Expression and localization of heat shock proteins in rat basophilic leukemia cells: differential modulation by degranulation,thermal or oxidative stress

BACKGROUND: Rat basophilic leukemia (RBL-2H3) cells are well characterized in terms of morphological and biochemical changes upon activation, and have been extensively used as a model system for studying the mechanisms of the immediate hypersensitivity reaction. To investigate whether overexpression of heat shock/stress proteins (HSP) is involved in the mast cell-dependent reactivity, we examined the adaptive responses of RBL-2H3 cells to classical stress conditions such as heat shock or oxidative injury produced by an aqueous extract of tobacco smoke. METHODS: HSP were determined by flow cytometry and immunocytochemistry. Degranulation was confirmed as the release of beta-hexosaminidase, determined spectrophotometrically, and by electron microscopy experiments. RESULTS: We found that RBL-2H3 cells respond to heat shock or oxidative injury by the synthesis of both the inducible 72 kDa HSP (Hsp70), and the oxidation-specific 32 kDa heme oxygenase (HO)-1. Heat shock induced mainly Hsp70 in a cell growth-dependent manner, whereas oxidative stress induced mainly HO-1 in a cell growth-independent manner. However, heat shock or oxidative stress had no significant effects on degranulation. CONCLUSION: Stress-mediated synthesis of HSP was not associated with RBL-2H3 degranulation and likewise, degranulation did not induce HSP.     

染料木黄酮诱导人卵巢癌HO-8910细胞凋亡的实验研究

目的研究染料木黄酮对人HO-8910卵巢癌细胞的抑制效应和诱导凋亡作用,探讨其机制。方法将不同浓度的染料木黄酮作用于人HO-8910卵巢癌细胞,利用MTT法检测其有效作用浓度,分别采用瑞氏染色、TUNEL、流式细胞仪观察和检测HO-8910细胞凋亡情况。结果MTT法测得其IC50值为28.63 mg/L,浓度为8~128 mg/L的染料木黄酮具有诱导人HO-8910卵巢癌细胞凋亡的作用,且随药物作用时间延长凋亡率逐渐增加。结论染料木黄酮具有抗卵巢癌作用,其重要作用机制之一是诱导人HO-8910卵巢癌细胞凋亡。     

Heme oxygenase1 (HSP-32) is induced in myelin-phagocytosing Schwann cells of injured sciatic nerves in the rat

Schwann cells participate in myelin phagocytosis in the early stage of Wallerian degeneration, prior to the recruitment of macrophages. This is the first report that Schwann cells induce heme oxygenase-1 (HO-1), a 32-kDa heat shock protein, only when they have transformed into myelin-phagocytosing cells from myelinating cells (days 2-3) immediately after crush injury of rat sciatic nerves. Double immunofluorescent labelling for HO-1 and transferrin receptors revealed that HO-1-immunoreactive Schwann cells also expressed transferrin receptors suggesting activation of iron metabolism. The transient induction of HO-1 in Schwann cells may contribute to the adaptive function in an altered environment when the cells have lost contact with axons, and may play a crucial role in the ensuing regeneration.     

Identification of Genes Differentially Expressed in Cultured Human Osteoblasts Versus Human Fibroblasts by DNA Microarray Analysis

Little is known about patterns of gene expression from cells populating the connective tissues. This study investigated the possible variance of gene expression profile between human osteoblasts (HO) and human fibroblasts (HF) in vitro, using DNA microarray technology. Clustering identification was used to compare expression patterns between HO and HF for biological significance. Our results showed that genes encoding the extracellular matrix or apoptosis-related proteins tended to be expressed in greater abundance in HO, while more proteolysis-related proteins were expressed in higher level in HF. Significant differences in expression were also noted with genes related to signaling pathways. To confirm the array results, three genes (periostin, MFG-E8, MMP-10) were selected and analyzed independently by RT-PCR and northern blot. The results were found consistent with the array data in HO and HF. The present findings suggest that HO and HF differ not only phenotypically but in the expression level of tissue specific genes to assure the turnover and homeostasis of their respective tissues.     

