阳离子脂质体介导增强型绿色荧光蛋白质粒转染骨骼肌卫星细胞

背景：骨骼肌卫星细胞是一种具有多向分化能力的全能干细胞,存在于骨骼肌间质中,对缺血、缺氧有一定的耐受力,是干细胞工程中重要来源细胞。 目的：为联合基因工程细胞心肌成形治疗初步探讨较为简便、经济的骨骼肌卫星细胞体外培养方法,建立一种简单、高效的转染骨骼肌卫星细胞的方法及探讨转染后基因表达的特点。 方法：分离、培养兔大腿骨骼肌卫星细胞,用CKK-8法测定其生长曲线。根据质粒和脂质体不同比例分组,用脂质体介导增强型绿色荧光蛋白质粒(plasmid enhanced green fluorescent protein,pEGFP)转染骨骼肌卫星细胞。测定各组转染效率及目标基因表达特点。 结果与结论：成功分离培养骨骼肌卫星细胞及转染pEFGF。在合适的质粒和脂质体比例下,转染效率可达35%以上。目标蛋白在转染12 h内开始表达,48-72 h表达最强,1周后逐渐减弱,2周后仍可观察到其表达。阳离子脂质体可介导pEGFP高效转染骨骼肌卫星细胞,转染效率与质粒、脂质体比例密切相关,目标基因表达随时间改变。     

酸性成纤维细胞因子基因体外转染肌卫星细胞

背景：研究报道外源性酸性成纤维细胞生长因子既可调节肌卫星细胞增殖和分化,又具有预防运动终板退变及肌萎缩的作用。 目的：通过酸性成纤维细胞因子基因转染大鼠骨骼肌卫星细胞,检测目的基因转染肌卫星细胞效果及基因表达情况,探讨建立有效预防运动终板退变及肌萎缩种子细胞可行性。 方法：取Wistar成年大鼠后肢肌肉,差速贴壁培养法分离纯化肌卫星细胞,观察细胞生长特性并做免疫组织化学鉴定;取第2代细胞,LipofectamineTM2000 Reagent转染试剂介导,将重组真核表达质粒pEGFP-N1-aFGF转染肌卫星细胞为实验组;阴性对照组转染空载质粒pEGFP-N1;空白对照组仅加入转染试剂。转染后24-72 h和传代后分别用倒置荧光显微镜观察细胞绿色荧光蛋白表达情况,计算转染效率。转染细胞行Western Blot检测酸性成纤维细胞因子表达。提取转染后72 h 细胞总RNA, 实时荧光定量PCR检测酸性成纤维细胞因子基因mRNA表达。 结果与结论：分离纯化细胞经免疫组织化学鉴定为肌卫星细胞。荧光显微镜观察到细胞转染6 h后即有绿色荧光发出,荧光强度和表达细胞总数在72 h达到高峰,传代后仍可观察到绿色荧光蛋白表达。实时荧光定量PCR证实目的基因mRNA表达水平远远高于对照组,Western Blot检测实验组有大量酸性成纤维细胞因子产生。提示酸性成纤维细胞基因转染肌卫星细胞可表达基因产物,有望作为基因工程种子细胞预防失神经支配后运动终板退变及肌萎缩。     

rhbFGF转染人皮肤成纤维细胞的研究

将重组人碱性成纤维细胞生长因子(rhbFGF)基因克隆入真核表达载体pEGFP-N1质粒增强型绿色荧光蛋白(EGFP)基因上游，并在其5′端加上白介素-4的信号肽序列，形成融合基因，利用脂质体转染原代培养的成纤维细胞，荧光显微镜和蛋白质印迹杂交检测rhbFGF-EGFP融合蛋白的表达，细胞计数法观察bFGF对细胞生长的影响。结果表明成功构建了pEGFP-rhbFGF质粒，并经脂质体介导有效地转入原代培养的人皮肤成纤维细胞内。用G418筛选纯化转基因后的细胞能够向胞外分泌rhbFGF活性物质，明显促进成纤维细胞自身的增殖。表明bFGF基因转染能够稳定有效地促进成纤维细胞的增殖。     

