【摘要】：研究背景 新生兒腦損傷目前尚無(wú)及時(shí)有效治療手段，MSCs是目前最有希望的治療新生兒腦損傷的細(xì)胞治療手段。但有關(guān)MSCs治療腦損傷仍處于研究階段。 目的 體外分離、鑒定人臍帶間充質(zhì)干細(xì)胞（hWJ-MSCs），觀察hWJ-MSCs在體外的擴(kuò)增效率、不同擴(kuò)增代數(shù)之間的生長(zhǎng)差異，觀察hWJ-MSCs在體外模擬炎性環(huán)境下的生物學(xué)表型改變，并將hWJ-MSCs與中樞神經(jīng)系統(tǒng)主要的免疫細(xì)胞-神經(jīng)小膠質(zhì)細(xì)胞在炎癥模型環(huán)境下進(jìn)行共培養(yǎng)，觀察兩種細(xì)胞之間的相互調(diào)控作用，為進(jìn)一步研究MSCs的顱內(nèi)移植治療提供研究基礎(chǔ)。 方法 為以上目的，本研究分為以下3部分。 1. hWJ-MSCs的分離、體外擴(kuò)增及鑒定 收集北京軍區(qū)總醫(yī)院正常足月剖宮產(chǎn)新生兒臍帶組織5根。所有臍帶組織在離體后立即放置入無(wú)菌環(huán)境保存，并在1-6h內(nèi)處理完畢以保證分離細(xì)胞活力。臍帶組織在無(wú)菌環(huán)境下剔除臍動(dòng)、靜脈，物理方法剪碎，隨后用胰酶+膠原酶-Ⅱ聯(lián)合消化法及貼壁法分離純化并體外擴(kuò)增hWJ-MSCs。采用流式細(xì)胞術(shù)對(duì)hWJ-MSCs細(xì)胞表面標(biāo)記CD14、CD31、CD73、CD105、CD90、HLA-ABC、HLA-DR鑒定。并采用體外成軟骨、成脂肪分化試劑盒觀察細(xì)胞分化潛力。將兩種分離方法所得細(xì)胞分別傳代，觀察細(xì)胞形態(tài)及細(xì)胞周期變化。 2.觀察toll-like receptor4（TLRs）在hWJ-MSCs上的表達(dá)及其信號(hào)通路對(duì)hWJ-MSCs生物學(xué)功能的影響 采用流式細(xì)胞術(shù)及qRT-PCR方法檢測(cè)TLR4在hWJ-MSCs上的表達(dá)情況。采用TLR4激活劑-LPS體外刺激hWJ-MSCs，在LPS刺激不同時(shí)間點(diǎn)觀察hWJ-MSCs細(xì)胞形態(tài)變化、細(xì)胞增殖及生長(zhǎng)周期變化。收集不同LPS刺激時(shí)間（24h、48h、72h）后的hWJ-MSCs細(xì)胞，采用trizol試劑提取總RNA，采用逆轉(zhuǎn)錄試劑盒及SYRB Green試劑盒檢測(cè)hWJ-MSCs的細(xì)胞因子（IL-1β、IL-1α、IL-2、IL-4、IL-6、IL-8、IL-10、IL-12、IL-13、indoleamine2,3-dioxygenase [IDO]-1、TNF-α、IFN-γ、MMP-2和MMP-9）在LPS刺激條件下的表達(dá)改變情況。收集細(xì)胞同時(shí)收集細(xì)胞培養(yǎng)上清，觀察細(xì)胞因子蛋白表達(dá)水平差異，進(jìn)一步確認(rèn)qRT-PCR結(jié)果。 3. hWJ-MSCs與小鼠源小膠質(zhì)神經(jīng)細(xì)胞系BV-2體外共培養(yǎng)研究 將體外分離所得的hWJ-MSCs與BV-2細(xì)胞在體外按不同比例進(jìn)行共培養(yǎng)，觀察兩種細(xì)胞的共培養(yǎng)體系在LPS+/LPS-下不同時(shí)間點(diǎn)兩種細(xì)胞各自的生物學(xué)變化。采用qRT-PCR檢測(cè)共培養(yǎng)條件下hWJ-MSCs表達(dá)IL-1β、IL-6、 IL-8及MMP-2差異；同時(shí)檢測(cè)BV-2細(xì)胞M2細(xì)胞表型基因Arginase1、Arginase2、Mrc1和Ym1的表達(dá)改變，觀察BV-2選擇性激活（M2型）標(biāo)記是否受到hWJ-MSCs的影響。使用NO檢測(cè)試劑盒檢測(cè)共培養(yǎng)體系中BV-2細(xì)胞NO分泌差異，觀察直接接觸共培養(yǎng)時(shí)hWJ-MSCs對(duì)LPS誘導(dǎo)BV-2細(xì)胞向M2型轉(zhuǎn)化的影響。 結(jié)果 1.成功分離并鑒定hWJ-MSCs 臍帶組織在無(wú)菌條件下經(jīng)剪成碎片貼壁培養(yǎng)皿生長(zhǎng)或進(jìn)一步消化獲得單細(xì)胞懸液接種培養(yǎng)瓶，均可見長(zhǎng)梭形類似成纖維細(xì)胞樣細(xì)胞生長(zhǎng)，細(xì)胞經(jīng)擴(kuò)增純化，體外最多可成功擴(kuò)增至20代。在20代之前細(xì)胞細(xì)胞可保持原代生長(zhǎng)形態(tài)。流式細(xì)胞術(shù)鑒定細(xì)胞表面標(biāo)記結(jié)果顯示所得細(xì)胞CD14、CD31、HLA-DR陰性， CD73、CD105、CD90、HLA-ABC陽(yáng)性，體外分化實(shí)驗(yàn)提示所得細(xì)胞體外誘導(dǎo)成軟骨、成脂肪分化成功。依據(jù)貼壁生長(zhǎng)形態(tài)、細(xì)胞表面標(biāo)記及分化潛力，符合國(guó)際細(xì)胞療法協(xié)會(huì)有關(guān)間充質(zhì)干細(xì)胞的定義。提示采用貼壁法、胰酶-膠原酶聯(lián)合消化法均成功獲得臍帶間充質(zhì)干細(xì)胞。所得細(xì)胞在3-7代間經(jīng)PI染色法觀察細(xì)胞周期，細(xì)胞均處于G0/G1期，極少數(shù)細(xì)胞處于增殖分裂期，具有干細(xì)胞特性。未見死亡、晚期凋亡細(xì)胞。 2.鑒定TLR4為hWJ-MSCs表面的功能性受體 流式細(xì)胞術(shù)和qRT-PCR分析揭示TLR4在hWJ-MSCs上的表達(dá)。