本文選題：腎臟 + 缺血再灌注損傷�。� 參考：《山西醫(yī)科大學》2014年碩士論文

【摘要】：1.目的 1.1研究骨髓間充質(zhì)干細胞的形態(tài)及生長特征；并對其進行鑒定。 1.2建立腎臟缺血再灌注損傷模型并移植SPIO標記BMSC（s經(jīng)腎動脈及腎包膜下兩種移植方法）；再利用MR活體示蹤BMSCs，T2及T2*map圖像定量分析。 1.3綜合評價BMSCs移植對腎臟缺血再灌注損傷的治療療效。 2.材料與方法 2.1實驗對象 SD大鼠40只,雄性，5-6w,170-200g(山西醫(yī)科大學實驗動物中心提供)。 2.2實驗方法 MRI檢查方法：SD大鼠分別于造模前、造模后1d、1w、2w、3w行磁共振掃描（采用SiemensVerio3.0T磁共振成像儀，水平臥位,頭先進,行軸位T2WI、T2*WI、多回波T2序列及T2*序列掃描）。 細胞鑒定方法：采用流式細胞儀檢測，，CellQuestPlot軟件分析數(shù)據(jù)；采用常規(guī)免疫熒光染色，用激光共聚焦顯微鏡觀察。 SD大鼠造模3w后計算體重增長率、下腔靜脈采血測定腎臟功能，取材進行組織切片及HE染色。 3.結(jié)果 3.1體外實驗 采用全骨髓貼壁法成功分離BMSCs，免疫熒光法及流式細胞分析法鑒定細胞表型：CD34(-)、CD45（-）、CD44(+)、CD90(+)；BMSCs能夠成功地被誘導分化為成骨、成脂細胞。 3.2MRI活體示蹤 （1）腎臟IRI后MRI表現(xiàn)：損傷前，T2WI圖像雙腎大小基本對稱、信號一致，而腎臟缺血再灌注損傷后左腎體積增大、信號增高；（2）T2*WI序列在體示蹤SPIO標記BMSCs：1d時，兩種不同方法低信號區(qū)（鐵信號）均局限在注射局部，隨著時間推移，腎動脈組左腎出現(xiàn)斑片狀低信號區(qū)，而腎包膜下組低信號區(qū)仍部分局限于包膜下。（3）損傷組與治療組（包括腎動脈組、腎包膜下組）T2值：損傷后，三組T2值均升高，最高點位于1d；隨著時間推移，T2值有下降趨勢，但兩組治療組下降幅度較損傷組大；在同一時間點，治療組T2值低于損傷組，但兩組治療組之間差異不明顯。損傷組與兩組治療組（腎動脈組、腎包膜下組）T2*值：損傷組T2*值趨勢比較穩(wěn)定，推測腎臟缺血再灌注損傷對T2*值影響不大；兩組治療組T2*值最低點均位于2w，在同一時間點，兩組治療組T2*值低于損傷組，但兩組治療組之間變化趨勢不一致。 3.3評價療效 （1）比較三組體重增長率，由高到低依次為：對照組治療組損傷組；（2）比較三組腎臟功能，尿素氮及肌酐升高水平由高到低為：損傷組治療組對照組，治療組間差異有統(tǒng)計學意義，腎動脈組較腎包膜下組水平低；（3）比較HE染色結(jié)果：對照組腎臟組織結(jié)構(gòu)基本正常，損傷組大量腎小管上皮細胞腫脹、空泡變性，治療組腎小管上皮細胞壞死程度較損傷組輕。 4.結(jié)論 4.1采用全骨髓貼壁法可以成功分離、培養(yǎng)BMSCs。 4.2SPIO標記的BMSCs能夠靶向遷移到腎臟缺血再灌注損傷區(qū)。 4.3MRIT2WI及T2map圖像能夠顯示腎臟缺血再灌注損傷后改變，T2*WI及T2*map圖像能夠活體示蹤SPIO標記的BMSCs，但是兩種移植方法無明顯差異。 4.4從體重增長率、腎臟功能及組織HE染色綜合分析，BMSCs對腎臟缺血再灌注損傷有一定修復作用。
[Abstract]:1. purposes
1.1 to study the morphology and growth characteristics of bone marrow mesenchymal stem cells and identify them.
