Effect of atorvastatin upon platelet activation in hypercholesterolemia, evaluated by flow cytometry

Hyperlipidemia is a well established risk factor for cardiovascular disease and atherothrombotic events, in which platelet activation also plays a significant role. However, very few studies have addressed platelet activation in hypercholesterolemia, the potential effect of lipid lowering drugs upon platelet hyperfunction, and the question of whether changes in the latter are correlated to normalization of plasma lipids. This study used whole blood flow cytometry to assess in vivo and in vitro platelet activation in a group of 33 patients with hypercholesterolemia, and also the ex vivo effect of atorvastatin (20 mg/day) upon such activation. A control group of 40 normolipidemic volunteers matched in terms of age, sex and added risk factors to the patient group was used. The results showed that hypercholesterolemic patients had in vivo a significantly greater percentage of GPIIb/IIIa- and phosphatidylserine-positive platelets compared with the control group (4.62+/-3.51% and 2.58+/-1.19% versus 2.73+/-1.08% and 1.54+/-0.68%, respectively). In vitro response of CD62 expression to thrombin was also greater in the patients than in the controls (92.51+/-6.00% versus 89.63+/-10.72%, p<0.05). Atorvastatin therapy normalized platelet hyperfunction in the patients studied and reduced GPIIb/IIIa response to ADP (from 82.65+/-6.43% to 75.84+/-4.89%, p<0.01). A significant correlation can be seen between such normalization and the decrease in plasma levels of total and LDL cholesterol.     

Effect of acetaminophen on expression and activity of rat liver multidrug resistance-associated protein 2 and P-glycoprotein

We evaluated the effect of acetaminophen (APAP), given as a single, 1g/kg body weight dose, on expression and activity of rat liver multidrug resistance-associated protein 2 (Mrp2) and P-glycoprotein (P-gp), two major canalicular drug transporters. The studies were performed 24h after administration of the drug. APAP induced an increase in plasma membrane content of Mrp2 detected by western blotting, consistent with increased detection of the protein at the canalicular level by immunoflourescence microscopy. In vivo biliary excretion of dinitrophenyl-S-glutathione, a well known Mrp2 substrate, was slightly but significantly increased by APAP, agreeing well with upregulation of the transporter. Basal biliary excretion of oxidized glutathione, an endogenous Mrp2 substrate, was also increased by APAP, likely indicating increased hepatic synthesis as a result of APAP-induced oxidative stress followed by accelerated canalicular secretion mediated by Mrp2. APAP also increased the expression of P-gp detected by western blotting and immunofluorescence microscopy as well as the in vivo biliary secretory rate of digoxin, a model P-gp substrate. Because specific APAP-conjugated metabolites are Mrp2 substrates, we postulate that induction of Mrp2 by APAP may represent an adaptive mechanism to accelerate liver disposition of the drug. In addition, increased Mrp2-mediated elimination of oxidized glutathione may be essential in maintaining the redox equilibrium in the hepatocyte under conditions of APAP-induced oxidative stress.     

盐酸水提法制备丹参水溶性有效部位群及其对成骨细胞活性测定

目的探讨丹参水溶性有效部位群(salvia miltiorrhiza utility aqueous extract,DUAE,丹参骨宝2号)的制备工艺,质量控制及其对成骨细胞活性测定。方法采用盐酸水提法处理丹参药材,制得丹参骨宝2号,用高效液相法(HPLC)进行含量测定;通过PNPP检测法比较DUAE对SD大鼠成骨细胞(osteoblast,OB)的碱性磷酸酶(ALP)表达水平的影响,评价丹参骨宝2号药物机制。结果用盐酸水提法制备的丹参骨宝2号与传统水提方法比较,丹参素含量提高3·28倍,为(11·50±0·52)mg·g-1生药,丹酚酸B含量减低1·46倍,为(16·40±1·22)mg·g-1生药,且两者含量均在1%~1·5%,原儿茶醛含量增加1·46倍,为(0·233±0·02)mg·g-1生药。传统水提法制备的DUAE药量为1mg生药·L-1时仅提高成骨细胞ALP活性5·5%±0·3%。丹参骨宝2号药量为0·25mg生药·L-1时提高成骨细胞ALP活性达52%±10%。结论用盐酸水提法制备的丹参骨宝2号中含有较高的丹参素和一定比例的丹酚酸B、原儿茶醛,且成分含量稳定,质量可控,对成骨细胞活性测定显示药效好,为研究丹参抗骨质疏松新制剂提供有益资料。     

