白介素11拮抗剂对实验性小鼠肺纤维化的作用

目的: 观察拮抗白介素11(IL-11)对博来霉素(BLM)诱导的实验性小鼠肺纤维化的作用。方法: 将120只雄性C57BL/6小鼠随机分为正常对照组、IL-11拮抗剂组、BLM组和BLM+IL-11拮抗剂组(每组各30只)。BLM组和BLM+ IL-11拮抗剂组小鼠一次性气管注射BLM(1.5 mg/kg)诱导肺纤维化。于造模当日开始,IL-11拮抗剂组和BLM+IL-11拮抗剂组小鼠每间隔3 d尾静脉注射IL-11拮抗剂IL-11 Rα FC(2.5 mg/kg)。观察各组小鼠生存状态。于造模后第21日取肺组织进行HE染色、Masson染色以及Ashcroft评分评价肺纤维化程度。通过碱水解法测定肺组织中羟脯氨酸(HYP)的含量;采用Real-time PCR和Western blot检测肺组织中Collagen I、Collagen III和α-SMA 的基因和蛋白表达;采用酶联免疫吸附法测定肺组织中TGF-β1含量。结果: 与正常对照组相比,BLM可降低小鼠存活率(P＜0.05),破坏肺组织结构,导致大量胶原沉积,显著升高HYP含量、肺组织中Collagen I、Collagen III和α-SMA的基因和蛋白表达(P＜0.05),以及TGF-β1含量(P＜0.05)。而IL-11 Rα Fc处理可改善肺纤维小鼠的生存率,减轻肺组织病理学改变以及胶原沉积,减少肺组织中HYP含量(P＜0.05),下调肺组织中Collagen I、Collagen III和α-SMA的基因和蛋白表达(P＜0.05),以及TGF-β1含量(P＜0.05)。结论: IL-11拮抗剂可减轻BLM诱导的小鼠肺纤维化,为临床治疗肺纤维化提供了新思路。     

不同剂量博莱霉素诱导小鼠肺纤维化的差异及动态变化

目的探讨博来霉素诱导小鼠肺纤维化最佳剂量和方法。方法 126只8周龄雄性ICR小鼠,随机分成一次性大剂量模型和多次小剂量模型。一次性大剂量模型分为200 mg/(kg.bw)BLM组、150 mg/(kg.bw)BLM组、100 mg/(kg.bw)BLM组及阴性对照组(DN组),每组18只,分别经尾静脉一次性注射BLM 200、150、100mg/(kg.bw)及生理盐水10 mL/(kg.bw),各组分别于第7、14、21天各处死6只。多次小剂量模型分为每日10 mg/(kg.bw)BLM组及阴性对照组(N组),分别经尾静脉注射BLM 10 mg/(kg.bw)及生理盐水10 mL/(kg.bw),每天1次,连续注射14 d,两组分别于第14、21、28天各处死6只。留取肺组织,观察肺组织病理改变,检测Ⅲ型胶原的含量,观察小鼠体重及生存率。结果①在一次性大剂量模型中,BLM各剂量组肺泡炎症评分及肺纤维化评分与正常组相比,除100 mg/(kg.bw)BLM组和150 mg/(kg.bw)BLM组在第7天的模型差异无显著性外(P>0.05),其余各组差异均有显著性(P<0.05);各个剂量组Ⅲ型胶原的表达面积与正常组相比,除100 mg/(kg.bw)BLM组在第7天的模型差异无显著性外(P>0.05),其余各组均较正常组高(P<0.05),各个剂量组分别在第21天达到高峰,以200 mg/(kg.bw)BLM组第21天组Ⅲ型胶原的表达面积最高;该模型小鼠各剂量组死亡率为0。②在多次小剂量模型中,各组的肺泡炎症与肺纤维化程度与正常组相比差异均有显著性(P<0.05);各组Ⅲ型胶原的表达也均高于正常组(P<0.05),且随着时间的延长呈进行性增加,在第28天达到高峰;该模型小鼠共死亡11只,死亡率为30.56%。结论在本实验中,以尾静脉一次性注射BLM 200 mg/(kg.bw)后第21天诱导建立的ICR小鼠肺纤维化模型成模最好,其小鼠死亡率低,操作简单,有效安全方便的特点使之有希望成为一种复制肺纤维化的理想模型。     

