亚低温对脊髓损伤后反应性星形胶质细胞增生的影响

目的：观察在不同温度条件下脊髓星形胶质细胞划痕损伤活化后的形态和活性改变,以探讨亚低温对脊髓损伤后反应性星形胶质细胞增生的影响。方法：体外原代培养新生SD大鼠脊髓星形胶质细胞,以划痕实验制备反应性星形胶质细胞。亚低温选择33℃,细胞培养48 h。实验分为对照组、划痕组、亚低温组和划痕+亚低温组。各组在相应的时间点观察细胞形态,采用免疫荧光染色方法检测Nestin阳性率,MTT比色法观察细胞活性,PI染色方法观察细胞凋亡程度。结果：与对照组和亚低温组相比,划痕组和划痕+亚低温组细胞胞体肥大,周围突起增多、延展以及胞浆丰富,细胞生长率明显升高。与划痕组相比,划痕+亚低温组细胞变化减慢,周围突起减少,细胞长入划痕处所需时间增加,细胞Nestin阳性率、PI阳性率和细胞生长率明显降低,各结果差异显著（P<0.01）。结论：划痕损伤后星形胶质细胞活化为反应性星形胶质细胞并会增生,亚低温明显抑制脊髓反应性星形胶质细胞的活化增生,并可以抑制星形胶质细胞的凋亡。     

电针对脊髓损伤星形胶质细胞增生及其NGF表达的影响

目的研究脊髓损伤后电针治疗对星形胶质细胞增生及其内源性神经生长因子(nerve growth factor,NGF)表达的影响.方法选用成年雌性Wistar大鼠,随机分为3组.A组为正常对照组,B组、C组为下胸段脊髓不完全损伤.B组损伤后不治疗,C组损伤后给予督脉电针治疗.损伤后3 d、1 、2或4周应用免疫组化染色分别观察损伤脊髓胶质原纤维酸性蛋白(glial fibroblast acid protein,GFAP)和NGF表达的变化.结果 B组术后3 d,GFAP阳性细胞明显增多, 2周后开始减少,4周时仍有较多的阳性细胞;C组GFAP阳性细胞明显少于B组,1周时达高峰.脊髓损伤后NGF表达呈逐渐增加的趋势.C组NGF的表达明显高于B组,且一直保持在较高水平.NGF阳性细胞大部分与GFAP阳性细胞形态相似.结论电针治疗能减少星形胶质细胞增生,促进内源性NGF的合成,从而创造了有利于神经再生的微环境.     

MiR-144通过靶向抑制GRK5表达促进脊髓星形胶质细胞活化

已知miR-144与细胞活化和增殖有关,然而其具体分子机制尚不明确。本研究发现miR-144通过靶向GRK5促进脊髓星形胶质细胞的活化。运用real-time PCR检测脊髓损伤和正常大鼠的脊髓组织及其脊髓星形胶质细胞中miR-144的表达,发现与正常的组织和细胞相比,miR-144在脊髓损伤组织和星形胶质细胞中的表达水平显著降低;Western印迹检测到脊髓损伤大鼠的星形胶质细胞中GFAP蛋白的表达显著低于正常大鼠,而GRK5蛋白的表达高于正常大鼠;MTT分析结果显示转染miR-144可显著提高脊髓损伤大鼠的星形胶质细胞活性,但对细胞增殖无明显作用;酶活性试剂盒分析发现miR-144显著提高了SOD和GSH活性;生物学信息分析和萤光素酶报告基因检测结果显示miR-144能靶向结合GRK5,并下调GRK5的表达;MiR-144 mimic转染或miR-144 mimic与pcDNA-GRK5共转染脊髓损伤的星形胶质细胞,发现miR-144转染能通过激活NF-κB通路消除pcDNA-GRK5引起的细胞活化抑制。综上所述,miR-144通过靶定结合癌基因GRK5来促进脊髓星形胶质细胞细胞的活化。     