In vitro and in vivo metabolism and detection of 3‐HO‐PCP,a synthetic phencyclidine,in human samples and pooled human hepatocytes using high resolution mass spectrometry

The new psychoactive substance (NPS) 3‐HO‐PCP, a phencyclidine (PCP) analog, was detected in a law enforcement seizure and in forensic samples in Denmark. Compared with PCP, 3‐HO‐PCP is known to be a more potent dissociative NPS, but no toxicokinetic investigations of 3‐HO‐PCP are yet available. Therefore, 3‐HO‐PCP was quantified in in vivo samples, and the following were investigated: plasma protein binding, in vitro and in vivo metabolites, and metabolic targets. All samples were separated by liquid chromatography and analyzed by mass spectrometry. The unbound fraction in plasma was determined as 0.72 ± 0.09. After in vitro incubation with pooled human hepatocytes, four metabolites were identified: a piperidine‐hydroxyl‐and piperidine ring opened N‐dealkyl‐COOH metabolite, and O‐glucuronidated‐ and O‐sulfate‐conjugated metabolites. In vivo, depending on the sample and sample preparation, fewer metabolites were detected, as the O‐sulfate‐conjugated metabolite was not detected. The N‐dealkylated‐COOH metabolite was the main metabolite in the deconjugated urine sample. in vivo analytical targets in blood and brain samples were 3‐HO‐PCP and the O‐glucuronidated metabolite, with 3‐HO‐PCP having the highest relative signal intensity. The drug levels of 3‐HO‐PCP quantified in blood were 0.013 and 0.095 mg/kg in a living and a deceased subject, respectively. The 3‐HO‐PCP concentrations in deconjugated urine in a sample from a living subject and in post‐mortem brain were 7.8 and 0.16 mg/kg, respectively. The post mortem results showed a 1.5‐fold higher concentration of 3‐HO‐PCP in the brain tissue than in the post mortem blood sample.     

Molecular signalling of a novel curcumin derivative versus Tadalafil in erectile dysfunction

The efficacy of a novel curcumin derivative (NCD) versus tadalafil in erectile signalling was assessed. Ten control male rats and 50 diabetic male rats were used and divided into the following: diabetic (DM), curcumin (CURC), NCD, tadalafil and NCD combined with tadalafil rat groups. Cavernous tissue gene expression of heme oxygenase‐1 (HO‐1), Nrf2, NF‐B and p38, enzyme activities of heme oxygenase (HO) and nitric oxide synthase (NOS), cGMP and intracavernosal pressure (ICP)/mean arterial pressure (MAP) were assessed. Results showed that 12 weeks after induction of diabetes, erectile dysfunction (ED) was confirmed by the significant decrease in ICP/MAP, a significant decrease in cGMP, NOS, HO enzyme activities, a significant decrease in HO‐1 gene and a significant increase in NF‐?β, p38 genes. Administration of all therapeutic interventions led to a significant increase in ICP/MAP, cGMP levels, a significant increase in HO‐1 and NOS enzymes, a significant increase in HO‐1, and Nrf2 gene expression, and a significant decrease in NF‐?β, p38 gene expression. NCD or its combination with tadalafil showed significant superiority and more prolonged duration of action. In conclusion, a tendency was observed that CURC and NCD have high efficacy and more prolonged duration of action in enhancing erectile function.     

