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Asymptomatic bile leakage developed in a woman following percutaneous liver biopsy. The case is remarkable because features of cholestasis were not present at the time of biopsy, the bile leak occurred through the needle tract in the liver, billous ascites occurred without signs of acute peritonitis, and the complication was the initial manifestation of extrahepatic biliary obstruction due to encasement of the common bile duct by histiocytic lymphoma.

The major route of protoporphyrin elimination is biliary secretion. To clarify the nature of the secretory process, maximal canalicular secretion of protoporphyrin was determined under basal conditions and after treatment with various choleretics. The maximal secretion of protoporphyrin under basal conditions was 0.07 +/- 0.01 micrograms.min-1.100 g body wt-1. Infusion of physiological amounts of sodium taurocholate increased protoporphyrin secretion 13-fold (0.90 +/- 0.02), primarily by increasing the biliary protoporphyrin concentration. Biliary protoporphyrin secretion tended to plateau in spite of a continued rise in both biliary bile acid secretion and concentration. Infusion of sodium dehydrocholate increased protoporphyrin secretion, but to only 35% of that achieved by sodium taurocholate. Ethacrynic acid and phenobarbital increased bile flow over controls but failed to enhance protoporphyrin transport. Thus, canalicular secretion of protoporphyrin was maximally enhanced by micelle-forming bile acids and unaffected by nonbile acid choleretics. The observed limitation of protoporphyrin secretion may be related to achievement of a canalicular transport maximum or to a toxic effect of protoporphyrin on the transport process.

The pathogenesis of liver disease in protoporphyria has been presumed to result from the hepatic deposition of protoporphyrin. To examine the effects of protoporphyrin on hepatic bile flow and histopathology, studies were performed employing an isolated, in situ, rat liver perfusion system. Rat livers in the control group were perfused with 0-80 mumol sodium taurocholate/h. Rat livers in the experimental group were perfused with sodium taurocholate and (a) sufficient quantities of protoporphyrin to produce maximal canalicular secretion and (b) perfusate protoporphyrin concentrations of 0.01, 0.1, and 1 muM. The administration of protoporphyrin sufficient to achieve maximal canalicular secretion was found to significantly reduce bile flow in rats infused with 0, 40, and 80 mumol sodium taurocholate/h. Linear regression analysis defined the relationship between bile flow and biliary bile acid secretion and showed that the bile acid-independent fraction of bile flow was reduced (P < 0.01). Bile acid-dependent flow was unaffected and there was no significant difference in biliary bile acid secretion rates between control and protoporphyrin-perfused livers. Perfusion of rat livers with varying concentrations of protoporphyrin demonstrated the reduction of bile flow was dose-related. Analysis of perfusate enzyme activity did not reveal abnormalities that could account for the cholestasis. Studies to evaluate the effect of protoporphyrin on regional hepatic hemodynamics were inconclusive. Histopathological studies of control and protoporphyrin-perfused rat livers did not show abnormalities on light microscopy. However, canalicular dilatation, distortion, and loss of microvilli were present in the protoporphyrin-perfused livers examined by transmission electron microscopy. Although ultraviolet microscopy showed diffuse fluorescence of the hepatocytes and canaliculi of protoporphyrin-perfused livers, the deposition of protoporphyrin in amorphous or crystalline forms was notably absent in studies with polarizing and transmission electron microscopy. These studies provide evidence that protoporphyrin has hepatotoxic properties that affect the canalicular secretory apparatus. The mechanism(s) responsible for the injury require further clarification.

Models to explain the pathophysiology of protoporphyria have been based on fluxes of protoporphyrin between plasma and liver for which no quantitative data exist. The present studies employed isolated, in situ, recirculating and nonrecirculating rat liver perfusions to define the kinetics of hepatic uptake and biliary secretion of protoporphyrin. Livers from Sprague-Dawley rats were perfused with Krebs-Henseleit-3% albumin solution (bile acid-free) to which protoporphyrin was added. In nonrecirculation studies, hepatic protoporphyrin extraction was determined from 5 sec to 3 min. In recirculation studies, protoporphyrin perfusate disappearance and biliary secretion were determined at 5 and 10 min intervals, respectively. Rate constants derived from compartmental analysis of recirculation studies correlated with early measured values obtained during nonrecirculation; however, a dose-related decrease in the influx rate constant was evident. The overall disappearance of protoporphyrin from perfusate followed first-order kinetics but was neither monoexponential nor saturable. Sinusoidal cells contained less than 8% of the protoporphyrin extracted by the liver. Only 0.1% to 4% of the extracted protoporphyrin was secreted into bile. Therefore, canalicular secretion appeared to be the rate-limiting step in hepatic protoporphyrin disposal. Extrapolations based on these data provide a theoretical framework to appreciate the generation of plasma protoporphyrin concentrations. Within this framework, it may be inferred that if plasma protoporphyrin concentration is in excess of that predicted from total protoporphyrin excretion, a defect in hepatic protoporphyrin clearance exits.

To further define the pathogenesis of protoporphyric liver disease, perfused rat livers received varying doses of protoporphyrin, and aberrations of hepatic ultrastructure and function were correlated. Results indicated that the relative canalicular volume was equally increased in all protoporphyrin-perfused livers; however, bile flow was only minimally diminished at the smallest protoporphyrin dose employed. Protoporphyrin injured microvilli at the sinusoidal pole and reduced the surface density of the endoplasmic reticulum in a dose-related fashion. No crystalline or amorphous material was detectable, and only slight mitochondrial distortions occurred. Thus, cholestasis did not correlate with canalicular dilatation, the presence of crystalline material, or mitochondrial changes. Liver plasma membrane abnormalities appeared to correlate with functional defects and support a direct hepatotoxicity. Long-term protoporphyrin overload studies are needed to assess differences between hepatic and erythroid (parenteral) sources of protoporphyrin overproduction.

To clarify the pathogenesis of protoporphyrin-induced cholestasis, liver surface membrane enzyme activities were determined after (a) isolated rat liver perfusion with protoporphyrin administered by bolus (1.0 mumol) or bolus plus constant infusion (1.0 mumol + 0.05 mumol/min) and (b) combination of liver surface membrane with protoporphyrin (0.9-53.4 nmol/ml) in vitro. The perfusion studies showed that protoporphyrin significantly inhibited Na+, K+- and Mg+2-adenosine triphosphatase activities. In vitro, these adenosine triphosphatase activities and the 5'-nucleotidase activity were inhibited linearly by protoporphyrin up to a concentration of approximately 18 mumol/ml; thereafter, enzyme activity was maintained. Greater inhibition of the adenosine triphosphatase activities occurred with protoporphyrin than was reported for chlorpromazine at similar molar concentrations. This effect was independent of the quantity of membrane protein analyzed and was not reversible with a 50:1 dilution. The inhibitory effect of protoporphyrin on surface membrane enzyme activities was also nonselective. Although the hepatotoxic effects of protoporphyrin may be more generalized, the present data underscore protoporphyrin's toxic interaction with liver surface membranes.