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Objectives Hepatocellular carcinoma is the fourth leading cause of cancer deaths in the world. Conventional methods of cancer therapy are either invasive or have undesirable side effects. Therefore, exploring new therapeutic strategies to control the progression of hepatocellular carcinoma, such as cell-based therapies, is a key issue for prolonging patient survival. In this study, we aimed to evaluate tumor suppressive effects of mesenchymal stem cells on the in vivo progression of hepatocellular carcinoma in murine model. Materials and methods Hepatocellular carcinoma was induced in 40 rats with diethylnitrosamine. Rats were divided into 4 groups: 1 group injected with diethylnitrosamine only, 1 group injected with diethylnitrosamine and 1 dose of rat bone marrowderived mesenchymal stem cells, 1 group injected with diethylnitrosamine and 2 doses of rat bone marrowderived mesenchymal stem cells, and 1 group was injected with diethylnitrosamine and 3 doses of rat bone marrow-derived mesenchymal stem cells. Rats were killed after 1 month of dose 3. Liver specimens were histopathologically examined, and serum samples were examined for liver function and cytokines. Results Histopathological examination revealed that mesenchymal stem cell transplant induced liver regeneration. It also improved liver function as revealed by decreased levels of alanine and aspartate aminotransferase. Mesenchymal stem cells also repaired the immunopathology of the liver environment, as it decreased levels of interleukin 2 and 10, tumor necrosis factor α, and interferon γ. Conclusions Mesenchymal stem cell infusion significantly enhanced hepatic structure and function of livers in a rat hepatocellular carcinoma model.

Introduction: Malignant ascites results from imbalance between protein in the peritoneal cavity and absorption of fluids via the lymphatic system. More than 20 interleukins (ILs) are known to play an important role in the protection against tumors. Materials and Methods: Ascitic fluid IL-1B, IL-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ levels were assessed in 45 patients with liver cirrhosis and ascites as judged by histopathological and ultrasonographic findings. They were divided into 2 groups according to presence of hepatic focal lesions. Ten patients with focal hepatic lesions were randomly selected and subjected to analysis of serum levels of IL-2 and IL-10. Results: Ascitic fluid IL-4, IL-6 and IL-10 levels were found to be significantly higher in patients with hepatocellular carcinoma (HCC) than patients with cirrhosis. TNF-α, and IFN-γ were also found to be higher in HCC than patients with cirrhosis but with no significance. On the other hand, there was no significant difference in levels of IL-1B and IL-2 between the 2 groups. Ascitic fluid IL-2 and IL-10 levels were found to be higher in ascitic fluid than in serum of the same patients. Conclusion: Ascitic fluid levels of IL-4, IL-6 and IL-10 are higher in HCC patients than patients with cirrhosis alone. Levels of ascitic fluid IL-2 and IL-10 proved to be a better prognostic tool than their levels in sera of the same patients. To conclude, patients with cirrhosis may be subjected to scheduled examination of ascitic fluid cytokines to predict transformation into HCC.

BACKGROUND The liver is one of the major target organs for which cell-based therapies are very promising. The limitations of various cellular therapies, including bone marrow (BM)-derived mesenchymal stem cells (MSCs), urges the exploration of stem cell sources more suitable for transplantation. Human umbilical cord blood (HUCB) can overcome these drawbacks with a favorable reparative outcome. OBJECTIVES The aim of this study was to evaluate the therapeutic potential of MSCs in 2 groups of chronic liver injury experimental models. MATERIAL AND METHODS Propagation and characterization of MSCs isolated from cord blood (CB) samples were performed and differentiation into osteogenic, adipogenic and hepatogenic lineages was induced. The 1st experimental model group (80 mice) included a negative control, a pathological control and 60 mice infected with Schistosoma mansoni (S. mansoni) and transplanted with MSCs. The 2nd experimental model group (30 hamsters) included 10 healthy hamsters serving as a negative control and 20 hamsters injected with repeated doses of carbon tetrachloride (CCl4) to induce liver fibrosis; 10 of them were treated with an intrahepatic (IH) injection of 3 × 106 MSCs and the other 10 were untreated pathological controls. Mice and hamsters were sacrificed 12 weeks post-transplantation and their liver sections were stained immunohistochemically for the detection of human hepatocyte-like cells. Moreover, the sections were examined for the levels of fibrosis. RESULTS In both models, the transplantation of CB-derived MSCs (CB-MSCs) resulted in the engraftment of the fibrotic livers with newly formed hepatocytes, as evidenced by positive immunohistochemistry staining with human Hepatocyte Paraffin 1 (Hep Par 1), alpha-fenoprotein (AFP), cytokeratin 18 (CK18), cytokeratin 7 (CK7), and OV6 monoclonal antibody. The transplanted liver sections showed markedly reduced hepatic fibrosis with a significantly lower fibrotic index, as well as significantly improved liver functions compared to the pathological control (p < 0.001). CONCLUSIONS This data provides hope that human CB-MSCs can be utilized as multipotent stem cells with unlimited potentiality in regenerative medicine and supports the concept of cellular therapy for the cure of hepatic fibrosis.

