복강 내 간세포이식에서 비실질간세포 동시이식의 효과
The effect of cotransplantation of nonparenchymal liver cells on the survival of intraperitoneally transplanted hepatocytes
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목적 : 간세포이식에서 복강은 대량의 세포를 안전하게 투여할 수 있다는 점에서 급성 간부전 치료를 위한 매력적인 이식 장소이지만, 복강 자체는 간세포 생착에 적합한 환경은 아니다. 간세포는 부착성장을 하는 세포로서 세포외 간질이나 비실질간세포와의 상호작용이 생존에 중요한 것으로 알려져 있다. 본 연구에서는 복강 내 이식된 간세포가 얼마나 빨리 기능을 발현하는지, 또한 비실질간세포를 동시에 이식함으로써 간세포 생존을 연장시킬 수 있는지 알아보고자 하였다. 방법 : 빌리루빈 결합 효소에 결함이 있어 선천적으로 황달이 발생하는 Gunn 쥐의 복강 내로 정상 Wistar 쥐의 간세포를 단독(kg당 109개, 1군, 10마리) 또는 비실질간세포(kg당 5×108개, 2군, 10마리)와 동시에 이식하였다. 대조를 위하여 Gunn 쥐의 간세포 및 비실질간세포를 Gunn 쥐의 복강 내로 이식하여 비교하였다(n=5). 이식 후 6시간, 1일, 4일, 1주, 2주, 3주, 4주, 5주, 6주에 담즙, 혈액, 복강 내 조직을 채취하였다. 담즙으로 결합형 빌리루빈이 배설되는지에 대해서 고작위액체크로마토그래피로 확인하였고, 혈청 빌리루빈의 농도를 측정하여 변화를 관찰하였다. 복강 내 간세포 존재는 형태학적 관찰, 면역조직화학염색, 제자리부합법으로 확인하였다.결과 : 복강 내 간세포이식 후 6시간째 담즙에서 결합형 빌리루빈이 나타나기 시작하였고, 4일째 정점을 이룬 후 점차 감소하였다. 담즙 내 검출량은 간세포만을 투여 받았던 1군이나 비실질간세포를 동시에 투여 받았던 2군 간에 차이가 없었으나, 검출기간은 1군이 2주까지 검출된 데 비하여 2군은 4주까지 검출되었다. 혈청 총 빌리루빈은 1군이 이식 전 7.05±0.24 mg/dL에서 이식 2주째 5.98±0.46 mg/dL로 15.1% 유의하게 감소하였고(p=0.0395), 2군은 이식 전 6.92±0.20 mg/dL에서 2주째 5.62±0.35 mg/dL로 18.9% (p=0.0036), 3주째 6.08±0.37 mg/dL로 12.1% 감소되었다(p=0.0478). 형태학적 관찰에서는 알부민 mRNA 발현세포가 1군에서 2주까지만 관찰된 반면 2군에서는 4주까지 관찰이 가능하였다.
Abstract
Background : Transplantation of primary hepatocytes (PH) has been shown to provide metabolic support during acute liver failure. However, PH are known to be subject to necrosis in the peritoneal cavity. This is because cell-cell interaction plays an important role in their survival, but the peritoneal cavity can not provide such an environment. We tried to improve the survival of PH by simultaneously transplanting nonparenchymal liver cells (NPL). Methods : PH from normal Wistar rats, either alone (109 cells/kg, group 1, n=10) or mixed with NPL (5×108 cells/kg, group 2, n=10) were transplanted into the peritoneal cavity of hyperbilirubinemic Gunn rats which are congenitally devoid of bilirubin glucuronidation. Liver cells from Gunn rats were transplanted as a control. Results : Bilirubin glucuronides (BG) were detected in the bile of both group 1 and 2 rats collected at 6 hours after transplantation, and reached peak levels in 4 days. However, in the third and fourth week, BG could be detected only in group 2 animals. The serum bilirubin levels were decreased by 12.1～18.9% of basal levels in the second and third week for group 2 rats, but decreased by 15.1% only in the second week for the group 1 rats. Using in situ hybridization, albumin mRNA positive cells could be detected until the fourth week for the group 2 animals, but only until the second week for the group 1 rats. Conclusions : PH start functioning in a short time after intraperitoneal transplantation and simultaneous transplantation of NPL with PH can prolong the survival and function of transplanted hepatocytes.(Korean J Med 72:162-171, 2007)
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