Interaction between porcine granulosa and thecal cells in steroidogenesis in an amnion dual chamber culture system

Objective: To study the interaction between granulosa and thecal cel1s in steroidogenesis in an am-nion dual chamber in comparison with the cellulose dual chamber.Method: A dual chamber culture system was designed and prepared with amnion membrane from human term placenta. The isolated porcine granulosa and thecal cells were grown on both sides of the amnion membrane, with granulosa cells in the inner chamber and thecal cells in the outer chamber. The concentrations of estradiol (E,), progesterone (P) and testosterone (T) in the culture media were mea-sured by radioimmunoassay.Results: The growth of both cells and their steroidogenic function were more active in amnion dual chamber system than in cellulose dual chamber system: (1) more T produced by thecal cells in the outer chamber, passing into inner chamber through the amnion membrane. T was used by granulosa cells as the substrate of aromatization, so that granulosa cells produced more E2 (up to 2 435 pmol/L); (2) the production of P (52. 5 μmol/L) and T (10. 2μmol/L) by granulosa cells cultured in the amnion mem-brane dual chamber system were also higher.Conclusions:The dual chamber system made of amnion mpmbrane showed better effect in studying steroidogenesis than with cellulose dual chamber system, and can be used as a model for studying paracrine regulatory interaction between granulosa and thecal cells in vitro.

Amnion epithelial cells——a novel therapy for ischemic stroke?

Stroke is a leading cause of death and disability and new therapies are desperately needed. Given the complex nature of ischemic brain injury, it has been postulated that cell-based therapies may be useful. However, cell resources, invasive extraction procedures, immunological rejection, tumorigenesis and ethical challenges make it unlikely that many stem cell types could serve as a practical source for therapy. By contrast, these issues do not pertain to human amnion epithelial cells（h AECs）, which are placenta-derived stem cells. We recently assessed the effects of systemically delivered hAECs on stroke outcome using four animal models of stroke. We demonstrated that when injected intravenously after ischemia onset, hAECs migrate preferentially to the spleen and injured brain to limit apoptosis and inflammation, and attenuate early brain infiltration of immune cells, progression of infarction and systemic immunosuppression and to ultimately ameliorate functional deficits. When administration of hAECs is delayed by 1-3 days poststroke, long-term functional recovery can still be enhanced in young and aged mice of either sex. Moreover, our proof-of-principle findings suggest that h AECs are effective at limiting post-stroke infarct development in non-human primates. Overall, the results suggest that hAECs could be a viable clinical stroke therapy.

Human amnion tissue injected with human umbilical cord mesenchymal stem cells repairs damaged sciatic nerves in rats

Human umbilical cord mesenchymal stem cells,incorporated into an amnion carrier tubes,were assessed for nerve regeneration potential in a rat nerve defect model.Damaged nerves were exposed to human amnion carriers containing either human umbilical cord mesenchymal stem cell （cell transplantation group）or saline（control group）.At 8,12,16 and 20 weeks after cell implantation,the sciatic functional index was higher in the cell transplantation group compared with the control group.Furthermore,electrophysiological examination showed that threshold stimulus and maximum stimulus intensity gradually decreased while compound action potential amplitude gradually increased.Hematoxylin-eosin staining showed that regenerating nerve fibers were arranged in nerve tracts in the cell transplantation group and connective tissue between nerve tracts and amnion tissue reduced over time.Gastrocnemius muscle cell diameter,wet weight and restoration ratio were increased.These data indicate that transplanted human umbilical cord mesenchymal stem cells,using the amnion tube connection method,promote restoration of damaged sciatic nerves in rats.