LPS體外刺激hWJ-MSCs成功引發(fā)細(xì)胞應(yīng)激性反應(yīng)，檢測(cè)TLR4信號(hào)通路相關(guān)炎性因子（IL-1β、IL-1α、IL-6、IL-8）基因表達(dá)差異，提示所檢測(cè)炎性因子基因均在刺激72h后表達(dá)明顯上調(diào)，但I(xiàn)L-12在LPS刺激條件后均表現(xiàn)為明顯的下調(diào)，未檢測(cè)到IL-2、IL-4、IL-10、IL-13、TNF-α、IFN-γ的表達(dá)。對(duì)MMP-2和MMP-9基因表達(dá)情況的檢測(cè)提示MMP-2在LPS刺激后72h出現(xiàn)明顯上調(diào)，但MMP-9表達(dá)在LPS刺激后出現(xiàn)短暫（48h）的下調(diào)，，但在72h后，MMP-9表達(dá)上升至與陰性對(duì)照大致持平的水平。此外，MSC免疫抑制分子IDO1、IDO-2、IFN-β和Cox2基因表達(dá)情況的檢測(cè)結(jié)果顯示：IDO1和IFN-β在LPS刺激條件下出現(xiàn)明顯上調(diào)，IDO1表達(dá)上調(diào)在72h內(nèi)上調(diào)呈現(xiàn)為雙峰，在48h檢測(cè)表達(dá)為低谷。Cox2的表達(dá)同樣受到LPS的誘導(dǎo)，但僅在LPS刺激72h后出現(xiàn)明顯的上調(diào)；此外，在我們觀察的3個(gè)時(shí)間點(diǎn)未發(fā)現(xiàn)IDO-2表達(dá)變化。該結(jié)果顯示LPS可修飾hWJ-MSCs的免疫抑制功能。 3.hWJ-MSCs與BV-2體外共培養(yǎng)時(shí)的相互調(diào)節(jié)作用 在體外共培養(yǎng)條件下，hWJ-MSCs和BV-2兩種細(xì)胞均為貼壁生長(zhǎng)。在LPS刺激條件下，hWJ-MSCs可顯著減弱LPS對(duì)BV-2的刺激作用；與hWJ-MSCs共培養(yǎng)時(shí)：如無(wú)LPS刺激，BV-2僅分泌極低量的NO；但LPS刺激條件下，hWJ-MSCs對(duì)BV-2分泌NO有明顯的抑制作用，且與細(xì)胞共培養(yǎng)比例有關(guān)；qRT-PCR分析發(fā)現(xiàn)：當(dāng)兩種細(xì)胞比例維持在1:1時(shí)，hWJ-MSCs可明顯上調(diào)BV-2細(xì)胞的M2細(xì)胞表型標(biāo)記Arginase1表達(dá)；BV-2與hWJ-MSCs共培養(yǎng)條件下，hWJ-MSCs表達(dá)IL-1β、IL-1α、IL-6、IL-8和MMP-2明顯受到LPS、BV-2細(xì)胞以及共培養(yǎng)細(xì)胞比例三種因素的影響：其中IL-1β表達(dá)表現(xiàn)為僅受到LPS的影響（LPS-上調(diào)，LPS+抑制）；MMP-2僅在LPS-共培養(yǎng)72h后出現(xiàn)上調(diào)，其他情況下均表現(xiàn)為抑制狀態(tài)；IL-6僅在LPS+共培養(yǎng)24h內(nèi)上調(diào)，其他時(shí)間、環(huán)境下均表現(xiàn)為下調(diào)；而IL-8的表達(dá)影響主要與時(shí)間有關(guān)，在共培養(yǎng)24h內(nèi)（LPS-/LPS+）均表現(xiàn)為上調(diào)，而在其他時(shí)間內(nèi)表現(xiàn)為下調(diào)。與共培養(yǎng)中BV-2細(xì)胞相比，hWJ-MSCs受到的調(diào)節(jié)更為復(fù)雜，與多種環(huán)境因素有關(guān)。 結(jié)論 1.貼壁法以及胰酶-膠原酶Ⅱ聯(lián)合消化法均能成功分離hWJ-MSCs，體外標(biāo)準(zhǔn)培養(yǎng)條件下，可體外傳代20代，提示hWJ-MSCs為有限傳代的MSCs； 2.基因及蛋白水平檢測(cè)提示MSCs表達(dá)TLR4，經(jīng)LPS體外刺激模型試驗(yàn)進(jìn)一步提示TLR4在hWJ-MSCs為一功能性受體，但與其他成體組織來(lái)源MSCs比較，hWJ-MSCs對(duì)LPS的刺激反應(yīng)顯遲鈍，需要LPS刺激72小時(shí)才能出現(xiàn)一系列基因表達(dá)上調(diào)（IL-1β、IL-1α、IL-6、IL-8、IDO1、MMP-2、TLR4、CD14）和下調(diào)（I L-12、MMP-9），且LPS通過(guò)上調(diào)IDO1、MMP-2可調(diào)節(jié)hWJ-MSC免疫功能、遷移能力； 3. hWJ-MSCs可誘導(dǎo)BV-2向M2型轉(zhuǎn)化，這一過(guò)程需要細(xì)胞間直接接觸；體外hWJ-MSCs與BV-2存在相互調(diào)節(jié)作用。
[Abstract]:Research background
There is no timely and effective treatment for brain injury in newborns. MSCs is the most promising treatment for brain injury in newborns. However, MSCs treatment of brain injury is still at the stage of study.
objective
