miR-455-5p靶向SOCS3抑制RSV感染致气道上皮炎症反应的机制研究

摘要 目的：揭示miR-455-5p对呼吸道合胞病毒（RSV）感染致气道上皮细胞炎症反应的作用机制。方法：qRT-PCR检测30例健康体检儿童（健康组）、RSV感染轻症组（n=41）和重症组（n=31）患儿血清miR-455-5p水平。将16HBE细胞分为Control组、NC-agomir组、miR-455-5p-agomir组、NC-antagomir组、miR-455-5p-antagomir组。使用Lipofectamine 3000转染16HBE细胞后培养48 h后,分为Blank组、NC组（转染了NC-agomir的细胞）、RSV+NC-agomir组、RSV+miR-455-5p-agomir组,用RSV病毒液感染RSV+NC-agomir组和RSV+miR-455-5p-agomir组16HBE细胞,Blank组和NC组16HBE细胞正常培养。用CCK-8法和EdU法检测细胞增殖、TUNEL法检测细胞凋亡、ELISA法检测上清液中TNF-α、IL-6、IL-8水平,qRT-PCR检测miR-455-5p和SOCS3的mRNA水平,Western blot检测SOCS3、IFN-α、STAT1、STAT2、p-STAT1和p-STAT2的蛋白水平。结果：与健康组相比,轻症组和重症组患儿的血清miR-455-5p水平降低（P<0.05）。与轻症组相比,重症组的血清miR-455-5p水平降低（P<0.05）。与Control组和NC-agomir组相比,miR-455-5p-agomir组16HBE细胞的miR-455-5p水平、相对细胞活力和EdU阳性率升高（P<0.05）,TUNEL阳性率降低（P<0.05）,上清液中的TNF-α、IL-6和IL-8水平降低（P<0.05）,SOCS3 mRNA和蛋白水平降低（P<0.05）,IFN-α蛋白、STAT1和STAT2磷酸化水平升高（P<0.05）。与Control组和NC-antagomir组相比,miR-455-5p-antagomir组16HBE细胞的miR-455-5p水平、相对细胞活力和EdU阳性率降低（P<0.05）,TUNEL阳性率升高（P<0.05）,上清液中的TNF-α、IL-6和IL-8水平升高（P<0.05）,SOCS3 mRNA和蛋白水平升高（P<0.05）,IFN-α蛋白、STAT1和STAT2磷酸化水平降低（P<0.05）。与Blank组和NC组相比,RSV+NC-agomir组16HBE细胞的miR-455-5p水平、相对细胞活力和EdU阳性率降低（P<0.05）,TUNEL阳性率升高（P<0.05）,上清液中的TNF-α、IL-6和IL-8水平升高（P<0.05）,SOCS3 mRNA和蛋白水平升高（P<0.05）,IFN-α蛋白、STAT1和STAT2磷酸化水平降低（P<0.05）。与RSV+NC-agomir组相比,RSV+miR-455-5p-agomir组的miR-455-5p水平、相对细胞活力和EdU阳性率升高（P<0.05）,TUNEL阳性率降低（P<0.05）,上清液中的TNF-α、IL-6和IL-8水平降低（P<0.05）,SOCS3 mRNA和蛋白水平降低（P<0.05）,IFN-α蛋白、STAT1和STAT2磷酸化水平升高（P<0.05）。结论：miR-455-5p在RSV感染患儿血清中下调,上调miR-455-5p通过抑制SOCS3的转录和表达从而激活RSV感染的16HBE细胞中IFN-α介导的抗病毒反应。     