肺纤维化形成中整合素相关激酶的表达与上皮-间充质细胞转化

目的观察整合素相关激酶(ILK)在博莱霉素致小鼠肺纤维化中的动态表达,探讨其在肺纤维化中与上皮-间充质细胞转化(EMT)的关系。方法将32只小鼠随机分成正常对照组,博莱霉素(BLM)致肺纤维化组。后者以博莱霉素腹腔注射致肺纤维化,正常对照组仅在相同处理条件下注射生理盐水。治疗的第28天和40天处死动物取出肺组织,用苏木精-伊红(HE)和Massontrichrome染色,分别观察实验动物肺炎症和纤维化的程度,以样本碱水解法检测肺组织中羟脯氨酸的量,逆转录-聚合酶链反应(RT-PCR)检测ILK mRNA表达、免疫组化检测不同实验组小鼠ILK,并同时检测间充质细胞标记物α-平滑肌肌蛋白(α-SMA)和上皮细胞标记物上皮钙粘附素(E-Cad)的表达。结果组织学结果显示小鼠肺组织胶原含量在第28d明显增加,第40d达高峰,羟脯氨酸测定动态增高。与正常对照组比较,肺纤维化组中ILK、α-SMA蛋白表达增高(P<0.01),E-Cad的蛋白表达降低(P<0.01)。RT-PCR结果显示肺纤维化组ILKmRNA的表达显著增高(P<0.01)。ILK的表达不仅与羟脯氨酸的含量成正相关(r分别为0.867、0.886,P均小于0.05),也与α-SMA/E-Cad比值成正相关(r分别为0.830、0.844,P均小于0.05);α-SMA与E-Cad成负相关(r分别为-0.820、-0.833,P均小于0.05)。结论ILK在小鼠肺纤维化中表达与纤维化的程度呈正相关增高,表明其可能是肺纤维化形成的促进因子,且由于ILK与α-SMA/E-Cad比值成正相关,推测ILK在肺纤维化形成中的机制可能与促进EMT有关。     

EGFR 对博莱霉素诱导小鼠肺纤维化中上皮- 间质转分化的影响

目的:探讨表皮生长因子受体(EGFR)在肺内的表达对博莱霉素(BLM)诱导小鼠肺纤维化中上皮-间质转分化的影响。方法:将40只4～6周龄C57BLB/c雄性小鼠随机分为正常对照组(气管滴入PBS),纤维化组(气管滴入BLM 3 mg/kg),EGFRRNAi组(气管滴入BLM 3 mg/kg+气管滴入siRNA 20μl)和RNAi阴性对照组(气管滴入BLM 3 mg/kg+气管滴入siRNA阴性对照20μl)。实验第10天处死小鼠,收获肺组织,检测羟脯氨酸含量;采用逆转录-聚合酶链反应(RT-PCR)法检测EGFR和α平滑肌肌动蛋白(α-SMA)mRNA的表达;肺组织切片行HE染色观察肺组织病理改变,免疫组化染色检测EGFR和α-SMA表达。结果:纤维化组EGFR和α-SMA两者的mRNA和蛋白表达均较正常对照组显著增加;RNAi组肺病理损伤较纤维化组减轻,气道上皮下胶原沉积及肺羟脯氨酸含量减少(P<0.05),肺组织EGFR和α-SMA两者的mRNA和蛋白表达均较纤维化组显著下降(P<0.05)。结论:在博来霉素诱导的肺纤维化中EGFR RNAi抑制EGFR活化,下调α-SMA的表达,减轻了博莱霉素诱导的肺纤维化病理改变。其抑制肺纤维化病理过程可能与其抑制上皮-间质转分化(EMT)有关。     

博莱霉素静脉注射制备小鼠肺间质纤维化模型

目的经尾静脉一次性注射博莱霉素(BLM)复制小鼠肺间质纤维化动物模型,并观察模型的肺组织病理学变化。方法8周龄雌性C57BL/6小鼠66只,随机分为BLM80组18只、BLM150组19只、BLM300组19只和对照组10只,分别经尾静脉一次性注射BLM80、150、300 mg/kg和生理盐水。结果①BLM80组和BLM150组小鼠最低体重分别为BLM注射前的84%和65%。②BLM80组、BLM150组、BLM300组和对照组小鼠的生存率分别为:100%、43%、0和100%。③BLM150组BLM注射后14 d、28 d,右肺羟脯氨酸含量分别为738±46 nmol、886±83 nmol,与对照组(360±75 nmol)比较差异均有显著性意义(P<0.01)。④BLM150组小鼠BLM注射28 d,在胸膜下及血管周围形成广泛、稳定、明显的纤维化改变。BLM150组与BLM80和对照组比较,肺纤维化病理评分分别呈明显增高(P值均小于0.001)。结论C57BL/6小鼠经尾静脉一次性注射BLM150mg/kg可以制备肺间质纤维化动物模型。     