In vitro isolation and identification of human umbilical cord mesenchymal stem cells (hWJ-MSCs), the amplification efficiency of hWJ-MSCs in vitro, the growth difference between different amplification algebras, and the biological phenotypic changes of hWJ-MSCs in the simulated inflammatory environment were observed, and the main immune cell neuroglia cells of the central nervous system and the central nervous system were in the inflammatory model. Co culture was carried out in the environment to observe the interaction between the two cells, and provide a basis for further study of MSCs's intracranial transplantation.
Method
For the above purpose, this study is divided into the following 3 parts.
Isolation, in vitro amplification and identification of 1. hWJ-MSCs
5 neonates with normal term caesarean section of General Hospital of Beijing Military Region were collected. All umbilical cord tissues were stored in aseptic environment immediately after in vitro and processed in 1-6h to ensure the vitality of the isolated cells. The umbilical tissue was removed from the umbilical cord tissue under the aseptic environment, the vein, and the physical methods were cut down, and then combined with pancreatin and collagenase - II. The separation and purification and in vitro amplification of hWJ-MSCs. by flow cytometry were used to identify the surface of hWJ-MSCs cells by flow cytometry on CD14, CD31, CD73, CD105, CD90, HLA-ABC, HLA-DR. The cell differentiation potential was observed by the chondrogenic and adipose differentiation kit in vitro. The cell morphology and cell were observed by two separate methods. Periodic changes.
2. to observe the expression of Toll-like receptor4 (TLRs) on hWJ-MSCs and the effect of signal transduction pathway on hWJ-MSCs biological function.
The expression of TLR4 on hWJ-MSCs was detected by flow cytometry and qRT-PCR. TLR4 activator -LPS was used to stimulate hWJ-MSCs in vitro. The morphological changes, cell proliferation and growth cycle of hWJ-MSCs cells were observed at LPS at different time points. Take the total RNA, use the reverse transcriptase kit and the SYRB Green kit to detect the cell factors of hWJ-MSCs (IL-1 beta, IL-1 alpha, IL-2, IL-4, IL-6, IL-8, and IL-10) in the stimulus conditions. Collect cells and collect cell culture supernatant and observe cells The level of factor protein expression was further confirmed by qRT-PCR results.
Co culture of 3. hWJ-MSCs and mouse microglia cell line BV-2 in vitro