碱性成纤维细胞生长因子基因修饰骨骼肌卫星细胞自体移植急性心肌梗死组织的血管新生北大核心

背景:前期研究发现,骨骼肌卫星胞移植能够诱导心肌梗死区新生血管形成,缩小梗死面积,改善其心功能,但整体效果并不太理想。目的:观察碱性成纤维细胞生长因子基因修饰骨骼肌卫星细胞在急性心肌梗死区的存活及对心肌梗死区血管新生的影响。方法:将18只新西兰大白兔随机分为3组,实验组、对照组结扎冠状动脉左前降支,构建急性心肌梗死动物模型;空白对照组只穿线,不结扎。造模成功即刻,实验组于局部梗死心肌内注射DAPI标记的碱性成纤维细胞生长因子基因修饰自体骨骼肌卫星细胞悬液50μL,对照组注射等量DAPI标记的自体骨骼肌卫星细胞。细胞移植4周后取标本,观察心肌梗死区骨骼肌卫星细胞存活及成纤维细胞生长因子基因表达情况,免疫组织化学染色检查心肌梗死区新生血管形成情况。结果与结论:(1)空白对照组未见DAPI标记的细胞,对照组及实验组缺血心肌区域均可见大量DAPI标记的骨骼肌卫星细胞,实验组还可见大量EGFP-碱性成纤维细胞生长因子融合蛋白绿色荧光表达;(2)实验组、对照组新生微血管密度多于空白对照组(P<0.05),实验组新生微血管密度多于对照组(P<0.05);(3)结果表明,碱性成纤维细胞生长因子修饰骨骼肌卫星细胞可在急性心肌梗死区存活,促进心肌梗死区血管新生。     

运动对骨骼肌卫星细胞的影响及作用机制

骨骼肌卫星细胞在骨骼肌生长发育、损伤修复以及骨骼肌重塑等生理病理过程中具有重要的作用。适宜的运动训练可活化卫星细胞,促进卫星细胞增殖并向成肌细胞分化。本文就骨骼肌卫星细胞的起源、形态特征和特异性的标记以及运动训练调控骨骼肌卫星细胞活化、增殖、分化的作用机制进行综述。     

大鼠骨骼肌卫星细胞的原代培养和鉴定

目的采用组织块培养技术探索大鼠骨骼肌卫星细胞的原代培养方法。方法以成年SPF级Sprague-Dawley大鼠为研究对象,采用组织块培养法获取大鼠骨骼肌卫星细胞,并与C2C12成肌细胞进行比较,对两种细胞进行形态学研究及采用免疫荧光和免疫组织化学法测定两种细胞α-actin蛋白和Desmin蛋白的表达及分布,从而对骨骼肌卫星细胞进行鉴定。结果通过组织块培养法获取的细胞增殖旺盛,分化良好。免疫细胞荧光和免疫组织化学实验结果显示,α-actin蛋白和Desmin蛋白在两种细胞胞浆中均有分布。结论用组织块培养法获取的骨骼肌卫星细胞具有良好的增殖与分化能力,用此种方法可培养出高纯度的骨骼肌卫星细胞。     

碱性成纤维细胞生长因子基因转染对人牙周韧带细胞的作用

背景：主要来源于因正畸或阻生拔除的健康牙培养而成的人牙周韧带细胞已成为牙周组织工程理想的种子细胞来源之一。 目的：了解采用重组腺相关病毒作为载体介导碱性成纤维细胞生长因子基因转染对体外培养的人牙周韧带细胞增殖及细胞周期的影响。 方法：体外培养人牙周韧带细胞, 经重组腺相关病毒作为载体介导碱性成纤维细胞生长因子基因转染,分为3组：对照组、空载病毒组、碱性成纤维细胞生长因子转染组。用RT-PCR,Western blot检测人牙周韧带细胞在转染前后碱性成纤维细胞生长因子基因和蛋白的表达。应用细胞生长曲线、四甲基偶氮唑蓝比色法观察细胞生长的优化作用;采用流式细胞术测定细胞周期分布的变化。 结果与结论：对照组、空载病毒组未检测到碱性成纤维细胞生长因子mRNA和蛋白表达;碱性成纤维细胞生长因子转染组碱性成纤维细胞生长因子mRNA和蛋白水平均有表达,细胞的生长速度明显增快,细胞周期G₀/G₁期减少,S期细胞数增多。各组间比较差异有显著性意义(P < 0.05)。     

