促红细胞生成素干预早产儿脑损伤机制的研究进展

早产儿脑损伤可继发多种严重并发症,其严重的神经系统后遗症已成为新的临床、公共卫生和社会问题.目前研究发现促红细胞生成素(erythropoietin,EPO)能减轻早产儿脑损伤,对损伤的脑神经细胞有保护和修复作用,并可能提高远期神经预后.EPO发挥神经保护作用的主要机制是促进早产儿脑血管成熟、缓解血管痉挛,促进血管新生,在缺血部位建立侧枝循环改善脑缺血情况;对缺血缺氧(hypoxic ischemic,HI)后神经元凋亡和炎性坏死的双重阻断作用;抗氧化效果;减少谷氨酸毒性;营养神经及促进神经元的增值、树突增多、功能增强等.EPO对早产儿脑损伤后的多重保护作用为早产儿脑损伤的治疗提供了临床应用前景.     

重组人促红细胞生成素对脑缺血再灌注小鼠海马VEGF表达的影响

目的探讨重组人促红细胞生成素(recombinant human erythropoietin,rHuEPO)对脑缺血再灌注小鼠血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的影响。方法 96只昆明小鼠随机分为3组,每组32只:假手术组(Sham);rHuEPO治疗组;缺血再灌注组(I/R)。每组依据缺血后再灌注不同时间点再分6h、24h、3d、7d五个小组。免疫组化和Westernblot检测各组小鼠VEGF的表达,TUNEL法检测海马细胞凋亡情况。结果缺血再灌注组海马VEGF蛋白表达量和凋亡细胞数明显高于假手术组(<0.05),与缺血再灌注组相比,rHuEPO治疗组VEGF蛋白表达量明显升高(<0.05),凋亡细胞数则明显少于缺血再灌注组(相似文献   

重组人促红细胞生成素对人脐静脉内皮细胞PCNA表达的影响

目的研究重组人促红细胞生成素（rHuEPO）对人脐静脉内皮细胞（HUVEC）增殖的影响。方法从脐静脉体外分离培养HUVEC，采用细胞培养及免疫组织化学方法检测不同剂量rHuEP0（10U／ml、20U／ml、40U／m1）对HUVEC增殖细胞核抗原（PCNA）表达的影响。结果对组照及10U／ml、20U／ml、40U／mlrHuEP0实验组内皮细胞PCNA阳性表达百分率分别为（31±3）％、（40±5）％、（68±6）％和（65±5）％，与对照组相比，rHuEPO作用后内皮细胞PCNA表达明显增强（P〈0．05），且随rHuEPO剂量提高作用增强，呈剂量效应关系。结论rHuEP0可促进体外培养内皮细胞增殖。     

重组人促红细胞生成素对小鼠胚胎干细胞向心肌细胞分化的影响

目的初步观察外源性细胞因子重组人促红细胞生成素（NEPO）对小鼠胚胎干细胞（ESC）向心肌细胞分化的影响。方法将NEPO加到细胞培养基中配成终浓度分别为0．5、1．O、5．0U／ml的诱导分化液，分别在分化第8、12天观察ESC自发及在rhEPO诱导情况下心肌细胞分化率；在分化第7天应用RT-PCR检测自发分化组及NEPO诱导组心肌早期转录因子GATA-4、Nkx2．5mRNA的相对表达。结果在分化第8、12天rhEPO诱导（1．0U/ml）心肌细胞分化率分别显著高于对应分化天数的自发心肌细胞分化率（0．11±0．02对0．05±0．01，P〈0．05；0．86±0．02对0．65±0．05，P〈0．05）；与自发分化组比较．NEPO干预（1．0U／ml）上调GATA-4、Nkx2．5mRNA的相对表达。结论外源性rhEPO早期干预能够促进ESC向心肌细胞分化。     

