Dynamic Analysis of Hepatic Microcirculation and Hepatic Function by In vivo-Fluorescence Microscopy -Application to the Study of Pathogenesis and Treatment of Liver Diseases.

本研究建立了活体荧光显微镜技术，可在2000倍以上放大观察肝脏微循环，以阐明酒精、药物和病毒致肝损伤过程中细胞功能的动态变化。对正弦血流进行连续定量测量。红细胞和白色细胞的正弦速度。肝小叶微循环的特点是：中央周围区（PC）血流量大于门静脉周围区（PP）血流量，从PP到PC区血流量有一个梯度（PP>PC）。此外，本研究还观察了乙醇对肝脏微循环的影响，以探讨微循环障碍在酒精性肝损伤中的作用。低剂量酒精使窦血流量和窦血氧合增加，但PC区部分窦血氧合下降，对酒精产生异质性反应。与此相反，高剂量酒精显着降低了窦血液的氧合，特别是在PC区，导致缺氧区域的数量增加。此外，荧光素钠的研究表明，从肝细胞到胆小管，特别是在PC区的排泄障碍。这些数据表明，高剂量酒精产生小叶内分流和局部缺氧，导致肝功能障碍。在灌流肝脏中，结果表明，酒精引起血管收缩在窦前区通过机制涉及细胞内钙。总之，在这项科学研究中开发的方法不仅是阐明肝脏疾病病理生理学的有用工具，而且是治疗和预防肝脏疾病的有用工具。

The in vivo fluorescence microscopy was developed to visualize the microcirculation (over 2000X) of the liver to elucidate the dynamic change of cellular function in the process of hepatic injury caused by alcohol, drug and virus. The continuous and quantitative measurement was performed for the sinusoidal blood flow. Sinusoidal velocity of red and white blood cells. Oxygen saturation of the sinusoidal blood and so on. The microcirculation of the liver lobule was characterized as follows: The sinusoidal blood flow was faster in the pericentral(PC) region than in periportal(PP) region; there was a gradient of the oxygen saturation of sinusoidal blood from PP to PC area(PP>PC). In addition, the effect of ethanol on the hepatic microcirculation was evaluated in order to clarify the involvement of microcirculatory disturbance in the mechanism of alcoholic liver injury. Low dose alcohol raised both the sinusoidal blood flow and oxygenation of sinusoidal blood, yet the oxygenation of sinusoidal blood fell in some sinusoids of PC region, giving a heterogenic response to alcohol. In contrast, high dose alcohol markedly reduced the oxygenation of sinusoidal blood particularly in PC region, resulting in an increased number of hypoxic regions. Furthermore, sodium fluorescein study using this system showed an impairment in the excretion from hepato-cytes to bile canaliculi particularly in PC area. These data indicate that high dose alcohol produces intralobular shunting and local hypoxia, leading to hepatic dys-function. In the perfused liver, it was demonstrated that alcohol caused a vaso-constriction at presinusoidal region via mechanism involving intracellular calcium. In conclusion, the methodology developed in this scientific research is a useful tool not only for the ellucidation of pathophysiology of the liver disease, but for the treatment and prevention of liver disease.