胚胎干细胞源性表皮干细胞在肾被囊微环境中分化潜能的研究

目的探讨胚胎干细胞源性表皮干细胞在肾被囊微环境中的分化情况,为研究其在不同微环境中的分化潜能和稳定性及寻找新的皮肤工程种子细胞奠定基础。方法129小鼠E14胚胎干细胞与人羊膜共培养4d,定向诱导其分化为呈β1整合素、CK15和CK19阳性的表皮样干细胞克隆,无菌手术下移植入129小鼠双侧肾被囊内,术后4周、6周和8周取材。对移植后细胞的分化情况进行形态学和CEA和CK18免疫组织化学观察。结果小鼠胚胎干细胞源性表皮干细胞在129小鼠肾被囊内4周,分化为由单层或复层上皮样细胞构成的管状和泡状结构,种植6周和8周,除上述结构外,可见角化复层扁平上皮、汗腺样、皮脂腺样及毛囊样等结构。免疫组化结果汗腺样结构呈CEA和CK18阳性。结论研究结果表明小鼠胚胎干细胞源性表皮干细胞在肾被囊微环境下,可具有分化为角化复层上皮、汗腺样、皮脂腺样及毛囊样结构的潜能。

羊膜与人胚半月板细胞构建复合物的实验研究

目的研究羊膜作为半月板组织工程支架材料应用的可行性。方法采用胶原酶阶段消化法获取人胚半月板纤维软骨细胞,取经TGF-β1扩增的第3代细胞与羊膜进行复合培养,进行MTT、光镜、扫描电镜观察。结果纤维软骨细胞于羊膜上牢固粘附,3天开始增殖,2周时已有大量细胞生长于羊膜上。结论羊膜表现出良好的细胞吸附性和体外生物相容性,半月板细胞可在其上粘附、增殖,因此可作为半月板组织工程细胞支架材料。

Neuronal-like differentiation of single versus multiple treatments with human amnion-derived mesenchymal stem cells induced by basic fibroblast growth factor

BACKGROUND： Cultures from multiple portions of umbilical cord blood mesenchymal stem cells have been shown to undergo more rapid proliferation and attachment than single portions. OBJECTIVE： To observe growth of basic fibroblast growth factor （bFGF）-induced cultures of human amnion-derived mesenchymal stem cells （AMSCs） and differentiation into neuronal-like cells. DESIGN, TIME AND SETTING： Comparative observation. The study was performed at the Laboratory of Microbiology and Immunology, Basic Medical School of Zhengzhou University from January to May 2008. METHODS： Amnia from full-term, uterine-incision delivery were donated by 12 healthy women. AMSCs were obtained by cell separation and culture techniques, and were passaged and induced by bFGF. From the third passage, a total of 1 mLAMSCs, at a density of 1.0 × 10^4/mL, was separately harvested from six samples, which served as group A. A total of 1 mL AMSCs, at a density of 1.0 × 10^4/mL, was harvested separately from the remaining six samples, which served a group B. A total of 0.5 mL from the six samples of group A and 0.5 mL from the six samples of grot, B were combined to form group C. MAIN OUTCOME MEASURES： Differences in cell quantity among the three groups were compare by cell quantification and 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） analysis. Expression of a glial cell marker, neuron-specific enolase, and nestin was detected in the three groups by immunocytochemistry. RESULTS： Cell quantification and MTT analysis of live cells, as well as AMSC absorbance, were significantly greater in group C compared with groups A and B at 18 days of culture （P 〈 0.05）, anc no significant difference was observed between groups A and B. Glial fibrillary acidic protein, neuron-specific enolase, and nestin were expressed in all groups following bFGF induction. CONCLUSION： Mixed AMSC cultures promoted proliferation, and bFGF-induced AMSCs differentiated into neuronal-like cells.

Effect of amnion membrane transplantation on corneal neovascularization in 10 patients with alkali burn

By observing clinical cases, we studied the curative effect of amnion membrane transplantation on decreasing corneal neovascularization (CNV). It was a non-randomized retrospective case-control study. Among 17 cases (21 eyes) of third-degree alkali burns from 2007 to 2010, 10 cases (12 eyes) were performed with amnion membrane transplantation operation, and others were not. Amnion membrane transplantation was performed at the 3rd day after burn in the treatment group. Areas of CNV in double groups were measured at the 14th day and 60th day after burn. Area of CNV in the treatment group was (66.207±7.251)mm2 at the 14th day after burn, and was 18.27% lower than that in the control group. Area of CNV in the treatment group was (120.046±13.812)mm2 at the 60th day after burn, and was 11.35% lower than that in the control group. There was both statistical significance (P<0.05). Amnion membrane transplantation operation can inhibit the growth of corneal neovascularization induced by alkali burn.

