HSP60/10 chaperonin systems are inhibited by a variety of approved drugs,natural products,and known bioactive molecules

All living organisms contain a unique class of molecular chaperones called 60?kDa heat shock proteins (HSP60 – also known as GroEL in bacteria). While some organisms contain more than one HSP60 or GroEL isoform, at least one isoform has always proven to be essential. Because of this, we have been investigating targeting HSP60 and GroEL chaperonin systems as an antibiotic strategy. Our initial studies focused on applying this antibiotic strategy for treating African sleeping sickness (caused by Trypanosoma brucei parasites) and drug-resistant bacterial infections (in particular Methicillin-resistant Staphylococcus aureus – MRSA). Intriguingly, during our studies we found that three known antibiotics – suramin, closantel, and rafoxanide – were potent inhibitors of bacterial GroEL and human HSP60 chaperonin systems. These findings prompted us to explore what other approved drugs, natural products, and known bioactive molecules might also inhibit HSP60 and GroEL chaperonin systems. Initial high-throughput screening of 3680 approved drugs, natural products, and known bioactives identified 161 hit inhibitors of the Escherichia coli GroEL chaperonin system (4.3% hit rate). From a purchased subset of 60 hits, 29 compounds (48%) re-confirmed as selective GroEL inhibitors in our assays, all of which were nearly equipotent against human HSP60. These findings illuminate the notion that targeting chaperonin systems might be a more common occurrence than we previously appreciated. Future studies are needed to determine if the in vivo modes of action of these approved drugs, natural products, and known bioactive molecules are related to GroEL and HSP60 inhibition.     

Investigation of B,C-ring truncated deguelin derivatives as heat shock protein 90 (HSP90) inhibitors for use as anti-breast cancer agents

On the basis of deguelin, a series of the B,C-ring truncated surrogates with N-substituted amide linkers were investigated as HSP90 inhibitors. The structure activity relationship of the template was studied by incorporating various substitutions on the nitrogen of the amide linker and examining their HIF-1α inhibition. Among them, compound 57 showed potent HIF-1α inhibition and cytotoxicity in triple-negative breast cancer lines in a dose-dependent manner. Compound 57 downregulated expression and phosphorylation of major client proteins of HSP90 including AKT, ERK and STAT3, indicating that its antitumor activity was derived from the inhibition of HSP90 function. The molecular modeling of 57 demonstrated that 57 bound well to the C-terminal ATP-binding pocket in the open conformation of the hHSP90 homodimer with hydrogen bonding and pi-cation interactions. Overall, compound 57 is a potential antitumor agent for triple-negative breast cancer as a HSP90 C-terminal inhibitor.     

细叶远志皂苷在Aβ23-35诱导SH-SY5Y 细胞氧化损伤中的作用及机制研究

为研究细叶远志皂苷(tenuifolin,TEN)在Aβ25-35诱导SH-SY5Y细胞氧化损伤中的作用,并探讨其作用机制。建立Aβ25-35诱导的细胞损伤模型,细叶远志皂苷以及自噬抑制剂3-MA进行干预,显微镜观察细胞形态变化,试剂盒检测细胞氧化应激水平,RT-qPCR和Westernblot检测细叶远志皂苷以及自噬抑制剂干预前后Beclin-1、LC3、mTOR、AMPK和ULK1mRNA及蛋白水平变化。结果发现,TEN改善Aβ25-35诱导的SH-SY5Y细胞形态损伤和细胞活力下降;降低ROS和MDA浓度,并提高SOD、GSH-Px及过氧化氢酶的活性;增加AMPK和ULK1的表达,减少mTOR的表达及增加Beclin-1和LC3-Ⅱ/Ⅰ的表达水平。而加入3-MA会拮抗TEN的作用。总之,TEN可能通过调控AMPK/mTOR/ULK1通路,增加Beclin-1及LC3-Ⅱ/Ⅰ蛋白水平激活自噬,进而改善Aβ25-35诱导的细胞形态损伤和细胞活力下降,提高细胞抗氧化应激能力,发挥神经保护作用。     