银杏内酯注射液联合巴曲酶治疗急性脑梗死的临床研究

目的探讨银杏内酯注射液联合巴曲酶治疗急性脑梗死的临床疗效。方法选取2015年2月—2019年2月在河南医学高等专科学校附属医院治疗的急性脑梗死患者78例,根据用药的不同分为对照组(39例)和治疗组(39例)。对照组静脉滴注巴曲酶注射液,首次10 BU加入生理盐水100 mL,隔日5 BU加入生理盐水100 mL,1次/2 d;治疗组在对照组基础上静脉滴注银杏内酯注射液,10 mL加入生理盐水250 mL,1次/d。两组患者均治疗14 d。观察两组患者临床疗效,同时比较治疗前后两组患者NIHSS评分、m RS评分、MoCA评分、SF-36评分、ADL评分、BI指数,血清血红素氧合酶1(HO1)、单核细胞趋化蛋白-1(MCP-1)、亲环素A(CyPA)、和肽素(CPP)、神经胶质纤维酸性蛋白(GFAP)和基质金属蛋白酶-9(MMP-9)水平,双侧大脑中动脉动脉峰流速(Vp)、平均流速(Vm)、流速差值(DVp、DVm),及血液红细胞压积(HCT)、全血黏度(WBV)、纤维蛋白原(FIB)和血浆黏度(PV)水平。结果治疗后,对照组临床有效率为82.05%,显著低于治疗组的97.44%,两组比较差异有统计学意义(P0.05)。治疗后,两组患者NIHSS和m RS评分显著降低(P0.05),MoCA评分、SF-36评分、ADL评分和BI指数评分显著升高(P0.05),且治疗组NIHSS、m RS、MoCA、SF-36、ADL和BI指数评分明显好于对照组(P0.05)。治疗后,两组血清HO1、MCP-1、Cy PA、CPP、GFAP、MMP-9水平均明显下降(P0.05),且治疗组HO1、MCP-1、CyPA、CPP、GFAP、MMP-9水平明显低于对照组(P0.05)。治疗后,两组患者双侧大脑中动脉Vp、Vm均增高(P0.05),DVp、DVm明显降低(P0.05),且治疗组脑血流动力学水平明显好于对照组(P0.05)。治疗后,两组患者HCT、WBV、FIB及PV水平明显降低(P0.05),且治疗组这些血液流变学指标明显低于对照组(P0.05)。结论银杏内酯注射液联合巴曲酶注射液治疗急性脑梗死有利于促进患者脑神经功能恢复,改善机体细胞因子水平,促进机体血液流变学及脑血流动力学指标的改善。     

脑络通胶囊联合丁苯酞治疗急性脑梗死的临床研究

目的探讨脑络通胶囊联合丁苯酞氯化钠注射液治疗急性脑梗死的临床疗效。方法选取2018年2月—2019年2月在朝阳市第二医院治疗的急性脑梗死患者94例,根据住院号分为对照组(47例)和治疗组(47例)。对照组患者静脉滴注丁苯酞氯化钠注射液,100 mL/次,2次/d;治疗组在对照组基础上给予脑络通胶囊,1.0 g/次,3次/d。两组患者均治疗2周。观察两组患者临床疗效,同时比较治疗前后两组患者NIHSS、MoCA、m RS和SF-36评分,血清血红素氧合酶1(HO1)、新蝶呤(Npt)、肽素(CP)、血小板激活因子(PAF)、亲环素A(CyPA)和可溶性肿瘤坏死因子相关凋亡诱导配体(sTRAIL)水平,亦及红细胞压积(HCT)、血浆黏度(PV)、全血黏度(WBV)、纤维蛋白原(FIB)、动态阻力(DR)、相对平均血流量(Qmean)、脑血管周围阻力(R)、平均血流速度(Vmean)和特性阻抗(ZCV)改变。结果治疗后,对照组临床有效率为80.85%,显著低于治疗组的97.87%,两组比较差异有统计学意义(P0.05)。治疗后,两组患者NIHSS评分和m RS评分均显著降低(P0.05),而MoCA评分和SF-36评分均明显升高(P0.05),且治疗组患者这些评分明显好于对照组(P0.05)。治疗后,两组患者血清HO1、Npt、CPP、PAF、CyPA、sTRAIL水平均明显降低(P0.05),且治疗组患者这些血清学指标明显低于对照组(P0.05)。治疗后,两组患者HCT、PV、WBV、FIB、ZCV和R水平均明显下降,而Qmean、DR和Vmean均显著升高(P0.05),且治疗组患者这些指标水平明显好于对照组(P0.05)。结论脑络通胶囊联合丁苯酞治疗急性脑梗死可促进神经功能恢复、血液流变学改善,以及提高生活质量。