Mona M. Nosseir

Immunohistochemical expression of CD95 (Fas), c‐myc and epidermal growth factor receptor in hepatitis C virus infection, cirrhotic liver disease and hepatocellular carcinoma

Gene product expression in normal and chronic hepatitis C virus infection was determined in an attempt to improve our understanding of the molecular events leading to the development of cirrhosis and liver carcinoma. Activation of CD95 (Fas) causes apoptosis of cells and liver failure in mice and has been associated with human liver disorders. c‐myc is involved in cell proliferation and EGFR in regeneration of cells. The material of the current study included 50 cases of chronic hepatitis C (CHC) (and negative hepatitis B virus infection), 29 cases of liver cirrhosis and HCV (LC), and 19 cases of hepatocellular carcinoma and HCV (HCC) admitted to the Theodor Bilharz Research Institute (TBRI) during the years 2003–2004. Ten wedge liver biopsies – taken during laparoscopic cholecystectomy – were included in the study as normal controls. Laboratory investigations, including liver function tests, serological markers for viral hepatitis and serum alpha fetoprotein level (α‐FP), were determined for all cases. Histopathological study and immunohistochemistry using monoclonal antibodies for CD95, c‐myc and EGFR were also done. In CHC cases, the histological activity index (HAI) revealed more expression of Fas antigen in liver tissues with active inflammation than in those without active inflammation (p<0.01). EGFR and c‐myc act synergistically in liver tumorigenesis. Upregulation of Fas in chronic hepatitis C infection and of c‐myc & EGFR in malignant transformation was concluded from this study. c‐myc expression may obstruct the induction of apoptosis of HCC cells and lead to uncontrolled cell growth.

Immunological and parasitological parameters after treatment with dexamethasone in murine Schistosoma mansoni

This work aimed to evaluate the effect of diphenyl dimethyl bicarboxylate (DDB) and dexamethasone alone and in combination with praziquantel on various parasitological, immunological and pathological parameters reflecting disease severity and morbidity in murine schistosomiasis. DDB and dexamethasone had no effect on worm burden but altered tissue egg distribution. This indicates that, under the schedule used, neither drug interfered with the development of adult worms or oviposition, but both can modulate liver pathology. Dexamethasone resulted in a greater reduction in granuloma size than did DDB. Dexamethasone-treated mice also showed lower levels of serum gamma interferon (IFN-γ), interleukin-12 (IL-12) and IL-4, together with higher IL-10 levels, than infected untreated control animals. These data suggest that dexamethasone is a convenient and promising coadjuvant agent that results in decreased morbidity in murine schistosomiasis.

Pharmacodynamics of pentoxifylline and/or praziquantel in murine schistosomiasis mansoni.

Pentoxifylline (PTX) was proved to exert both anti‐inflammatory and anti‐fibrotic effects, and was used therapeutically in this experimental model to investigate its role alone or with praziquantel (PZQ) in Schistosoma mansoni‐infected mice, and to explore its impact on the tissue expression of transforming growth factor‐β1 (TGF‐β1). S. mansoni‐infected mice were divided into seven groups: Control untreated (I), treated with curative dose of PZQ, 500 mg/kg/day for 2 consecutive days (II), or subcurative dose, 100 mg/kg/day for 2 consecutive days (III), treated with PTX (10 mg/kg/day for 5 days/wk) alone for 4 weeks (IV) or in addition to subcurative dose of PZQ (V), and treated with PTX alone for 8 weeks (VI) or in addition to subcurative dose of PZQ (VII). All animals were killed 10 weeks post infection. Parasitological assessment of worm burden, tissue egg load and oogram pattern was carried out. The degree of granulomatous fibrosis and eosinophilic cell population was quantified in Sirius‐red‐stained sections and tissue transforming growth factor beta‐1 expression was estimated immunohistochemically. Serum ALAT and GGT, as well as hepatic content of reduced GSH, were measured. The results revealed the highest percent of worm reduction and dead ova in groups (II) and (VII) accompanied by significant diminution in granulomatous parameters, collagen content and TGF‐β1 tissue expression. Moreover, treatments with PTX and/or PZQ ameliorated the liver functions. In conclusion, prolonged treatment with PTX has a potent anti‐fibrogenic role especially when used in the early stages of infection, with limited toxic effects on schistosome worms and eggs. Thus, PTX can be used as an adjuvant therapeutic tool with anti‐helminthic drugs in the treatment of human schistosomiasis.

