间充质干细胞来源外泌体对视网膜新生血管影响的研究进展

视网膜新生血管形成是许多视网膜疾病的病理特征,例如早产儿视网膜病变和糖尿病视网膜病变,可导致严重的视力丧失甚至失明。抑制视网膜新生血管形成是治疗这些视网膜疾病的治疗策略。目前,已存在几种抑制视网膜新生血管形成的治疗策略,包括激光封闭、抑制血管内皮生长因子(VEGF)以及干细胞的移植等。随着干细胞研究的深入,发现干细胞治疗尽管潜力极大,但亦存在如移植细胞的低生存力,先天异质性等技术障碍,目前研究发现来源于间充质干细胞(MSCs)的外泌体具有与MSCs相似的功能,且尺寸小、易于通过生物膜,为细胞治疗提供了一种新思路,本文就外泌体对视网膜新生血管疾病的最新进展作一综述。     

间充质干细胞外泌体对眼部疾病的治疗作用

间充质干细胞(mesenchymal stem cells,MSCs)是一种来源于多种组织,具有多向细胞分化潜能和自我细胞更新的多能干细胞,可对多种眼部疾病发挥治疗作用。研究表明,MSCs的生物学功能与其分泌的外泌体(MSC-Exo)有密切联系。相较而言,MSC-Exo的生物性能更加稳定,且不具有成瘤性,因此有望成为MSCs生物学功能的有效替代物。本文对MSC-Exo在眼部疾病中的应用进展讨论,以期为MSC-Exo在眼部疾病的治疗提供思路和借鉴。     

间充质干细胞源性外泌体治疗干眼的研究进展

外泌体是一种细胞外囊泡,通过生物分子传递改变受体细胞的生化特性,并在细胞通讯中发挥作用。其中,间充质干细胞源性外泌体(MSC-Exos)是由间充质干细胞分泌的双层脂质囊泡,具有与间充质干细胞(MSCs)相似的功能,并携带蛋白质、脂质和核酸等生物活性物质,可作为细胞间通讯的媒介参与免疫调节、组织损伤修复和促进血管生成等多种重要生理过程,近年来在治疗免疫炎性疾病、缺血性疾病等相关领域成为研究的热点。本文从MSC-Exos的生物学功能出发,针对干眼不同发病机制如炎症反应、神经及组织修复等,阐述其治疗干眼的可能机制,以期为MSC-Exos在干眼治疗中的应用奠定基础,并为未来的临床应用提供参考依据。     

间充质干细胞源性外泌体对前部缺血性视神经病变患者的治疗潜力

前部缺血性视神经病变(anterior ischemic optic neuropathy,AION)是继青光眼后成人永久性视神经相关视力丧失的第二大常见原因,目前尚无确切有效的治疗方案。既往研究发现间充质干细胞源性外泌体(mesenchymal stem cells-exosomes,MSC-exosomes)与MSCs非常相似地改善缺血后神经组织的损伤,并诱导与增强血管神经发生相关的长期神经保护作用。外泌体是细胞-细胞间交流的一种重要通讯途径,因此可作为小分子物质传递载体信号,并通过不同机制作用于靶细胞,从而成为眼前部缺血性视神经病变潜在的有效治疗工具。本文对MSC-exosomes的生物发生及其功能,以及MSC-exosomes在前部缺血性视神经性病变中的治疗机制进行综述。     

人脐带间充质干细胞外泌体保护人视网膜色素上皮细胞抵抗高糖诱导的细胞凋亡

目的 人脐带间充质干细胞(h UMSCs)外泌体在保护人视网膜色素上皮(RPE)细胞抵抗高糖诱导的细胞凋亡中的作用。方法 实验研究。超速离心法收集h UMSCs外泌体,通过透射电镜和纳米颗粒追踪分析其形态结构;采用CCK-8和Ed U实验检测外泌体处理后的人RPE在高糖条件下的增殖情况;通过蛋白免疫印迹(western blot)检测周期蛋白和凋亡蛋白的变化;运用流式细胞仪检测h UMSCs外泌体在高糖诱导凋亡率中的作用。结果高浓度糖培养环境通过促进线粒体介导的细胞凋亡而抑制了ARPE-19的增殖;来自h UMSCs的外泌体(h UMSC-exo)能促进人RPE细胞的增殖及保护ARPE-19抵抗高糖诱导的细胞凋亡。结论 h UMSCs外泌体能够保护人RPE细胞抵抗高糖诱导的细胞凋亡。     