重组人促红细胞生成素抑制内皮细胞凋亡的实验研究

目的：探讨重组人促红细胞生成素（Recombinant human eryfhropoietin,rHuEPO)对脂多糖（Lipopolysaccharide,LPS)诱导的人脐静脉内皮细胞（Human umbilical vein endotheilial cells,HUVECs)凋亡的影响。方法：实验分为四组;第一组仅用LPS处理;第二组预先用rHuEPO孵育后再用LPS处理;第三组只用rHuEPO处理;对照组用无血清的M199培养。用流式细胞仪DNA分析检测细胞凋亡。结果：与对照组相比,LPS处理后的细胞DNA亚二倍体含量显著增加（P＜0．05）,预先用rHuEPO处理后则可显著降低LPS的这种作用（P＜0．05）。结论：rHuEPO抑制了LPS诱导的内皮细胞凋亡,这种保护作用可能是rHuEPO诱导内皮细胞生长和血管生成的一个重要因素。     

水凝胶携带重组人促红细胞生成素对大鼠深Ⅱ度烫伤创面愈合的影响

目的:探讨水凝胶携带重组人促红细胞生成素(rHuEPO)对深Ⅱ度烫伤创面愈合的影响。方法:建立18只SD大鼠深Ⅱ度烫伤模型,随机分为三组,每组6只,分别给予PBS、水凝胶、水凝胶+rHuEPO治疗处理,分别于烫伤即刻、14天、21天测量创面面积,并计算创面愈合指数;治疗21天后取烫伤组织进行组织切片、HE染色和免疫组化检测,比较三组标本上皮化程度、肉芽组织生成和血管形成情况,并进行组织学评分;比较治疗21天后各组血管内皮生长因子(VEGF)的表达。结果:治疗21天,水凝胶+rHuEPO组组织学评分高于水凝胶组和PBS组,表现为较多的肉芽组织形成和血管生成(P<0.05),创面愈合指数大于PBS组和水凝胶组(P<0.05),VEGF表达量高于PBS组和水凝胶组(P<0.05)。结论:应用水凝胶携带rHuEPO治疗深Ⅱ度烫伤可以增加烫伤组织VEGF表达,促进血管生成、肉芽组织生长、内皮增生和迁移,从而促进烫伤愈合。     

重组人促红细胞生成素对人脐静脉内皮细胞CyclinD1表达的影响

目的探讨重组人促红细胞生成素(rHuEPO)对人脐静脉内皮细胞(HUVEC)细胞周期蛋白CyclinD1的影响。方法采用细胞培养及免疫组织化学方法检测不同剂量(10、20、40U/mL)rHuEPO作用下HUVEC中CyclinD1表达的变化。结果对组照及10、20、40U/mlrHuEPO实验组内皮细胞CyclinD1表达阳性百分率(%)分别为26±3、41±5、69±3、65±4,与对照组相比,rHuEPO作用后HUVEC中CyclinD1表达明显增强(P﹤0.05),且随rHuEPO剂量增加作用增强,呈剂量效应关系。结论rHuEPO能明显促进体外培养内皮细胞CyclinD1表达,提示rHuEPO具促内皮细胞增殖作用。     

重组人促红细胞生成素对大鼠皮肤深Ⅱ°烫伤创面愈合的影响

目的研究重组人促红细胞生成素(rHuEPO)对大鼠皮肤深Ⅱ°烫伤愈合的影响。方法健康SD大鼠随机分成低剂量组(50U/mL rHuEPO)、中剂量组(100U/mL rHuEPO)、高剂量组(200U/mL rHuEPO)和生理盐水(NS)对照组,建立深Ⅱ°烫伤动物模型,并于伤后3、5、7、14、21d取创面皮肤标本,HE染色和免疫组织化学方法染色分别检测创面愈合情况及微血管密度。结果与对照组相比,rHuEPO治疗组创面愈合时间缩短,同一时相内大鼠烫伤愈合率高于对照组,微血管密度明显增加。结论rHuEPO能促进深Ⅱ°烫伤创面微血管形成,加速创面愈合。     

