Focal intestinal lipogranulomatous lymphangitis in 10 dogs.

OBJECTIVES: To describe the clinical and pathological features of canine focal lipogranulomatous lymphangitis, to evaluate its underlying infectious cause and to compare it with human Crohn's disease. METHODS: Retrospective review of case records with a histopathological diagnosis of focal lipogranulomatous lymphangitis. Bacterial and fungal colonisation was evaluated using fluorescence in situ hybridisation and histochemical staining, respectively. A comparison with Crohn's disease was performed by a human pathologist. RESULTS: Ten dogs were evaluated. The historical complaints were predominantly chronic diarrhoea (10/10) and vomiting (5/10). The biochemical abnormalities included hypoalbuminaemia (6/10) and hypocobalaminaemia (4/6). Abdominal sonography revealed a thickened distal ileum±ileocolic junction. Colonoscopy showed a swollen caecal ostium and oedematous caecum in 7/10 dogs. A stenotic ileo-colic opening prevented endoscopic intubation in all dogs. Histology from the resected lesions revealed granulomatous inflammation involving the muscularis and serosa. Fluorescence in situ hybridization demonstrated invasive bacteria in 2/10 dogs. Post-resection, all dogs received metronidazole and tapering immunosuppressive doses of prednisolone. Remission (median 17 months) was achieved in 8/10 dogs. CLINICAL SIGNIFICANCE: Focal lipogranulomatous lymphangitis is a rare and severe form of canine inflammatory bowel disease with preferential localisation to the ileum and the ileocolic junction. An underlying infectious aetiology was not identified.

In vivo infectivity of liver extracts after resolution of hepadnaviral infection following therapy associating DNA vaccine and cytokine genes.

DNA-based vaccination appears of promise for chronic hepatitis B immunotherapy, although there is an urgent need to increase its efficacy. In this preclinical study, we evaluated the therapeutic benefit of cytokine (IL-2, IFN-γ) genes co-delivery with DNA vaccine targeting hepadnaviral proteins in the chronic duck hepatitis B virus (DHBV) infection model. Then, we investigated the persistence of replication-competent virus in the livers of apparently resolved animals. DHBV carriers received four injections of plasmids encoding DHBV envelope and core alone or co-delivered with duck IL-2 (DuIL-2) or duck IFN-γ (DuIFN-γ) plasmids. After long-term (8 months) follow-up, viral covalently closed circular (ccc) DNA was analysed in duck necropsy liver samples. Liver homogenates were also tested for in vivo infectivity in neonatal ducklings. Co-delivery of DuIFN-γ resulted in significantly lower mean viremia starting from week 21. Viral cccDNA was undetectable by conventional methods in the livers of 25% and 57% of animals co-immunized with DuIL-2 and DuIFN-γ, respectively. Interestingly, inoculation of liver homogenates from 7 such apparently resolved animals, exhibiting cccDNA undetectable in Southern blotting and DHBV expression undetectable or restricted to few hepatocytes, revealed that three liver homogenates transmitted high-titre viremia (3-5×10(10) vge/mL) to naïve animals. In conclusion, our results indicate that IFN-γ gene co-delivery considerably enhances immunotherapeutic efficacy of DNA vaccine targeting hepadnaviral proteins. Importantly, we also showed that livers exhibiting only minute amounts of hepadnaviral cccDNA could induce extremely high-titre infection, highlighting the caution that should be taken in occult hepatitis B patients to prevent HBV transmission in liver transplantation context.

Hyperphosphataemia sensitizes renally impaired rats to the profibrotic effects of gadodiamide.