MiR-144通过靶向抑制GRK5表达促进脊髓星形胶质细胞活化北大核心CSCD

尽管miR-144与细胞活化、增殖有关,但其对脊髓星形胶质细胞的作用尚不明确。本文旨在探究正常和脊髓损伤(spinal cord injury,SCI)组织和细胞中miR-144表达,以及miR-144能否通过靶向调节G蛋白偶联受体激酶5(GRK5)上调脊髓星形胶质细胞活化。实时定量PCR(RT-q PCR)和Western印迹结果揭示,与正常的组织/细胞相比,miR-144在脊髓损伤组织和星形胶质细胞中的表达水平显著降低,而GRK5的表达升高;脊髓损伤大鼠的星形胶质细胞中胶质原纤维酸性蛋白(GFAP)的表达显著低于正常大鼠,而GRK5蛋白的表达高于正常大鼠;MTT分析结果显示,转染miR-144可显著提高脊髓损伤大鼠的星形胶质细胞活性,但对细胞增殖无明显作用;酶活性分析发现,miR-144显著提高SOD和GSH活性;萤光素酶报告基因检测结果证明,miR-144能靶向结合GRK5,下调GRK5表达。miR-144 mimic转染或miR-144 mimic与pc DNA-GRK5共转染脊髓损伤的星形胶质细胞后,我们发现,miR-144转染可通过激活NF-κB通路消除GRK5对细胞活化的抑制作用。综上所述,miR-144通过靶向抑制GRK5促进脊髓星形胶质细胞的活化。     

MiR-144通过靶向抑制GRK5表达促进脊髓星形胶质细胞活化北大核心CSCD

尽管miR-144与细胞活化、增殖有关,但其对脊髓星形胶质细胞的作用尚不明确。本文旨在探究正常和脊髓损伤(spinal cord injury,SCI)组织和细胞中miR-144表达,以及miR-144能否通过靶向调节G蛋白偶联受体激酶5(GRK5)上调脊髓星形胶质细胞活化。实时定量PCR(RT-q PCR)和Western印迹结果揭示,与正常的组织/细胞相比,miR-144在脊髓损伤组织和星形胶质细胞中的表达水平显著降低,而GRK5的表达升高;脊髓损伤大鼠的星形胶质细胞中胶质原纤维酸性蛋白(GFAP)的表达显著低于正常大鼠,而GRK5蛋白的表达高于正常大鼠;MTT分析结果显示,转染miR-144可显著提高脊髓损伤大鼠的星形胶质细胞活性,但对细胞增殖无明显作用;酶活性分析发现,miR-144显著提高SOD和GSH活性;萤光素酶报告基因检测结果证明,miR-144能靶向结合GRK5,下调GRK5表达。miR-144 mimic转染或miR-144 mimic与pc DNA-GRK5共转染脊髓损伤的星形胶质细胞后,我们发现,miR-144转染可通过激活NF-κB通路消除GRK5对细胞活化的抑制作用。综上所述,miR-144通过靶向抑制GRK5促进脊髓星形胶质细胞的活化。     

成年大鼠吻侧迁移流的神经发生

目的探讨成年SD大鼠吻侧迁移流(rostral migratory stream,RMS)的神经发生。方法成年6周龄SD大鼠被处死。矢状位切片,免疫组织化学染色观察RMS区微管相关蛋白双皮质素(doublecortin,DCX)及胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)的表达及DCX/GFAP、DCX/p-CREB的共表达情况。结果在RMS区的垂直臂、肘部、水平臂均有DCX阳性细胞和GFAP阳性细胞,RMS区有GFAP/DCX和p-CREB/DCX共表达细胞。结论吻侧迁移流有广泛的神经元前体细胞及星形胶质细胞标记物表达;迁移神经元标记物可表达于星形胶质细胞,且神经元的迁移受到CREB信号通路的调控。     

胰蛋白酶消化强度优化获得高纯度体外培养的星形胶质细胞

本文旨在观察胰蛋白酶消化对体外培养的星形胶质细胞纯度的影响,优化星形胶质细胞培养方法。常规分离新生Sprague Dawley(SD)大鼠大脑皮质,分别用0.25%胰蛋白酶消化20、30和40 min制备单细胞悬液并接种细胞。细胞长满瓶底时进行恒温摇床振荡,前两个不同消化时间组再分为常规消化的对照组和二次胰蛋白酶消化组进行传代纯化。倒置相差显微镜观察细胞生长情况,MTT法检测细胞增殖,GFAP免疫荧光分析星形胶质细胞纯度并观察其形态,流式细胞术分析细胞凋亡。结果显示,胰蛋白酶消化20 min的细胞在原代培养9 d长满瓶底。而延长胰蛋白酶消化时间至30 min,细胞增殖更快,培养7 d即可铺满瓶底,且星形胶质细胞形态正常,纯度达(70.2±4.0)%,较20 min组有显著性提高(P0.05)。40 min组虽然星形胶质细胞纯度也较20 min组有所提高,但细胞增殖缓慢且损伤明显。在恒温摇床振荡结束后,进行二次胰蛋白酶消化可减少传代后杂细胞数量。二次胰蛋白酶消化组第一代(P1)的GFAP阳性率普遍高于各自对照组,其中30 min+二次胰蛋白酶消化组GFAP阳性率为(98.1±1.7)%,相当于20 min+对照组P3水平,且二者的凋亡率无显著性差异。以上结果表明,胰蛋白酶消化30 min+二次消化能有效提高星形胶质细胞纯度,缩短原代培养及纯化时间,是体外快速获得高纯度星形胶质细胞的有效方法。     