Cyclooxygenase-2 expression on urothelial and inflammatory cells of cystoscopic biopsies and urine cytology as a possible predictive marker for bladder carcinoma

Cyclooxygenase‐2 (COX‐2) is a key inducible enzyme involved in the production of prostaglandins. It contributes to human carcinogenesis by various mechanisms. The aim of the current study was to elucidate the possible involvement of COX‐2 in human bladder carcinoma by examining its expression on both urothelial and inflammatory cells in tissue biopsies and urine cytology samples of different urinary bladder lesions. A total of 65 patients were included in the study and were selected from cases admitted to Urology Department, Theodor Bilharz Research Institute (TBRI), Giza, Egypt. They represented seven control cases with almost normal‐looking bladder tissue; pure chronic cystitis (n=12); premalignant lesions (18) in the form of squamous metaplasia (n=8) or urothelial dysplasia (n=10) as well as transitional cell carcinoma (TCC) (n=18), and squamous cell carcinoma (SqCC) (n=10). Immunohistochemistry of formalin‐fixed, paraffin‐embedded tissue sections and urine cytology samples was performed for all cases using COX‐2 (H‐62): sc‐7951, a rabbit polyclonal antibody. The study revealed positive COX‐2 expression on the urothelial and inflammatory cells of cystoscopic biopsies from all cases of pure chronic cystitis, squamous metaplasia and SqCC compared with 42.8% and 71.4% of normal controls, respectively. The score of urothelial COX‐2 expression was sequentially up‐regulated from normal to chronic cystitis (either pure or associated with premalignant changes) (p<0.05) to malignant changes (p<0.05). However, the inflammatory cellular expression was down‐regulated with malignant transformation compared with chronic cystitis (p<0.05). In TCC, COX‐2 was over‐expressed on both urothelial and inflammatory cells in advanced tumors. Urine cytology samples were positive for COX‐2 in a comparable manner to that observed in cystoscopic biopsies. Accordingly, the results of the current study have provided new information in two aspects: First, is the possibility of using the differential COX‐2 expression on both inflammatory and urothelial cells as markers for premalignant or malignant transformation; second, besides cystoscopy, urine cytology was found to have a high sensitivity for COX‐2 expression and hence proved to be valuable in malignancy as a non‐invasive substitute for cystoscopy.

Hematopoietic stem cell mobilization into the peripheral circulation in patients with chronic liver diseases

The present study was aimed to investigate and compare the kinetics of bone marrow‐derived hematopoietic stem cells (BMHSC) migration in the peripheral blood and liver in response to liver injury in patients with chronic liver disease (CLD).

Does granulocyte-colony stimulating factor administration induce damage or repair response in schistosomiasis?

AIM To introduce Granulocyte-colony stimulating factor (G-CSF) as a new therapeutic modality for schistosomiasis through stem cell mobilization, immunomodulation or fibrosis remodeling. METHODS In this study, a 5 d course of human recombinant G-CSF (100 μg/kg sc) was applied to Schistosoma mansoni-infected mice at different stages of disease (5 d before infection as well as 3, 5 and 7 wk post-infection). The animals were sacrificed at 10 d as well as 4, 6 and 8 wk post infection. Mice were examined for: (1) Total leukocyte count which is an accepted surrogate marker for the stem cell mobilization into the circulation; (2) Egg count in intestine and liver tissue to assess the parasitic load; and (3) Histopathological changes in Hx/E and Masson trichrome stained sections as well as collagen content in Sirius red-stained liver sections to determine the severity of liver fibrosis. RESULTS Mice developed leukocytosis. The egg load and the number of granulomas were not affected by the G-CSF treatment but there was an obvious change in the composition of granulomas towards an increased cellularity. Moreover, fibrosis was significantly decreased in treated groups compared to untreated animals (collagen content either preinfection or at 3 and 5 wk post infection: 5.8 ± 0.5, 4.7 ± 0.5, 4.0 ± 0.7 vs 8.2 ± 0.9; P ≤ 0.01). CONCLUSION Although G-CSF did not cause direct elimination of the parasite, it enhanced granulomatous reaction and reduced the fibrosis. Further investigation of the underlying mechanisms of these two actions is warranted.