BACKGROUND AND PURPOSE: Hyperphosphataemia is common in patients with nephrogenic systemic fibrosis (NSF). NSF has been linked to administration of gadolinium (Gd) chelates (GCs) and elevated serum phosphate levels accelerate the release of Gd from linear, non-ionic GCs but not macrocyclic GCs. Hence, we determined whether hyperphosphataemia is a cofactor or risk factor for NSF by investigating the role of hyperphosphataemia in renally impaired rats. EXPERIMENTAL APPROACH: Firstly, the clinical, pathological and bioanalytical consequences of hyperphosphataemia were investigated in subtotal nephrectomized (SNx) Wistar rats following i.v. administration of the non-ionic, linear GC gadodiamide (5 × 2.5 mmol·kg(-1) ·day(-1) ). Secondly, the effects of several GCs were compared in these high-phosphate diet fed rats. Total Gd concentration in skin, femur and plasma was measured by inductively coupled plasma mass spectrometry (ICP-MS) and free Gd(3+) in plasma by liquid chromatography coupled to ICP-MS. Relaxometry was used to measure dissociated Gd in skin and bone. KEY RESULTS: Four out of seven SNx rats fed a high-phosphate diet administered gadodiamide developed macroscopic and microscopic (fibrotic and inflammatory) skin lesions, whereas no skin lesions were observed in SNx rats treated with saline, the other GCs and free ligands or in the normal diet, gadodiamide-treated group. Unlike the other molecules, gadodiamide gradually increased the r(1) relaxivity value, consistent with its in vivo dissociation and release of soluble Gd. CONCLUSIONS AND IMPLICATIONS: Hyperphosphataemia sensitizes renally impaired rats to the profibrotic effects of gadodiamide. Unlike the other GCs investigated, gadodiamide gradually dissociates in vivo. Our data confirm that hyperphosphataemia is a risk factor for NSF.

[Protein-losing enteropathy of non neoplastic origin in the dog: a retrospective study of 34 cases]. / Les entéropathies exsudatives d'origine non néoplastique du chien: étude rétrospective de 34 cas.

This study reports the results in 34 dogs with protein-losing enteropathy (PLE). Rottweilers and Yorkshire Terriers were overrepresented. Most obvious clinical signs and laboratory findings were diarrhea (91 %), weight loss (74 %), anorexia (56 %), lethargy (51 %), ascites (18 %), ascites without gastrointestinal signs (9 %), hypoalbuminemia (<12 g/l, 65 %) and hypocholesterolemia (51 %). Ultrasonographic findings included intestinal wall thickening in 71 %, abnormal echogenicity in 68 %, and no abnormalities in 29 %. The most important endoscopic findings were diffuse and profound dilation of villi in 35 %, a more granular appearing mucous membrane in 32 % and a "rice-grain" appearing villus dilation in 12 %. In 21 % of the dogs the mucous membrane was considered unremarkable. Histologically, 62 % had lymphangiectasia, in 86 % of these associated with moderate to severe inflammatory infiltrates, and in 71 % associated with dilatation of intestinal crypts. In 68 % of the dogs lesions in intestinal crypts were found, 35 % of these without lymphangiectasia. In 12 % of the dogs the histological findings failed to explain intestinal protein loss. This study shows the important prevalence of crypts' lesions in protein-losing enteropathy.

Circulating fibrosis markers, eosinophil cationic protein and eosinophil protein X in patients with Wuchereria bancrofti infection: association with clinical status.

We measured the concentrations of several circulating fibrosis markers (type I collagen I, type III procollagen, hyaluronan) and eosinophil granule proteins (ECP and EPX) in lymphatic filariosis patients to investigate their relationship with clinical, parasitological and immunological data. This study was conducted in Polynesian patients with various stages of the disease (acute lymphangitis, chyluria, hydrocoele, elephantiasis), a closely related microbial lymphangitis and endemic controls. We observed modifications of the different markers in this pathology. Serum type I collagen and PIIINP were decreased. Serum hyaluronan, linked to perilymphatic granulomatous inflammation, was significantly increased in acute lymphangitis and elephantiasis patients. Serum ECP was also increased, at the limit of significance in our sample, in elephantiasis patients. These two last markers, already validated in another helminth disease, schistosomiasis, have potential interest in terms of follow-up of morbidity in these parasitic diseases.

Ultrasound detection of spontaneous hepato-cellular carcinomas in X/myc bitransgenic mice.

AIMS: To evaluate trans-abdominal ultrasound for the detection of Hepatocellular carcinoma (HCC) in a bitrasgenic murine (X/myc) model using a commercially available high-frequency ultrasound unit. METHODS: Sixty-one female animals were included in this study. These animals were submitted to a single ultrasound examination of the liver under general anesthesia (isoflurane), and then euthanized. Results of ultrasound were compared with necropsy and histopathology. RESULTS: The lesions demonstrated a fairly consistent aspect (oval- or round-shaped, well-defined hypoechoic homogeneous lesions), and lesions as small as 2 mm were identified. For detection of hepatic nodules per mouse the sensitivity was 75%, the specificity was 100% and the accuracy was 88.5%. For detection of hepatic focal lesions per lesions the overall sensitivity was 60%, the specificity was 97%, and the accuracy was 75.9%. Contrast-enhanced harmonic ultrasound imaging did not improve the identification of the lesions in our experimental conditions. CONCLUSION: High-frequency ultrasound appears to be an efficient tool allowing new possibilities to use this animal model and evaluate new therapies in longitudinal studies, which are much more powerful.