Mesenchymal stromal cells from human perinatal tissues: From biology to cell therapy

Cell-based regenerative medicine is of growing interest in biomedical research. The role of stem cells in this context is under intense scrutiny and may help to define principles of organ regeneration and develop innovative therapeutics for organ failure. Utilizing stem and progenitor cells for organ replacement has been conducted for many years when performing hematopoietic stem cell transplantation. Since the first successful transplantation of umbilical cord blood to treat hematological malignancies, non-hematopoietic stem and progenitor cell populations have recently been identified within umbilical cord blood and other perinatal and fetal tissues. A cell population entitled mesenchymal stromal cells (MSCs) emerged as one of the most intensely studied as it subsumes a variety of capacities: MSCs can differentiate into various subtypes of the mesodermal lineage, they secrete a large array of trophic factors suitable of recruiting endogenous repair processes and they are immunomodulatory.Focusing on perinatal tissues to isolate MSCs, we will discuss some of the challenges associated with these cell types concentrating on concepts of isolation and expansion, the comparison with cells derived from other tissue sources, regarding phenotype and differentiation capacity and finally their therapeutic potential.

腺病毒载体携带标记基因EGFP转染人羊膜上皮细胞

背景:人羊膜上皮细胞具有干细胞的某些性能,可诱导分化为角膜样上皮细胞,但在体外培养时无法示踪,不能有效地监测其细胞转归。目的:探讨利用腺病毒载体将标记基因EGFP转染入人羊膜上皮细胞的可行性及感染效率。方法:pD C316-mC MV-EGFP腺病毒包装后,腺病毒载体携带标记基因EGFP转染体外培养的人羊膜上皮细胞,并扩增培养,观察转染后人羊膜上皮细胞与未转染人羊膜上皮细胞的形态差异,并在荧光显微镜下观察转染基因的表达,用流式细胞仪检测转染细胞的细胞周期及EGFP阳性表达率。结果与结论:(1)多数转染细胞筛选出的抗性克隆以及培养的克隆细胞与正常体外培养的人羊膜上皮细胞在形态上无显著差异;(2)瞬时转染后48 h的人羊膜上皮细胞中增强型绿色荧光蛋白阳性表达率最高达99.01%;(3)转染后的人羊膜上皮细胞的细胞周期有所减慢,说明利用腺病毒载体能将标记基因EGFP转染入人羊膜上皮细胞,可更直观地观察到人羊膜上皮细胞在体外的进一步转归。

不同冻存方法对羊膜超微结构和上皮细胞活性的影响（英文）

背景:羊膜冻存方法众多,对羊膜超微结构和生物活性的影响不一,目前尚无有效的冻存方法。目的:比较不同冻存方法对羊膜超微结构和活性影响的研究,探寻更为理想的冻存方法。方法:将新鲜羊膜采用深低温和玻璃化冻存法保存,分别于冻存后3,6个月复苏羊膜,以新鲜羊膜组织为对照组,比较羊膜的超微结构差异、羊膜上皮细胞离体氧分压和乳酸脱氢酶活性。结果与结论:不同冻存方法保存的羊膜超微结构有明显改变,但玻璃化冻存对其超微结构的影响相对较小;与新鲜羊膜相比较,深低温冻存组3,6个月羊膜的乳酸脱氢酶灰度值和氧分压明显降低(P<0.05);玻璃化冻存组6个月后的羊膜乳酸脱氢酶灰度值和氧分压明显降低(P<0.05),而玻璃化冻存组3个月后的上述检测结果与新鲜羊膜相比差异无显著性意义(P>0.05)。结果证实,羊膜的玻璃化冻存技术优于深低温冻存技术,不仅维持了羊膜的超微结构,而且保持了羊膜上皮细胞的功能和活性。

新鲜羊膜移植治疗翼状胬肉的观察及护理

目的探索适合胬肉切除羊膜移植术后的护理措施,提高手术成功率。方法对143例(168眼)因翼状胬肉而施行新鲜羊膜移植手术的患者进行观察及护理。结果143例患者均未发生术后感染,溃烂、排斥反应等,愈合良好。结论手术前后的护理及正确的出院健康指导是手术成功的保证。

