Mn-SOD修饰脂质干细胞通过隧道纳米管转运线粒体对糖尿病性ED氧化应激损伤的修复机制

The mechanisms involved in diabetes associated erectiel dysfunction (ED) are complicated. Our previous studies found that oxidative stress in the corpus cavernosum of diabetic rats played a important role in the development of diabetic ED and anti-oxidative therapy partly repaired the pathological injury.It has been demonstrated that intracavernous injection of adipose tissue-derived stem cells (ADSCs) partly improved erectile function in several ED animal models. However, both the anti-oxidative stress therapy and ADSCs injection have some limitations. Recently, tunneling nanotubule (TNT) has been found between cells. Endothelial progenitor cells (EPCs) transport mitochondria to the other cells in oxidative stress status to protect them. We hypothesize that intracavernous injection of ADSCs not only replace the injured cells, but also transport mitochondria to the endothelial cells and smooth muscle cells which are in oxidative stress status within the corpus cavernosum of diabetic rats. With this anti-oxiative effects, ADSCs could cure the injured cells in corpus cavernosum and improve erectile function of diabetic rats. Manganese-superoxide dismutase (Mn-SOD)is a kind of anti-oxidative enzyme wihtin the mitochondria. Mn-SOD gene modified ADSCs could express high level of Mn-SOD and improve the anti-oxidative effects. The proposed project may open a new window for ADSCs improving erectile function.

课题组前期研究发现，糖尿病性勃起功能障碍（DED）大鼠阴茎海绵体局部存在明显线粒体氧化应激损伤，抗氧化剂和脂质干细胞（ADSCs）均可以部分改善DED病理损害和勃起功能，但是两种治疗都存在一定局限。最近研究发现，细胞之间存在隧道纳米管（TNT），内皮祖细胞将线粒体等细胞器和其他物质通过TNT转运到处于应激状态的细胞内，提高应激细胞的防御抵抗力。锰超氧化物岐化酶（Mn-SOD）是存在于线粒体中的抗氧化酶。我们拟使用Mn-SOD基因修饰的ADSCs治疗DED，观察Mn-SOD/ADSCs是否可以通过TNT转运线粒体和高表达的Mn-SOD到阴茎海绵体局部内皮细胞和平滑肌细胞内，帮助靶细胞对抗氧化应激损伤，重新建立自由基代谢平衡，从而获得更好的治疗DED效果。干细胞通过TNT，"直接注射""细胞保护剂"到阴茎海绵体局部靶细胞内，治疗DED的作用机制有十分重要的研究价值。

干细胞治疗糖尿病性ED的相关研究近期较多，但多数均从干细胞局部替代或干细胞旁分泌机制研究。本研究从另外一个角度，即纳米隧道管（TNTs），来探讨干细胞治疗糖尿病性ED的作用机制。. 本研究发现，高糖基化终产物可以使内皮细胞出现氧化应激损伤，线粒体功能下降，细胞呼吸功能下降，凋亡增加。体外培养脂肪干细胞（ADSCs），并转染Mn-SOD-EGFP或EGFP，并经过鉴定，过表达Mn-SOD与线粒体共定位。与ADSCs共培养，可以对抗高糖基化终产物培养内皮细胞的氧化应激损伤，可提高损伤内皮细胞NO产生能力。活细胞工作站连续拍摄其动态变化，Mn-SOD-ADSCs与内皮细胞之间形成了TNTs结构，并且Mn-SOD-ADSCs向处于氧化应激损伤的内皮细胞通过TNT结构，转运线粒体，帮助内皮细胞抗氧化作用。DED大鼠经Mn-SOD-ADSC治疗后可改善其勃起功能，提高海绵体组织内平滑肌以及内皮的含量，增加肌纤维、减少胶原的量，减轻DED大鼠海绵体组织内的氧化应激损伤，提高DED大鼠阴茎海绵体组织内NO、cGMP的水平，提高eNOS、cNOS的活性，降低iNOS的活性，增加线粒体数量和对抗线粒体肿胀。. 干细胞通过TNTs结构，像打针一样，直接将正常干细胞内线粒体“注射”到氧化应激损伤的内皮细胞中，发挥抗氧化作用，这是干细胞治疗糖尿病性ED的一个新的机制，有别于以往的细胞替代学说和细胞旁分泌因子学说。对于干细胞治疗机制研究是一个十分重要的补充。