Effect of a combination of clevudine and emtricitabine with adenovirus-mediated delivery of gamma interferon in the woodchuck model of hepatitis B virus infection.

Our aim was to evaluate the antiviral effect of a combination of two nucleoside reverse transcriptase inhibitors, emtricitabine (FTC) and clevudine (L-FMAU), with the addition of an adenovirus-driven delivery of recombinant gamma interferon (IFN-gamma) in the woodchuck model of hepatitis B virus infection. Six woodchuck hepatitis virus (WHV)-infected woodchucks received L-FMAU (10 mg/kg) plus FTC (30 mg/kg) intraperitoneally for 8 weeks; six other animals received in addition an intravenous injection of a recombinant adenovirus vector expressing woodchuck IFN-gamma (Ad-IFN) at weeks 4 and 8. In the control group, two animals received Ad-IFN alone, two received adenovirus vector expressing the green fluorescent protein reporter gene, and one remained untreated. In less than 2 weeks, all woodchucks that received L-FMAU plus FTC showed a rapid and marked inhibition of viral replication, with a 4-log(10) drop in serum WHV DNA. In two animals, viremia remained suppressed for several months after the end of treatment. Similarly, a dramatic decrease in intrahepatic replicative intermediates of viral DNA was observed in the L-FMAU/FTC-treated groups. The additional administration of Ad-IFN led to increased inflammation in the liver but did not enhance the antiviral effect of the L-FMAU/FTC combination. In conclusion, therapies combining L-FMAU and FTC in WHV-infected woodchucks resulted in a potent and sustained antihepadnaviral effect both in the liver and in the blood circulation. However, no extra benefit of adding IFN-gamma gene transduction to the L-FMAU/FTC combination could be detected.

Influence of medium composition, static and stirred conditions on the proliferation of and matrix protein expression of bovine articular chondrocytes cultured in a 3-D collagen scaffold.

Interest in chemical and physical modifications of culture conditions and composition, as a way to improve engineered cartilage, has grown over the last decade. To address some of these aspects, articular bovine chondrocytes seeded in collagen sponges (2.3x10(6) cells/cm(3), whose growth and metabolism have been previously reported) were grown under static or stirred conditions (orbital shaker at 30 rpm), in either 10% FCS-supplemented or serum-free media (1% ITS+1mM cysteine). Under stirred conditions, we observed a 2-fold increase in both cell proliferation and sulphated glycosaminoglycan deposition after 1 month of culture, compared to static conditions, and after 3 months, a more homogeneous distribution of both cells and neomatrix in the constructs. During the first month of culture, the substitution of FCS by ITS led to low cell proliferation and poor neomatrix deposition but, after 2 months a steep increase was observed with ITS for these two parameters that reached, after 3 months the levels observed with FCS. Aggrecan was the more abundant component at both gene and protein levels, whereas the collagenous network formed was looser than with FCS. In conclusion, the use of these simple culture conditions should improve, in long-term culture, the quality of the cartilage construct.

Inhibition of hepadnaviral replication by polyethylenimine-based intravenous delivery of antisense phosphodiester oligodeoxynucleotides to the liver.

Antisense oligodeoxynucleotides (ODNs) appear as attractive anti-hepatitis B virus (HBV) agents. We investigated in vivo, in the duck HBV (DHBV) infection model, whether linear polyethylenimine (lPEI)-based intravenous delivery of the natural antisense phosphodiester ODNs (O-ODNs) can prevent their degradation and allow viral replication inhibition in the liver. DHBV-infected Pekin ducklings were injected with antisense O-ODNs covering the initiation codon of the DHBV large envelope protein, either in free form (O-ODN-AS2) or coupled to lPEI (lPEI/O-ODN-AS2). Following optimization of lPEI/O-ODN complex formulation, complete O-ODN condensation into a homogenous population of small (20-60 nm) spherical particles was achieved. Flow cytometry analysis showed that lPEI-mediated transfer allowed the intrahepatic delivery of lPEI/O-ODN-AS2 to increase three-fold as compared with the O-ODN-AS2. Following 9-day therapy the intrahepatic levels of both DHBV DNA and RNA were significantly decreased in the lPEI/O-ODN-AS2-treated group as compared with the O-ODN-AS2-treated, control lPEI/O-ODN-treated, and untreated controls. In addition, inhibition of intrahepatic viral replication by lPEI/O-ODN-AS2 was not associated with toxicity and was comparable with that induced by the phosphorothioate S-ODN-AS2 at a five-fold higher dose. Taken together, our results demonstrate that phosphodiester antisense lPEI/O-ODN complexes specifically inhibit hepadnaviral replication. Therefore we provide here the first in vivo evidence that intravenous treatment with antisense phosphodiester ODNs coupled to lPEI can selectively block a viral disease-causing gene in the liver.