间充质干细胞来源外泌体（MSCs-exo）对大鼠角膜移植排斥反应的抑制作用

目的　观察间充质干细胞来源外泌体（mesenchymal stems cells derived exosomes,MSCs-exo）对大鼠角膜移植排斥反应的抑制作用。方法　制作大鼠异体角膜移植排斥反应模型,随机分为4组:空白对照组、MSCs结膜下治疗组、PBS结膜下注射组、MSCs-exo结膜下治疗组。术后第3天开始裂隙灯下观察角膜植片排斥情况,并对MSCs-exo进行染色及示踪观察其结局。结果　空白对照组角膜植片存活时间为（9.67±0.56）d,MSCs结膜下治疗组为（12.33±0.76）d,PBS结膜下注射组为（9.50±0.34）d,MSCs-exo结膜下治疗组为（16.5±1.15）d。PBS结膜下注射组角膜植片存活时间较空白对照组差异无统计学意义（P>0.05）,而MSCs结膜下治疗组与MSCs-exo结膜下治疗组角膜植片存活时间较PBS结膜下注射组均明显延长（均为P<0.05）,MSCs-exo结膜下治疗组植片存活时间长于MSCs结膜下治疗组（P<0.05）。空白对照组大鼠免疫组织化学染色结果显示植片部分厚度比植床角膜厚度显著增厚,上皮层及基质层内见大量CD4+细胞聚集。而MSCs-exo结膜下治疗组植片中CD4+细胞比空白对照组明显减少。MSCs-exo结膜下注射后72 h时,仅有少量外泌体可在结膜下被观测到。进一步将观测时间前推至注射后24 h,发现大量外泌体存在于结膜下,而角膜组织和前房内也可见到外泌体的定位表达。 结论　结膜下注射MSCs-exo可以显著延长角膜植片存活时间,外泌体治疗可能成为无细胞治疗的新方法。     

外泌体在糖尿病视网膜病变的机制研究进展

外泌体是广泛存在于人体，由机体细胞产生并分泌的纳米级细胞外囊泡，能相对稳定地存在于各种生物组织和体液中，并携带特定的miRNA、蛋白质、转录因子等多种信息分子，参与调控体内多种疾病的病理生理过程。近年来随着外泌体在各学科研究的不断深入，其在眼科学领域的研究也迅速开展，目前发现外泌体在糖尿病视网膜病变、年龄相关性黄斑变性、自身免疫性葡萄膜炎、角膜病及青光眼等多种疾病中发挥重要作用。随着生活水平的提高，全世界糖尿病视网膜病变致盲人数逐年增加，而糖尿病视网膜病变机制研究未明，近年许多研究发现外泌体在其中发挥着重要作用，本文对外泌体在糖尿病视网膜病变的发生、发展机制的最新研究进展进行综述。     

视网膜微血管周细胞来源的外泌体对糖尿病小鼠视网膜血管功能障碍的影响

目的 探讨视网膜微血管周细胞(RMVPCs)来源的外泌体(Exos)通过调控血管内皮细胞的生物学功能进而对糖尿病视网膜血管功能障碍的影响。方法 通过超速离心法提取并鉴定人视网膜微血管周细胞(HRMVPCs)来源的Exos(HRMVPCs-Exos),并对Exos进行PKH67染色后，与人脐静脉血管内皮细胞(HUVECs)孵育进行内吞实验，此后，体外实验分为Control组(5.55 mmol·L^-1葡萄糖)、高糖(HG)组(30.00 mmol·L^-1葡萄糖)和HG+HRMVPCs-Exos组，各组HUVECs进行相应处理后，应用CCK-8、细胞迁移、流式细胞术以及成管实验等方法检测HUVECs的增殖、迁移以及血管的生成。体内实验分为NC组、链脲佐菌素(STZ)组和STZ+HRMVPCs-Exos组，每组5只健康雄性C57BL/6J小鼠，各组小鼠进行相应处理后，进行眼底照相、OCT、荧光素眼底血管造影(FFA)、视网膜血管密度以及血管渗漏检测。结果 HRMVPCs可分泌Exos,可以进入HUVECs中。与Control组相比，HG组和HG+H...     

