Geraci J P, Mariano M S
Department of Environmental Health, School of Public Health and Community Medicine, University of Washington, Seattle 98195.
Radiat Res. 1993 Nov;136(2):205-13.
Cellular changes that occur within the liver before and during liver fibrosis have been studied. At various times (0-90 days) after gamma irradiation (0-25 Gy) of the whole liver, hepatocytes and liver nonparenchymal cells were isolated by enzymatic dissociation of the liver and differential centrifugation. Differential cell counts were done to quantify the yield of hepatocytes and nonparenchymal cells. Hepatocyte function in vitro was assayed by the uptake of rose bengal and compared with the clearance of the dye in vivo. Intracellular alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were used to estimate the effect of radiation on the phenotypic expression of hepatocytes. Gene expression of transforming growth factor beta 1 (TGF-beta 1), a cytokine which has been found to play a role in hepatic fibrosis, was measured by 32P-cDNA:mRNA hybridization of poly(A+) mRNA isolated from purified populations of hepatocytes and nonparenchymal cells. Morphologically, hemorrhages were evident in perfused livers 28 days after 25 Gy whole-liver irradiation. This injury was preceded by a significant increase in liver nonparenchymal cells at 2 weeks, the earliest time after irradiation at which measurements were made. Significant increases in liver nonparenchymal cells occurred at doses greater than 10 Gy and persisted throughout the follow-up period after irradiation. The lowest dose studied (5 Gy) inhibited both normal growth of the liver and the associated increase in the number of hepatocytes. Doses greater than 10 Gy resulted in a dose-dependent decline in liver mass and hepatocytes. Viable hepatocytes isolated from irradiated dysfunctional livers, with respect to clearance of rose bengal, were functionally normal in vitro. A dose-dependent decline in the ALT and AST contents of liver cell suspensions and homogenates was observed but the relative decline in ALT was greater, resulting in lower ALT/AST ratios. This decline in the ALT/AST ratios occurred at doses (< 15 Gy) which caused no loss of hepatocytes, suggesting possible conversion of ALT-rich periportal hepatocytes to ALT-poor hepatocytes due to changes in the microenvironment and/or aging of the cell population. TGF-beta 1 mRNA was detected mainly in nonparenchymal cells, and radiation preferentially enhanced the TGF-beta 1 gene expression in these cells. These data suggest that radiation-induced alterations in liver nonparenchymal cell populations may be responsible for microvascular fibrosis, which results in a cascade of pathological events which lead to hepatocyte loss.
已经对肝纤维化之前和期间肝脏内发生的细胞变化进行了研究。在对整个肝脏进行γ射线照射（0 - 25 Gy）后的不同时间（0 - 90天），通过肝脏的酶解和差速离心分离肝细胞和肝脏非实质细胞。进行差异细胞计数以量化肝细胞和非实质细胞的产量。通过孟加拉玫瑰红摄取来测定体外肝细胞功能，并与体内染料清除率进行比较。使用细胞内丙氨酸转氨酶（ALT）和天冬氨酸转氨酶（AST）来评估辐射对肝细胞表型表达的影响。通过对从纯化的肝细胞和非实质细胞群体中分离的多聚腺苷酸（poly(A+)）mRNA进行32P - cDNA:mRNA杂交，测量在肝纤维化中起作用的细胞因子转化生长因子β1（TGF - β1）的基因表达。形态学上，在25 Gy全肝照射28天后，灌注肝脏中出现明显出血。在照射后最早进行测量的时间点，即2周时，肝脏非实质细胞显著增加先于这种损伤。在大于10 Gy的剂量下，肝脏非实质细胞显著增加，并在照射后的随访期内持续存在。研究的最低剂量（5 Gy）抑制了肝脏的正常生长以及肝细胞数量的相关增加。大于10 Gy的剂量导致肝脏质量和肝细胞呈剂量依赖性下降。从照射后的功能失调肝脏中分离出的有活力的肝细胞，就孟加拉玫瑰红清除率而言，在体外功能正常。观察到肝细胞悬液和匀浆中ALT和AST含量呈剂量依赖性下降，但ALT的相对下降更大，导致ALT/AST比值降低。ALT/AST比值的这种下降发生在导致肝细胞无损失的剂量（< 15 Gy）下，这表明由于微环境变化和/或细胞群体老化，富含ALT的门周肝细胞可能转变为ALT含量低的肝细胞。TGF - β1 mRNA主要在非实质细胞中检测到，并且辐射优先增强这些细胞中的TGF - β1基因表达。这些数据表明，辐射诱导的肝脏非实质细胞群体改变可能是微血管纤维化的原因，这会导致一系列病理事件，进而导致肝细胞丢失。
