案例教学法结合PBL在肾内科见习教学中的应用

目的:探讨案例教学法结合PBL应用于肾内科临床见习教学中的效果。方法:研究对象为在我院肾内科见习的46名学生,随机分为实验组和对照组,分别采用案例教学法结合PBL和传统教学法,用问卷调查和见习报告分数进行评价。结果:案例教学法结合PBL在教学满意度及见习报告得分等方面显著高于传统教学法。结论:案例教学法结合PBL相较传统教学法,提高了学生的学习兴趣及综合能力,值得推广。     

缓释异搏定和开搏通对高血压病纤溶活性的影响

采用发色底物法测定了50例Ⅰ、Ⅱ期高血压病(EH)患者组织型纤溶酶原激活剂(t-PA)及其抑制物(PAI-1)活性,并与25例正常人对照.其中21例EH患者接受缓释异搏定治疗,19例EH患者接受开搏通治疗,共12周.结果显示:EH患者t-PA活性下降(P<0.05),PAI-1活性升高(P<0.05).缓释异搏定和开搏通有效降压12周后,t-PA活性升高,PAI-1活性降低.提示EH患者纤溶活性下降,缓释异搏定和开搏通在有效降压的同时能改善高血压患者的纤溶活性.     

Alternative Gnas gene products have opposite effects on glucose and lipid metabolism

Gnas is an imprinted gene with multiple gene products resulting from alternative splicing of different first exons onto a common exon 2. These products include stimulatory G protein alpha-subunit (G(s)alpha), the G protein required for receptor-stimulated cAMP production; extralarge G(s)alpha (XLalphas), a paternally expressed G(s)alpha isoform; and neuroendocrine-specific protein (NESP55), a maternally expressed chromogranin-like protein. G(s)alpha undergoes tissue-specific imprinting, being expressed primarily from the maternal allele in certain tissues. Heterozygous mutation of exon 2 on the maternal (E2m-/+) or paternal (E2+/p-) allele results in opposite effects on energy metabolism. E2m-/+ mice are obese and hypometabolic, whereas E2+/p- mice are lean and hypermetabolic. We now studied the effects of G(s)alpha deficiency without disrupting other Gnas gene products by deleting G(s)alpha exon 1 (E1). E1+/p- mice lacked the E2+/p- phenotype and developed obesity and insulin resistance. The lean, hypermetabolic, and insulin-sensitive E2+/p- phenotype appears to result from XLalphas deficiency, whereas loss of paternal-specific G(s)alpha expression in E1+/p- mice leads to an opposite metabolic phenotype. Thus, alternative Gnas gene products have opposing effects on glucose and lipid metabolism. Like E2m-/+ mice, E1m-/+ mice had s.c. edema at birth, presumably due to loss of maternal G(s)alpha expression. However, E1m-/+ mice differed from E2m-/+ mice in other respects, raising the possibility for the presence of other maternal-specific gene products. E1m-/+ mice had more severe obesity and insulin resistance and lower metabolic rate relative to E1+/p- mice. Differences between E1m-/+ and E1+/p- mice presumably result from differential effects on G(s)alpha expression in tissues where G(s)alpha is normally imprinted.     

Immunohistochemical Expression of Components of the Akt-mTORC1 Pathway is Associated with Hepatocellular Carcinoma in Patients with Chronic Liver Disease

Activation of the Akt-mTORC1 signaling pathway was evaluated in premalignant and hepatocellular carcinoma (HCC) lesions by assessing the expression of pS6, an Akt effector, and PTEN, an Akt suppressor. Methods: Immunohistochemical staining for pS6 and PTEN was performed on liver tissue from 52 patients with cirrhosis, with and without HCC. Two pathologists independently evaluated pS6 staining on a semiquantitative scale and categorized PTEN staining as present or absent. Results: In the HCC group, pS6 staining was greatest in HCC, followed by dysplasia, and benign cirrhotic tissue (P < 0.001). pS6 staining was greater in cirrhotic tissue from patients with HCC compared to cirrhosis in patients without HCC (P = 0.03). PTEN staining in tumor was absent in 8/33 (24%) cases. Loss of PTEN expression was more common in patients with higher tumor stage, compared to those with stage 1 tumors (P = 0.04). Conclusion: Immunohistochemical evidence of activation of the Akt-mTORC1 pathway is associated with HCC.     

