Bruno Heyd

A twenty-year history of alveolar echinococcosis: analysis of a series of 117 patients from eastern France.

Objectives Alveolar echinococcosis of the liver is a very rare and severe parasitic disease due to the growth of the larva of Echinococcus multilocularis. The aim of this paper was to describe a 20‐year study of the epidemiological, clinical and therapeutic aspects of alveolar echinococcosis in eastern France. Design One hundred and seventeen consecutive cases, diagnosed and followed in our liver unit, were studied from 1972 to 1993. Methods Data from 85 patients followed since 1983 (period B) were compared to data from a first series of 32 patients (period A) collected from 1972 to 1982; 1983 was chosen as the cut‐off year because of the numerous changes that occurred in the diagnosis, follow‐up and treatment of the disease at this time, in particular the introduction of parasitostatic benzimidazoles. Results The results of patient follow‐up were evaluated in December 1997. The cumulative prevalence was 2.5 per 100 000 persons in period A whereas it reached 6.6 per 100 000 in period B. The annual incidence in period B was 7.3 on average, compared with 2.7 in period A. Twenty‐nine per cent of patients from period B were asymptomatic at the time of diagnosis compared with 10% in period A. This change was correlated with less advanced liver lesions, and was related to the extensive use of abdominal ultrasound, and from 1987, serological screening. Curative resections were performed in 24% of the cases in period B versus only 3% in period A. From 1986, liver transplantations were performed in eight patients from period A and 13 patients from period B. In period B, palliative surgery was frequently replaced by radiological non‐operative procedures to treat abscesses and jaundice. From 1982, 73 patients received benzimidazoles for a period of time ranging from 4 to 138 months. Stabilization of the lesions was observed in two‐thirds of the patients. Episodes of jaundice or digestive haemorrhage due to portal hypertension were 31.5 and 11 times less frequent respectively in patients from period B compared with period A. Actuarial survival at 5 years improved from 67% in period A to 88% in period B in patients of similar age. Conclusions Radical changes in the diagnosis and the management of alveolar echinococcosis have occurred during the last decade. Together they have contributed to an improvement in the status of the patients affected by this very severe parasitic disease. Eur J Gastroenterol Hepatol 12:327‐336

Gene expression in human hepatocytes in suspension after isolation is similar to the liver of origin, is not affected by hepatocyte cold storage and cryopreservation, but is strongly changed after hepatocyte plating

Isolated primary human hepatocytes are a well accepted system for evaluating pharmacological and toxicological effects in humans. However, questions remain regarding how culturing affects the liver-specific functions of the hepatocytes. In addition, cryopreservation could also potentially affect the differentiation state of the hepatocytes. The first aim of the present study was to compare gene expression in freshly isolated primary hepatocytes to that of the liver of origin and to evaluate the expression changes occurring after cryopreservation/thawing, both when maintained in suspension and after plating. The second aim of the present study was to evaluate gene expression in hepatocytes after cold storage of suspensions up to 24 h compared with freshly isolated hepatocytes in suspension. Our results show that the gene expression in freshly isolated human hepatocytes in suspension after isolation is similar to that of the liver of origin. Furthermore, gene expression in primary human hepatocytes in suspension is not affected by hepatocyte cold storage and cryopreservation. However, the gene expression is profoundly affected in monolayer cultures after plating. Specifically, gene expression changes were observed in cultured relative to suspensions of human hepatocytes that are involved in cellular processes such as phase I/II metabolism, basolateral and canalicular transport systems, fatty acid and lipid metabolism, apoptosis, and proteasomal protein recycling. An oxidative stress response may be partially involved in these changes in gene expression. Taken together, these results may aid in the interpretation of data collected from human hepatocyte experiments and suggest additional utility for cold storage and cryopreservation of hepatocytes.

Experience of liver transplantation for incurable alveolar echinococcosis: a 45-case European collaborative report.