间充质干细胞治疗早期糖尿病视网膜病变研究进展

糖尿病视网膜病变(DR)是糖尿病的主要并发症之一,其病变主要包括微血管病变和神经元功能的损害.治疗早期DR的研究从未间断,包括药物、生物制剂等,其中间充质干细胞(MSCs)作为一种具有广阔应用前景的组织工程细胞得到越来越多的关注.MSCs具有多向分化及免疫调节等能力,MSCs治疗DR可能有多种机制,通过MSCs体外培养和动物体内实验的研究发现,不同途径移植MSCs对DR进行早期干预后,其对DR的抗炎及神经保护等作用可改善DR视网膜功能,延缓DR的发展.就研究进展作一综述.     

Protective effect of human umbilical cord mesenchymal stem cell-derived exosomes on rat retinal neurons in hyperglycemia through the brain-derived neurotrophic factor/TrkB pathway

AIM: To explore whether human umbilical cord mesenchymal stem cell (hUCMSC)-derived exosomes (hUCMSC-Exos) protect rat retinal neurons in high-glucose (HG) conditions by activating the brain-derived neurotrophic factor (BDNF)-TrkB pathway. METHODS: hUCMSC-Exos were collected with differential ultracentrifugation methods and observed by transmission electron microscopy. Enzyme-linked immunosorbent assays (ELISAs) was used to quantify BDNF in hUCMSC-Exos, and Western blot was used to identify surface markers of hUCMSC-Exos. Rat retinal neurons were divided into 4 groups. Furthermore, cell viability, cell apoptosis, and TrkB protein expression were measured in retinal neurons. RESULTS: hUCMSCs and isolated hUCMSC-Exos were successfully cultured. All hUCMSC-Exos showed a diameter of 30 to 150 nm and had a phospholipid bimolecular membrane structure, as observed by transmission electron microscopy. ELISA showed the BDNF concentration of hUCMSCs-Exos was 2483.16±281.75. hUCMSCs-Exos effectively reduced the apoptosis of retinal neuron rate and improved neuron survival rate, meanwhile, the results of immunofluorescence verified the fluorescence intensity of TrKB in neurons increased. And all above effects were reduced by treated hUCMSCs-Exos with BDNF inhibitors. hUCMSC-Exos effectively reduced the apoptosis rate of retinal neurons by activating the BDNF-TrkB pathway in a HG environment. CONCLUSION: In the HG environment, hUCMSC-Exos could carry BDNF into rat retinal neurons, inhibiting neuronal apoptosis by activating the BDNF-TrkB pathway.     

炎症调控因子在糖尿病视网膜病变新生血管形成中的作用

糖尿病视网膜病变是常见的致盲性眼病,其发病机制复杂,至今仍尚未得到完全阐述。多项研究已表明,慢性炎症反应在糖尿病视网膜病变中占据重要位置。近年研究发现,许多“热点”炎症因子,如白细胞介素(IL)-6、IL-1β、肿瘤坏死因子-α(TNF-α)、单核细胞趋化蛋白-1(MCP-1)等通过复杂交织的炎症径路,参与疾病发生的核心环节。特别是在病理性新生血管发生时,上述炎症因子与血管生成因子相互促进,极大地增加了病情的严重程度。本文就炎症机制在糖尿病视网膜病变中的作用及其与新生血管的关系研究进展作一综述。     

Protective effects of umbilical cord mesenchymal stem cell exosomes in a diabetic rat model through live retinal imaging