The technique of stem cells or hepatocytes transplantation has recently improved in order to bridge the time before whole-organ liver transplantation. In the present study, unfractionated bone marrow stem cells (BMSCs) were harvested from the tibial and femoral marrow compartments of male mice, which were cultured in Dulbecco's modified Eagle's medium (DMEM) with and without hepatocyte growth factor (HGF), and then transplanted into Schistosoma mansoni-infected female mice on their 8th week post-infection. Mice were sacrificed monthly until the third month of bone marrow transplantation, serum was collected, and albumin concentration, ALT, AST, and alkaline phosphatase (ALP) activities were assayed. On the other hand, immunohistopathological and immunohistochemical changes of granuloma size and number, collagen content, and cells expressing OV-6 were detected for identification of liver fibrosis. BMSCs were shown to differentiate into hepatocyte-like cells. Serum ALT, AST, and ALP were markedly reduced in the group of mice treated with BMSCs than in the untreated control group. Also, granuloma showed a marked decrease in size and number as compared to the BMSCs untreated group. Collagen content showed marked decrease after the third month of treatment with BMSCs. On the other hand, the expression of OV-6 increased detecting the presence of newly formed hepatocytes after BMSCs treatment. BMSCs with or without HGF infusion significantly enhanced hepatic regeneration in S. mansoni-induced fibrotic liver model and have pathologic and immunohistopathologic therapeutic effects. Also, this new therapeutic trend could generate new hepatocytes to improve the overall liver functions.

Elkhafif N, El Baz H, Hammam O, Hassan S, Salah F, Mansour W, Mansy S, Yehia H, Zaki A, Magdy R. CD133+ human umbilical cord blood stem cells enhance angiogenesis in experimental chronic hepatic fibrosis. APMIS 2010. The in vivo angiogenic potential of transplanted human umbilical cord blood (UCB) CD133+ stem cells in experimental chronic hepatic fibrosis induced by murine schistosomiasis was studied. Enriched cord blood-derived CD133+ cells were cultured in primary medium for 3 weeks. Twenty-two weeks post-Schistosomiasis infection in mice, after reaching the chronic hepatic fibrotic stage, transplantation of stem cells was performed and mice were sacrificed 3 weeks later. Histopathology and electron microscopy showed an increase in newly formed blood vessels and a decrease in the fibrosis known for this stage of the disease. By immunohistochemical analysis the newly formed blood vessels showed positive expression of the human-specific angiogenic markers CD31, CD34 and von Willebrand factor. Few hepatocyte-like polygonal cells showed positive expression of human vascular endothelial growth factor and inducible nitric oxide synthase. The transplanted CD133+ human stem cells primarily enhanced hepatic angiogenesis and neovascularization and contributed to repair in a paracrine manner by creating a permissive environment that enabled proliferation and survival of damaged cells rather than by direct differentiation to hepatocytes. A dual advantage of CD133+ cell therapy in hepatic disease is suggested based on its capability of hematopoietic and endothelial differentiation.

The first solid phase synthesis of the important anthelmintic praziquantel is described. The synthesis is rapid and efficient. The method may be extended to the synthesis of libraries of urgently needed replacements for this drug.

Background This research was carried out to develop a reliable monoclonal antibody (MoAb)-based sandwich enzyme linked immunosorbent assay (ELISA) for the diagnosis of active Fasciola gigantica infection in both serum and stool for comparative purposes. Methods From a panel of MoAbs raised against F. gigantica excretory/secretory antigens (ES Ags), a pair (12B/11D/3F and 10A/9D/10G) was chosen due to its high reactivity and strict specificity to F. gigantica antigen by indirect ELISA. Results The two MoAbs were of the IgG1 and IgG2a subclasses, respectively. Using SDS-PAGE and EITB, the selected MoAbs recognized 83, 64, 45 and 26 kDa bands of ES Ags. The lower detection limit of ELISA assay was 3 ng/ml. In stool, the sensitivity, specificity and diagnostic efficacy of ELISA was 96%, 98.2 and 97.1%; while in serum they were 94%, 94.6% and 94.3%, respectively. Moreover, a positive correlation was found between ova count in stool of F. gigantica infected patients and the OD readings of ELISA in both stool and serum samples (r = 0.730, p < 0.01 and r = 0.608; p < 0.01, respectively). Conclusions These data showed that the use of MoAb-based sandwich ELISA for the detection of F. gigantica coproantigens in stool specimens was superior to serum samples; it provides a highly efficient, non-invasive technique for the diagnosis of active F. gigantica infection.