Value of reelin for assessing hepatic fibrogenesis in a group of Egyptian HCV infected patients

Abstract Background: Development of non-invasive markers that can predict the stages of hepatic fibrosis without resorting to repeated liver biopsies is still an important goal to evaluate the effectiveness of antifibrotic treatment. The present work investigates the value of the assessment of peripheral circulating reelin, in which the liver represents its prime source, as a marker for monitoring hepatic fibrogenesis. Methods: Seventy-four cases with chronic hepatitis positive for serum HCV RNA and 15 healthy volunteers were enrolled in this study. Assessment of reelin in the harvested serum and in 64 corresponding liver biopsies using immunofluorescence technique was done. The results were evaluated in relation to the stages and quantitative morphometric analysis of hepatic fibrosis as well as the serum levels of the validated biomarker hyaluronic acid. Results: Significant correlation was detected between the levels of serum reelin and the semiquantitative assessment of reelin immunoreactivity in liver tissue, the stages of hepatic fibrosis, the morphometrically determined collagen and serum hyaluronic acid with a correlation coefficient of 0.675, 0.623, 0.479, 0.772, respectively with p<0.001. The sensitivity and the specificity of reelin for the determination of advanced (F2+F3) and significant fibrosis (F2–F4) were nearly comparable to the result of hyaluronic acid. In addition the area under curve (AUC) were 0.859, 0.871 for the reelin versus 0.878, and 0.891 for the hyaluronic acid. Conclusions: In conclusion serum reelin may be considered an additional useful parameter for monitoring the progression of hepatic fibrosis in HCV-infected patients specially in those with active rheumatological conditions which result in an increase in serum hyaluronic acid.

Spotlight on the three main hepatic fibrogenic cells in HCV-infected patients: Multiple immunofluorescence and ultrastructure study

ABSTRACT The present work deals with the simultaneous ultrastructure and triple immunofluorescence study of the three main hepatic fibrogenic cells, hepatic stellate cell, myofibroblast (MF), and fibroblast, in a group of hepatitis C virus (HCV) RNA positive patients, as their exact interrelation behavior in vivo with the progress of hepatic fibrosis is still inadequate. In this study, for the first time, cells having the morphological characteristic of MF and not bone marrow fibrocytes were revealed in liver portal vessels. This necessitates the reevaluation of the available knowledge concerning bone marrow fibrocyte. Also, the distribution, cellular interrelations, and the fate of MF were highlighted.

Parasitological, histopathological, and immunohistochemical assessment of nitric oxide synthase inhibitor: aminoguanidine versus albendazole in the treatment of experimental murine toxocariasis

Objectives The study was intended to evaluate the effect of aminoguanidine compared with that of albendazole on mice infected with eggs of Toxocara canis by parasitological, histopathological, and immunohistochemical studies. Background T. canis is a widely distributed parasite. The nematode can cause toxocariasis in man . The infection takes place after swallowing fully embryonated eggs with larvae inside. Materials and methods The study was conducted on 117 albino mice classified into four groups: GI (the control group with subgroups Ia, Ib, and Ic); GII (infected with T. canis eggs); GIII (infected with T. canis eggs and treated with albendazole); and GIV (infected with T. canis eggs and treated with aminoguanidine). Sera from different groups of mice were collected for nitrite assay. The lung and brain tissues were taken for larval recovery and histopathological and immunohistochemical [inducible nitric oxide synthase (iNOS)] studies. Results The mean larval count decreased significantly from 7th to 45th day post infection (d.p.i), with albendazole and aminoguanidine, which was more effective in decreasing larval count. Aminoguanidine treatment caused early improvement in histopathological lesions initiating from the second d.p.i.; however, albendazole improvement was observed on the seventh d.p.i. The grade of iNOs expression in GII and GIII was high as compared with that in GIV. There was significantly positive correlation between serum nitrite and the grade of iNOS expression, especially on early d.p.i only in groups II and III. Conclusion Both albendazole and aminoguanidine treatment for T. canis -infected mice caused decrease in larval count and improvement in histopathological lesions. Aminoguanidine was more effective with early response and decreased tissue damage.