重组反转录病毒碱性成纤维细胞生长因子基因转染对人骨髓基质细胞成骨的影响

背景：国内外有关成纤维细胞生长因子基因转染促血管和促肌肉生长的研究较多,而成纤维细胞生长因子基因促成骨的研究未见报道。 目的：观察重组反转录病毒retrovirus pLXSN/碱性成纤维细胞生长因子基因转染对人骨髓基质细胞成骨能力的影响。 方法：从健康志愿者全骨髓中分离培养人骨髓基质细胞,体外扩增纯化后分为4组：①retrovirus pLXSN/碱性成纤维细胞生长因子组：培养液中加入碱性成纤维细胞生长因子基因重组反转录病毒。②retrovirus pLXSN组：培养液中加入反转录病毒空载体。③阳性对照组：培养液中添加地塞米松、抗坏血酸和β-甘油磷酸钠。④空白对照组：不给予特殊处理。 结果与结论：经多次换液传代,人骨髓基质细胞呈均一梭形形态。处理后retrovirus pLXSN/碱性成纤维细胞生长因子组与阳性对照组细胞形态逐渐趋于扁平,突起减少。免疫组织化学染色见retrovirus pLXSN/碱性成纤维细胞生长因子组碱性成纤维细胞生长因子表达明显强于其他3组。retrovirus pLXSN/碱性成纤维细胞生长因子和阳性对照组可引起细胞碱性磷酸酶活性增高和矿化结节及骨胶原形成。提示基因重组反转录病毒成纤维细胞生长因子转染对人骨髓基质细胞成骨能力具有促进作用。     

重组腺相关病毒对大鼠神经干细胞的体外转染及其对细胞生长的影响

目的 研究重组腺相关病毒（recombinant adeno-associated virus，rAAV）载体对原代培养的神经干细胞（neural stemcells，Nsc）的体外转染及其对细胞增殖、分化和迁移能力的影响。方法 取新生24h内的Wistar大鼠的海马进行神经干细胞原代培养，用不同滴度的rAAV为载体，以增强型绿色荧光蛋白（enhanced green fluorescent protein，EGFP）基因作为报告基因进行体外转染NSC，通过观察有绿色荧光的NSC的数量，测定rAAV对NSC的体外转染情况；用MTT法测定不同病毒滴度下rAAV对NSC增殖能力的影响，用免疫组化法鉴定NSC、神经元及神经胶质细胞，并在倒置荧光显微镜下直接测定细胞的迁移距离。结果 rAAV对NSC的体外转染效率随着病毒滴度的增加而提高，在转染后的第11天表达水平最高，用MOI为10^4、10^5、10^6的rAAV转染后第11天的转导率分别是9．81％、56．30％、64．67％；不同滴度rAAV转染NSC后不同时间点的MTT测定A值随着病毒滴度的增加而明显减小，差异均有统计学意义；但不同滴度rAAV转染NSC，其分化为神经元和神经胶质细胞的比率以及迁移的距离未见差异。结论 较高滴度（MOI为10^5和10^6）的rAAV可以在体外有效地转染神经干细胞，且不影响神经干细胞的分化和迁移能力，但对其增殖能力有明显抑制，并表现为病毒滴度依赖性。     

新生大鼠骨骼肌卫星细胞的原代培养和鉴定

目的：本文建立了一种肌卫星细胞的纯化培养方法，并对其进行了鉴定。方法：①肌卫星细胞的分离培养：取3日龄SD大鼠，取大腿肌组织，用Hanks液冲洗，剪碎后加入0．5％胶原酶Ⅱ，0．1％透明质酸酶，37℃消化20分钟。400目尼龙网过滤，190g离心清洗3次。加入1％链酶蛋白酶(pronase)37℃消化20分钟。     

H2O2诱导骨骼肌卫星细胞凋亡及法舒地尔的保护作用

背景：骨骼肌卫星细胞是成体骨骼肌中位于肌细胞膜和基膜之间具有增殖分化潜能的肌源性干细胞,研究表明骨骼肌卫星细胞的有效性及安全性,但是移植后的干细胞成活率极低,极大的限制了骨骼肌卫星细胞的应用。 目的：观察过氧化氢(H₂O₂)对大鼠骨骼肌卫星细胞凋亡的影响以及法舒地尔的保护作用。 方法：取体外培养的骨骼肌卫星细胞,随机分为正常对照组,H₂O₂组,H₂O₂⁺法舒地尔组(法舒地尔组),采用流式细胞仪检测细胞凋亡率,ELISA法检测白细胞介素4、肿瘤坏死因子a的浓度,Western blot检测Bax蛋白的表达。 结果与结论：同H₂O₂组相比,法舒地尔组凋亡发生率明显下降(P < 0.05),Bax蛋白水平表达明显降低(P < 0.05),白细胞介素4、肿瘤坏死因子a的分泌显著减少(P < 0.05)。结果提示,法舒地尔抑制Rho激酶信号途径发挥抗凋亡保护作用,其机制可能与减少Bax蛋白的表达有关。      