1.2 the model of renal ischemia-reperfusion injury was established and SPIO labeled BMSC was transplanted (two kinds of transplantation methods under renal artery and renal capsule), and the quantitative analysis of BMSCs, T2 and T2*map images was traced by MR in vivo.
1.3 to evaluate the therapeutic effect of BMSCs transplantation on renal ischemia-reperfusion injury.
2. materials and methods
2.1 experimental objects
40 SD rats, male, 5-6W, 170-200g (provided by Shanxi Medical University experimental animal center).
2.2 experimental method
MRI examination method: SD rats were performed by magnetic resonance scanning (using SiemensVerio3.0T magnetic resonance imaging instrument, horizontal position, head advanced, axis T2WI, T2*WI, multiple echo T2 sequence and T2* sequence) after model building, 1D, 1W, 2W, 3W.
Cell identification methods: flow cytometry, CellQuestPlot software analysis data, routine immunofluorescence staining, laser confocal microscopy observation.
The weight gain rate of SD rats was calculated after modeling 3W, and the renal function was measured by inferior vena cava blood collection. Tissue sections and HE staining were taken.
3. results
3.1 in vitro experiment
BMSCs was successfully separated by full bone marrow adherence, and the cell phenotype was identified by immunofluorescence and flow cytometry: CD34 (-), CD45 (-), CD44 (+), and CD90 (+); BMSCs could be successfully induced to differentiate into osteoblast and lipocyte.
3.2MRI living body tracer
(1) MRI expression after IRI: before injury, the size of the two kidneys in the T2WI image was basically symmetrical and the signal was consistent, but the volume of the left kidney increased and the signal increased after the renal ischemia-reperfusion injury. (2) when the T2*WI sequence traced the SPIO marker BMSCs:1d, the two different methods of the low signal region (iron signal) were limited to the injection part, and the renal artery group went on with time. There was a low signal area in the left kidney, while the low signal area in the subcapsular group was still partially confined to the capsule. (3) the T2 value of the injury group and the treatment group (including the renal artery group and the renal subcapsular group): after the injury, the three groups were all increased and the highest point was in 1D; the T2 value decreased as the time went on, but the decrease of the two groups was larger than that of the injury group. At the same time point, the value of T2 in the treatment group was lower than that of the injury group, but the difference between the two groups was not obvious. The T2* value of the injured group and the two groups (renal artery group and subcapsular group): the trend of T2* value in the injured group was more stable, and the effect of renal ischemia reperfusion injury on the value of T2* was not significant; the lowest T2* value of the two groups was in the same one in the same group. At the time point, the T2* value of the two groups was lower than that of the injury group, but the trend of change between the two groups was not consistent.
3.3 evaluation of curative effect
(1) the weight growth rate of the three groups was compared from high to low to the control group, and (2) the renal function of the three groups was compared with the high to low levels of urea nitrogen and creatinine. The difference between the treatment group and the control group was statistically significant, and the renal artery group was lower than the renal subcapsular group; (3) the results of HE staining were compared: A large number of renal tubular epithelial cells were swollen and vacuolar degeneration in the injured group. The necrosis degree of renal tubular epithelial cells in the treatment group was lighter than that in the injury group.
4. conclusion
4.1 the whole bone marrow adherent method can be successfully used to isolate BMSCs..
4.2SPIO labeled BMSCs can migrate to renal ischemia-reperfusion injury area.
4.3MRIT2WI and T2map images can show changes in renal ischemia-reperfusion injury, and T2*WI and T2*map images can trace SPIO labeled BMSCs in vivo, but there is no significant difference between the two methods of transplantation.
4.4 comprehensive analysis of body weight growth rate, renal function and tissue HE staining indicated that BMSCs had certain restorative effect on renal ischemia-reperfusion injury.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R699.2;R445.2

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