螺旋弹簧和橡皮链拉尖牙向远中的临床效果研究

目的研究Ni-Ti螺旋弹簧和橡皮链施力后,尖牙远中移动速率、龈沟液(gingival crevicular fluid,GCF)中碱性磷酸酶(alkaline phosphatase,ALP)活性,探讨不同持续张力的正畸效果和生物学效应。方法用OPAK直丝弓矫治技术托槽,排齐整平后,采用0.018×0.025″SS的Ni-Ti方丝进行尖牙远中移动。然后,用初始力150g的Ni-Ti螺旋弹簧和橡皮链牵引尖牙向远中,21d后,采用游标卡尺测定牙齿移动距离,Beckman全自动生化分析仪检测龈沟液中碱性磷酸酶(GCF-ALP)的活性。结果螺旋弹簧和橡皮链施力21d后,尖牙向远中移动距离差异有显著性(P<0.05)。在螺旋弹簧和橡皮链持续牵引力作用下,GCF-ALP活性随时间推移逐渐增加,6个月时达到最高水平,与加力前比较,差异有显著性(P<0.05);6个月后,GCF-ALP开始下降,12个月时,橡皮链加力侧GCF-ALP活性基本恢复到加力前的水平。但在6和12个月时,GCF-ALP活性在螺旋弹簧施力侧显著高于橡皮链施力侧,差异有显著性(P<0.01)。结论螺旋弹簧拉尖牙向远中移动速率较快,产生的生物学效应较强而持久。     

Acute and subchronic oral toxicity assessment of the herbal formula Kai-Xin-San

Ethnopharmacological relevance

Kai-Xin-San (KXS) is a famous traditional Chinese medicine (TCM) formula. It has been used in the treatment of diseases including neurasthenia, Alzheimer's disease and neurosis.

Aim of the study

To provide information on the potential toxicity of KXS, we evaluated the acute and subchronic toxicity in rodents.

Materials and methods

In acute study, a single administration of KXS was given orally to mice at doses ranging from 19.67 to 60.04 g/kg. In the sub-chronic oral toxicity study, KXS was administered to rats at 0, 1, 3 and 9 g/kg for 13 weeks. Moreover, 30 days of post treatment (withdrawal study) was conducted. Mortalities, clinical signs, body weight changes, food and water consumption, haematological and biochemical parameters, gross findings and organ weights were monitored during the study period.

Results

In the sub-chronic study in rats, daily oral administration of KXS at the dose of 9 g/kg/day result in significant increase in WBC, lymphocyte, alkaline phosphatase, blood sugar and significant decrease in bodyweight, serum Cre, CK and CHO at the last week of treatment. Recovery except for the body weight was observed after 30 days of post treatment.

Conclusions

KXS is relatively safe for oral medication. The LD₅₀ of KXS was over 32.59 g/kg for mice. The no-observed-adverse-effect-level (NOAEL) was considered to be 19.67 g/kg/day for rats.     

生长板软骨细胞促进骨髓基质干细胞血管内皮生长因子的表达

目的观察骨髓基质干细胞(BMSCs)在生长板软骨细胞旁分泌作用下血管内皮生长因子(VEGF)的表达规律及其与成骨分化的相关性。方法大鼠BMSCs与生长板软骨细胞进行间接共培养,培养终末期做细胞化学染色,定量测定碱性磷酸酶(ALP)活性,用RT-PCR方法半定量检测VEGFmRNA的表达。结果生长板软骨细胞持续高表达VEGF。BMSCs随共培养时间的延长,ALP活性升高,BMSCs的VEGF的表达也逐渐增强。培养液加入两种分泌型VEGF中和抗体后,VEGF表达趋势不变,ALP活性仍为升高趋势,也不影响培养终末期钙化结节的形成。培养终末期BMSCs的CD31和CD34均阴性。结论BMSCs成骨分化过程中VEGF的表达符合成骨细胞分化基因的表达规律,与成骨细胞特征性基因的表达趋势一致,体外条件共培养条件下,中和VEGF后并不能阻碍BMSCs的成骨分化。     

Indocyanine green loaded hyaluronan-derived nanoparticles for fluorescence-enhanced surgical imaging of pancreatic cancer