Background. Alveolar echinococcosis (AE) of the liver is a rare and severe parasitic disease. It behaves like a slow-growing liver cancer, and liver transplantation (LT) has been proposed in advanced cases since 1985. The aim of this retrospective study was to collect all AE transplant cases in Europe, analyze the results, and specify the usefulness of LT for this unusual indication. Methods. A questionnaire was sent to 83 LT centers from July 1996 to December 1999. Results. Sixty-five centers responded: 45 AE patients (mean age, 45.8 years) underwent an LT procedure at 16 LT centers. The mean interval between diagnosis and LT was 5 years. One patient died during the hepatectomy phase. Five-year survival was 71%. Five-year survival without recurrence was 58%. The nine early deaths were mostly related to bacterial or fungal infections, or both, in patients in bad condition when LT was performed. Six patients had a graft AE reinfection. Five late deaths were related directly to ongoing AE. In the other cases, benzimidazole (BZM) therapy seemed to stabilize AE residues. Conclusions. This unique experience indicates that LT is feasible for life-threatening AE. Specific management is needed to optimize the results: earlier decision for LT in incurable symptomatic biliary AE, pre- and post-LT BZM therapy, meticulous pre-LT evaluation to identify extrahepatic extension, and an immunosuppressive regimen kept to a minimum.

Effects of clofibric acid on mRNA expression profiles in primary cultures of rat, mouse and human hepatocytes

The mRNA expression profile in control and clofibric acid (CLO)-treated mouse, rat, and human hepatocytes was analyzed using species-specific oligonucleotide DNA microarrays (Affymetrix). A statistical empirical Bayes procedure was applied in order to select the significantly differentially expressed genes. Treatment with the peroxisome proliferator CLO induced up-regulation of genes involved in peroxisome proliferation and in cell proliferation as well as down-regulation of genes involved in apoptosis in hepatocytes of rodent but not of human origin. CLO treatment induced up-regulation of microsomal cytochrome P450 4a genes in rodent hepatocytes and in two of six human hepatocyte cultures. In addition, genes encoding phenobarbital-inducible cytochrome P450s were also up-regulated by CLO in rodent and human hepatocyte cultures. Up-regulation of phenobarbital-inducible UDP-glucuronosyl-transferase genes by CLO was observed in both rat and human but not in mouse hepatocytes. CLO treatment induced up-regulation of L-fatty acid binding protein (L-FABP) gene in hepatocytes of both rodent and human origin. However, while genes of the cytosolic, microsomal, and mitochondrial pathways involved in fatty acid transport and metabolism were up-regulated by CLO in both rodent and human hepatocyte cultures, genes of the peroxisomal pathway of lipid metabolism were up-regulated in rodents only. An up-regulation of hepatocyte nuclear factor 1alpha (HNF1alpha) by CLO was observed only in human hepatocyte cultures, suggesting that this trans-activating factor may play a key role in the regulation of fatty acid metabolism in human liver as well as in the nonresponsiveness of human liver to CLO-induced regulation of cell proliferation and apoptosis.

Indications and results of liver transplantation for Echinococcus alveolar infection: an overview.

