A SIMPLIFIED MODEL OF HYPOXIC INJURY IN PRIMARY CULTURED RAT HEPATOCYTES

The Anaeropack system for cell culture, which was originally designed for the growth of anaerobic bacteria, was used to produce a hypoxic at mosphere for cultured hepatocytes. We measured changes in the oxygen a nd carbon dioxide concentrations and the atmospheric temperature in an airtight jar We also measured changes in the pH of the medium during hypoxia to assess the accuracy of this system. Moreover, we used three durations (2, 3, and 4 h) of hypoxia and 8 h of reoxygenation in cult ured rat hepatocytes, and then measured the lactate dehydrogenase (LDH ), ketone body concentration (acetoacetate + beta-hydroxybutyrate), an d the ketone body ratio (KBR: acetoacetate/beta-hydroxybutyrate) in th e medium in order to assess the suitability of this system as a model for reperfusion following liver ischemia. The oxygen concentration dro pped to 1% or less within 1 h. The concentration of carbon dioxide ros e to about 5% at 30 min after the induction of the hypoxic conditions; and was maintained at this level for 5 h. No effect of tile reaction heat produced by the oxygen absorbent in the jar was recognized. The e xtent of cell injury produced by changing the hypoxic parameters was s atisfactorily reflected by the KBR, the ketone body concentration, and the LDH activity released into the medium. Because this model can dup licate the conditions of the hepatocytes during revascularization foll owing ischemic liver, and the Anaeropack system for cell culture is ea sy to manipulate, it seems suitable for the experimental study of hypo xic injury and revascularization in vitro.