针灸联合逍遥散合半夏厚朴汤治疗肝郁脾虚型抑郁症患者伴胃肠功能紊乱的临床疗效

摘要 目的：探讨针灸联合逍遥散合半夏厚朴汤治疗肝郁脾虚型抑郁症患者伴胃肠功能紊乱的临床疗效。方法：选取我院2020年5月到2023年5月收治的80例抑郁症伴胃肠功能紊乱患者作为研究对象,中医证型均为肝郁脾虚型,分为观察组与对照组,40例。对照组患者采取逍遥散合半夏厚朴汤治疗,观察组在对照组基础上增加针灸治疗,对比两组患者临床疗效,分别在治疗前后应用汉密尔顿抑郁量表（HAMD）、负性认知加工偏向问卷（NCPBQ）评价抑郁症状和负性认知情况,并对比治疗前后肿瘤坏死因子-α（TNF-α）、C反应蛋白（CRP）、白细胞介素-6（IL-6）表达水平和胃动素（motilin,MTL）、生长抑素（SS）、胃泌素（GAS）相关血清肠胃激素表达水平,并采用世界卫生组织生活质量-100量表（WHOQOL-100）评估生活质量。结果：观察组治疗总有效率较对照组高（P＜0.05）;治疗前两组患者HAMD、NCPBQ评分对比无差异（P＞0.05）,治疗后两组患者均降低,且观察组较对照组低（P＜0.05）;治疗前两组患者TNF-α、CRP、IL-6相关炎症因子和MTL、SS、GAS相关胃肠激素水平对比无明显差异（P＞0.05）,治疗后两组患者TNF-α、CRP、IL-6、SS、GAS水平均降低,且观察组较对照组低,MTL水平升高,观察组较对照组高（P＜0.05）;两组患者治疗后生活质量相关评分均升高,且观察组较对照组高（P＜0.05）。结论：针灸联合逍遥散合半夏厚朴汤治疗肝郁脾虚型抑郁症伴胃肠功能紊乱疗效显著,可减轻患者抑郁情绪和负性认知,降低炎症反应,改善胃肠激素水平,提升患者生活质量。     

基于经会阴实时三维超声评估生物反馈电刺激联合盆底肌锻炼治疗产后盆底功能障碍的临床疗效

摘要 目的：探讨经会阴实时三维超声评估生物反馈电刺激联合盆底肌锻炼治疗产后盆底功能障碍的临床疗效。方法：选择2020年9月至2022年9月我院收治的96例产后盆底功能障碍患者,根据随机数字表法将患者分为两组,对照组（48例）采用盆底肌锻炼治疗,研究组（48例）采用生物反馈电刺激联合盆底肌锻炼治疗。治疗前后采用经会阴实时三维超声检查,对比两组治疗前后的盆底功能障碍调查表（PFDI-20）、盆底障碍影响简易问卷7（PFIQ-7）评分、静息和Valsalva动作状态下的肛提肌超声参数。分析肛提肌超声参数与PFDI-20、PFIQ-7评分的相关性。结果：两组治疗后PFDI-20、PFIQ-7评分,静息和Valsalva动作状态下肛提肌裂孔前后径、肛提肌裂孔左右径、肛提肌裂孔面积较治疗前降低（P＜0.05）,静息时肛提肌厚度较治疗前增加（P＜0.05）。研究组治疗后PFDI-20、PFIQ-7评分,静息和Valsalva动作状态下肛提肌裂孔前后径、肛提肌裂孔左右径、肛提肌裂孔面积低于对照组（P＜0.05）,静息时肛提肌厚度大于对照组（P＜0.05）。静息和Valsalva状态下肛提肌裂孔前后径、肛提肌裂孔左右径、肛提肌裂孔面积与PFDI-20、PFIQ-7评分呈正相关（P＜0.05）,静息状态肛提肌厚度与PFDI-20、PFIQ-7评分呈负相关（P＜0.05）。结论：经生物反馈电刺激联合盆底肌锻炼治疗后肛提肌裂孔大小较治疗前降低,肛提肌厚度较治疗前增加,且与PFDI-20、PFIQ-7评分改善有关,经会阴实时三维超声可客观、有效评价产后盆底功能障碍患者的治疗效果。     

Molecular bases for human complement C7 polymorphisms,C7*3 and C7*4

Complement C7 is one of the components of membrane attack complex (MAC) generated by the terminal complement cascade. C7 protein is polymorphic and most of its polymorphisms have been identified using isoelectric focusing (IEF), which detects protein charge differences. To date, the molecular bases of the polymorphisms detected by IEF have not been determined. In this paper, we describe the structural bases of two C7 IEF-detected polymorphisms, C7*3 and C7*4, both of which are common in Asian populations. C7*3 resulted from substitution of cysteine (Cys) at amino acid residue 106 by charged arginine (Arg; C106R), while charged lysine (Lys) at amino acid residue 398 was replaced by neutral glutamine (Gln; K398Q) in C7*4. As C7*3 is hypomorphic, it is important to study its possible associations with diseases such as immunological disorders and infections. We present genetic bases for this C7 polymorphism, which we determined using polymerase chain reaction (PCR)-based genotyping, a simple and accurate method suitable for large-scale studies.     