The in vivo angiogenic potential of transplanted human umbilical cord blood (UCB) CD133(+) stem cells in experimental chronic hepatic fibrosis induced by murine schistosomiasis was studied. Enriched cord blood-derived CD133(+) cells were cultured in primary medium for 3 weeks. Twenty-two weeks post-Schistosomiasis infection in mice, after reaching the chronic hepatic fibrotic stage, transplantation of stem cells was performed and mice were sacrificed 3 weeks later. Histopathology and electron microscopy showed an increase in newly formed blood vessels and a decrease in the fibrosis known for this stage of the disease. By immunohistochemical analysis the newly formed blood vessels showed positive expression of the human-specific angiogenic markers CD31, CD34 and von Willebrand factor. Few hepatocyte-like polygonal cells showed positive expression of human vascular endothelial growth factor and inducible nitric oxide synthase. The transplanted CD133(+) human stem cells primarily enhanced hepatic angiogenesis and neovascularization and contributed to repair in a paracrine manner by creating a permissive environment that enabled proliferation and survival of damaged cells rather than by direct differentiation to hepatocytes. A dual advantage of CD133(+) cell therapy in hepatic disease is suggested based on its capability of hematopoietic and endothelial differentiation.

Background Bladder cancer is the most prevalent cancer in Egypt and in most African countries. Two distinct types of bladder cancer exist in Egypt, namely the bilharzial and non-bilharzial types. Nitric oxide (NO) exerts several functions in benign and malignant human tissues. Many studies have proved significantly higher urinary NO levels, as well as presence of inducible nitric oxide synthase (iNOS) in bladder tumors. Because tissue iNOS is responsible for the production of urinary and serum NO, it is possible that urine from bladder cancer patients includes high levels of NO. Aim of the work To study serum and urinary levels of NO as well as iNOS expression in bladder tissue of patients with different chronic schistosomal and non-schistosomal bladder lesions including malignancy to determine the possibility of using them as biological markers for early diagnosis and prediction of the clinical out come of bladder tumors. Patients and methods Sixty five patients; 40 with malignancy and 25 with chronic cystitis, were included in the study. Seventeen individuals with apparently normal urothelium served as controls. NO was assayed in urine and serum using the modified Griss reaction technique. Expression of iNOS in bladder tissue was determined by indirect immunohistochemical techniques. Results All controls were negative for NO in both serum and urine. The rate of positivity of NO in sera of the malignant group was significantly higher than in the chronic cystitis group ( p p p Conclusion Urinary and serum NO levels as well as iNOS expression in bladder tissue could be used as tumor markers for diagnosis of bladder cancer. Urinary NO could be also used as an early non-invasive predictive marker for the tumor behavior and prognosis.

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.

To control spread of Schistosma mansoni infection, rapid investigation of infected Biomphalaria Alexandrina snails that surveyed from any suspected area of a field, must be evaluated before shedding of cercariae. Detection of S. mansoni antigens in snails hemolymph was exposed to be an accurate and rapid method for assessment of infected snails. In the present study, two murine monoclonal antibodies (MABs) against soluble egg antigen (SEA) were employed for detecting S. mansoni in the hemolymph of 143 infected B. Alexandrina snails and 88 normal control snails, by using an enzyme linked immunosorbent assay (sandwich ELISA). Also, 63 snails and 56 snails of B. glabrata and Bulinus truncularia respectively were examined. The optical density (OD) values of SEA were measured in the hemolymph of infected snails. A total of 143 infected snails showed 100% percentage of positivety. Optical density at intervals 1, 2, 3 weeks post-exposure (prepatency) was measured and percentage positivity was 7%, 35.3%, 82.6%, respectively. It is also measured at early shedding snails (4 weeks, i.e. early patency) and at shedding (5, 6 weeks or maximal shedding) and percentage positivity was 87.5%, 96.2%, 89.5%, respectively. Also, after stop shedding (late patency) the percentage positivity was 15%.The hemolymph of normal control group of snails were all negative against SEA. Hemolymph of B.trunculata and B. glabrata infected snails showed 0% positivity against SEA. Polymorphic DNA (RAPD-PCR) technique was used for molecular identification of the infected snails at the time of 2 weeks post-infection and at the time of snails shedding. Conclusion The use of monoclonal antibodies for early detection of S. mansoni infected B. Alexandrina snails, is considered as an accurate, rapid and specific method to facilitate the control of infected snails that randomly surveyed from different areas of a suspected fields.

The first solid phase synthesis of the important anthelmintic praziquantel is described. The synthesis is rapid and efficient. The method may be extended to the synthesis of libraries of urgently needed replacements for this drug.