干细胞因子和粒细胞集落刺激因子动员自身骨髓干细胞治疗缺血再灌注肾损伤

背景：骨髓干细胞可分化为肾脏组织固有细胞、修复损伤肾组织。正常情况下,外周血干细胞数目较少,骨髓干细胞动员剂可提高外周血干细胞数目。 目的：观察动员自身骨髓干细胞对缺血再灌注损伤肾脏修复作用及对缺氧诱导因子1α系统的影响,分析骨髓干细胞修复损伤肾脏的机制。 方法：SD大鼠随机分为4组。对照组不作处置;模型组制备肾缺血再灌注模型;治疗组给予缺血再灌注模型大鼠皮下注射骨髓干细胞动员剂干细胞因子200 μg/(kg•d)及粒细胞集落刺激因子50 μg/(kg•d),治疗对照组给予正常大鼠皮下注射与治疗组相同的药物,连续5 d。于术后5,10,17,24,31 d观察大鼠肾脏病理改变、骨髓干细胞表面抗原标记CD34⁺细胞表达以及缺氧诱导因子1α、血管内皮生长因子、血红素加氧酶1的表达变化。 结果与结论：①联合应用骨髓干细胞动员剂能明显增加损伤肾组织骨髓干细胞的数量,减轻肾组织损伤程度。②骨髓干细胞能促进肾组织缺氧诱导因子1α系统的表达,缺氧诱导因子1α系统及其靶基因产物血管内皮生长因子、血红素加氧酶1表达增加是骨髓干细胞促进急性肾损伤修复的可能机制之一。③骨髓干细胞动员剂对缺氧诱导因子1α系统的表达有一定的增强作用。     

丙二醛、ET-1、VEGF在动脉粥样硬化形成过程中的动态变化

目的：探讨家兔动脉粥样硬化形成过程中丙二醛、内皮素-1(Endothelin-1，ET-1)、血管内皮生长因子(VEGF)的动态变化。方法：雄性新西兰家兔40只，随机分为正常对照组及动脉粥样硬化模型组，分别于实验的第4、8、12周末，采集兔耳中央动脉空腹血，测定血清中丙二醛含量；同时随机处死每组兔各5-8只，取主动脉弓段及胸段，采用宏观和微观相结合的方法，定性及定量观察病变进展情况。同时用免疫组化法观察ET-1、VEGF的表达特征。结果：(1)动脉粥样硬化组动脉斑块随时间延长而逐渐加大。(2)动脉粥样硬化组血清丙二醛含量显著高于对照组水平；且随时间延长动脉粥样硬化组丙二醛水平逐渐上升。(3)斑块内ET-1、VEGF阳性表达的程度及范围逐渐增加。结论：脂质过氧化(lipid peroxide,LPO)、ET-1、VEGF在动脉粥样硬化病变过程中持续发挥一定作用，共同促进动脉粥样硬化的发生发展。     

激光心肌血运重建术结合转VEGF基因对缺血心肌侧枝循环的影响

为明确激光心肌血运重建术 (Transmyocardiallaserrevascularization ,TMLR)与转血管内皮生长因子 (Vascularen dothelialgrowthfactor,VEGF)基因相结合能否促进侧枝循环建立 ,我们在猪的急性心肌缺血模型上 ,用冠脉造影观察了TMLR、TMLR +VEGF对缺血心肌侧枝循环的影响。结果显示 ,与术后即刻或单纯缺血组相比 ,TMLR(Rentrop记分 1 4± 0 9)、TMLR +VEGF(Rentrop记分 2± 0 5 )均能显著促进侧枝循环建立 (P均 <0 0 5 ) ;TMLR +VEGF组较TMLR组侧枝有增多的趋势。分时间点统计显示 ,TMLR +VEGF组 4周为 2± 0 8,8周为 2± 0 0 ,TMLR组 4周为 2± 0 0 ,8周为 1± 1 0。其中TMLR +VEGF组 4周、8周和TMLR组 4周结果无差异 ,而TMLR组 8周与其他各组相比均呈下降的趋势。提示与TMLR相比 ,TMLR结合转VEGF基因在一定程度上有助于侧枝循环的稳定     