Respiratory system dynamical mechanical properties: modeling in time and frequency domain

The mechanical properties of the respiratory system are important determinants of its function and can be severely compromised in disease. The assessment of respiratory system mechanical properties is thus essential in the management of some disorders as well as in the evaluation of respiratory system adaptations in response to an acute or chronic process. Most often, lungs and chest wall are treated as a linear dynamic system that can be expressed with differential equations, allowing determination of the system’s parameters, which will reflect the mechanical properties. However, different models that encompass nonlinear characteristics and also multicompartments have been used in several approaches and most specifically in mechanically ventilated patients with acute lung injury. Additionally, the input impedance over a range of frequencies can be assessed with a convenient excitation method allowing the identification of the mechanical characteristics of the central and peripheral airways as well as lung periphery impedance. With the evolution of computational power, the airway pressure and flow can be recorded and stored for hours, and hence continuous monitoring of the respiratory system mechanical properties is already available in some mechanical ventilators. This review aims to describe some of the most frequently used models for the assessment of the respiratory system mechanical properties in both time and frequency domain.     

A rapid PCR-based method for genetically mapping ESTs

A simple, semi-automatable procedure was developed for converting expressed sequence tags (ESTs) into mappable genetic markers. The polymerase chain reaction is used to amplify regions immediately 5′ or 3′ to the coding regions of genes in order to maximise sequence variability between alleles. Fragment length and nucleotide substitution polymorphisms among amplified alleles can be detected using either ethidium bromide staining or automated laser-based fluorescence. A 6% non-denaturing acrylamide gel, analysed with an ABI 377 DNA sequencer, proved capable of resolving homoduplexes and heteroduplexes formed between amplified alleles containing nucleotide substitutions as well as resolving allelic length differences. With this approach 75% of 60 ESTs from a range of Pinus species could be genetically mapped in each of three pedigrees from P. radiata and P. taeda. Furthermore, three or four alleles were detected in each pedigree for 42% of the EST markers. Received: 4 January 2000 / Accepted: 26 May 2000     

Overexpression of polo-like kinase 1 and its clinical significance in human non-small cell lung cancer

Polo-like kinase 1 is a serine/threonine kinase which plays an essential role in mitosis and malignant transformation. The aim of this study was to investigate the prognostic significance of polo-like kinase 1 expression and determine its possibility as a therapeutic target in non-small cell lung cancer. Semi-quantitative RT-PCR assay was performed to detect polo-like kinase 1 mRNA expression in non-small cell lung cancer cells or tissues. Immunohistochemistry was performed to detect polo-like kinase 1 protein expression in 100 non-small cell lung cancer tissue samples, and the associations of polo-like kinase 1 expression with clinicopathological factors or prognosis of non-small cell lung cancer patients were evaluated. RNA interference was employed to inhibit endogenous polo-like kinase 1 expression and analyzed the effects of polo-like kinase 1 inhibition on the malignant phenotypes of non-small cell lung cancer cells including growth, apoptosis, radio- or chemoresistance. Also, the possible molecular mechanisms were also investigated. The levels of polo-like kinase 1 mRNA expression in non-small cell lung cancer cell lines or tissues were significantly higher than those in normal human bronchial epithelial cell line or corresponding non-tumor tissues. High polo-like kinase 1 expression was significantly correlated with advanced clinical stage, higher tumor classification and lymph node metastasis of non-small cell lung cancer patients (P = 0.001, 0.004 and 0.001, respectively). Meanwhile, high polo-like kinase 1 protein expression was also an independent prognostic molecular marker for non-small cell lung cancer patients (hazard ratio: 2.113; 95% confidence interval: 1.326-3.557; P = 0.017). Polo-like kinase 1 inhibition could significantly inhibit in vitro and in vivo proliferation, induce cell arrest of G₂/M phase and apoptosis enhancement in non-small cell lung cancer cells, which might be activation of the p53 pathway and the Cdc25C/cdc2/cyclin B1 feedback loop. Further, inhibition of polo-like kinase 1 could enhance the sensitivity of non-small cell lung cancer cells to taxanes or irradiation. Thus, polo-like kinase 1 might be a prognostic marker and a chemo- or radiotherapeutic target for non-small cell lung cancer.     