骨骼肌减少症与运动训练对肌卫星细胞影响的研究现状及展望

BACKGROUND: The phenomenon of atrophy or reduction of muscle, causing degenerative changes of muscle functions, appears along with age. Sports training, in which muscle satellite cells are of great importance, is beneficial to increase in muscle mass and improvement of muscle function.  OBJECTIVE:To summarize regulatory mechanism of satellite cells in skeletal muscle mass; changes of satellite muscle cells in the degenerative process of muscle mass and strength; declining and reverse effects of sports training intervention; situations and problems of current research and prospective of the future.  METHODS: A computer-based online search was conducted in PubMed database by using the key words of “sarcopenia, skeletal muscle, satellite cells” from 1986 to 2015. The language was limited to English. The eligible papers were further analyzed and reviewed. RESULTS AND CONCLUSION: A total of 168 papers were screened. Finally, 39 papers were selected according to the titles and objectives. Skeletal muscle atrophy is shown as II type muscle fiber atrophy, and the II type muscle fiber satellite cell content decreases simultaneously. Exercise is beneficial to increase muscle mass and improve muscle function in older people. Both resistance and endurance trainings can increase the skeletal muscle, especially the II muscle fiber satellite cell content with a further increase in the satellite cell activation and proliferation. The number and activation degree of satellite cells are related to muscle aging, and satellite cells and proliferation factors regulate muscle cell formation. Therefore, future researches should not only focus on the increase of satellite cell bank, but also explore effective ways to promote the activation of satellite cells, such as exercise training, nutrition and drugs. 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

生长因子对骨骼肌损伤与修复的作用

BACKGROUND: Growth factors involved in the regulation of cellular processes play an important role in the wound healing and tissue regeneration. OBJECTIVE:To elaborate the role of a variety of cellular processes involving growth factors in the repair of skeletal muscle injury, and to provide the references for the treatment and rehabilitation strategies, and the synthesis of biomaterials with growth factor for the skeletal muscle after injury. METHODS: A computer-based online search was conducted in PubMed, Mendeley, Google Scholar, and CNKI databases from 1995 to November 2015 to screen the relevant literatures using the keywords “skeletal muscle, damage repair, insulin-like growth factor, epidermal growth factor, growth factor”. Data screening, processing, and summary were performed. RESULTS AND CONCLUSION: Fifty-one eligible literatures were included. Exercise training promotes the repair and regeneration of the injured skeletal muscle cells and the recovery of the function by activating satellite cells in the sarcolemma and basement membrane to produce the numerous myoblasts. The repair involves the complex biological process regulated by growth factors. Exogenous growth factors up-regulate the mRNA expression of endogenous growth factors, stimulate the proliferation of the myoblasts, accelerate the fusion between myotubes and muscle fibers, promote the repair of skeletal muscle injury, inhibit the formation of scars, thereby enhancing the healing quality. 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

20 W微波照射骨骼肌卫星细胞的增殖与分化：兔骨折内固定模型研究

BACKGROUND:Low power microwave irradiation has been shown to promote the healing of fractures with internal fixation; however, its action mechanisms on the skeletal muscle around the fracture site are unclear. OBJECTIVE:To study the effects of low power microwave irradiation (20 W) on the proliferation ability of skeletal muscle satellite cells in a rabbit model of femoral fracture with internal fixation. METHODS:Forty male New Zealand rabbits were used to establish femoral fracture followed by internal fixation models, and then were equally randomized into spontaneous recovery and microwave treatment groups. Low power microwave irradiation (20 W) was given for 30 consecutive days in the microwave treatment group on day 4 after modeling, while no microwave irradiation was given in the spontaneous recovery group. Rabbit thigh muscles adjacent to the implant were obtained to isolate skeletal muscle satellite cells. Immunohistochemical staining, hematoxylin-eosin staining and quantitative RT-PCR were used to evaluate the ability of the proliferation and differentiation of skeletal muscle satellite cells. RESULTS AND CONCLUSON: Hematoxylin-eosin staining showed that there was no significant difference in the morphology and histology of skeletal muscle tissues between the spontaneous recovery and microwave treatment groups. However, the relative mRNA expression of MyoG in the cultured skeletal muscle satellite cells in vitro and the number of α-sarcometric actin-postive cells in the microwave treatment group were significantly increased compared with the spontaneous recovery group (P < 0.05). The proliferative ability of skeletal muscle satellite cells was inhibited at the early stage, but not at the later stage. Our results suggest that low power microwave irradiation (20 W) can promote the proliferation and differentiation of skeletal muscle satellite cells around the implant in a rabbit model of femoral fracture with internal fixation, and thereby confirm the efficacy and safety of low power microwave irradiation for the internal fixation of fractures. 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