The role of duck hepatitis B virus and aflatoxin B1 in the induction of oxidative stress in the liver.

The aim of our study was to use the Pekin duck model to investigate the interactions between hepadnaviral infection and aflatoxin B1 (AFB1) exposure including the role of both factors in the induction of oxidative stress in the liver. AFB1 exposure of duck hepatitis B virus (DHBV) infected Pekin ducks induced a significant increase in viral replication associated with an intense biliary ductular cells proliferation. Interestingly, extremely high levels of AFB1-DNA adducts (40-120 pmol AFB1-Fapy/mg DNA) and AFB1-albumin adducts (1,500-3,000 pg AFB1-lys Eq/mg albumin) were detected in duck liver and serum respectively, as compared to other animal species exposed to a similar AFB1 dose. DHBV infection was found to induce a non-significant increase in AFB1-albumin adduct levels in duck serum. During the treatment duration there was no effect on formation of oxidative base damage within DNA and no effect on oxidative lipid peroxidation following either viral infection or AFB1 exposure. In terms of hepatic antioxidant enzymes (catalase, superoxide dismutase (SOD), glutathione peroxidase) a significant increase in SOD activity occurred following AFB1 exposure, but not DHBV infection, but this was observed only after the cessation of treatment, when biliary ductular cells proliferation was reduced.

Antiviral activity of beta-L-2',3'-dideoxy-2',3'-didehydro-5-fluorocytidine in woodchucks chronically infected with woodchuck hepatitis virus.

The L-nucleoside analog beta-L-2',3'-dideoxy-2',3'-didehydro-5-fluorocytidine (beta-L-Fd4C) was first shown to exhibit potent activity against hepatitis B virus (HBV) in tissue culture and then to significantly inhibit viral spread during acute infection in the duck HBV model (F. Le Guerhier et al., Antimicrob. Agents Chemother. 44:111-122, 2000). We have therefore examined its antiviral activity in a mammalian model of chronic HBV infection, the woodchuck chronically infected with woodchuck hepatitis virus (WHV). Side-by-side comparison of beta-L-Fd4C and lamivudine administered intraperitoneally during short-term and long-term protocols demonstrated a more profound inhibition of viremia in beta-L-Fd4C-treated groups. Moreover, beta-L-Fd4C induced a marked inhibition of intrahepatic viral DNA synthesis compared with that induced by lamivudine. Nevertheless, covalently closed circular (CCC) DNA persistence explained the lack of clearance of infected hepatocytes expressing viral antigens and the relapse of WHV replication after drug withdrawal. Liver histology showed a decrease in the inflammatory activity of chronic hepatitis in woodchucks receiving beta-L-Fd4C. An electron microscopy study showed the absence of ultrastructural changes of hepatic mitochondria, biliary canaliculi, and bile ducts. However, a loss of weight was observed in all animals, whatever the treatment, as was a transient skin pigmentation in all woodchucks during beta-L-Fd4C treatment. There was no evidence that lamivudine or beta-L-Fd4C could prevent the development of hepatocellular carcinoma with the protocols used. These results indicate that beta-L-Fd4C exhibits a more potent antiviral effect than lamivudine in the WHV model but was not able to eradicate CCC DNA and infected cells from the liver at the dosage and with the protocol used.

Hepatic fibrosis in rheumatoid arthritis patients treated with methotrexate: application of a new semi-quantitative scoring system.