AIM: To assess the protective effect of human umbilical cord mesenchymal stem cell exosomes (hucMSC-Exs) in a diabetic rat model by using a variety of retinal bioassays. METHODS: hucMSCs were subjected to differential ultracentrifugation for the collection of exosomes, and transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA) using a NanoSight analysis system and Western blotting (WB) were used to analyze the expression of surface marker proteins such as CD63, CD9 and Calnexin. Streptozotocin (STZ) was injected into the intraperitoneal cavity to establish a diabetic model. Rats were divided into a normal group, diabetic group and hucMSC-Ex group. Fundus fluorescein angiography (FFA), optical coherence tomography (OCT) and other live imaging methods were used to observe the fundus of the rats. Finally, the eyeballs of rats from each group were collected for hematoxylin-eosin (HE) staining to further analyze the retinal structure. RESULTS: Through TEM, NTA and WB, we successfully isolated hucMSC-Exs. Subsequent FFA and OCT confirmed that hucMSC-Exs effectively prevented early retinal vascular damage and thickening of the retina. Finally, HE staining of rat retinal sections revealed that exosomes effectively alleviated retinal structure disruption caused by diabetes. CONCLUSION: hucMSC-Exs have a protective effect on the retina in diabetic rat through FFA, OCT and HE staining.     

Transfection with CXCR4 potentiates homing of mesenchymal stem cells in vitro and therapy of diabetic retinopathy in vivo

AIM: To investigate the effect of the overexpression of C-X-C chemokine receptor type 4 (CXCR4) on homing of mesenchymal stem cells (MSCs) in vitro and therapeutic effects of diabetic retinopathy (DR) in vivo. METHODS: MSCs were infected by lentivirus constructed with CXCR4. The expression of CXCR4 was examined by immunofluorescence, Western blot, and quantitative polymerase chain reaction. CXCR4-overexpressing MSCs were cultured in vitro to evaluate their chemotaxis, migration, and apoptotic activities. CXCR4-overexpressing MSCs were intravitreally injected to observe and compare their effects in a mouse model of DR. The histological structure of DR in rats was inspected by hematoxylin and eosin staining. The expression of rhodopsin, neuron-specific enolase (NSE), and inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α was examined by Western blot and immunohistochemical analyses. RESULTS: The transduction of MSCs by lentivirus was effective, and the transduced MSCs had high expression levels of CXCR4 gene and protein. Improved migration activities were observed in CXCR4-overexpressing MSCs. Further, reduced retinal damage, upregulation of rhodopsin and NSE protein, and downregulation of inflammatory cytokines IL-6 and TNF-α were observed in CXCR4-overexpressing MSCs in vivo. CONCLUSION: The homing of MSCs can be enhanced by upregulating CXCR4 levels, possibly improving histological structures of DR. CXCR4-overexpressing MSCs can be a novel strategy for treating DR.     

自体干细胞移植术后糖尿病视网膜病变的变化分析

目的：观察自体干细胞移植术后糖尿病视网膜病变变化情况。

方法：自体干细胞移植术后糖尿病患者58例116眼经直接或间接检眼镜及眼底荧光血管造影(FFA)确定为无糖尿病视网膜病变者18眼,轻度非增殖期糖尿病视网膜病变(轻度NPDR)41眼,中度非增殖期糖尿病视网膜病变(中度NPDR)51眼,重度非增殖期糖尿病视网膜病变(重度NPDR)6眼,随访6～12mo,观察视网膜病变变化情况。

结果：患者视力、视网膜病变治疗的总有效率为84.4%、76.7%,其中重度非增殖期糖尿病视网膜病变组视力及视网膜病变治疗的有效率均显著低于轻、中度非增殖期糖尿病视网膜病变组及无视网膜病变组,差异有显著统计学意义(P<0.05)。

结论：自体干细胞移植术后稳定的血糖水平及胰岛功能的改善可能有助于稳定或延缓视网膜病变进展,其长期效果有待进一步深入研究。     

氧化应激与糖尿病视网膜病变

糖尿病视网膜病变（DR）是严重的糖尿病眼部并发症之一。研究表明，氧化应激和应激激活的信号通路与DR有着密切关系。高糖通过多种机制刺激活性氧类物质的产生，从而生长因子和细胞因子的基因表达。正常情况下，过多的活性氧物质可被抗氧化防御系统清除，但持续性高血糖情况下过多的活性氧物质可导致细胞损害，包括视网膜内皮细胞、周细胞和视网膜节细胞。现就氧化应激在DR发生发展中所起的作用做一综述。