本文關鍵詞： 應用 組織 工程技術(shù) 提高 脂肪 體細胞 存活率 研究　出處：《浙江大學》2011年碩士論文　論文類型：學位論文

【摘要】：背景：自體脂肪組織作為人體組織的填充材料被廣泛的運用。雖然脂肪組織在人體內(nèi)來源相對豐富,但大量的脂肪組織移植來源仍然受限。所以具有分化、增殖潛能的脂肪干細胞和脂肪前體細胞作為脂肪組織工程的種子細胞引起了關注。脂肪前體細胞是從脂肪組織分離出的脂肪干細胞分化出的早期脂肪細胞,通過誘導具有分化增殖的潛能。外源性血管內(nèi)皮細胞生長因子(vascular endothelial growth factor, VEGF)可促進移植組織內(nèi)的血管增生和脂肪細胞分化增殖,但外源性VEGF需要持續(xù)作用于組織才能起到較好效果。透明質(zhì)酸鈉(HA)是葡糖醛酸-N-乙酰氨基葡糖為雙糖單位組成的直鏈高分子多糖,廣泛存在于組織細胞基質(zhì)中；具有優(yōu)良的生物相容性和生物可降解性。HA的大分子網(wǎng)狀結(jié)構(gòu)通過與水形成氫鍵可結(jié)合大量的水,可以維持局部VEGF濃度,同時其獨特的三維結(jié)構(gòu)使之成為理想的種子細胞的支架材料。 目的：以透明質(zhì)酸支架為載體,復合小鼠脂肪前體細胞、VEGF植入小鼠體內(nèi),通過檢測組織血管形成脂肪分化情況,評價該復合載體材料對脂肪前體細胞移植存活率的影響。 方法：將小鼠3T3-L1脂肪前體細胞復蘇、傳代培養(yǎng)后,以脂肪分化誘導劑誘導8天。然后分為4組注射入小鼠皮下：實驗組(誘導后脂肪細胞、PBS緩沖液、透明質(zhì)酸、VEGF)、HA對照組(誘導后脂肪細胞、PBS緩沖液、透明質(zhì)酸)、VEGF對照組(誘導后脂肪細胞、PBS緩沖液、VEGF)和空白組(誘導后脂肪細胞、PBS緩沖液)。術(shù)后2,4,8周取材,仔細剝離取出移植物,觀察色澤、質(zhì)地。電子天平測量質(zhì)量。標本分別進行HE組織染色和蘇丹Ⅲ染色。體外PCR實驗檢測。 結(jié)果：8周取材見小鼠背部皮下脂肪組織淡黃色,質(zhì)軟,容易剝離。實驗組、,HA對照組均取到標本。HE染色切片表現(xiàn)為移植物外周為纖維包膜,空白對照組可見部分壞死脂肪細胞,炎性細胞浸潤。脂肪細胞大小不一,存活較少。實驗組存活脂肪細胞較多,壞死灶較少,細胞較大排列均一,可見微血管生長。蘇丹Ⅲ染色可見對照組紅染脂肪較少,排列不規(guī)則,大小不一,有局部纖維化。實驗組紅染然脂肪較多,壞死灶較少,細胞較大排列均一。應用SPSS 16.0軟件對2周,4周,8周各組重量進行方差分析,2周、4周VEGF對照組與空白組無統(tǒng)計學差異,其余各組均有統(tǒng)計學差異。8周實驗組與其他三組有統(tǒng)計學差異,三組間沒有統(tǒng)計學差異。體外PCR實驗結(jié)果提示各組PPAR-γ、C-EBPα條帶密度均相近。說明在體外各組表達無明顯差異。 結(jié)論：應用組織工程技術(shù),以透明質(zhì)酸支架復合小鼠3T3-L1脂肪前體細胞、VEGF注射充填可以延長VEGF持續(xù)作用時間,促進新生脂肪組織血管形成,提高小鼠脂肪前提細胞誘導后移植的存活率。
[Abstract]:Background: autogenous adipose tissue is widely used as the filling material of human body tissue. Although adipose tissue is relatively abundant in human body, a large number of adipose tissue transplantation sources are still limited. The proliferation potential of adipose stem cells and adipose progenitor cells as seed cells for adipose tissue engineering have attracted much attention. Adipose progenitor cells are early adipocytes derived from adipose stem cells isolated from adipose tissue. Exogenous vascular endothelial growth factor (VEGF) and vascular endothelial growth factor were induced to have the potential of differentiation and proliferation. VEGF can promote vascular proliferation and adipocyte differentiation and proliferation in grafts. Hyaluronic acid sodium hyaluronate (HA) is a straight-chain polymer polysaccharide composed of glucuronic acid -N-acetyl glucosamine as a disaccharide unit. It is widely found in tissue and cell matrix. The macromolecular reticular structure with excellent biocompatibility and biodegradability. Ha can maintain local VEGF concentration by forming hydrogen bonds with water. At the same time, its unique three-dimensional structure makes it an ideal scaffold for seed cells. Objective: using hyaluronic acid scaffold as carrier, combined with mouse adipocyte precursor cells (VEGF) were implanted into mice, and adipose differentiation was detected by tissue angiogenesis. To evaluate the effect of the composite carrier material on the survival rate of fat precursor cell transplantation. Methods: the 3T3-L1 adipose progenitor cells were resuscitated and cultured for 8 days, then divided into 4 groups: experimental group (adipocytes). PBS buffer, hyaluronic acid VEGFU HA control group (induced adipocyte buffer), hyaluronic acid (hyaluronic acid) PBS buffer group (induced adipocyte buffer solution). VEGF) and blank group (induced adipocyte PBS buffer). The grafts were removed and the color was observed 8 weeks after operation. Texture. Measurement of quality by electronic balance. Specimens were stained by HE and Sudan 鈪，

本文編號：1461531