Effects of skeletal muscle regeneration on the proliferation potential of satellite cells

Skeletal muscle satellite cells are myogenic stem cells that function to repair damaged muscle fibers. Participation of satellite cells in a regeneration response following muscle injury results in a significant reduction in their cumulative proliferation potential. The magnitude of the reduction is proportional to the number of regeneration responses in which the cells participate.     

组织激肽释放酶1转染骨髓间充质干细胞的病毒感染复数选择

背景：血管舒缓素即组织激肽释放酶1,是组织激肽释放酶-激肽系统重要组成部分之一。有研究表明血管舒缓素通过促进血管新生,抑制心肌炎症产生等发挥其对心血管系统的保护作用,但并未从干细胞诱导分化方面进行探讨。 目的：实验利用已构建的腺病毒载体,将其携带的血管舒缓素基因转染到大鼠骨髓间充质干细胞上,观察其是否成功转染及转染率如何。 方法：以腺病毒作为载体,将目的基因血管舒缓素转染到大鼠骨髓间充质干细胞上,并采用荧光显微镜、四甲基偶氮唑盐法和流式细胞技术观察病毒对细胞的转染效果,以确定最佳的病毒感染复数。 结果与结论：荧光显微镜下观察到腺病毒携带的血管舒缓素目的基因成功转染到大鼠骨髓间充质干细胞上;流式细胞仪结果显示转染率跟病毒感染复数值的大小有关系,当病毒感染复数为150时,转染率为80.8%;四甲基偶氮唑盐法结果提示,病毒感染复数为200时,细胞生长受到明显的抑制。结果证实,腺病毒介导的血管舒缓素能成功转染到大鼠骨髓间充质干细胞上,且最佳的病毒感染复数为150。 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

Implantation of muscle satellite cells overexpressing myogenin improves denervated muscle atrophy in rats

This study evaluated the effect of muscle satellite cells (MSCs) overexpressing myogenin (MyoG) on denervated muscle atrophy. Rat MSCs were isolated and transfected with the MyoG-EGFP plasmid vector GV143. MyoG-transfected MSCs (MTMs) were transplanted into rat gastrocnemius muscles at 1 week after surgical denervation. Controls included injections of untransfected MSCs or the vehicle only. Muscles were harvested and analyzed at 2, 4, and 24 weeks post-transplantation. Immunofluorescence confirmed MyoG overexpression in MTMs. The muscle wet weight ratio was significantly reduced at 2 weeks after MTM injection (67.17±6.79) compared with muscles injected with MSCs (58.83±5.31) or the vehicle (53.00±7.67; t=2.37, P=0.04 and t=3.39, P=0.007, respectively). The muscle fiber cross-sectional area was also larger at 2 weeks after MTM injection (2.63×10³±0.39×10³) compared with MSC injection (1.99×10³±0.58×10³) or the vehicle only (1.57×10³±0.47×10³; t=2.24, P=0.049 and t=4.22, P=0.002, respectively). At 4 and 24 weeks post-injection, the muscle mass and fiber cross-sectional area were similar across all three experimental groups. Immunohistochemistry showed that the MTM group had larger MyoG-positive fibers. The MTM group (3.18±1.13) also had higher expression of MyoG mRNA than other groups (1.41±0.65 and 1.03±0.19) at 2 weeks after injection (t=2.72, P=0.04). Transplanted MTMs delayed short-term atrophy of denervated muscles. This approach can be optimized as a novel stand-alone therapy or as a bridge to surgical re-innervation of damaged muscles.     

