The development of cell therapy for liver fibrolysis and prevention of carcinogenesis using bone marrow cell transplantaion

我们研究了骨髓细胞（BMC）移植对已建立的肝纤维化的影响。将绿色荧光蛋白（GFP）小鼠的骨髓细胞经尾静脉移植到四氯化碳（CCl_4）处理4周的C57 BL 6小鼠体内，再用CCl_4处理4周（共8周）。天狼星红和绿色荧光蛋白（GFP）染色清楚显示迁移的BMCs与纤维沿着存在，抗MMP-9抗体和原位杂交显示MMP-9的强表达。双荧光免疫组化显示MMP-9在GFP阳性细胞表面表达。电影原位酶谱分析显示，在门静脉周围区与MMP-9阳性的BMC的位置相一致的明胶酶活性强。BMC移植后4周，与单独用CCl_4处理的小鼠相比，通过肝脏羟脯氨酸含量评估，小鼠的肝纤维化显著减少。Liv 8阴性的BMCs亚群负责这种纤维溶解作用。总之，与单独用CCl_4治疗的小鼠相比，连续注射CCl_4的BMC移植小鼠在BMC移植后具有减少肝纤维化和显著提高存活率的作用。这一发现为肝纤维化的治疗提供了新的思路。还已知预防纤维化可减少致癌作用。因此，这些结果可能表明，纤维溶解骨髓细胞移植导致预防肝癌的发生。

We investigated the effect of bone marrow cell (BMC) transplantation on established liver fibrosis. BMCs of green fluorescent protein (GFP) mice were transplanted into 4-week carbon tetrachloride (CCl_4)-treated C57BL6 mice through the tail vein, and the mice were treated for 4 more weeks with CCl_4 (total, 8 weeks). Sirius red and GFP staining clearly indicated migrated BMCs existing along with fibers, with strong expression of matrix metalloproteinase (MMP)-9 shown by anti-MMP-9 antibodies and in situ hybridization. Double fluorescent immunohistochemistry showed the expression of MMP-9 on the GFP-positive cell surface. Film in situ zymographic analysis revealed strong gelatinolytic activity in the periportal area coinciding with the location of MMP-9-positive BMCs. Four weeks after BMC transplantation, mice had significantly reduced liver fibrosis, as assessed by hydroxyproline content of the livers, compared to that of mice treated with CCl_4 alone. Subpopulation of Liv8-negative BMCs was responsible for this fibrolytic effect. In conclusion, mice with BMC transplants with continuous CCl_4 injection had reduced liver fibrosis and a significantly improved survival rate after BMC transplantation compared with mice treated with CCl_4 alone. This finding introduces a new concept for the therapy of liver fibrosis. Also it has been known that prevention of fibrosis may reduce the carcinogenesis. Thus these results may suggest that fibrolysis bone marrow cell transplantation leads prevention of hepato-carcinogenesis.