Species of Borrelia distinguished by restriction site polymorphisms in 16S rRNA genes

Abstract Three phyletic groups of Borrelia associated with Lyme disease, B. burgdorferi, B. garinii and group VS461 can be distinguished from each other and other species of Borrelia by Bfa I restriction site polymorphisms in PCR amplified 16S rRNA genes. One strain isolated from an Ixodes pacificus tick in California that was previously unclassifiable was distinguishable from B. burgdorferi by an Mnl I restriction site polymorphism.     

Solvent‐dependent conformation of a regioselective amylose carbamate: Amylose‐2‐acetyl‐3,6‐bis(phenylcarbamate)

Six amylose‐2‐acetyl‐3,6‐bis(phenylcarbamate) (AAPC) samples ranging in weight‐average molar mass M_w from 1.8 × 10⁴ g mol^?1 to 1.1 × 10⁶ g mol^?1 have been prepared from enzymatically synthesized amylose samples. Static light scattering, small‐angle X‐ray scattering, sedimentation equilibrium, and viscosity measurements were made for the samples in 1,4‐dioxane (DIOX), 2‐ethoxyethanol (2EE), and 2‐butanone (MEK) all at 25°C to determine particle scattering functions, z‐average radii of gyration, intrinsic viscosities, as well as M_w. The data were analyzed in terms of the wormlike cylinder model mainly to yield the helix pitch per residue h and the Kuhn segment length λ^?1, which corresponds to twice of the persistence length. The latter parameters (λ^?1) in 2EE (11 nm) and MEK (12 nm) are quite smaller than those for amylose tris(phenylcarbamate) (ATPC) in the same solvent (16 nm in 2EE and 18 nm in MEK) whereas those for AAPC (21 nm) and ATPC (22 nm) in DIOX are essentially the same as each other. This indicates that the chain stiffness of AAPC is more strongly influenced by the solvents since the number of intramolecular H‐bonds of AAPC is more changeable than that for ATPC. © 2012 Wiley Periodicals, Inc. Biopolymers 97:1010–1017, 2012.     

Cell-free synthesis of membrane proteins: Tailored cell models out of microsomes

Incorporation of proteins in biomimetic giant unilamellar vesicles (GUVs) is one of the hallmarks towards cell models in which we strive to obtain a better mechanistic understanding of the manifold cellular processes. The reconstruction of transmembrane proteins, like receptors or channels, into GUVs is a special challenge. This procedure is essential to make these proteins accessible to further functional investigation. Here we describe a strategy combining two approaches: cell-free eukaryotic protein expression for protein integration and GUV formation to prepare biomimetic cell models. The cell-free protein expression system in this study is based on insect lysates, which provide endoplasmic reticulum derived vesicles named microsomes. It enables signal-induced translocation and posttranslational modification of de novo synthesized membrane proteins. Combining these microsomes with synthetic lipids within the electroswelling process allowed for the rapid generation of giant proteo-liposomes of up to 50 μm in diameter. We incorporated various fluorescent protein-labeled membrane proteins into GUVs (the prenylated membrane anchor CAAX, the heparin-binding epithelial growth factor like factor Hb-EGF, the endothelin receptor ETB, the chemokine receptor CXCR4) and thus presented insect microsomes as functional modules for proteo-GUV formation. Single-molecule fluorescence microscopy was applied to detect and further characterize the proteins in the GUV membrane. To extend the options in the tailoring cell models toolbox, we synthesized two different membrane proteins sequentially in the same microsome. Additionally, we introduced biotinylated lipids to specifically immobilize proteo-GUVs on streptavidin-coated surfaces. We envision this achievement as an important first step toward systematic protein studies on technical surfaces.