The hWJ-MSCs and BV-2 cells isolated in vitro were co cultured in different proportion in vitro, and the biological changes of two cells in the co culture system of two cells at different time points in LPS+/LPS- were observed. The difference of hWJ-MSCs expression of IL-1 beta, IL-6, IL-8 and MMP-2 under the co culture condition of qRT-PCR, and the M2 of BV-2 cell M2 were detected. The expression of cell phenotypic gene Arginase1, Arginase2, Mrc1 and Ym1 were changed, and the effect of hWJ-MSCs was observed by the selective activation of BV-2 (M2 type) markers. The difference of BV-2 cell NO secretion in the co culture system was detected by NO detection kit, and the effect of hWJ-MSCs on LPS induced transformation was observed when co culture was directly exposed to co culture.
Result
1. successful separation and identification of hWJ-MSCs
Under the aseptic condition, the cell growth of a spindle like cell like cell can be grown by the growing or further digestion of a single cell suspension culture bottle under the aseptic condition. The cells can be amplified and purify to 20 generations in vitro. The cell cells can maintain the original growth form before the 20 generation. Cell surface labeling results showed that the cells CD14, CD31 and HLA-DR were negative, CD73, CD105, CD90, HLA-ABC positive. The differentiation experiment in vitro suggested that the cells were induced into cartilage in vitro, and the adipose differentiation was successful. It was suggested that the umbilical cord mesenchymal stem cells were successfully obtained by the adherence method and the combined digestion of pancreatin collagenase and collagenase. The cell cycle was observed by PI staining between the 3-7 generations. The cells were in the G0/G1 stage, and a few cells were in the stage of proliferation and division, and they had the characteristics of stem cells. No death and late apoptotic cells were not found.
2. identification of TLR4 as a functional receptor on the surface of hWJ-MSCs
Flow cytometry and qRT-PCR analysis revealed that the expression of TLR4 on hWJ-MSCs stimulated hWJ-MSCs in vitro to trigger the cell stress response, and to detect the difference in the expression of the TLR4 signaling pathway related inflammatory factors (IL-1 beta, IL-1 a, IL-6, IL-8), suggesting that the detected inflammatory factor genes were obviously up-regulated after the stimulation of 72h, but IL-12 was in the stimulus bar. The expression of IL-2, IL-4, IL-10, IL-13, TNF-, and IFN- y was not detected. The detection of the expression of MMP-2 and MMP-9 gene indicated that MMP-2 was obviously up-regulated after LPS stimulation, but the MMP-9 expression was down down after the stimulation, but the expression rose to the same level as the negative control. In addition, the detection results of the expression of IDO1, IDO-2, IFN- beta and Cox2 genes of MSC immunosuppressive molecules showed that IDO1 and IFN- beta were up regulated under the stimulus of LPS, and the up regulation of IDO1 expression was in Shuangfeng. The expression of IDO-2 was not detected at the 3 time points observed. The results showed that LPS could modify the immunosuppressive function of hWJ-MSCs.
