic) prostate cancer cell lines were established by treating cells with the demethylating agent 5-aza-2'-deoxycytidine (5-Aza) followed by DNA microarray analysis. Comparative genome-wide methylation analyses of 5-Aza-treated versus untreated cells identified 297 genes overexpressed in P69 and 191 genes overexpressed in M12 cells. 102 genes were upregulated in both benign and metastatic cell lines. In addition, our analyses identified the PITX2 gene as a master regulator upstream of the AR and IGF-IR genes. The PITX2 promoter was semi-methylated in P69 cells but fully methylated (i. e., silenced) in M12 cells. Epigenetic regulation of PITX2 during the course of the disease may lead to orchestrated control of the AR and IGF signaling pathways. In summary, our results provide new insights into the epigenetic changes associated with progression of prostate cancer from an organ confined, androgen-sensitive disorder to an aggressive, androgen-insensitive disease.     

Sex related differences in platelet TxA2 generation

Platelet lipid composition, c arachidonic acid (AA) metabolism by platelets (stimulated with thrombin), serum thromboxane (Tx)B2 production and plasma lipid composition were investigated in 53 healthy females (18-45 years) and 65 males (19-45 years) with similar dietary habits. In males, serum TxB2 production and cholesterol platelet membrane levels were found significantly higher (p less than 0.001 and p less than 0.05) than in females. No differences were observed between the two groups in the AA conversion through cyclo-oxygenase and lipoxygenase pathways or in the platelet phospholipid fatty acid composition. These findings indicate that in males the platelet proaggregatory capacity is greater than in females and the higher platelet TxB2 production does not depend on a larger AA availability or on enzyme activation for its conversion. The increased TxB2 production may be, at least in part, induced by functional differences such as a different membrane cholesterol content inducing, in its turn, an increased microviscosity and/or higher number of platelet receptors for thrombin.     

Phytochemical and biological investigation of Aristolochia maurorum L

Aristolochia maurorum L. of Jordanian origin has been investigated phytochemically, quantitatively, and biologically. Three atypical alkaloids, namely aristolochic acid I (1), aristolochic acid II (2) and aristolochic acid IIIa (3), have been isolated and identified. Of these known 1-phenanthrenecarboxylic acids, 2 and 3 are reported for the first time from this species. The identified compounds 1-3 were first evaluated biologically as cytotoxic agents against the brine shrimp lethality test (BST), in which compound 1 was found to be the most potent (LC50, 4.9 microg/mL). The antiplatelet activity of the methanolic extracts, the acidic fractions of aerial and root parts, and the identified compounds 1-3 were evaluated using an automatic platelet aggregometer and coagulation tracer (APACT 2). Using external reference standards, and a reverse-phase isocratic method, the distribution of aristolochic acid I and aristolochic acid II in different plant parts of Aristolochia maurorum L. during flowering stage was analyzed by PDA-HPLC. A quantitative comparison between two previously reported extraction methods was also made. Roots were found to be the main storage of aristolochic acid I and aristolochic acid II during flowering stage with about 0.22 and 0.108% (w/w), respectively.