Sensitive HPLC-APCI-MS method for the determination of cyclovirobuxine D in human plasma

A sensitive high performance liquid chromatography-atmospheric pressure chemical ionisation-mass spectrometry (HPLC-APCI-MS) assay for determination of cyclovirobuxine D (CVB-D) in human plasma using mirtazapine as internal standard (I.S.) was established. After adjustment to a basic pH with sodium hydroxide, plasma was extracted by ethyl acetate and separated by high performance liquid chromatography (HPLC) on a reversed-phase C(18) column with a mobile phase of 30 mM ammonium acetate buffer solution containing 1% formic acid-methanol (48:52, v/v). CVB-D was determined with atmospheric pressure chemical ionisation-mass spectrometry (APCI-MS). HPLC-APCI-MS was performed in the selected-ion monitoring (SIM) mode using target ions at [M+H](+)m/z 403.4 for CVB-D and [M+H](+)m/z 266.2 for I.S. Calibration curves were linear over the range 10.11-4044 pg/ml. The lower limit of quantification was 10.11 pg/ml. The intra- and inter-run variability values were less than 9.5 and 12.4%, respectively. The mean plasma extraction recovery of CVB-D was in the range of 85.3-92.8%. The method was successfully applied to determine the plasma concentrations of CVB-D in Chinese volunteers.     

Crystal structures of human NUDT5 reveal insights into the structural basis of the substrate specificity

Human NUDT5 (hNUDT5) is an ADP-ribose pyrophosphatase (ADPRase) belonging to the Nudix hydrolase superfamily. It presumably plays important roles in controlling the intracellular level of ADP-ribose (ADPR) to prevent non-enzymatic ADP-ribosylation by hydrolyzing ADPR to AMP and ribose 5'-phosphate. We report here the crystal structures of hNUDT5 in apo form, in complex with ADPR, and in complex with AMP with bound Mg2+. hNUDT5 forms a homodimer with substantial domain swapping and assumes a structure more similar to Escherichia coli ADPRase ORF209 than human ADPRase NUDT9. The adenine moiety of the substrates is specifically recognized by the enzyme via hydrogen-bonding interactions between N1 and N6 of the base and Glu47 of one subunit, and between N7 of the base and Arg51 of the other subunit, providing the molecular basis for the high selectivity of hNUDT5 for ADP-sugars over other sugar nucleotides. Structural comparisons with E. coli ADPRase ORF209 and ADPXase ORF186 indicate that the existence of an aromatic residue on loop L8 in ORF186 seems to be positively correlated with its enzymatic activity on APnA, whereas hNUDT5 and ORF209 contain no such residue and thus have low or no activities on APnA.     

YAP regulates periodontal ligament cell differentiation into myofibroblast interacted with RhoA/ROCK pathway

During orthodontic tooth movement (OTM), periodontal ligament cells (PDLCs) receive the mechanical stimuli and transform it into myofibroblasts (Mfbs). Indeed, previous studies have demonstrated that mechanical stimuli can promote the expression of Mfb marker α-smooth muscle actin (α-SMA) in PDLCs. Transforming growth factor β1 (TGF-β1), as the target gene of yes-associated protein (YAP), has been proven to be involved in this process. Here, we sought to assess the role of YAP in Mfbs differentiation from PDLCs. The time-course expression of YAP and α-SMA was manifested in OTM model in vivo as well as under tensional stimuli in vitro. Inhibition of RhoA/Rho-associated kinase (ROCK) pathway using Y27632 significantly reduced tension-induced Mfb differentiation and YAP expression. Moreover, overexpression of YAP with lentiviral transfection in PDLCs rescued the repression effect of Mfb differentiation induced by Y27632. These data together suggest a crucial role of YAP in regulating tension-induced Mfb differentiation from PDLC interacted with RhoA/ROCK pathway.     

Perivascular adipose tissue‐derived stromal cells contribute to vascular remodeling during aging

Aging is an independent risk factor for vascular diseases. Perivascular adipose tissue (PVAT), an active component of the vasculature, contributes to vascular dysfunction during aging. Identification of underlying cell types and their changes during aging may provide meaningful insights regarding the clinical relevance of aging‐related vascular diseases. Here, we take advantage of single‐cell RNA sequence to characterize the resident stromal cells in the PVAT (PVASCs) and identified different clusters between young and aged PVASCs. Bioinformatics analysis revealed decreased endothelial and brown adipogenic differentiation capacities of PVASCs during aging, which contributed to neointimal hyperplasia after perivascular delivery to ligated carotid arteries. Mechanistically, in vitro and in vivo studies both suggested that aging‐induced loss of peroxisome proliferator‐activated receptor‐γ coactivator‐1 α (PGC1α) was a key regulator of decreased brown adipogenic differentiation in senescent PVASCs. We further demonstrated the existence of human PVASCs (hPVASCs) and overexpression of PGC1α improved hPVASC delivery‐induced vascular remodeling. Our finding emphasizes that differentiation capacities of PVASCs alter during aging and loss of PGC1α in aged PVASCs contributes to vascular remodeling via decreased brown adipogenic differentiation.     