慢病毒载体Pax6-EGFP构建及转染人骨髓间充质干细胞

BACKGROUND:Pax6 gene plays an important role in eye development and differentiation, and to study how it regulates the differentiation of human bone marrow mesenchymal stem cells (BMSCs), gaining the BMSCs stably over-expressing Pax6 is crucial, which is also the basis of stem cell replacement therapy. OBJECTIVE:To construct a lentivirus vector containing Pax6 and detect the expression of Pax6 in transfected human BMSCs. METHODS:Pax6 gene was extracted using PCR. After its connection with lentivirus vector pHIV-EGFP, it was then packaged by 293T cells. The human BMSCs were transfected with recombinant lentivirus Pax6-EGFP as well as lentivirus vector pHIV-EGFP, which was considered negative control group. The cellular morphology was observed by a fluorescence microscope, and the mRNA expression of Pax6 was detected by real-time PCR. RESULTS AND CONCLUSION:The recombinant lentivirus Pax6-EGFP was constructed successfully with a titer of 3×109 pfu/L. After the transfection, both the green fluorescent protein and Pax6 gene were expressed detected using fluorescence microscope and real-time PCR, showing that the method of lentiviral transfection is a safe and effective way to modify BMSCs.     

超声靶向微泡破坏技术介导报告基因对骨骼肌基因转染及不同给药途径的作用评价

目的 探讨超声靶向微泡破坏(UTMD)对小鼠骨骼肌基因转染的有效性,比较不同给药途径及报告质粒转染效率的差异.方法 实验随机分为单纯质粒注射组(P)、质粒+微泡组(P+MB)、质粒+超声辐照组(P+US)、质粒+微泡+超声辐照组(P+UTMD)4组.将绿色荧光蛋白(GFP)质粒或红色荧光蛋白(RFP)质粒分别与微泡(或生理盐水对照)混合,将混合物分别予以局部注射(单侧鼠胫骨前肌)和尾静脉注射,进行或不进行超声辐照,辐照频率为3 MHz、声强为1.0 W/cm2,辐照2 min,7d后处死小鼠,分别取各组小鼠的肝、心、骨骼肌组织制备冰冻切片.应用激光共聚焦显微镜检测各组小鼠骨骼肌组织荧光蛋白质粒GFP、RFP的转染效率;分析P+UTMD组小鼠肝脏、心脏组织中的荧光素酶活性;苏木精-伊红(HE)染色观察P+UTMD组小鼠肝、心、骨骼肌组织的病理学改变;对比P+UTMD组小鼠局部肌肉注射和尾静脉注射时不同的给药途径下肝、心、骨骼肌组织基因转染的效果.结果 单纯质粒注射组骨骼肌组织每个视野只有少数细胞表达RFP和GFP,表达率为(1.83±1.21)％、(1.18±0.25)％,P+MB组和P+US组的RFP、GFP表达率与单纯质粒注射的效果相当[(2.33±1.39)％、(3.42±1.09)％和(3.48±0.18)％、(2.52±0.33)％];而P+UTMD组RFP、GFP的表达率为(23.96±2.13)％、(25.69±1.98)％,显著高于其他3组(P＜0.05).P+UTMD组小鼠部分肝脏和心脏中均有较微弱的荧光表达,HE染色未发现UTMD对肝、心、骨骼肌组织造成明显的损伤,完整性良好,无感染、出血、水肿及细胞死亡.直接肌肉局部注射质粒联合UTMD能显著增强小鼠骨骼肌局部的基因转染,显著优于尾静脉注射(P＜0.05).结论 UTMD可有效促进基因转移,局部注射联合UTMD能显著增强骨骼肌局部基因转染,是一种安全、高效的基因转染新技术.     

Probucol对碱性成纤维细胞生长因子和过氧化氢促大鼠血管平滑肌细胞增殖的影响

目的:研究probucol对bFGF和H₂O₂促大鼠血管平滑肌细胞(VSMC)增殖的影响。方法:采用MTT、细胞计数和[3H]-TdR掺入法观察probucol对bFGF和H₂O₂促VSMC增殖的影响。结果:①Probucol抑制bFGF和H₂O₂刺激VSMC增殖和DNA合成,且呈剂量依赖性。Probucol+bFGF和probucol+H₂O₂组与bFGF和H₂O₂组比较,细胞计数、A值和[3H]-TdR掺入量分别下降了40.0%、39.1%、45.5%和46.9%、45.0%、39.5%(P＜0.05,P＜0.01)。②bFGF和H₂O₂刺激前给予probucol预处理24h能显著抑制VSMC增殖和DNA合成(P＜0.05),而bFGF和H₂O₂刺激后24h给予probucol则无明显影响(P＞0.05)。结论:Probucol能显著抑制bFGF和H₂O₂刺激的VSMC增殖和DNA合成,但对bFGF和H₂O₂预刺激诱导的细胞增殖无抑制作用。