BackgroundAlveolar echinococcosis (AE) of the liver, caused by the larval stage of the fox tapeworm Echinococcus multilocularis, has the characteristics of a slow-growing liver cancer. It is one of the rare parasitic diseases for which a parasitolytic drug is not yet available, and AE is lethal in the absence of appropriate therapeutic management. Complete surgical resection of the parasite at an early stage of infection provides favourable prospects for cure, but, due to a long clinical latency, many cases are diagnosed at an advanced stage, so that partial liver resection can be performed in only 35% of patients. Benzimidazole (BZM) treatment is given in inoperable cases but these compounds are only parasitostatic, and lifelong therapy is required. During the past 20 years some centres have considered liver transplantation (LT) for the treatment of incurable AE.MethodsOur review summarizes the results of this experience based on a series of 47 European patients who received transplants between 1985 and 2002, tries to specify the real place of LT for AE, and underlines the measures that could be undertaken in the future to improve the results.ResultsFive-year survival was 71%. Five-year survival without recurrence was 58%. Major technical difficulties related either to previous laparotomies or to the loco-regional involvement were observed. The nine early deaths concerned AE patients with a long past-history of symptomatic AE (iterative cholangitis, secondary biliary cirrhosis). Five late deaths were directly related to ongoing AE, located in the brain in three cases, a very rare AE location that was not investigated before LT in these patients.ConclusionsIn general, the pre-LT screening for distant AE metastases appeared insufficient in this series. Heavy immunosuppressive schemes, absence or delayed re-introduction of BZM after LT have clearly played a role in this unfavourable course. This unique experience indicates that, despite major technical difficulties, LT for incurable AE is feasible and could be discussed in very symptomatic cases. Before LT, interventional radiology should be preferred to repeated laparotomies. Pre-LT and post-LT BZM treatment is mandatory. A careful evaluation of possible distant metastases should be done before the decision for LT is made. After LT, the possibility of an ongoing AE must be permanently kept in mind. This could be reduced by lightening the immunosuppressants, carefully following the specific circulating antibodies, and applying a systematic radiological evaluation, not only to the graft but also to the lungs and the brain.

Tissue collection, transport and isolation procedures required to optimize human hepatocyte isolation from waste liver surgical resections. A multilaboratory study

Abstract: Background: The European Center for Validation of Alternative Methods (ECVAM) has funded a prevalidation study in three laboratories (France, USA and UK) on the use of human hepatocyte cultures to predict cytochrome P‐450 induction.

Effects of Andrographis paniculata extract and Andrographolide on hepatic cytochrome P450 mRNA expression and monooxygenase activities after in vivo administration to rats and in vitro in rat and human hepatocyte cultures.

The expression of cytochrome P450 (CYP) is regulated by both endogenous factors and foreign compounds including drugs and natural compounds such as herbs. When herbs are co-administrated with a given drug in modern medicine it can lead to drug-herb interaction that can be clinically significant. The ability of Andrographis paniculata extract (APE) and Andrographolide (AND), the most medicinally active phytochemical in the extract, to modulate hepatic CYP expression was examined in vivo in rats and in vitro in rat and human hepatocyte cultures. After in vivo administration, APE at dose levels of 0.5 g/kg/day (i.e. 5 mg/kg/day AND equivalents) and at 2.5 g/kg/day (i.e. 25 mg/kg/day AND equivalents) and AND at dose levels of 5 and 25 mg/kg/day significantly decreased CYP2C11 activity. In primary cultures of rat and human hepatocytes, treatment with AND 50 microM and APE-containing 50 microM AND also resulted in significant decreases in CYP2C expression and activity. In addition, in human hepatocytes, treatment with APE and AND 50 microM resulted in a decrease in CYP3A expression and activity. In conclusion, this study suggests that AND and APE could cause herb-drug interactions in humans through modulation of CYP2C9 and CYP3A4 expression and activities.

Early detection of pancreatic cancer in patients with chronic pancreatitis: diagnostic utility of a K-ras point mutation in the pancreatic juice.

OBJECTIVE:Point mutations of the K-ras oncogene at codon 12 have been described several months before the onset of pancreatic cancer in isolated cases of chronic pancreatitis (CP). The aim of this study was to evaluate the interest of a prospective follow-up of patients with CP and K-ras mutations at codon 12 in the detection of early pancreatic cancer.METHODS:From February 1996 to March 1998, 36 patients (mean age 52.6 yr, 31 men, five women) with CP (alcoholic: 61.1%, pancreas divisum: 5.6%, autoimmune: 5.6%, unknown origin: 27.7%) were included and then prospectively monitored (median duration of 22 months) for detection of pancreatic carcinoma. K-ras point mutations were examined by two-step polymerase chain reaction combined with restriction enzyme digestion in pancreatic juice collected during endoscopic retrograde pancreatography.RESULTS:Ten patients (27.8%) were positive for K-ras mutation. Patients with and without the mutation were not different with respect to age and sex ratio. K-ras mutations were homogeneously distributed according to the etiology (alcoholic vs nonalcoholic) and morphological characteristics (ductal stricture or mass vs none) of CP. A pancreatic carcinoma was discovered at an invasive stage in two patients, respectively at 7 and 17 months after disclosure of a K-ras mutation, versus none in patients without the mutation (p < 0.02).CONCLUSIONS:Presence of a K-ras gene mutation is not rare in patients with CP and represents an increased risk of developing pancreatic cancer. However, its utility for the detection of early pancreatic cancer remains doubtful in clinical practice.