DYRK1A inhibition suppresses STAT3/EGFR/Met signalling and sensitizes EGFR wild‐type NSCLC cells to AZD9291

DYRK1A is considered a potential cancer therapeutic target, but the role of DYRK1A in NSCLC oncogenesis and treatment requires further investigation. In our study, high DYRK1A expression was observed in tumour samples from patients with lung cancer compared with normal lung tissues, and the high levels of DYRK1A were related to a reduced survival time in patients with lung cancer. Meanwhile, the DYRK1A inhibitor harmine could suppress the proliferation of NSCLC cells compared to that of the control. As DYRK1A suppression might be effective in treating NSCLC, we next explored the possible specific molecular mechanisms that were involved. We showed that DYRK1A suppression by siRNA could suppress the levels of EGFR and Met in NSCLC cells. Furthermore, DYRK1A siRNA could inhibit the expression and nuclear translocation of STAT3. Meanwhile, harmine could also regulate the STAT3/EGFR/Met signalling pathway in human NSCLC cells. AZD9291 is effective to treat NSCLC patients with EGFR‐sensitivity mutation and T790 M resistance mutation, but the clinical efficacy in patients with wild‐type EGFR remains modest. We showed that DYRK1A repression could enhance the anti‐cancer effect of AZD9291 by inducing apoptosis and suppressing cell proliferation in EGFR wild‐type NSCLC cells. In addition, harmine could enhance the anti‐NSCLC activity of AZD9291 by modulating STAT3 pathway. Finally, harmine could enhance the anti‐cancer activity of AZD9291 in primary NSCLC cells. Collectively, targeting DYRK1A might be an attractive target for AZD9291 sensitization in EGFR wild‐type NSCLC patients.     

住院患者导管相关性血流感染病原菌分布与耐药分析

目的 探讨住院患者血管导管相关性血流感染的病原菌分布及耐药情况,为医院感染的控制提供依据.方法 对2011年1月至2012年2月住院患者中导管相关性血流感染病例的643株细菌的耐药性进行统计分析.结果 导管相关性血流感染发生率为4.1％,其中多重耐药细菌感染发生率为3.0％,分离病原菌以革兰阴性杆菌为主,占55.4％,革兰阳性球菌占43.2％.革兰阴性杆菌以肺炎克雷伯菌、大肠埃希菌为主,对头孢哌酮/舒巴坦、哌拉西林/他唑巴坦、阿米卡星耐药率低,鲍曼不动杆菌对亚胺培南耐药率增高;革兰阳性球菌以耐甲氧西林凝固酶阴性葡萄球菌为主,未出现耐万古霉素肠球菌.结论 加强细菌耐药检测,合理使用抗菌药物,对减少医院导管相关性血流感染的发生和耐药菌株产生有重要意义.     

Calpastatin Gene (CAST) Is Not Associated with Late Onset Sporadic Parkinson’s Disease in the Han Chinese Population

Recent studies point to an association between the late-onset sporadic Parkinson’s disease (PD) and single nucleotide polymorphisms (SNPs) rs1559085 and rs27852 in Ca²⁺-dependent protease calpain inhibitor calpastatin (CAST) gene. This finding is of interest since loss of CAST activity could result in over activated calpain, potentially leading to Ca²⁺ dysregulation and loss of substantia nigra neurons in PD. We explored the association between CAST SNPs and late-onset sporadic PD in the Han Chinese population. The study included 615 evaluable patients (363 male, 252 female) with PD and 636 neurologically healthy controls (380 male, 256 female) matched for age, gender, ethnicity, and area of residence. PD cases were identified from the PD cohort of the Chinese National Consortium on Neurodegenerative Diseases (www.chinapd.cn). A total of 24 tag-SNPs were genotyped capturing 95% of the genetic variation across the CAST gene. There was no association found between any of the polymorphisms and PD in all models tested (co-dominant, dominant-effect and recessive-effect). Similarly, none of the common haplotypes was associated with a risk for PD. Our data do not support a significant association between the CAST gene polymorphisms and late onset sporadic PD in the Han Chinese population.     

CoCl2预处理对大鼠海马神经元急性低氧后Na+、K+电流的影响

目的：观察氯化钴(COCl2)预处理对急性低氧后海马神经元电压门控性Na^ 、K^ 电流的影响。方法：原代培养大鼠海马神经元,分为COCl2预处理和非处理组,采用膜片钳全细胞记录技术,检测急性低氧后海马神经元钠电流(INa)、钾电流(Ik)的变化。结果：急性低氧后,海马神经元INa、Ik电流幅度明显降低,INa阈值右移,而经CoCl2预处理的海马神经元INa、Ik电流的降低幅度明显减轻。结论：COCl2预处理减轻急性低氧所致的INa、Ik电流变化,对神经元有明显的保护作用。     

Long-Term Existence of SARS-CoV-2 in COVID-19 Patients: Host Immunity,Viral Virulence,and Transmissibility

Virologica Sinica - COVID-19 patients can recover with a median SARS-CoV-2 clearance of 20 days post initial symptoms (PIS). However, we observed some COVID-19 patients with existing...     

模拟微重力条件下大鼠肝细胞三维培养支架材料的筛选（简报）

组织工程是一门新兴的边缘学科,它是利用体外培养的人体功能细胞与适当的细胞外基质或支架材料相结合,然后将其移植到体内病损部位以期达到修复目的。微重力组织工程(Microgravity Tissue Engineering)是近年来由美国空间生物技术研究人员开创的一个独特研究领域,其核心技术是