三种人胎盘组织来源间充质干细胞的生物学特性

背景:胎盘组织结构复杂,包括胎儿侧的羊膜、绒毛膜以及母体侧的蜕膜等。尽管一些文献已经报道这3种组织来源间充质干细胞具有一些共有的生物学特性,但是对于来自同一胎盘不同组织来源间充质干细胞是否存在不同的生物学特性,目前仍缺乏定量的研究分析。目的:探讨人胎盘不同组织来源间充质干细胞的生物学特性,包括分化潜能和免疫调节功能。方法:利用酶消化法从1例男婴胎盘组织分离羊膜来源间充质干细胞、绒毛膜来源间充质干细胞和蜕膜来源间充质干细胞。系统性研究这3种间充质干细胞的生物学特性,包括细胞形态、表面标志物表达、核型分析、成脂、成骨分化能力以及调节性T细胞的增殖能力。结果与结论:(1)3种细胞均呈成纤维细胞形态,均表达间充质干细胞的表面标志物:高表达CD73、CD90和CD105,低表达CD14、CD19、CD34、CD45和HLA-DR;(2)羊膜、绒毛膜间充质干细胞核型与胎儿相同,蜕膜间充质干细胞核型与母体相同;(3)3种间充质干细胞成脂分化能力存在显著性差异,依次为蜕膜间充质干细胞>绒毛膜间充质干细胞>羊膜间充质干细胞(P <0.05);相反,羊膜间充质干细胞的成骨分化能力明显高于蜕膜间充质干细胞(P <0.05);(4)羊膜间充质干细胞和绒毛膜间充质干细胞促进调节性T细胞增殖的能力显著高于蜕膜间充质干细胞;(5)结果表明,人胎盘羊膜、绒毛膜、蜕膜组织来源间充质干细胞具有不同的核型、成脂、成骨分化潜能和免疫调节能力,为不同种子细胞治疗相关疾病达到最佳效果提供了一定的参考。

葛根芩连汤联合人羊膜间充质干细胞移植治疗糖尿病早期视网膜病变

背景:研究发现人羊膜间充质干细胞可以向胰岛素分泌细胞分化,还具有免疫调节作用。临床证实葛根芩连汤有明确降低血糖的作用。目的:探讨葛根芩连汤联合人羊膜间充质干细胞移植对糖尿病大鼠早期视网膜病变形态学、相关血清学指标及血管内皮生长因子基因表达的影响。方法:以腹腔注射链脲佐菌素方式制作糖尿病模型,将造模成功的42只Wistar大鼠随机分为模型组、人羊膜间充质干细胞移植组和联合组(人羊膜间充质干细胞移植+葛根芩连汤),移植后4周测定糖尿病大鼠血糖、血清胰岛素水平;移植后4周苏木精-伊红染色镜下观察糖尿病大鼠视网膜的病理形态;免疫组织化学方法检测视网膜组织CD45的表达水平;RT-PCR法检测视网膜血管内皮生长因子m RNA的表达。结果与结论:1与模型组比较,人羊膜间充质干细胞移植组和联合组大鼠血糖降低,血清胰岛素浓度增加,联合组改善更为明显,组间比较差异有显著性意义(P<0.05);2模型组大鼠有明显视网膜水肿、层次紊乱,细胞排列不规则;与模型组比较,人羊膜间充质干细胞移植组大鼠视网膜病变相对轻微,联合组视网膜病变明显好转;3模型组视网膜CD45的表达明显增加,人羊膜间充质干细胞移植组CD45的表达量明显小于模型组(P<0.05),但大于联合组(P<0.05);4联合组和人羊膜间充质干细胞移植组血管内皮生长因子m RNA表达低于模型组,联合组降低幅度更大,差异有显著性意义(P<0.05);5结果表明,葛根芩连汤联合人羊膜间充质干细胞移植可能通过抑制CD45和血管内皮生长因子在视网膜组织表达,从而改善糖尿病视网膜病损严重程度。