神经干细胞移植显著改善脑外伤小鼠的神经功能

目的探讨鼠源性神经干细胞(neural stem cells,NSCs)静脉移植对脑外伤小鼠海马的治疗作用。方法30只BALB/c小鼠随机分为3组,假手术组(10只)、脑外伤组(10只)和NSCs移植组(10只)。NSCs移植7d后,根据小鼠脑损伤NSS(Neurological Severity Score)评分系统进行评分。TUNEL法检测各组小鼠海马细胞凋亡情况。免疫组化和Westernblot检测各组小鼠海马半胱氨酸蛋白酶-3(cysteinyl aspirate specificproteinase3,caspase-3)蛋白的表达。结果NSCs移植7d后,NSS评分显示NSCs移植组的神经功能明显改善(<0.001)。TUNEL法检测结果发现,脑外伤组小鼠海马凋亡细胞百分数明显高于假手术组(<0.05),但经过NSCs移植后,凋亡率显著下降。免疫组化和Westernblot结果显示,脑外伤组小鼠海马caspase-3蛋白表达量明显高于假手术组(<0.05),但NSCs移植组小鼠海马caspase-3表达明显降低(<0.05)。结论静脉NSCs移植能PPPP够显著改善脑外伤小鼠的神经功能,其机制可能与降低caspase-3的表达有关。     

人脂肪间充质干细胞来源的外排体促进大鼠创伤性脑损伤后神经功能恢复

目的研究人脂肪间充质干细胞(h AMSCs)来源的外排体对创伤性脑损伤(TBI)的治疗作用及其可能的机制。方法分离健康成人脂肪MSCs,通过超滤法提取外排体。将大鼠分成:假手术组,PBS对照组,MSC治疗组,exosomes治疗组。于TBI建模24 h后,治疗组分别沿损伤边缘区局部注射,PBS 30μL,MSC 2×10~5个细胞/只,exosomes 25μg总蛋白量/只,总体积30μL。在建模前和TBI后1、3、7、10、13、16和30 d测试所有大鼠的m NSS评分和前肢踩空试验。3和7d处死大鼠,提取大鼠脑组织总RNA,实时定量PCR检测大鼠炎性因子TNF-α和IL-1β的表达,30 d处死大鼠,tunel-neun双标免疫荧光检测TBI后神经元凋亡。结果外排体的治疗显著促进TBI后的神经功能的恢复,治疗效果与MSC治疗效果相当,其机制可能是通过抑制大鼠TBI后急性炎性反应,减少神经元凋亡。结论人脂肪间充质干细胞来源的外排体促进脑外伤后神经功能的恢复,这将为临床提供一种新的更安全的TBI治疗手段。     

脑外伤小鼠海马血管内皮生长因子表达的变化

目的探讨脑外伤小鼠海马血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的变化。方法 40只Balb/c小鼠随机分为2组,假手术组(10只)和脑外伤组(30只)。脑外伤组依据脑外伤的不同时间点再分6h、1d、3d三个小组,每组10只。免疫组化和Western blot检测各组小鼠海马VEGF蛋白的表达。RT-PCR方法检测各组小鼠海马VEGF mRNA的表达变化。结果免疫组化和Western blot结果发现,脑外伤组小鼠海马VEGF蛋白表达量明显高于假手术组(<0.05);RT-PCR结果也发现,脑外伤组小鼠海马VEGF mRNA表达量明显高于假手术组(<0.05)。结论脑外伤小鼠海马VEGF表达明显升高。     

大鼠创伤性脑损伤后星形胶质细胞的变化

目的：探讨大鼠创伤性脑损伤后星形胶质细胞的形态学变化及GFAP和NOS的表达情况。方法：采用大鼠自由落体脑损伤模型，伤后1、3、7d取脑切片，行Nissl染色以及GFAP免疫组化和NADPH—d组化单标记及双标记染色。结果：损伤区周围皮质GFAP阳性细胞胞体增大、突起增粗增长，GFAP阳性细胞数量与正常侧及对照组相比，伤后1d即有明显增加，伤后3d、7d数量持续增加；损伤侧海马CAI～3区和DG各层GFAP阳性细胞排列紊乱，胞体增大、突起增粗增长，GFAP阳性细胞数量与正常侧及对照组相比则无明显变化。损伤区周围皮质、损伤侧海马NOS阳性细胞数量明显增加。伤后3d损伤区周围皮质和损伤侧海马中GFAP与NOS双标细胞分别占GFAP阳性细胞的14．2％和13．4％左右。结论：大鼠创伤性脑损伤后大量的星形胶质细胞活化、GFAP表达增加并且部分转化为NOS阳性细胞，提示其参与了脑组织的损伤与修复过程。     