Interaction between 3.hWJ-MSCs and BV-2 co cultured in vitro
Under the conditions of co culture in vitro, both hWJ-MSCs and BV-2 cells were adherent growth. Under the LPS stimulation, hWJ-MSCs could significantly weaken the stimulation of LPS to BV-2; when co cultured with hWJ-MSCs, BV-2 only secreted a very low amount of NO without LPS stimulation. QRT-PCR analysis showed that when the proportion of two cells was maintained at 1:1, hWJ-MSCs could obviously increase the expression of M2 cell phenotypic marker Arginase1 expression in BV-2 cells; BV-2 and hWJ-MSCs co culture conditions, hWJ-MSCs expressed IL-1 beta, IL-1 alpha, IL-6, and three kinds of co culture cells. Influence of factors: IL-1 beta expression was only affected by LPS (LPS- up, LPS+ inhibition); MMP-2 was up-regulated only after LPS- co culture 72h, and all other cases were inhibited; IL-6 was up only in LPS+ co culture 24h, and the other time, under the environment, was down-regulation, and IL-8 expression was mainly related to time. Both in co cultured 24h (LPS-/LPS+) showed up regulation and down regulated at other times. Compared with BV-2 cells in co culture, the regulation of hWJ-MSCs was more complex and related to a variety of environmental factors.
conclusion
1. adherent wall method and pancreatin collagenase II combined digestion method can successfully separate hWJ-MSCs. Under the standard culture conditions, the 20 generation can be passed to the 20 generation, suggesting that hWJ-MSCs is the MSCs of the limited passages.
The detection of 2. gene and protein level suggested that MSCs expressed TLR4. The stimulation model test of LPS in vitro further hinted that TLR4 was a functional receptor in hWJ-MSCs, but compared with other adult tissue sources MSCs, hWJ-MSCs had a slow response to LPS, and a series of gene expression up-regulated (IL-1 beta, IL-1 alpha, IL-6, and IL-6) needed LPS stimulation for 72 hours. IDO1, MMP-2, TLR4, CD14) and down regulation (I L-12, MMP-9), and LPS can regulate the immune function and migration ability of the hWJ-MSC by upregulating IDO1.
3. hWJ-MSCs can induce BV-2 to transform into M2, which requires direct contact between cells. HWJ-MSCs and BV-2 interact in vitro.
【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2013
【分類號(hào)】：R722.1