Vasoactive intestinal peptide in rats with focal cerebral ischemia enhances angiogenesis

We studied the effect of vasoactive intestinal peptide (VIP) on angiogenesis in the ischemic boundary area after focal cerebral ischemia. Adult male Sprague–Dawley rats underwent middle cerebral artery occlusion for 2 h. A single dose of VIP was given via i.c.v. injection at the beginning of reperfusion. Immunohistochemistry and Western blotting were performed to assay angiogenesis and brain levels of vascular endothelial growth factor (VEGF) protein, respectively. In addition, the expression of VEGF and its receptors (flt-1 and flk-1), as well as endothelial proliferation, was measured using rat brain microvascular endothelial cells. Immunohistochemical analyses revealed significant (P<0.05) increases in the numbers of bromodeoxyuridine (BrdU) positive endothelial cells and microvessels at the boundary of the ischemic lesion in rats treated with VIP compared with rats treated with saline. Western blotting analysis showed that treatment with VIP significantly (P<0.05) raised VEGF levels in the ischemic hemisphere. In addition, treatment with VIP increased flt-1 and flk-1 immunoreactivity in endothelial cells. In vitro, incubation with VIP significantly (P<0.01) increased the proliferation of endothelial cells and induced the expression of VEGF, flt-1 and flk-1 in endothelial cells. The stimulatory effect of VIP on the proliferation of endothelial cells was significantly (P<0.01) inhibited by SU5416, a selective inhibitor of VEGF receptor tyrosine kinase. Our data suggest that treatment with VIP enhances angiogenesis in the ischemic brain, and this effect may be mediated by increases in levels of VEGF and its receptors.     

Overexpression of VEGF-A induces neovascularization and increased vascular leakage in rabbit eye after intravitreal adenoviral gene transfer

Aims: The aim of this study was to determine dose–response effects of vascular endothelial growth factor A as delivered using an adenoviral vector on vascular growth and pathological changes in the rabbit eye. Moreover, we wanted to develop a large animal model for angioproliferative diseases in the eye. Methods: Seventeen New Zealand White rabbits were injected with adenoviral vascular endothelial growth factor‐A (AdVEGF‐A) intravitreally with different doses (10⁹–10¹¹ vp). Controls were injected with an empty virus (AdCMV). Some animals had a combination of AdVEGF‐A and AdsKDR (a soluble form of the VEGF receptor‐2). Animals were killed 6 days after the gene transfer. On the basis of these results, 14 rabbits were injected intravitreally with AdVEGF‐A or adenoviral LacZ (AdLacZ) with 10¹⁰ vp in a volume of 0.1 mL. Animals were killed 3, 6, 14 and 28 days after the gene transfer, eyes were removed and analysed histologically. Results: In enzyme‐linked immunosorbent assay (ELISA) analysis, human VEGF‐A was present in vitreous humour in all VEGF‐A transduced eyes. The amount of VEGF‐A showed a dose‐dependent increase with the AdVEGF‐A dose and was the highest 6 days after the gene transfer. Histologic analyses revealed an increased capillary area and density in the AdVEGF‐A eyes when compared with the AdLacZ eyes (P < 0.05). In the AdVEGF‐A/AdsKDR eyes the average capillary area was not increased compared with AdLacZ eyes. Conclusion: This model could be useful for large animal studies regarding the pathogenesis of neoangiogenesis and for the development of new therapeutic strategies for angioproliferative diseases of the eye. Our results establish the key role of VEGF‐A in the induction of neovascularization and pathological changes in the rabbit eye.     