OBJECTIVE: Evaluation of hepatic lesions in patients treated with methotrexate (MTX) generally used the Roenigk histological score. However, the sensitivity of the method for hepatic fibrosis assessment has been discussed. The semi-quantitative histological scoring system (SSS) offers a sensitive and specific evaluation of liver fibrosis. Both scores have been evaluated in liver biopsies of patients with rheumatoid arthritis. METHODS: Seventy-four liver biopsies were obtained in 57 rheumatoid arthritis patients before initiation of MTX (group 1, 38 cases), in cases of a persistently high level of transaminases during 1 yr of treatment (group 2, 10 cases) and after a MTX total dose of 2 g (group 3, 26 cases). Eleven biopsies of groups 1 and 3 originated from the same patient in 11 cases. Specimens were examined blindly by two anatomopathologists. The three groups were compared with an ANOVA. Sequential biopsies performed in 11 patients were compared with the Wilcoxon paired test. RESULTS: The Roenigk score and the SSS were significantly correlated (P<0.0001). Only a mild fibrosis was found in 33.8% (25/74) of the biopsies with the Roenigk score. Liver fibrosis, graded as mild (48.6%), moderate (41.8%) or severe (4%), was demonstrated in 94.6% (70/74) of the biopsies with the SSS. The Roenigk score and the SSS of the three patient groups were not statistically significantly different. The scores did not progress in the 11 patients who had serial biopsies. CONCLUSION: SSS is much more sensitive than the Roenigk score for the evaluation of hepatic fibrosis. However, SSS did not show progression of hepatic fibrosis in patients with rheumatoid arthritis treated with MTX.

Characterization of the antiviral effect of 2',3'-dideoxy-2', 3'-didehydro-beta-L-5-fluorocytidine in the duck hepatitis B virus infection model.

A novel L-nucleoside analog of deoxycytidine, 2',3'-dideoxy-2', 3'-didehydro-beta-L-5-fluorocytidine (beta-L-Fd4C), was recently shown to strongly inhibit hepatitis B virus (HBV) replication in the 2.2.15 cell line. Therefore, its antiviral activity was evaluated in the duck HBV (DHBV) infection model. Using a cell-free system for the expression of the DHBV polymerase, beta-L-Fd4C-TP exhibited a concentration-dependent inhibition of dCTP incorporation into viral minus-strand DNA with a 50% inhibitory concentration of 0.2 microM which was lower than that of other tested deoxycytidine analogs, i.e. , lamivudine-TP, ddC-TP, and beta-L-FddC-TP. Further analysis showed that beta-L-Fd4C-TP is likely to be a competitive inhibitor of dCTP incorporation and to cause premature DNA chain termination. In primary duck hepatocyte cultures infected in vitro, beta-L-Fd4C administration exhibited a long-lasting inhibitory effect on viral DNA synthesis but could not clear viral covalently closed circular DNA (CCC DNA). Results of short-term antiviral treatment in experimentally infected ducklings showed that beta-L-Fd4C exhibited the most potent antiviral effect, followed by beta-L-FddC, lamivudine, and ddC. Longer administration of beta-L-Fd4C induced a sustained suppression of viremia (>95% of controls) and of viral DNA synthesis within the liver. However, the persistence of trace amounts of viral CCC DNA detected only by PCR was associated with a recurrence of viral replication after drug withdrawal. In parallel, beta-L-Fd4C treatment suppressed viral antigen expression within the liver and decreased intrahepatic inflammation and was not associated with any sign of toxicity. Our data, therefore, demonstrate that in the duck model of HBV infection, beta-L-Fd4C is a potent inhibitor of DHBV reverse transcriptase activity in vitro and suppresses viral replication in the liver in vivo.

Long-term administration of interferon-alpha in non-responder patients with chronic hepatitis C: follow-up of liver fibrosis over 5 years.

In chronic hepatitis C, previous data have shown that short-term treatment with interferon-alpha (IFN-alpha) can reduce collagen deposition in the liver independently of the viral response. The aim of this work was to determine, in non-responder patients, the long-term effect of IFN-alpha on liver fibrosis according to the total administered dose and the fibrotic stage. Fibrosis was investigated on liver biopsies from 24 non-responder patients with chronic hepatitis C retreated with successive courses of IFN-alpha. The degree of liver fibrosis was assessed on three successive biopsies, performed before IFN-alpha treatment and 1 and 5 years later, in 13 and 11 patients, respectively, treated for less (mean: 7.5 months, 313 MU) and more (mean: 21.8 months, 791 MU) than 1 year. For each biopsy, fibrosis was assessed using a histological semiquantitative fibrosis scoring system and by morphometry after picrosirius red staining. Regardless of the dose and duration of IFN-alpha therapy, a slight decrease of fibrosis was observed in patients 5 years after starting treatment. In cirrhotic patients, a short treatment induced an improvement followed by a relapse of fibrosis in 57%, and only 43% of patients showed constant collagen regression over the 5 years of follow-up. On the contrary, after prolonged therapy, a progressive and significant decrease occurred throughout the follow-up period in all patients (P = 0.045). Long-term treatment with IFN-alpha is therefore associated with regression of liver fibrosis, particularly in cirrhotic patients. These promising results need to be confirmed in a larger series of patients.