【共引文獻(xiàn)】

相關(guān)期刊論文 前10條

1 沈延飛;李志軍;鄭繼生;李敏靜;董雷;蔡宛如;;“肺腸同治”對(duì)急性肺損傷大鼠肺和大腸組織P38表達(dá)影響[J];浙江中醫(yī)藥大學(xué)學(xué)報(bào);2013年08期

2 許鍵煒;申長(zhǎng)清;趙芳芳;舒莉萍;何志旭;;同源異體間充質(zhì)干細(xì)胞上清液干預(yù)大鼠佐劑性關(guān)節(jié)炎[J];貴陽(yáng)醫(yī)學(xué)院學(xué)報(bào);2013年04期

3 朱興陽(yáng);黃永明;徐偉龍;蘇海濤;劉軍;;間充質(zhì)干細(xì)胞治療骨性關(guān)節(jié)炎的研究進(jìn)展[J];廣東醫(yī)學(xué);2013年18期

4 闕雪梅;黃玉紅;孫明軍;;益生菌對(duì)炎癥性腸病的作用及機(jī)制[J];國(guó)際消化病雜志;2013年05期

5 許楓;;腦卒中后癲癇36例的臨床特征及其預(yù)后分析[J];內(nèi)蒙古醫(yī)學(xué)院學(xué)報(bào);2010年S3期

6 鄭潔;陳曉丹;鄭希;王順和;;體外磁鐵定向遷移SPIO標(biāo)記的間充質(zhì)干細(xì)胞對(duì)大鼠腦缺血治療的影響[J];基礎(chǔ)醫(yī)學(xué)與臨床;2013年10期

7 戚智鋒;師文娟;閆峰;張晨誠(chéng);劉克建;羅玉敏;;羥基紅花黃色素A對(duì)腦缺血-再灌注大鼠星形膠質(zhì)細(xì)胞活性的影響[J];中國(guó)腦血管病雜志;2013年09期

8 譚笑;汪年松;;干細(xì)胞治療糖尿病腎病的研究進(jìn)展[J];中國(guó)中西醫(yī)結(jié)合腎病雜志;2013年08期

9 楊雯;惠玲;呂同德;;人臍帶間充質(zhì)干細(xì)胞向血管內(nèi)皮細(xì)胞的定向分化及臨床應(yīng)用進(jìn)展[J];解放軍醫(yī)藥雜志;2013年10期

10 王海峰;;骨髓間充質(zhì)干細(xì)胞移植治療脊髓損傷研究[J];大家健康(學(xué)術(shù)版);2013年24期

相關(guān)會(huì)議論文 前1條

1 何鵬;胡忠玉;;Toll樣受體研究進(jìn)展[A];2013年中國(guó)藥學(xué)大會(huì)暨第十三屆中國(guó)藥師周論文集[C];2013年

相關(guān)博士學(xué)位論文 前10條

1 盧有勝;甲基強(qiáng)的松龍、Toll樣受體對(duì)激活CD4~+T細(xì)胞存活和增殖的影響[D];南京醫(yī)科大學(xué);2010年

2 張遷;樹突狀細(xì)胞分化成熟的表觀遺傳組學(xué)研究[D];第二軍醫(yī)大學(xué);2011年

3 張廣慧;大鼠嗅球?qū)δX內(nèi)神經(jīng)發(fā)生與軸突再生的作用研究[D];重慶醫(yī)科大學(xué);2011年

4 孫琳琳;電針對(duì)腦缺血再灌注模型大鼠炎性反應(yīng)相關(guān)信號(hào)的影響及機(jī)制研究[D];北京中醫(yī)藥大學(xué);2013年

