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To investigate the effects of culture conditions on the maintenance of metabolic functions of cultured hepatocytes, long-term hepatocyte culture lasting 20 days was performed under two different culture conditions, i.e. stationary cultures utilizing porous polymer (polyvinyl formal (PVF) resin) as a substratum and conventional monolayer dish cultures without PVF. Metabolic activities specific to hepatocytes were evaluated in terms of ammonia metabolism, urea synthesis, and albumin secretion. Concerning ammonia metabolic and urea synthetic activities, no significant differences in maintenance of these activities were found between the two culture conditions, and these activities rapidly decreased with the elapse of the culture period, especially during the early stage of the experiments. However, after day 10, these activities in the stationary cultures were maintained at a slightly more favorable level than in the monolayer cultures. On the other hand, compared with ammonia metabolism and urea synthesis, stable and well-maintained albumin secretion of hepatocytes (60% of the activity in day 1) was exhibited in the stationary culture experiments, despite that this particular activity under the monolayer culture condition gradually reduced to a very low level (5.7% of that on day 1) at the end of the culture. From the morphological observations, hepatocytes immobilized in the PVF resin revealed individual spherical shapes without forming multicellular aggregation, and it was suggested that this characteristic structure contributed to good albumin secretion of hepatocytes. In conclusion, the advantages of the hepatocyte culture technique utilizing PVF resin over the conventional dish culture in maintaining some representative metabolic function specific to hepatocytes were clarified.

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White blood cells roll spontaneously in venules of intact, noninflamed rat skin. We investigated noninvasively in two experimental series which leukocyte subtypes participate in this phenomenon and the possible involvement of E-selectin. Male Lewis rats were anesthetized with sodium pentobarbital, and intravital video microscopy was performed on postcapillary venules in the nail-fold of a hind leg. In series 1 acridine yellow was infused for 15 min (50 mg per kg intravenously) to stain the leukocyte nuclei in situ. With the use of fluorescence microscopy rolling leukocytes could be classified unequivocally as polymorphonuclear (granulocytes) or monomorphonuclear (lymphocytes/monocytes) by the shape of their nucleus. Irrespective of vessel depth beneath the skin surface (25-45 microm), most identified rolling leukocytes were classified as granulocytes (72%-100%; median 89%). This percentage was independent of total rolling leukocyte flux, systemic leukocyte count, or their in vitro differentiation pattern. In series 2, rats were treated with either a synthetic, highly selective E-selectin blocking peptide or a control peptide (intravenously, 12 mg peptide per kg bolus, followed by 50 mg per kg per h). E-selectin blockade significantly reduced the leukocyte rolling level to about 50% of baseline (p <0.01), whereas the rolling velocity increased (p <0.01); the control peptide had no effect. In summary, most of the leukocytes rolling spontaneously in postcapillary venules of intact rat skin are granulocytes, despite the absence of an acute inflammatory reaction. One of the adhesion molecules involved in this phenomenon is E-selectin.