人羊膜间充质干细胞体外大量扩增的方法

背景:一个胎盘羊膜的面积约600 cm2,羊膜来源间充质干细胞即取材于胎盘上的羊膜。目的:建立一种简单的体外分离培养人羊膜来源间充质干细胞的方法,并分析其生物学特性。方法:采用胰酶和直接贴壁相结合的方法分离获取人羊膜来源间充质干细胞并进行体外培养,观察细胞形态,分析传代第5代细胞生长曲线,检测第5代细胞表面标志表达和检测细胞周期。取传代第4代细胞行体外成骨细胞诱导和成脂诱导,冻存第4代细胞6个月后复苏,计数复苏后细胞存活率并绘制复苏细胞生长曲线。结果与结论:原代接种后第9天有少许细胞爬出,15 d左右细胞达80%-90%融合,细胞以梭形为主。细胞传代后,细胞形态均一,螺旋状排列。传代细胞潜伏期48 h,对数增殖期4 d左右,对数增殖期后进入平台期。间充质干细胞表面CD34、CD14、CD19、CD45、HLA-DR呈阴性表达,CD73、CD105、CD90呈阳性表达。茜素红染色及油红O染色阳性,证实具有向脂肪细胞、成骨细胞分化的能力。流式细胞术细胞周期检测,S期占28%。冻存复苏后细胞存活率达90%以上,且与未冻存传代细胞具有相同的生长特性。结果证实实验成功地建立了一种简化的人羊膜来源间充质干细胞大量扩增的方法。

人胎盘羊膜上皮细胞的分离鉴定及多向诱导分化

背景:人羊膜上皮细胞来源稳定,正逐渐成为广受关注的再生医学种子细胞来源。目的:建立人羊膜上皮细胞体外分离培养、成脂、成软骨、成骨诱导分化的方法。方法:通过胰蛋白酶消化法从人胎盘羊膜中分离羊膜上皮细胞,进行体外培养与鉴定,观察培养12 d内的细胞生长曲线。取P1代羊膜上皮细胞,分别进行成脂、成软骨、成骨诱导,以常规培养细胞为对照,诱导16 d后,分别进行油红O染色、Masson染色及碱性磷酸酶染色,同时采用荧光定量PCR检测诱导过程中成脂转录因子、Ⅱ型胶原蛋白、骨桥蛋白、碱性磷酸酶m RNA表达变化。结果与结论:(1)从人羊膜中分离出的羊膜上皮细胞,免疫荧光检测表达上皮细胞标记物CK19;(2)P1代细胞具有较强的分裂增殖能力,P2细胞与P1相比增殖能力略有下降,P3细胞增殖能力最差;(3)羊膜上皮细胞经成脂、成软骨、成骨诱导后,油红O染色有红色脂滴,Masson染色有亮蓝色软骨基质,碱性磷酸酶染色有红褐色钙结节;随着诱导时间的延长,成脂转录因子、Ⅱ型胶原蛋白、骨桥蛋白、碱性磷酸酶表达量升高;(4)结果表明,采用酶消化法可从人羊膜中分离得到羊膜上皮细胞,羊膜上皮细胞可向脂肪细胞、成软骨细胞、成骨细胞分化。

