骨髓基质干细胞S334转基因大鼠和SD大鼠眼内移植的初步研究

A Pilot Study of Bone Marrow Stromal Cells Intraocular Transplantation in the S334 Transgenic Rats and Sprague-Dawley Rats

目的:探讨骨髓基质干细胞（bone marrow stromal cell,BMSC）在新生S334转基因视网膜变性大鼠和（Sprague-Dawey）SD大鼠眼内生存、发育、分化的情况。方法:人BMSC培养在含10％胎牛血清的（a-Modified Eagle Medium,a-MEM）培养基上扩增;实验分四组;第一组:S334转基因鼠,细胞移植联合维甲酸（Retinoid acid）RA注射组;第二组:S334转基因大鼠,细胞移植组;第三组:SD大鼠,细胞移植联合RA注射组;第四组:SD大鼠,细胞移植组。以上每组各5只。 2μl细胞混悬液（约 4×104个细胞）注入生后1d大鼠的玻璃体腔。于生后14 d,生后 23 d处死动物,取出眼球作塑料切片,进行组织学分析。结果:本实验结果表明,BMSC移植到生后 1d S334杂合子转基因大鼠玻璃体腔,在外源性RA的作用下,可能参与宿主视网膜的后期发育,可见内核层细胞增多,整个神经视网膜增厚,但感光细胞层数不变。而无外源性RA移植组,在生后14 d时,可见移植细胞形成类似血岛样结构,而向神经分化的成分少。SD组,在联合RA注射时,生后23d将动物处死,作组织学分析,发现移植细胞在眼内能移行分化,也可见视网膜内核层细胞增多,同时由于RA的作用,可见感光细胞增生。在无RA作用时,可见宿主神经视网膜结构紊乱,移植细胞增生形成非典型性增生细胞团。

Purpose: To investigate the survival and differentiation of human bone marrow stromal cells (BMSC) intraocular transplantation in newborn S334 retinal degeneration transgenic rats and (Sprague-Dawley)SD rats.Methods: Human bone marrow stromal cells line was grown on the adhesive substrate in the condition media including a-Modified Eagle medium (a-MEM) /10% fetal bovine serum. The experiments were divided into four groups: Group 1: BMSC + (Retinoid Acid) RA transplanted in S334 transgenic rats( n = 5); Group 2: BMSC transplanted in S334 transgenic rats(n = 5); Group 3: BMSC +RA transplanted in SD rats(n = 5); Group 4: BMSC transplanted in SD rats( n = 5) . 2 μl cell suspension (about 4 × 104cells) was injected into the vitreous space in the transgenic rats and normal SD rats at Postnatal 1(P1) respectively. The right eyes were treated eyes and the left eyes were used as control. At P14 and P23, the rats were killed and enucleated for histological assays using plastic section.Results: In Group 1, the transplanted cells were well survived. They could continue to differentiate and participate in late-stage retinal development. The number of inner nuclear layer increased. Moreover, the host retina increased their thickness, but photoreceptor cells were not rescued from transplant. In Group 2, at P14, the BMSC continue to differentiate toward their linage cell fate and formed into hemorrhage island structures with few neurons if RA was not applied. Group 3, BMSC could survive, migrate. The number of inner nuclear layer increased also. In Group 4, it revealed that host retina structures were disorganized and transplant cells formed atypical proliferating mass. Conclusions: This pilot experiment indicated that bone marrow stromal cells could survival, differentiate and participate in the retinal development after transplanted into vitreous space in the new born transgenic rats and SD rats. Histological assays showed that transplanted cells integrated with inner nuclear layer of host retina. Thus, bone marrow stromal cells may be a useful vehicle for auto- transplantation for the therapy of variety of retinal degenerative disorders.