Enhanced duck hepatitis B virus gene expression following aflatoxin B1 exposure.

Epidemiological studies have suggested synergistic interactions between chronic hepatitis B virus (HBV) infection and aflatoxin B1 (AFB1) exposure in the etiology of hepatocellular carcinoma (HCC), although the molecular mechanisms of their interactions are still not understood. The aim of this study was to use the Pekin duck model to investigate the impact of AFB1 exposure on duck hepatitis B virus (DHBV) replication during the early stages of virus-carcinogen interactions. Six-week-old chronic DHBV-carrier or uninfected ducks were exposed to AFB1 for 5 weeks or treated with dimethylsulfoxide (DMSO) as a control. Animals were observed for 6 to 13 weeks after AFB1 treatment to study the influence of AFB1 exposure on DHBV replication and liver pathologies. Histological analysis showed more marked changes in the livers of AFB1-treated ducks, and these were enhanced by DHBV infection. A significant increase in serum and liver DHBV DNA level was observed in AFB1-treated ducks as compared with DMSO-treated controls. In addition, viral RNAs, in particular the pregenomic RNA that is the template of viral replication, and intrahepatic DHBV DNA replicative intermediates, were significantly increased by AFB1 treatment. Moreover, an overexpression and accumulation of DHBV large envelope (L) protein was observed in the hepatocytes of AFB1-exposed animals. The in vitro study has further confirmed an increase in intracellular viral DNA and in virus release in AFB1-treated primary duck hepatocytes. Taken together, our results indicate that AFB1 exposure leads to an increase in virus gene expression associated with intrahepatic accumulation of DHBV L protein and enhanced liver pathology.

Myofibroblasts in schistosomal portal fibrosis of man.

Myofibroblasts, cells with intermediate features between smooth muscle cells and fibroblasts, have been described as an important cellular component of schistosomal portal fibrosis. The origin, distribution and fate of myofibroblasts were investigated by means of light, fluorescent, immunoenzymatic and ultrastructural techniques in wedge liver biopsies from 68 patients with the hepatosplenic form of schistosomiasis. Results demonstrated that the presence of myofibroblasts varied considerably from case to case and was always related to smooth muscle cell dispersion, which occurred around medium-sized damaged portal vein branches. By sequential observation of several cases, it was evident that myofibroblasts derived by differentiation of vascular smooth muscle and gradually tended to disappear, some of them further differentiating into fibroblasts. Thus, in schistosomal pipestem fibrosis myofibroblasts appear as transient cells, focally accumulated around damaged portal vein branches, and do not seem to have by themselves any important participation in the pathogenesis of hepatosplenic schistosomiasis.

Echinococcus multilocularis: relationship between susceptibility/resistance and liver fibrogenesis in experimental mice.

To analyze collagen and other matrix protein deposits in experimental alveolar echinococcosis as well as the expression of lysyl oxidase, the enzyme that initiates the first steps in the pyridinoline cross-linking of collagen, and to establish a relationship between resistance/susceptibility to Echinococcus multilocularis larval growth and fibrogenesis, we compared AKR/J mice (susceptible to E. multilocularis infection) with NMRI mice (resistant hosts) in this study. Collagen deposits in the lesions were evaluated using a colorimetric method; the nature of matrix proteins involved in the periparasitic fibrosis and lysyl oxidase expression were assessed using immunostaining on tissue sections. The results obtained in this sequential study confirm that fibrogenesis is an important aspect of the host immune reaction against parasitic development and that both the extent and the course of matrix protein deposition differ in the liver of susceptible and resistant mice, respectively. The long-lasting expression of alpha-actin and lysyl oxidase by host cells in NMRI mice suggests that in this resistant strain, fibrosis was not only more developed but also more highly cross-linked and, thus, less sensitive to collagenases than in susceptible mice. A very strong expression of lysyl oxidase by parasitic cells was observed in both strains of mice; the observation that E. multilocularis itself has a role in lysyl oxidase cross-linking of host collagens can be hypothesized and would be a new example of parasite-host interplay.