人羊膜间充质干细胞移植减轻体外循环再灌注肺损伤

背景：近年来,间充质干细胞在器官或组织修复中展现了良好的前景,因此,基于间充质干细胞可塑性的细胞移植为体外循环后肺损伤的细胞再生及功能恢复提供了可能。目的：探讨外源性移植人羊膜间充质干细胞对犬体外循环缺血再灌注肺损伤的影响,从炎性因子角度初步探讨其作用机制。方法：健康成年杂种犬18只,随机分为空白组（建立体外循环模型,主动脉不阻断＋股静脉注射1 mL生理盐水）、对照组（体外循环＋主动脉阻断1h＋主动脉开放15 min后股静脉注射1 mL生理盐水）、实验组（体外循环＋主动脉阻断1 h＋主动脉开放15 min后股静脉注射含2×10~7人羊膜间充质干细胞的生理盐水1 mL）,每组6只。分别在体外循环前（T1）、主动脉开放后15 min（T2）、开放后1h（T3）、开放后2h（T4）、开放后3h（T5）5个时间点,各取动脉血2 mL计算氧合指数和呼吸指数,取静脉血8 mL分别用ELISA法检测血清肿瘤坏死因子α、基质金属蛋白酶9、白细胞介素8、白细胞介素10水平,Western Blot印迹法检测肺组织NF-κB蛋白表达,同时光镜下观察各组肺组织病理学变化。结果与结论：①与对照组比较,实验组氧合指数在人羊膜间充质干细胞移植后2,3 h显著升高,实验组呼吸指数在人羊膜间充质干细胞移植后1,2,3 h显著下降。②与对照组同时间点相比,实验组肿瘤坏死因子α、白细胞介素8以及基质金属蛋白酶9呈明显下降趋势,白细胞介素10有所回升。③与空白组相比,对照组开放后3 h肺组织NF-κB蛋白明显增强,静脉注入人羊膜间充质干细胞后肺组织NF-κB表达较对照组显著降低。④实验组在主动脉开放后,病理改变明显轻于对照组,可见少量间隙内渗出红细胞与中性粒细胞,微血管扩张、充血。⑤结果表明,人羊膜间充质干细胞静脉移植可通过下调促炎因子肿瘤坏死因子α、白细胞介素8,降低基质金属蛋白酶9水平,上调抗炎因子白细胞介素10水平,抑制肺组织NF-κB活化,减轻体外循环缺血再灌注相关肺损伤。

痰热清联合人羊膜间充质干细胞移植对输血相关急性肺损伤的保护

背景:随着细胞技术的提高,使得干细胞移植研究延伸到包括输血相关急性肺损伤在内的各个领域。目的:探讨痰热清注射液联合人羊膜间充质干细胞移植对输血相关急性肺损伤小鼠的保护作用及机制。方法:选取80只BALB/C雄性小鼠,随机分为对照组、模型组、痰热清注射液组、人羊膜间充质干细胞移植组及联合治疗组。除对照组外,其他4组制备输血相关急性肺损伤模型。经尾静脉注入痰热清注射液和人羊膜间充质干细胞悬液干预24 h后,镜下观察各组小鼠肺组织病理形态,计算肺组织湿干质量比,ELISA测定各组小鼠支气管肺泡灌洗液中白细胞介素10、白细胞介素1β、肿瘤坏死因子α水平,RT-PCR、Western blot检测肺损伤小鼠肺组织KGF-1基因和蛋白表达,TUNEL法检测肺组织细胞凋亡情况。结果与结论:(1)模型组小鼠肺组织出现肺泡间隔增厚、肺泡内纤维蛋白浸润、肺泡内出血、支气管壁增厚等病理变化,肺组织湿干质量比增高,人羊膜间充质干细胞移植组和痰热清注射液组以上病理变化较模型组略轻,肺组织湿干质量比低于模型组(P<0.05),联合治疗组肺组织湿干质量比明显低于模型组(P<0.01);(2)人羊膜间充质干细胞移植组和痰热清注射液组小鼠肺泡灌洗液中白细胞介素10、白细胞介素1β、肿瘤坏死因子α水平均低于模型组(P<0.05);联合治疗组上述各因子水平明显低于模型组(P<0.01),仍高于正常对照组(P<0.05);(3)联合治疗组KGF-1基因和蛋白的表达高于人羊膜间充质干细胞移植组和痰热清注射液组(P<0.05),人羊膜间充质干细胞移植组和痰热清注射液组高于模型组(P<0.05);(4)联合治疗组凋亡细胞数低于人羊膜间充质干细胞移植组和痰热清注射液组,人羊膜间充质干细胞移植组和痰热清注射液组低于模型组;(5)结果表明,痰热清注射液联合人羊膜间充质干细胞移植能提高输血相关急性肺损伤小鼠肺组织中KGF-1的表达,减轻肺部炎症反应及组织病理学损伤程度,减少肺组织细胞凋亡,对肺损伤具有保护作用。