博莱霉素与博莱霉素A5诱导小鼠肺组织损伤的差异

目的探讨静脉注射博莱霉素(BLM)和博莱霉素A5(BLM-A5)对小鼠肺组织损伤的不同作用。方法8周龄雌性C57BL/6小鼠49只,随机分为BLM组19只、BLM-A5组20只和对照组10只,分别经尾静脉一次性注射BLM 150 mg/kg、BLM-A5 150 mg/kg和生理盐水。观察每组小鼠体重、生存率,肺组织病理改变及肺组织羟脯胺酸的含量。结果①BLM组和BLM-A5组小鼠最低体重分别为静脉注射处置前的65.46%和66.87%,均较对照组明显降低(P值均小于0.001)。②BLM组、BLM-A5组和对照组小鼠的生存率分别为:43%、55%和100%。③BLM组小鼠注射处置后28 d,在胸膜下及血管周围形成广泛、稳定的间质纤维化病理改变,而BLM-A5组无明显纤维化形成。BLM组与BLM-A5组、正常组比较,肺纤维化病理评分均明显增高(P值均小于0.001)。④BLM-A5组和对照组注射处置后28 d,右肺羟脯氨酸含量分别为471.6±49.4 nmol和405.0±74.6 nmol,两组间无统计学差异。BLM组为978.4±106.1 nmol,与BLM-A5组、对照组比较分别存在显著性差异(P值均小于0.01)。结论静脉注射BLM与BLM-A5,对小鼠肺组织损伤的作用存在着明显的差异,BLM可导致肺间质形成广泛的纤维化,而BLM-A5仅引起轻微的肺组织损伤,不形成明显的间质纤维化。     

结缔组织生长因子在肺纤维化初期肺动脉中的表达

本研究观察了博莱霉素(bleomycin,BLM)诱导肺纤维化初期肺动脉压、肺动脉壁Ⅰ、Ⅲ型胶原的含量以及肺动脉壁结缔组织生长因子(connective tissue growth factor,CTGF)免疫阳性表达和分布.用气管内一次性滴注BLM(5 mg/kg体重)的方法复制肺纤维化动物模型大鼠;用右心漂浮导管技术检测肺动脉压;用天狼星红胶原纤维特异染色和偏振光观察肺动脉Ⅰ、Ⅲ型胶原;用免疫组织化学法检测肺动脉壁CTGF表达.结果显示:滴注BLM后第14天,大鼠肺动脉压高于对照组大鼠(P<0.05);肺动脉主干和肺内动脉壁Ⅰ、Ⅲ型胶原的染色面积大于对照组大鼠(P<0.05,P<0.01),肺动脉主干血管壁Ⅰ、Ⅲ型胶原染色面积的比值高于对照组大鼠(P<0.05);肺动脉主干和肺内动脉壁CTGF免疫染色面积均大于对照组大鼠,平均光密度也高于对照组大鼠(均P<0.05);增多的CTGF免疫阳性细胞主要分布在肺动脉的平滑肌层和内皮层.以上结果表明,在BLM致肺纤维化形成初期肺动脉高压和肺血管壁结构重塑过程中,肺动脉壁平滑肌层和内皮层CTGF表达增多,这可能是肺动脉高压维持和发展的机制之一.     