Pancreatic ductal adenocarcinoma is highly lethal and surgical resection is the only potential curative treatment for the disease. In this study, hyaluronic acid derived nanoparticles with physico-chemically entrapped indocyanine green, termed NanoICG, were utilized for intraoperative near infrared fluorescence detection of pancreatic cancer. NanoICG was not cytotoxic to healthy pancreatic epithelial cells and did not induce chemotaxis or phagocytosis, it accumulated significantly within the pancreas in an orthotopic pancreatic ductal adenocarcinoma model, and demonstrated contrast-enhancement for pancreatic lesions relative to non-diseased portions of the pancreas. Fluorescence microscopy showed higher fluorescence intensity in pancreatic lesions and splenic metastases due to NanoICG compared to ICG alone. The in vivo safety profile of NanoICG, including, biochemical, hematological, and pathological analysis of NanoICG-treated healthy mice, indicates negligible toxicity. These results suggest that NanoICG is a promising contrast agent for intraoperative detection of pancreatic tumors.     

Selective impairment of drug-metabolizing enzymes in pig liver during subchronic dietary exposure to aflatoxin B1.

Consequences of subchronic exposure to aflatoxin B1 (AFB1) on liver monooxygenase and transferase enzymes were compared in control pigs and pigs given 385, 867 or 1,807 microg AFB1/kg of feed for 4 weeks. Animals exposed to the highest dose of toxin developed clinical signs of aflatoxicosis, like liver fibrosis, hepatic dysfunction and decreased weight gain. This group had significantly lower levels of liver cytochrome P450, ethoxyresorufin O-deethylase (EROD) activity, testosterone metabolism, P450 1A and P450 3A protein expression. By comparison, mild degenerative hepatic changes, no hepatic dysfunction but a similar pattern of liver P450 enzymes activity without changes in P450 3A expression were observed in pigs exposed to 867 microg AFB1/kg of feed. Benzphetamine and aminopyrine N-demethylase activities were increased in pigs exposed to 867 or 1,807microg AFB1/kg of feed. Pigs exposed to 385 microg AFB1/kg of feed had low levels of EROD activity and all other biotransformation and clinical parameters remained at control levels. Aniline hydroxylase activity, P450 2C protein expression, UDP-glucuronosyl and glutathione S-transferase activities were unaffected at all doses of AFB1. In conclusion, P450 1A and P450 3A appear to be specific targets of AFB1 even if pig did not display clinical sign of liver toxicosis.     

Immunomagnetic tumor cell selection--implications for the detection of disseminated cancer cells

BACKGROUND: The optimal method for the detection of disseminated epithelial cancer cells has not yet been found. The standard method, using immunocytochemistry, offers a sensitivity of up to 10(-6). Molecular methods such as cytokeratin-19 RT-PCR are about 10 times as sensitive, but they are hampered by interference such as illegitimate gene expression. STUDY DESIGN AND METHODS: Immunomagnetic bead selection of epithelial cancer cells using conjugates directed against the human epithelial antigen (HEA) followed by immunocytochemistry testing was investigated in this trial. RESULTS: No cytokeratin-positive cells could be enriched from 56 control samples. In 104 clinical samples of bone marrow aspirations, PBPC collections, and venous blood obtained from breast cancer patients, the cytokeratin-positive rate increased significantly, from 29.9 percent before selection to 54.8 percent after enrichment. Even the yield of detected cancer cells was significantly higher after selection. Up to 2.5 x 10(8) MNCs were easily processed. However, the mean cancer cell recovery after HEA enrichment was only 24.4 percent. Subsequently, selected epithelial cells were successfully immunophenotyped by use of a double-stain technique detecting cytokeratin-positive cells and the urokinase-like plasminogen activator receptor. CONCLUSION: HEA bead selection in combination with the standard immunocytochemistry method is a powerful and specific tool for the detection of disseminated cancer cells without false-positive results. Furthermore, it delivers enough cells for subsequent investigations such as characterization studies.     

抗坏血酸加透明质酸对软骨钙化化骨的影响

本实验观察到抗坏血酸（ＶＣ）加透骨质酸（ＨＡ）对胚鸡肢体骨的生长、发育有促进作用．能提高碱性磷酸酶活性（ＡＪＰ），使骨钙含量、骨长度、骨重量均增加。ＨＡ能协助ＶＣ促进钙化化骨．本实验用维生素Ｄ３（ＶＤ３）作阳性对照。