长沙市不同污染程度区域桂花和香樟叶表面PM2.5吸附量及其影响因素

为阐释不同污染程度下城市绿化植物吸滞PM_2.5机理、解析污染物来源,应用气溶胶再发生器定量测定长沙市常见的2种园林绿化树种(桂花和香樟)植物叶片PM_2.5吸附量,同时应用原子力显微镜(AFM)观察了不同污染区(交通区、文教区、清洁区)植被的叶表面微形态特征,使用离子色谱仪测定样品中水溶性离子含量.结果表明: 污染程度与植物叶表面PM_2.5吸附量呈正相关,不同植物单位叶面积PM_2.5吸附量全年均值表现为交通区(0.56±0.04 μg·cm^-2)＞文教区(0.48±0.06 μg·cm^-2)＞清洁区(0.33±0.02 μg·cm^-2),植物单位叶面积PM_2.5吸附量季节变化为冬季(0.70±0.10 μg·cm^-2)＞春季(0.43±0.14 μg·cm^-2)＞秋季(0.39±0.12 μg·cm^-2)＞夏季(0.31±0.09 μg·cm^-2),桂花的单位叶面积PM_2.5吸附量大于香樟;污染程度轻的区域的植物叶片比较光滑,污染程度重的区域的叶片较粗糙,植物粗糙度排序为交通区(195.45±16.09 nm)＞文教区(176.99±8.45 nm)＞清洁区(131.88±12.98 nm);不同污染程度地区PM_2.5离子含量均表现为冬季最大,其次是春季和秋季,夏季最低;3个污染区PM_2.5离子成分均以Na⁺、NH₄⁺、Cl^-和Br^-这4种离子为主,不同程度污染区PM_2.5污染均以移动源污染为主.     

Mutation of the Apc1 homologue shattered disrupts normal eye development by disrupting G1 cell cycle arrest and progression through mitosis

The shattered1 (shtd1) mutation disrupts Drosophila compound eye structure. In this report, we show that the shtd1 eye defects are due to a failure to establish and maintain G1 arrest in the morphogenetic furrow (MF) and a defect in progression through mitosis. The observed cell cycle defects were correlated with an accumulation of cyclin A (CycA) and String (Stg) proteins near the MF. Interestingly, the failure to maintain G1 arrest in the MF led to the specification of R8 photoreceptor cells that undergo mitosis, generating R8 doublets in shtd1 mutant eye discs. We demonstrate that shtd encodes Apc1, the largest subunit of the anaphase-promoting complex/cyclosome (APC/C). Furthermore, we show that reducing the dosage of either CycA or stg suppressed the shtd1 phenotype. While reducing the dosage of CycA is more effective in suppressing the premature S phase entry in the MF, reducing the dosage of stg is more effective in suppressing the progression through mitosis defect. These results indicate the importance of not only G1 arrest in the MF but also appropriate progression through mitosis for normal eye development during photoreceptor differentiation.     

Peptides derived from Cdk5 activator p35, specifically inhibit deregulated activity of Cdk5

Normal Cdk5 activity, conferred mainly by association with its primary activator p35, is critical for normal function of the cell and must be tightly regulated. During neurotoxicity, p35 is cleaved to form p25, which becomes a potent and mislocalized hyperactivator of Cdk5, resulting in a deregulation of Cdk5 activity. p25 levels have been found to be elevated in Alzheimer's disease (AD) brain and overexpression of p25 in a transgenic mouse results in the formation of phosphorylated tau, neurofibrillary tangles and cognitive deficits that are pathological hallmarks of AD. p25/Cdk5 also hyperphosphorylates neurofilament proteins that constitute pathological hallmarks found in Parkinson's disease and amyotrophic lateral sclerosis. The selective targeting of p25/Cdk5 activity without affecting p35/Cdk5 activity has been unsuccessful. In this review we detail our recent studies of selective p25/Cdk5 inhibition without affecting p35/Cdk5 or mitotic Cdk activities. We found that a further truncation of p25 to yield a Cdk5 inhibitory peptide (CIP) can specifically inhibit p25/Cdk5 activity in transfected HEK cells and primary cortical neurons. CIP was able to reduce tau hyperphosphorylation and neuronal death induced caused by p25/Cdk5 and further studies with CIP may develop a specific Cdk5 inhibition strategy in the treatment of neurodegeneration.     

Heat shock pretreatment enhances porcine myoblasts survival after autotransplantation in intact skeletal muscle

Myoblast transplantation (MT) is a cell-based gene therapy treatment, representing a potential treat-ment for Duchenne muscular dystrophy (DMD), cardiac failure and muscle trauma. The rapid and mas-sive death of transplanted cells after MT is considered as a major hurdle which limits the efficacy of MT treatment. Heat shock proteins (HSPs) are overexpressed when cells undergo various insults. HSPs have been described to protect cells in vivo and in vitro against diverse insults. The aim of our study is to investigate whether HSP overexpression could increase myoblast survival after autotransplantation in pig intact skeletal muscle. HSP expression was induced by warming the cells at 42℃ for 1 h. HSP70 expression was quantified by Western blot and flow cytometry 24 h after the treatment. To investigate the myogenic characteristics of myoblasts, desmin and CD56 were analysed by Western blot and flow cytometry; and the fusion index was measured. We also quantified cell survival after autologous transplantation in pig intact skeletal muscle and followed cell integration. Results showed that heat shock treatment of myoblasts induced a significative overexpression of the HSP70 (P < 0.01) without loss of their myogenic characteristics as assessed by FACS and fusion index. In vivo (n=7), the myoblast survival rate was not significantly different at 24 h between heat shock treated and non- treated cells (67.69% ± 8.35% versus 58.79% ± 8.35%, P > 0.05). However, the myoblast survival rate in the heat shocked cells increased by twofold at 48 h (53.32% ± 8.22% versus 28.27% ± 6.32%, P < 0.01) and more than threefold at 120 h (26.33% ± 5.54% versus 8.79% ± 2.51%, P < 0.01). Histological analy-sis showed the presence of non-heat shocked and heat shocked donor myoblasts fused with host myoblasts. These results suggested that heat shock pretreatment increased the HSP70 expression in porcine myoblasts, and improved the survival rate after autologous transplantation. Therefore, heat shock pretreatment of myoblast in vitro is a simple and effective way to enhance cell survival after transplantation in pig. It might represent a potential method to overcome the limitations of MT treat-ment.     

