Establishment of simulation model for human drug metabolism by bioartificial liver

我们还在继续研究放射流生物人工肝。在该系统中培养肝细胞后，CYP3A4的表达显著增加。我们使用原表达CYP3A4并保持高功能的FLC细胞。在RFB中培养其他表达CYP3A4的肝细胞后，CYP3A4 mRNA的表达量远高于单层培养和FLC细胞。此外，我们在RFB中培养了引入谷氨酰胺合成酶基因和CYP3A4基因的GS3A4-HepG2。因此，我们确定在RFB培养下的这些细胞有效地代谢了铵。接下来，我们研究了其他几种肝细胞系是否适合RFB培养。结果表明，倍增时间短于24hr的细胞可以在RFB上培养。我们推测倍增时间超过24hr的细胞在附着于培养基质之前可能会被丢弃。我们在2005年的日本再生大会上展示了这些结果。另一方面，我们研究了除CYP3A4外，RFB和单层培养后CYP1A1、2D6、2A6、2C9和2C19的表达差异。因此，我们只得到了它们之间培养后2D6的表达差异。我们没有得到1A1的差异，这意味着与肿瘤发生相关的酶没有诱导。但RFB培养后，这些细胞株的尿毒症酶精氨酸琥珀酸裂解酶、氨甲酰对合成酶、精氨酸琥珀酸合成酶的表达增加，甲胎蛋白（AFP）的产生受到抑制。此外，我们也尝试在RFB上培养肾细胞，因为我们想将RFB应用于肝细胞综合征的治疗。其中，近端小管细胞（PTC）能在RFB上培养，功能增强。例如，在高糖/低氧培养PTC时，RFB增加了CYP27的表达，增加了细胞外基质的表达。这些结果表明，高糖和缺氧可能是诱发急性肾功能衰竭的必要条件。我们在2005年的再生大会和2006年的东京医科大学大会上展示了这些结果，jojima博士得到了东京医科大学的资助。最后，我们在2006年的《东京医科大学学报》上报道了这些结果。少

We have continued to study by using radial flow bio artificial liver. The expression of CYP3A4 was extremely increased after culturing liver cells in this system. We used FLC cells which originally expressed CYP3A4 and keep high function. After we cultured other liver cells that express CYP3A4 in RFB, CYP3A4 mRNA expressed much more than monolayer culture as well as FLC cells. Furthermore, we cultured GS3A4-HepG2 which introdued gene of glutamin synthetase and CYP3A4 in RFB. As a result, we determined that annmonium was metabolized effectively by those cells under RFB culture.Next, we investigated whether other several liver cell lines are suitable for RFB culture or not. We got the results that the cells which those doubling time were shorter than 24hr could culture on RFB. We speculated that the cells which doubling time was longer than 24hr might be thrown away before attaching on culture matrix. We presented these results at the Japanese congress of regeneration on 2005.On the othe … More r hand, we investigated the differences of expression of CYP1A1, 2D6, 2A6, 2C9, and 2C19 except for CYP3A4 after culturing on RFB or monolayer. As a result, we only got the differences of expression of 2D6 after culturing between them. We could not get the differences of 1A1, which means that enzyme associated with tumorgenesis did not induce.But, expression of arginosuccinate lyase, carbamoyl-p-synthetase, argininosuccinate synthetase that are uremic enzyme on those cell lines increased after RFB culture, and their production of alphafetoprotein (AFP) was inhibited.Furthermore, we also tried to culture kidney cells on RFB because we want to apply RFB for treatment of hepatocellular syndrome.Among them, proximal tubular cells (PTC) were capable of culture on RFB and their function also was increased. For example, expression of CYP27 was increased and expression extracellular matrix was increased on high glucose/hypoxic culture of PTC by using RFB. These results indicated that both high glucose and hypoxia might be necessary to induce acute renal failure. We presented these results at the congress of regeneration on 2005 and congress of Tokyo medical university on 2006, and Dr.Jojima got the grant from Tokyo medical university. Finally, we reported these results on Journal of Tokyo medical university on 2006. Less