老年大鼠神经组织星形胶质细胞的分选培养与炎性特征分析*

目的: 建立优化的年轻与老年大鼠神经组织星形胶质细胞的分离纯化方法,比较年轻大鼠与老年大鼠星形胶质细胞的形态、功能差异,探讨老化后星形胶质细胞的功能改变及其在衰老过程中发挥的可能机制。方法: 采用50%-35%的percoll密度梯度离心法分选年轻(2月龄)和老年(20月龄)SD大鼠的大脑与脊髓星形胶质细胞;每组细胞设置3个复孔,培养72 h后,采用免疫荧光检测星形胶质细胞特异性标志物胶质纤维酸性蛋白(GFAP),观察不同年龄阶段星形胶质细胞的形态特征;qPCR检测衰老标志(p16、p21)的表达,β-半乳糖苷酶染色检测星形胶质细胞的衰老情况;qPCR检测促炎因子(IL-1β、TNF-α)与抗炎因子(IL-10)的表达水平。结果: 采用50%-35%的percoll梯度分选得到的星形胶质细胞的数量多、活性好、纯度高达95%以上,可用于后续实验。与年轻大鼠神经组织的星形胶质细胞相比,分选自老年大鼠神经组织的星形胶质细胞在细胞形态上偏向激活态,突起较少;星形胶质细胞β-半乳糖苷酶染色阳性率升高,p16、p21表达也明显增多(P<0.01);老年大鼠神经组织的星形胶质细胞的促炎因子(IL-1β、TNF-α)表达升高(P<0.05),抗炎因子(IL-10)表达有所降低(P<0.05)。结论: 50%-35%的percoll梯度可以作为大鼠神经组织星形胶质细胞的分选纯化、原代培养的方法;随着年龄的增加,星形胶质细胞发生细胞老化,表现出促炎症表型,促进神经系统的炎性衰老,可能是神经系统老化及神经退行性疾病的机制之一。     

体外培养大鼠星形胶质细胞液压冲击损伤的研究

目的：研究大鼠星形胶质细胞液压冲击损伤后形态学及蛋白质组学表达变化。方法：建立体外培养星形胶质细胞液压冲击损伤模型。原代培养SD大鼠脑皮质星形胶质细胞,随机分为损伤组与对照组,对照组给予（0.2±0.01）MPa液压冲击损伤,损伤后不同时间点观察细胞形态学变化;双向凝胶电泳技术分析液压冲击损伤后蛋白质组学表达变化。结果：星形胶质细胞在液压冲击损伤后发生了显著的形态学改变,损伤后2h星形胶质细胞出现了细胞水肿、细胞皱缩、细胞连接断开和坏死,损伤后24h、48h细胞胞体肥大、突起增粗明显,部分区域细胞反应性增生明显。液压冲击损伤后,星形胶质细胞蛋白质表达谱发生了显著改变,损伤后有13个蛋白点表达发生显著改变,其中5种蛋白得到质谱鉴定,分别是肌动蛋白结合蛋白、破解蛋白、磷酸甘油酸变位酶1、NADH脱氢酶10亚基和膜联蛋白1。结论：液压冲击损伤能够引起星形胶质细胞发生显著的形态学改变和蛋白质谱表达改变,损伤后表达改变的蛋白质可能与星形胶质细胞的损伤后应激反应相关。     

治疗脊髓损伤之建立星形胶质细胞模型的研究进展

脊髓损伤(spinal cord injury,SCI)是一种极为复杂的破坏性疾病,一旦脊髓损伤发生,治疗棘手,对患者家庭、国家带来巨大的经济、社会负担。近年来,通过建立大鼠脊髓损伤细胞相关模型,对于脊髓损伤的病因病机治疗等方面有了进一步的认识,而星形胶质细胞模型的建立对脊髓损伤治疗有深远意义。研究发现,星形胶质细胞作为靶细胞通过血-脑脊液屏障直接或间接对脊髓损伤有双向调控作用。本文通过对近年来星形胶质细胞模型培养制备方案等研究进行总结,以期为建立一个客观化、定量化、可模拟化的星形胶质细胞模型提供指导对脊髓损伤的治疗提供新的思路。     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.