尾静脉移植羊膜间充质干细胞对缺血再灌注损伤后神经功能恢复的影响

背景:研究表明,MAG、OMgp等髓磷脂抑制物的过表达是早期神经元再生失败的主要原因。目的:探讨人羊膜间充质干细胞移植对缺血再灌注损伤大鼠神经功能恢复的影响。方法:SD大鼠60只,随机分为假手术组、缺血再灌注损伤组及人羊膜间充质干细胞移植组,每组20只。人羊膜间充质干细胞组于缺血再灌注损伤后24 h经尾静脉注入细胞浓度为1.0×108 L-1的人羊膜间充质干细胞悬液1 m L。移植后24 h,3 d及1,2,3周,采用Longa行为学评分、圆筒实验、水平梯行走实验及肢体对称性实验对各组大鼠进行行为学评估,观察神经功能恢复情况;移植1 d和1,2,3周后免疫荧光法观察细胞迁移和分布情况;移植后2周采用Western blot检测各组大鼠脑组织MAG及OMgp蛋白的表达;移植后3周采用TUNEL染色观察神经元凋亡情况。结果与结论:(1)移植后1周在损伤区周围可见红色阳性标记细胞,并且随移植时间的延长,阳性标记细胞逐渐增多;(2)与缺血再灌注损伤组比较,在移植后3 d及1,2,3周人羊膜间充质干细胞移植组大鼠神经功能得到明显改善(P<0.05);(3)与缺血再灌注损伤组比较,人羊膜间充质干细胞移植组MAG及OMgp蛋白的表达量均显著降低,差异有显著性意义(P<0.05);(4)与假手术组相比,缺血再灌注损伤组凋亡细胞数增多;人羊膜间充质干细胞移植组凋亡细胞数目较缺血再灌注损伤组下降;(5)结果表明,人羊膜间充质干细胞移植可能通过降低MAG及OMgp表达,减少神经元凋亡,进而促进缺血再灌注损伤大鼠的神经功能恢复。

生物衍生羊膜对大鼠跟腱粘连影响的实验研究

目的 研究生物衍生羊膜预防 SD大鼠跟腱粘连的效果 ,并为临床预防术后跟腱粘连提供实验数据。方法 以 2 8只 SD大鼠后肢跟腱为实验模型 ,切断并缝合跟腱 ,实验侧跟腱包裹生物衍生羊膜 ,对侧不包裹作为对照 ,术后 1、2、4、6、8和 12周分别取跟腱 ,大体及组织学观察跟腱的粘连、愈合情况。 结果 术后各时间点两侧跟腱吻合处炎性细胞浸润情况、跟腱愈合进程无明显差别 ,跟腱粘连的半定量评分表明实验侧和对照侧差异有统计学意义 ( P<0 .0 5 )。 结论 生物衍生羊膜能有效预防跟腱粘连 ,是一种较为理想的预防跟腱粘连的生物材料。

不同浓度人羊膜匀浆上清液培养大鼠许旺细胞的增殖

背景:许旺细胞是周围神经修复过程的重要细胞,而研究发现人羊膜细胞分泌的多种细胞因子能够促进许旺细胞增殖。目的:观察不同浓度人羊膜匀浆上清液对鼠许旺细胞(RSC96)生长的影响。方法:使用含体积分数20%胎牛血清的高糖DMEM培养基原代培养RSC96细胞株,传代至第2代用于实验研究。根据人羊膜匀浆上清液在培养基中的不同体积分数(0,10,15,20,25%)分组。结果与结论:人羊膜匀浆上清液的总蛋白浓度为675 mg/L,表皮生长因子、碱性成纤维生长因子和血管内皮生长因子浓度分别为(470.625±2.546),(4.121±0.026)和(0.172±0.002)ng/L。在培养第1-7天,10%和15%人羊膜匀浆上清液组的增殖率大于20%和25%人羊膜匀浆上清液组(P<0.05);10%、15%人羊膜匀浆上清液组显示出促进细胞增殖的作用,而20%、25%人羊膜匀浆上清液组显示出抑制细胞增殖的作用;各实验组的细胞活力与对照组接近(P>0.05)。提示人羊膜匀浆上清液低浓度时(10%和15%)具有促进RSC96增殖作用,高浓度时(20%和25%)抑制RSC96增殖。