Changes in liver fibrosis at the end of alpha interferon therapy and 6 to 18 months later in patients with chronic hepatitis C: quantitative assessment by a morphometric method.

BACKGROUND/AIMS: The aims of the study were to determine, in patients with chronic hepatitis C treated with alpha interferon: (i) changes in the morphometric evaluation of liver fibrosis at the end of treatment and 6, 12 and 18 months after treatment; (ii) the predictive value of histologic lesions for the response to treatment, in particular the predictive value of morphometric evaluation of liver fibrosis. METHODS: Seventy patients with chronic hepatitis C who participated in two trials of recombinant interferon alpha 2b treatment were studied. Liver specimens were obtained before and at the end of treatment and 6, 12 or 18 months later. Histologic lesions were assessed according to the Knodell system. Quantitative study of total fibrosis and of Disse space collagen was done by the computerized automated morphometric method. RESULTS: A significant decrease in morphometric Disse space collagen was observed at the end of treatment and 6 months later. This decrease was also observed, although it was not significant, 12 and 18 months after treatment. There was no relationship between this decrease and the biochemical and virological responses or the dose of interferon. The pretreatment Knodell activity score, but not the morphometric evaluation of fibrosis, was a significant predictor of sustained response. CONCLUSION: A decrease in Disse space collagen, as assessed by the sensitive morphometric method, was observed at the end of and 6 months after treatment. This observation is consistent with an anti-fibrogenetic effect of alpha interferon. Mild or moderate histologic activity was associated with a sustained response to therapy.

Liver fibrosis in unisexual murine Schistosomiasis: quantitative study and morphological changes in mice with chronic infection.

The main complication of Schistosomiasis is the development of liver fibrosis as a result of periovular granuloma in portal tract. This study was undertaken to evaluate the role of the worms in the liver, using the unisexual murine schistosomiasis model. Hepatic fibrosis and histopathological lesions were sequentially followed for 65 weeks in mice experimentally infected with male Schistosoma mansoni. Morphological study revealed that, from the 25th week post-infection, a diffuse fibrosis affected the main branches of the portal vascular system following the host inflammatory reaction, associated with the proliferation of myofibroblasts in situ. Quantitative methods confirmed that an increase of fibrotic deposit occurred during chronic unisexual infection from 25 to 50 weeks post-infection, as compared to controls (35.06 +/- 3.55% versus 27.41 +/- 2.56%) suggesting that antigenic substances secreted by adult schistosomes, in the absence of any eggs, might initiate periportal and perisinusoidal fibrous reaction. The cytokine production and the evolution of inflammation towards fibrosis involves stellate cells and myofibroblasts stimulation. This fibrotic process consequently traduces a reparative phenomenon of the liver.

Quantitative studies on liver fibrosis and alpha-smooth muscle actin expression in heroin abusers.

Lobular hepatic fibrosis and the presence of myofibroblasts were studied in heroin abusers, by quantitative automatic image analysis. Nineteen addicts (DA) and thirteen patients having stopped consumption (exDA) were compared to a non-addict group (CONTROL). Addicts, all anti-HIV and HBsAg negative, showed increased transaminase levels. Hepatitis C markers were ot available, at the time of biopsy. The surface of the centrolobular fibrosis, measured on picrosirius stained slides, was respectively 1.9 and 3.5 times larger in DA and exDA than in CONTROL (p < 0.0001). Immunolabelling with an alpha-smooth muscle actin antibody (alpha-SMA) revealed stellate cells in a perisinusoidal location, mainly in areas of matrix thickening in the space of Disse. Morphometric analysis of alpha-SMA expression showed significant differences between the three groups of patients, p < 0.0001 (CONTROL: 198.06 +/- 5.59 microns2; DA: 2227.91 +/- 88.02 microns2; exDA: 3469.10 +/- 154.98 microns2). The surface density of collagen and of alpha-SMA reactivity was also significantly different between these groups (p < 0.0001). These data strongly suggest that heroin is responsible for an early and progressive centrolobular liver fibrosis, occurring simultaneously with a myofibroblastic response. It might represent a reparative phenomenon arising from a direct vascular injury, leading to an impairment of blood-hepatocyte exchange.