The effect of Cyclosporine A on cardiomyocytes differentiation

Cyclosporine A (CsA) is a powerful immunosuppressive drug which significantly improved the success of organ transplantation; however, the major limiting factors for the drug's clinical use are its long and short term adverse effects. The present study was conducted to examine, in a dose-dependent manner, in a model of cardiogenesis, the effect of CsA on cardiomyocytes differentiation.     

术前TFF1、HSP70、ASPH与原发性肝癌患者手术切除术后早期复发的关系研究

摘要 目的：研究术前三叶因子1（TFF1）、热休克蛋白70（HSP70）、天冬氨酸-天冬酰胺β羟化酶（ASPH）与原发性肝癌（PHC）患者手术切除术后早期复发的关系。方法：选取2018年1月～2019年12月我院收治的83例PHC患者,均行手术切除治疗,术后均随访2年,根据是否复发分为复发组49例以及未复发组34例。比较两组术前TFF1、HSP70、ASPH水平差异,收集患者基线资料,采用多因素Logistic回归分析PHC患者手术切除术后早期复发的危险因素,采用受试者工作特征（ROC）曲线分析术前TFF1、HSP70、ASPH水平预测PHC患者手术切除术后早期复发的效能。结果：复发组术前TFF1、HSP70、ASPH水平均高于未复发组（P＜0.05）。复发组肿瘤直径≥5 cm、肿瘤数目为多发、有血管侵犯的患者比例高于未复发组（P＜0.05）。多因素Logistic回归分析结果显示：肿瘤直径≥5 cm、多发肿瘤、血管侵犯及术前TFF1、HSP70、ASPH高水平是PHC患者手术切除术后早期复发的危险因素（均OR＞1,P＜0.05）。ROC曲线分析结果显示：术前TFF1、HSP70、ASPH联合检测预测PHC患者手术切除术后早期复发的曲线下面积为0.815,明显高于上述三项指标单独检测的0.704、0.713、0.707。结论：术前TFF1、HSP70、ASPH与PHC患者手术切除术后早期复发密切相关,联合检测三项指标水平可能有助于预测PHC患者手术切除术后早期复发。     

早发性卵巢功能不全患者血清25-OH-VD、DBP、SIRT1与性激素和氧化应激的关系研究

摘要 目的：探讨早发性卵巢功能不全（POI）患者血清25-羟基维生素D（25-OH-VD）、邻苯二甲酸二丁酯（DBP）、沉默信息调节因子2相关酶1（SIRT1）与性激素和氧化应激的关系。方法：选取2019年1月～2020年12月攀枝花学院附属医院妇产科收治的97例POI患者（POI组）,另选取同期54名体检健康女性（对照组）。检测两组血清性激素[卵泡刺激素（FSH）、黄体生成素（LH）、雌二醇（E2）、抗缪勒管激素（AMH）]、氧化应激[超氧化物岐化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）、丙二醛（MDA）]、25-OH-VD、DBP、SIRT1水平。采用Pearson/Spearman相关系数分析POI患者血清25-OH-VD、DBP、SIRT1与性激素、氧化应激指标的相关性,Logistic回归分析血清25-OH-VD、DBP、SIRT1与POI的关系。结果：POI组FSH、LH、MDA、DBP水平高于对照组,E2、AMH、SOD、GSH-Px、25-OH-VD、SIRT1水平低于对照组（P＜0.05）。Pearson/Spearman相关系数显示,POI患者血清25-OH-VD、SIRT1与FSH、LH、MDA呈负相关,与E2、AMH、SOD、GSH-Px呈正相关（P＜0.05）;血清DBP与FSH、LH、MDA呈正相关,与E2、AMH、SOD、GSH-Px呈负相关（P＜0.05）。Logistic回归分析显示,调整混杂因素后,25-OH-VD（OR＝0.825,95％CI：0.741～0.919）、SIRT1（OR＝0.872,95％CI：0.810～0.938）是POI发生的保护因素,DBP（OR＝1.173,95％CI：1.074～1.282）是危险因素（P＜0.05）。结论：POI患者血清25-OH-VD、SIRT1水平下调,DBP水平上调,与性激素和氧化应激相关,可能成为POI的辅助预测因子。