Inhibitor of vascular endothelial growth factor receptor tyrosine kinase attenuates cellular proliferation and differentiation to mature neurons in the hippocampal dentate gyrus after transient forebrain ischemia in the adult rat

Neurogenesis in the adult hippocampal dentate gyrus is promoted by transient forebrain ischemia. The mechanism responsible for this ischemia-induced neurogenesis, however, remains to be determined. It has been suggested that there may be a close relationship between neurogenesis and the expression of vascular endothelial growth factor, an angiogenic factor. The purpose of the present study was to examine the relationship between vascular endothelial growth factor and cell proliferation in the dentate gyrus after transient forebrain ischemia. The mRNA expression of vascular endothelial growth factor was increased in the dentate gyrus on day 1 after ischemia. Immunohistochemical analysis on day 9 after ischemia, when a significant increase in cell proliferation was seen, showed that the cerebral vessel space in the subgranular zone of the dentate gyrus had not been affected by the ischemia. Neither were the vascular densities on days 1 and 3 after ischemia altered compared with those of non-operated naïve control rats. Furthermore, the distance from the center of the proliferative cells to the nearest cerebral vessel of ischemic rats was comparable to that of the sham-operated rats. We demonstrated that transient forebrain ischemia-induced cell proliferation and differentiation to mature neurons in the hippocampal dentate gyrus was attenuated by the i.c.v. administration of a vascular endothelial growth factor receptor tyrosine kinase inhibitor. These results suggest that vascular endothelial growth factor receptor at the early period of reperfusion may contribute to neurogenesis rather than to angiogenesis in the hippocampal dentate gyrus.     

Recombinant human endostatin reduces hypertrophic scar formation in rabbit ear model through down-regulation of VEGF and TIMP-1

BackgroundRecombinant human endostatin (Endostar) has been widely used to suppress angiogenesis in carcinoma patients. Hypertrophic scar (HS) tissue, much like a carcinoma, is often associated with angiogenesis. However, there have been few studies conducted on the effects of Endostar on HS or its mechanism.ObjectiveThis paper investigated the effects Endostar on the HS of rabbit ears and studied the effects of Endostar on VEGF and TIMP-1 expression.MethodsSixteen New Zealand white rabbits were used to establish HS models. Then, rabbit ears containing HS were randomly assigned to either the Endostar group or the control group. The changes of appearance and histology were evaluated using the naked eye, hematoxylin eosin staining, and a scar elevation index. The VEGF and TIMP-1 expressions were detected by immunohistochemical staining, RT-PCR, and western blot.ResultsThe thickness of the connective tissue in the Endostar group were thinner, the numbers of micro vessels and fibroblasts were fewer, and the collagen fibers were smoother. Moreover, the mRNA and protein expressions of VEGF and TIMP-1 in the Endostar group were significantly lower than those in the control group.ConclusionThe results suggested that Endostar reduced the formation of HS by down-regulation of VEGF and TIMP-1 expressions.     

富集自体骨髓干细胞移植与髓芯减压治疗缺血性股骨头坏死患者血清成纤维细胞生长因子2、缺氧诱导因子1α、血管内皮生长因子的变化

背景：缺血性股骨头坏死以股骨头血液供应不足为主要病因。髓芯减压术属于临床常用治疗手段,有研究发现采用自体骨髓干细胞移植能较好地改善髋关节功能,二者联合应用可促使患者尽早恢复。 目的：观察自体骨髓干细胞移植联合髓芯减压治疗早期缺血性股骨头坏死的疗效以及血清成纤维细胞生长因子2、缺氧诱导因子1α、血管内皮生长因子水平。 方法：选取2016年1月至2018年1月收治的62例早期缺血性股骨头坏死患者进行研究,参照随机数字表法分为研究组(n=31)和对照组(n=31),对照组给予细针多孔道髓芯减压术治疗,研究组在对照组基础上联合自体骨髓干细胞移植治疗,观察两组患者治疗前后Harris评分、目测类比评分、血清成纤维细胞生长因子2、缺氧诱导因子1α、血管内皮生长因子水平及不良反应发生情况。 结果与结论：①两组患者治疗后3,6,12个月Harris评分均高于治疗前(P < 0.05),研究组高于对照组(P < 0.05);②两组患者治疗后3,6,12个月目测类比评分均低于治疗前(P < 0.05),研究组治疗后3,6个月目测类比评分低于对照组(P < 0.05);③两组患者治疗后3,6,12个月血清成纤维细胞生长因子2水平高于治疗前(P < 0.05),研究组高于对照组(P < 0.05);缺氧诱导因子1α、血管内皮生长因子水平明显低于治疗前(P < 0.05),研究组低于对照组(P < 0.05);④两组患者治疗后不良反应发生率比较差异无显著性意义(P > 0.05);⑤结果表明自体骨髓干细胞移植联合髓芯减压治疗早期缺血性股骨头坏死,可有效改善髋关节功能,促进股骨头坏死区修复,减轻患者疼痛,提高成纤维细胞生长因子2水平,降低缺氧诱导因子1α、血管内皮生长因子水平。 ORCID: 0000-0001-9884-7530(张景义) 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     