冠状动脉介入术后腹膜后血肿的危险因素研究

目的探讨冠状动脉介入术(PCI)后腹膜后血肿(RPH)的发生率、临床特征和危险因素。方法对北京安贞医院心内科2000-01—2005-09共3729例经股动脉穿刺进行PCI术的病例进行回顾性分析,其中并发RPH的21例设为观察组,选择另外30例未发生RPH的病例设为对照组。采用单因素和多因素分析RPH发生的危险因素。结果RPH的发生率为0·6%。RPH最常见的临床特征为贫血(100%),其余还包括低血压(95%)、出汗(57%)、腹股沟痛(48%)、下腹痛(38%)和背痛(38%)。RPH的独立预测因素有3个:女性(OR=5·23,P<0·0001)、体表面积(BSA)<1·53m2(OR=7·11,P=0·005)和高部位股动脉穿刺(OR=5·42,P=0·001)。结论女性、低BSA和高部位股动脉穿刺是RPH发生的独立预测因素。     

Phospholipase C-linked receptors regulate the ATP-sensitive potassium channel by means of phosphatidylinositol 4,5-bisphosphate metabolism

In the COS7 cells transfected with cDNAs of the Kir6.2, SUR2A, and M(1) muscarinic receptors, we activated the ATP-sensitive potassium (K(ATP)) channel with a K(+) channel opener and recorded the whole-cell K(ATP) current. The K(ATP) current was reversibly inhibited by the stimulation of the M(1) receptor, which is linked to phospholipase C (PLC) by the G(q) protein. The receptor-mediated inhibition was observed even when protein kinase C (PKC) was inhibited by H-7 or by chelating intracellular Ca(2+) with 10 mM 1, 2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetate (BAPTA) included in the pipette solution. However, the receptor-mediated inhibition was blocked by U-73122, a PLC inhibitor. M(1)-receptor stimulation failed to inhibit the K(ATP) current activated by the injection of exogenous phosphatidylinositol 4,5-bisphosphate (PIP(2)) through the whole-cell patch pipette. The receptor-mediated inhibition became irreversible when the replenishment of PIP(2) was blocked by wortmannin (an inhibitor of phosphatidylinositol kinases), or by including adenosine 5'-[beta,gamma-imido]triphosphate (AMPPNP, a nonhydrolyzable ATP analogue) in the pipette solution. In inside-out patch experiments, the ATP sensitivity of the K(ATP) channel was significantly higher when the M(1) receptor in the patch membrane was stimulated by acetylcholine. The stimulatory effect of pinacidil was also attenuated under this condition. We postulate that stimulation of PLC-linked receptors inhibited the K(ATP) channel by increasing the ATP sensitivity, not through PKC activation, but most probably through changing PIP(2) levels.     

Inhibition of the NF-κB pathway by R65 ribozyme gene via adeno-associatedvirus serotype 9 ameliorated oxidized LDL induced human umbilical vein endothelial cell injury

Objective: NF-κB signaling plays a central role in the regulation of inflammatory responses in atherosclerosis. R65 ribozyme gene suppresses activation of NF-κB pathway, therefore we studied whether R65 gene therapy can ameliorate oxidized low-density lipoprotein (ox-LDL) induced human umbilical vein endothelial cells (HUVECs) injury. Methods and results: Recombinant adeno-associated virus serotype 9 (rAVV9) vector was used to transfect the R65 ribozyme gene (rAVV9-R65) into HUVECs then following ox-LDL stimulation, expression of NF-κB p65 and p50 subunits, inflammatory mediators and cell apoptosis were examined. First, rAVV9-enhanced green fluorescent protein (eGFP)-R65 at 1×10⁷ v.g./cell multiplicity of infection reached a long-lasting and significant increase in R65 gene expression. Second, ox-LDL treatment led to time- and dose-dependent activation of NF-κB pathway, and enhanced inflammatory response and cell death evidenced by increased expression of nuclear NF-κB p65 and p50 subunits, greater production of tumor necrosis factor α, interleukin-6 and von willebrand factor and 20.57% increasedapoptotic HUVECs. Third, over-expression ofR65 gene was 2-fold increased in HUVECs attenuated ox-LDL induced unclear accumulation and expression of p65 subunit and ameliorated inflammation and cell death (all P < 0.05). Conclusion: rAAV9-mediated R65 ribozyme gene transfection in cultured HUVECs effectively inhibits ox-LDL induced activation of NF-κB and production of inflammatory cytokines and prevents cell apoptosis.     

Detection of MGMT promoter methylation in glioblastoma using pyrosequencing

Recent clinical trials on patients with glioblastoma revealed that O6-Methylguanine-DNA methyltransferase (MGMT) methylation status significantly predicts patient’s response to alkylating agents. In this study, we sought to develop and validate a quantitative MGMT methylation assay using pyrosequencing on glioblastoma. We quantified promoter methylation of MGMT using pyrosequencing on paraffin-embedded fine needle aspiration biopsy tissues from 43 glioblastoma. Using a 10% cutoff, MGMT methylation was identified in 37% cases of glioblastoma and 0% of the non-neoplastic epileptic tissue. Methylation of any individual CpG island in MGMT promoter ranged between 33% and 95%, with a mean of 65%. By a serial dilution of genomic DNA of a homogenously methylated cancer cell line with an unmethylated cell line, the analytical sensitivity is at 5% for pyrosequencing to detect MGMT methylation. The minimal amount of genomic DNA required is 100 ng (approximately 3,000 cells) in small fine needle biopsy specimens. Compared with methylation-specific PCR, pyrosequencing is comparably sensitive, relatively specific, and also provides quantitative information for each CpG methylation.