MRI evaluation of axonal reorganization after bone marrow stromal cell treatment of traumatic brain injury

We treated traumatic brain injury (TBI) with human bone marrow stromal cells (hMSCs) and evaluated the effect of treatment on white matter reorganization using MRI. We subjected male Wistar rats (n = 17) to controlled cortical impact and either withheld treatment (controls; n = 9) or inserted collagen scaffolds containing hMSCs (n = 8). Six weeks later, the rats were sacrificed and MRI revealed selective migration of grafted neural progenitor cells towards the white matter reorganized boundary of the TBI‐induced lesion. Histology confirmed that the white matter had been reorganized, associated with increased fractional anisotropy (FA; p < 0.01) in the recovery regions relative to the injured core region in both treated and control groups. Treatment with hMSCs increased FA in the recovery regions, lowered T₂ in the core region, decreased lesion volume and improved functional recovery relative to untreated controls. Immunoreactive staining showed axonal projections emanating from neurons and extruding from the corpus callosum into the ipsilateral cortex at the boundary of the lesion. Fiber tracking (FT) maps derived from diffusion tensor imaging confirmed the immunohistological data and provided information on axonal rewiring. The apparent kurtosis coefficient (AKC) detected additional axonal remodeling regions with crossing axons, confirmed by immunohistological staining, compared with FA. Our data demonstrate that AKC, FA, FT and T₂ can be used to evaluate treatment‐induced white matter recovery, which may facilitate restorative therapy in patients with TBI. Copyright © 2011 John Wiley & Sons, Ltd.     

Neurovascular hypoxia after mild traumatic brain injury in juvenile mice correlates with heart–brain dysfunctions in adulthood

Aim

Retrospective studies suggest that mild traumatic brain injury (mTBI) in pediatric patients may lead to an increased risk of cardiac events. However, the exact functional and temporal dynamics and the associations between heart and brain pathophysiological trajectories are not understood.

Methods

A single impact to the left somatosensory cortical area of the intact skull was performed on juvenile mice (17 days postnatal). Cerebral 3D photoacoustic imaging was used to measure the oxygen saturation (sO₂) in the impacted area 4 h after mTBI followed by 2D and 4D echocardiography at days 7, 30, 90, and 190 post-impact. At 8 months, we performed a dobutamine stress test to evaluate cardiac function. Lastly, behavioral analyses were conducted 1 year after initial injury.

Results

We report a rapid and transient decrease in cerebrovascular sO₂ and increased hemoglobin in the impacted left brain cortex. Cardiac analyses showed long-term diastolic dysfunction and a diminished systolic strain response under stress in the mTBI group. At the molecular level, cardiac T-p38MAPK and troponin I expression was pathologic modified post-mTBI. We found linear correlations between brain sO₂ measured immediately post-mTBI and long-term cardiac strain after 8 months. We report that initial cerebrovascular hypoxia and chronic cardiac dysfunction correlated with long-term behavioral changes hinting at anxiety-like and memory maladaptation.

Conclusion

Experimental juvenile mTBI induces time-dependent cardiac dysfunction that corresponds to the initial neurovascular sO₂ dip and is associated with long-term behavioral modifications. These imaging biomarkers of the heart–brain axis could be applied to improve clinical pediatric mTBI management.     