转导骨桥蛋白短发夹状RNA预防动脉粥样硬化模型兔血管成形后的再狭窄

背景：血管成形后再狭窄严重限制了经皮冠状动脉介入治疗的应用和远期疗效。平滑肌细胞的表型转变,增殖是血管成形后再狭窄的重要机制。 目的：探讨利用球囊在体转导骨桥蛋白短发夹状RNA,通过抑制实验性动脉粥样硬化模型兔损伤血管部位骨桥蛋白的表达,预防血管成形后再狭窄。 方法：构建动脉粥样硬化模型兔20只,随机等分成空质粒组和OPN-shRNA质粒组,分别利用球囊在腹主动脉导入OPN-shRNA质粒和空载体。 结果与结论：2组兔球囊扩张后血管平滑肌层出现特异性绿色荧光,且随转染后时间的延长荧光强度逐渐降低,与空质粒组相比,OPN-shRNA质粒组兔扩张动脉的管腔面积明显增加,而斑块负荷明显减小。提示在动脉粥样硬化兔模型局部血管利用球囊导管能成功地转导OPN-shRNA质粒,被扩张血管的再狭窄程度减轻,血栓负荷减轻。这对于预防模型兔血管成形后再狭窄的发生具有十分重要的意义。 中国组织工程研究杂志出版内容重点：肾移植;肝移植;移植;心脏移植;组织移植;皮肤移植;皮瓣移植;血管移植;器官移植;组织工程全文链接：     

Hypoxia-induced therapeutic neovascularization in a mouse model of an ischemic limb using cell aggregates composed of HUVECs and cbMSCs

Cell transplantation for therapeutic neovascularization holds great promise for treating ischemic diseases. This work prepared three-dimensional aggregates of human umbilical vein endothelial cells (HUVECs) and cord-blood mesenchymal stem cells (cbMSCs) with different levels of internal hypoxia by a methylcellulose hydrogel system. We found that few apoptosis occurred in these cell aggregates, despite developing a hypoxic microenvironment in their inner cores. Via effectively switching on the hypoxia-inducible factor-1α-dependent angiogenic mechanisms, culturing the internally hypoxic HUVEC/cbMSC aggregates on Matrigel resulted in formation of extensive and persistent tubular networks and significant upregulation of pro-angiogenic genes. As the level of internal hypoxia created in cell aggregates increased, the robustness of the tubular structures developed on Matrigel increased, and expression levels of the pro-angiogenic genes also elevated. Transplantation of hypoxic HUVEC/cbMSC aggregates into a mouse model of an ischemic limb significantly promoted formation of functional vessels, improved regional blood perfusion, and attenuated muscle atrophy and bone losses, thereby rescuing tissue degeneration. Notably, their therapeutic efficacy was clearly dependent upon the level of internal hypoxia established in cell aggregates. These analytical results demonstrate that by establishing a hypoxic environment in HUVEC/cbMSC aggregates, their potential for therapeutic neovascularization can be markedly enhanced.