Comparison of clearance predictions using primary cultures and suspensions of human hepatocytes

Various incubation conditions of human hepatocytes were compared for their accuracy in predicting the in vivo hepatic clearance (CLH) of model compounds. The test compounds were the highly cleared, low protein bound naloxone (in vivo CLH = 25 ml min−1 kg−1; free fraction = 0.6), the medium clearance, highly protein bound midazolam (CLH = 12 ml min−1 kg−1; free fraction = 0.04) and the low clearance, highly protein bound bosentan (CLH = 3.9 ml min−1 kg−1; free fraction = 0.02). Each compound was tested in three ‘hepatocyte systems’, using resections from three donors, in the presence and absence of human serum. Those hepatocyte systems were: conventional primary cultures, freshly isolated suspensions and cryopreserved suspended hepatocytes. Except for a twofold overestimated CLH for bosentan from conventional primary cultures, and despite variable cryopreservation recoveries, similar predictions of CLH were recorded with all hepatocyte systems. Moreover, the CLH values obtained with cryopreserved suspended hepatocytes were similar to those obtained with freshly isolated suspensions. For midazolam and bosentan, the predicted in vivo CLH was markedly higher in the presence of serum, whereas serum had little influence on the scaled-up CLH of naloxone. In vivo, CLH was properly approached for naloxone and bosentan (particularly from experiments in the presence of serum), but it was strongly underestimated for midazolam (particularly in the absence of serum). Additional compounds need to be investigated to confirm the above findings as well as to assess why the clearances of some highly protein-bound compounds are still considerably underestimated.

Treatment of ulcerative colitis refractory to steroid therapy by oral microemulsion cyclosporine (Neoral)

Background Intravenous cyclosporine is active in 60% to 80% of patients with ulcerative colitis (UC) who failed to respond to intravenous corticosteroids. Several studies have suggested that cyclosporine in microemulsion form (Neoral) has some efficacy in this setting, but the optimal dose, blood level, time to response, and remission need to be better defined. The aim of this study was to evaluate the response to Neoral and its toxicity in active corticosteroid‐refractory UC. Methods Between March 2002 and August 2005, 20 courses of Neoral [initial dose, 2.3 mg/kg (range, 1.8 to 2.8 mg/kg) every 12 hours] were prescribed in 19 consecutive patients for a UC attack that did not respond to intravenous methylprednisolone. All patients received prophylaxis against Pneumocystis carinii. Results Response was obtained in 17 of 20 attacks (85%) after 3.5 days (range, 1 to 7). Remission was obtained in 15 of 20 attacks (75%) after 13 days (range, 2 to 30 days). Four responders relapsed and underwent colectomy 21 to 900 days after the start of Neoral. Overall, 14 of 19 patients (74%) were colectomy free after a median follow‐up of 8 months (range, 1 to 41 months). Cyclosporine blood levels were measured at fasting (C0) and 2 hours after Neoral administration (C2) in a subgroup of 10 responders. The results were 103 ng/mL (range, 32 to 240 ng/mL) for C0 and 761 ng/mL (183 to 1390 ng/mL) for C2. One severe bedridden patient with neonatal encephalopathy died. Main side effects observed were mild transient renal impairment (n = 2), hypertension (n = 1), cytomegalovirus infection (n = 2), and esophageal candidiasis (n = 1). Conclusions In active corticosteroid‐refractory UC, Neoral seems to have the same efficacy and toxicity as the intravenous form. Trough target cyclosporine blood levels should not exceed 100 ng/mL for C0 and 700 ng/mL for C2.