颅脑外伤后并发低钠血症的危险因素分析

目的探讨颅脑外伤患者并发低钠血症的发生机制及危险因素,以期为其早期预测及预防提供参考。方法回顾性分析2016年6月至2019年6月我院收治的185例中型和重型颅脑外伤患者的临床资料,包括导致低钠血症的不同病因、损伤类型、性别、格拉斯哥昏迷(GCS)评分、手术、脑水肿、颅底骨折和穿透性性损伤等;采用单因素χ^2检验和多因素Logistic回归分析探究颅脑外伤后并发低钠血症的危险因素。结果所有患者中,80例出现低钠血症,其中钠盐摄入不足、利尿剂过量使用47例,抗利尿激素分泌失调综合征19例,脑性耗盐综合征14例。低钠血症更多发生在脑挫裂伤、蛛网膜下腔出血和弥漫性轴索损伤患者中,差异具有统计学意义(P<0.05)。单因素χ^2检验结果显示,GCS评分(P=0.000)、脑水肿(P=0.000)、颅底骨折(P=0.000)、穿透性损伤(P=0.001)是颅脑外伤后并发低钠血症的相关因素。多因素Logistic回归分析结果显示,GCS评分(P=0.006)、脑水肿(P=0.006)、颅底骨折(P=0.000)、穿透性损伤(P=0.015)是颅脑外伤后并发低钠血症的危险因素。结论脑挫裂伤、蛛网膜下腔出血、弥漫性轴索损伤、GCS评分≤8分、脑水肿、颅底骨折和穿透性损伤的颅脑外伤患者更易发生低钠血症,应早期关注患者血清钠水平,明确病因及时纠正,防止病情恶化。     

Regional difference of reactive astrogliosis following traumatic brain injury revealed by hGFAP-GFP transgenic mice

Reactive astrocytes greatly influence the wound healing and neuronal regeneration processes following brain injury. However, the origin and fate of reactive astrocytes appear to be different depending on the type, severity and duration of brain injury. Using the cryogenic traumatic brain injury model, here we comprehensively addressed the regional differences of reactive astrocytes in the injured cortex. In the proximal region of injury site, NG2-expressing and cytoplasmic Olig2-labeled cells were densely localized 3 days after the injury. Next to this proximal layer, most of reactive astrocytes did not express NG2 but exhibited radial glia-like shape with elongated processes. Accordingly, they expressed the progenitor or radial glial markers, such as vimentin, brain lipid binding protein (BLBP) and the green fluorescent protein (GFP) under the control of the human GFAP (hGFAP) promoter. However, only few glial fibrillary acidic protein (GFAP) expressing astrocytes were found in this layer. Distal to the injury site, most of astrocytes strongly expressed GFAP with hypertonic morphology. At day 15 after injury, all layers expressing GFAP and other marker expressions disappeared, indicating the termination of reactive astrogliosis. Taken together, our data suggest that reactive astrogliosis occurs in a regionally segregated manner in the early phase of brain injury.     

急诊颅脑手术合并心脏支架术后动态监测血小板功能1例报告

重型颅脑创伤患者病情危重,病情进展极快,后期并发症多,治疗颇为复杂,病死率高。对于颅脑损伤合并冠心病支架术后长期服用双抗药物的患者,围手术期密切监测血小板聚集试验以平衡预防出血与心脏事件之间的矛盾极为关键。现报告动态监测血小板功能应用于急诊颅脑手术合并心脏支架术后患者1例。     

大鼠创伤性脑损伤后细胞凋亡及NOS阳性细胞的变化

目的：探讨大鼠创伤性脑损伤后不同时相皮质、海马、隔区细胞凋亡及NOS、ChAT阳性细胞的变化。方法：采用大鼠自由落体脑损伤模型，伤后1、2、3、4、5、7、10d取脑切片，经Nissl染色，用TUNEL法检测细胞凋亡，NADPH—d组化染色观察NOS阳性细胞，ChAT免疫组化染色观察隔区ChAT阳性细胞。结果：Nissl染色可见损伤侧海马CA2、CA3区锥体细胞层细胞消失或紊乱。损伤区周围皮质凋亡细胞伤后3d达到高峰；损伤侧海马凋亡细胞伤后5d达到高峰；损伤侧隔区凋亡细胞7d达到高峰。正常侧上述脑区各时相点均未见到凋亡细胞。损伤区周围皮质、损伤侧海马和隔区iNOS阳性细胞数量明显增加。损伤侧隔区ChAT阳性神经元也明显减少。结论：大鼠创伤性脑损伤后损伤区周围皮质和损伤侧海马、隔区细胞凋亡数量的变化与伤后时程有关。伤后细胞iNOS表达增加是导致细胞凋亡的因素。