5 謝梓菁;基于多元Granger因果分析的針刺治療中風(fēng)偏癱的機(jī)制研究[D];北京中醫(yī)藥大學(xué);2013年

6 申晶;骨髓源間充質(zhì)干細(xì)胞輸注促進(jìn)STZ誘導(dǎo)的糖尿病大鼠胰腺內(nèi)α細(xì)胞向β細(xì)胞的轉(zhuǎn)變：糖尿病治療的新模式[D];中國(guó)人民解放軍軍醫(yī)進(jìn)修學(xué)院;2013年

7 鄒蘇琪;成年斑馬魚視神經(jīng)損傷后軸突再生與髓鞘再建的研究[D];中國(guó)科學(xué)技術(shù)大學(xué);2013年

8 趙一萍;蒙古馬免疫相關(guān)基因表達(dá)研究及脾臟表達(dá)譜分析[D];內(nèi)蒙古農(nóng)業(yè)大學(xué);2013年

9 朱天琦;GDNF和NT-3雙基因誘導(dǎo)BMSCs分化為神經(jīng)樣細(xì)胞的實(shí)驗(yàn)研究[D];華中科技大學(xué);2013年

10 金文杰;米諾環(huán)素對(duì)70%肝切除小鼠術(shù)后遠(yuǎn)期學(xué)習(xí)記憶能力的影響及機(jī)制研究[D];南京醫(yī)科大學(xué);2013年

相關(guān)碩士學(xué)位論文 前10條

1 陸俊秀;血清IL-6、NSE及顱腦超聲對(duì)早產(chǎn)兒WMI的診斷價(jià)值以及GM-1對(duì)其的療效[D];南方醫(yī)科大學(xué);2012年

2 李東卿;經(jīng)肋間后動(dòng)脈介入移植骨髓基質(zhì)干細(xì)胞修復(fù)脊髓損傷的研究[D];南方醫(yī)科大學(xué);2012年

3 劉希偉;通心絡(luò)對(duì)急性腦梗死大鼠腦微血管變化的影響[D];北京中醫(yī)藥大學(xué);2013年

4 高平;骨髓間充質(zhì)干細(xì)胞（BMSCs）誘導(dǎo)分化為類神經(jīng)細(xì)胞移植治療大鼠脊髓損傷[D];山東大學(xué);2013年

5 孫忠文;2型糖尿病大鼠腦組織TIPE2和TNFAIP8的表達(dá)特征及其與Aβ1-40沉積的相關(guān)性[D];山東大學(xué);2013年

6 李雪;小膠質(zhì)細(xì)胞活化在孕期感染所致精神分裂癥大鼠模型中的研究[D];鄭州大學(xué);2013年

7 張惠莉;骨髓間充質(zhì)干細(xì)胞移植改善慢性腦缺血大鼠空間學(xué)習(xí)記憶能力并影響B(tài)DNF及P75NTR表達(dá)[D];鄭州大學(xué);2013年

8 李勝偉;大鼠肝門阻斷過(guò)程中捫摸對(duì)肝臟熱缺血再灌注損傷的影響及機(jī)制研究[D];鄭州大學(xué);2013年

9 吳芳芳;線粒體解耦聯(lián)蛋白2與抑郁癥的相關(guān)性研究[D];南京醫(yī)科大學(xué);2013年

10 馬云鵬;應(yīng)用大塊平鋪法對(duì)人臍帶間充質(zhì)干細(xì)胞分離、傳代、凍存及復(fù)蘇的研究[D];河北醫(yī)科大學(xué);2013年

本文編號(hào)：2151881