黄芪甲甙对实验性肺纤维化大鼠Cathepsin B表达的影响

目的:探讨黄芪甲甙时实验性肺纤维化大鼠肺组织中组织蛋白酶B(Cathepsin B,CB)表达的影响。方法:36只SD大鼠,随机分为对照组、模型组和干预组。模型组和干预组气管内注射博来霉素(BLM,5mg/kg)诱导肺纤维化,对照组在相同条件下给予生理盐水。第二天起干预组大鼠每天经胃管灌服0．1g/L黄芪甲甙2ml,其余两组相同条件下给予助溶剂羧甲基纤维素钠。治疗的第7d和28d,处死动物取出肺组织,进行病理学观察;测定28d肺组织中羟脯氨酸含量;用免疫组化和RT-PCR观察各组鼠肺组织CB蛋白及mRNA表达的水平。结果:病理学观察显示:与模型组比较,干预组肺泡炎和纤维化程度均减轻;模型组羟脯氨酸含量显著增高,而干预组与模型组比较羟脯氨酸含量明显降低;与模型组比较,干预组CB蛋白及mRNA表达显著降低。结论:黄芪甲甙可能通过押制CB的过度表达,对实验性大鼠肺纤维化具有良好的治疗作用。     

小剂量辣椒素抗小鼠肺纤维化的作用及其机制

目的: 观察小剂量辣椒素(Cap)抗小鼠肺纤维化的作用是否与其激动瞬时电位感受器香草酸受体1(TRPV1)有关。方法: 实验设对照(CON)组、博莱霉素(BLM)组、Cap(0.5、1、2 mg/kg)剂量组及Cap (2 mg/kg)+ TRPV1受体阻断剂SB-452533(2.5 mg/kg)剂量组,每组n=12。BLM(3.5 mg/kg)气管注射诱导肺纤维化小鼠模型。造模后连续皮下注射给药21 d。血浆降钙素基因相关肽(CGRP)浓度采用ELISA法快速测定。肺组织病理变化和胶原沉积分别采用HE染色、Masson染色和免疫组化进行检测。肺组织α-CGRP、β-CGRP、I 型胶原(collagen I)、III 型胶原(collagen III)、E钙粘蛋白(E-Cadherin)、紧密连接蛋白-1(ZO-1)、波形蛋白(Vimentin)、α-平滑肌肌动蛋白(α-SMA)、TRPV1、磷酸化ERK1/2(p-ERK1/2)和真核翻译起始因子3a(eIF3a)mRNA及蛋白表达分别采用qPCR和(或)Western blot法检测。结果: 与BLM组相比,小剂量Cap给药21 d后,病理检查发现小鼠肺纤维化明显减轻;肺组织胶原表达明显下降(P＜0.05或P＜0.01);肺泡上皮细胞上皮间质转分化(EMT)明显受到抑制(E-Cadherin和ZO-1的表达明显升高(P＜0.05或P＜0.01)而Vimentin和α-SMA的表达明显降低(P＜0.05或P＜0.01));TRPV1和CGRP表达水平明显升高(P＜0.05或P＜0.01)而ERK磷酸化水平和eIF3a表达水平明显降低(P＜0.05或P＜0.01)。但当使用TRPV1受体阻断剂阻断TRPV1后,小剂量Cap逆转肺泡EMT、改善肺纤维化的作用被显著取消,同时CGRP表达水平又明显降低(P＜0.01)而p-ERK1/2和eIF3a的表达水平又明显升高(P＜0.01)。结论: 小剂量Cap能够逆转肺泡上皮细胞EMT、缓解肺纤维化。其机制可能与其激动TRPV1、促进CGRP释放,进而抑制ERK磷酸化和下调eIF3a的表达有关。     

经鼻滴入博莱霉素致小鼠肺纤维化模型的建立及其病理演变

目的经鼻快速滴入博莱霉素复制小鼠肺纤维化模型,观察肺纤维化模型小鼠的病理形态学改变及其羟脯氨酸含量变化,鉴定肺纤维化模型的成功建立。方法经鼻滴入博莱霉素建立小鼠肺纤维化模型。分别于造模第14天和第28天后,取肺组织行HE染色和天狼猩红染色,取心、肝、脾、肾、脑组织行HE染色,光镜下观察组织病理学变化;造模第28天后用样本碱水解法检测肺组织中羟脯氨酸（HYP）的含量。结果①肺组织病理形态学改变：造模第14天和第28天后模型组小鼠肺泡炎及纤维化程度均明显高于阴性对照组,并且在造模第28天后肺纤维化程度进一步加重;②肺组织中HYP含量：与阴性对照组比较,模型组显著升高（P〈0.05）。结论经鼻滴入博莱霉素可以成功复制小鼠肺纤维化模型,肺纤维化模型小鼠HYP含量升高。     

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.