Protozoan parasites of the genus Leishmania cause a number of important human diseases. One of the key determinants of parasite infectivity and survival is the surface glycoconjugate lipophosphoglycan (LPG). In addition, LPG is shown to be useful as a transmission blocking vaccine. Since culture supernatant of parasite promastigotes is a good source of LPG, we made attempts to characterize functions of the culture supernatant, and membrane LPG isolated from metacyclic promastigotes of Leishmania major. The purification scheme included anion-exchange chromatography, hydrophobic interaction chromatography and cold methanol precipitation. The purity of supernatant LPG (sLPG) and membrane LPG (mLPG) was determined by SDS-PAGE and thin layer chromatography. The effect of mLPG and sLPG on nitric oxide (NO) production by murine macrophages cell line (J774.1A) was studied. Both sLPG and mLPG induced NO production in a dose dependent manner but sLPG induced significantly higher amount of NO than mLPG. Our results show that sLPG is able to promote NO production by murine macrophages.
Nitric Oxide/analysis/*biosynthesis ; Mice, Inbred BALB C ; Mice ; Macrophages/*drug effects/metabolism/parasitology ; Leishmania major/chemistry/pathogenicity/*physiology ; Glycosphingolipids/isolation & purification/*pharmacology ; Endotoxins/analysis ; Electrophoresis, Polyacrylamide Gel ; Culture Media ; Chromatography, Thin Layer/methods ; Cell Membrane/chemistry ; Cell Line ; Animals

This study investigated whether trinitroglycerine (TNG) as nitric oxide (NO) releasing agent had anti-leishmanial effects and mediated pathology in BALB/c mice infected with Leishmania major. Cutaneous leishmaniasis (CL), a zoonotic infection caused by leishmania protozoa is still one of the health problems in the world and in Iran. NO is involved in host immune responses against intracellular L. major, and leishmania killing by macrophages is mediated by this substance. Moreover, application of CL treatment with NO-donors has been recently indicated. In our study, TNG was used for its ability to increase NO and to modify CL infection in mice, in order to evaluate NO effects on lesion size and formation, parasite proliferation inside macrophages, amastigote visceralization in target organs, and NO induction in plasma and organ suspensions. Data obtained in this study indicated that TNG increased plasma and liver-NO, reduced lesion sizes, removed amastigotes from lesions, livers, spleens, and lymph nodes, declined proliferation of amastigotes, hepatomegaly, and increased survival rate. However, TNG reduced spleen-NO and had no significant effects on spelenomegaly. The results show that TNG therapy reduced leishmaniasis and pathology in association with raised NO levels. TNG had some antiparasitic activity by reduction of positive smears from lesions, livers, spleens, and lymph nodes, which could emphasize the role of TNG to inhibit visceralization of L. major in target organs.
Animal Structures/parasitology ; Animals ; Antiprotozoal Agents/chemistry/*therapeutic use ; Female ; Leishmania major/*drug effects/immunology ; Leishmaniasis, Cutaneous ; Macrophages/parasitology ; Mice ; Mice, Inbred BALB C ; Nitric Oxide/blood/metabolism/*pharmacology ; Nitroglycerin/*analogs & derivatives/*therapeutic use ; Severity of Illness Index ; Skin/pathology ; Survival Analysis

Infection of schistosomiasis japonica may eventually lead to liver fibrosis, and no effective antifibrotic therapies are available but liver transplantation. Hedgehog (HH) signaling pathway has been involved in the process and is a promising target for treating liver fibrosis. This study aimed to explore the effects of pentoxifylline (PTX) on liver fibrosis induced by schistosoma japonicum infection by inhibiting the HH signaling pathway. Phorbol12-myristate13-acetate (PMA) was used to induce human acute mononuclear leukemia cells THP-1 to differentiate into macrophages. The THP-1-derived macrophages were stimulated by soluble egg antigen (SEA), and the culture supernatants were collected for detection of activation of macrophages. Cell Counting Kit-8 (CCK-8) was used to detect the cytotoxicity of the culture supernatant and PTX on the LX-2 cells. The LX-2 cells were administered with activated culture supernatant from macrophages and(or) PTX to detect the transforming growth factor-β gene expression. The mRNA expression of shh and gli-1, key parts in HH signaling pathway, was detected. The mRNA expression of shh and gli-1 was increased in LX-2 cells treated with activated macrophages-derived culture supernatant, suggesting HH signaling pathway may play a key role in the activation process of hepatic stellate cells (HSCs). The expression of these genes decreased in LX-2 cells co-cultured with both activated macrophages-derived culture supernatant and PTX, indicating PTX could suppress the activation process of HSCs. In conclusion, these data provide evidence that PTX prevents liver fibrogenesis in vitro by the suppression of HH signaling pathway.
Animals ; Antigens, Helminth ; isolation & purification ; pharmacology ; Cell Culture Techniques ; Cell Differentiation ; drug effects ; Cell Line ; Culture Media, Conditioned ; chemistry ; pharmacology ; Gene Expression Regulation ; Hedgehog Proteins ; agonists ; antagonists & inhibitors ; genetics ; immunology ; Hepatic Stellate Cells ; cytology ; drug effects ; metabolism ; Humans ; Liver Cirrhosis ; metabolism ; parasitology ; prevention & control ; Macrophage Activation ; drug effects ; Macrophages ; cytology ; drug effects ; immunology ; Models, Biological ; Monocytes ; cytology ; drug effects ; metabolism ; Pentoxifylline ; pharmacology ; Phosphodiesterase Inhibitors ; pharmacology ; RNA, Messenger ; genetics ; immunology ; Schistosoma japonicum ; chemistry ; Signal Transduction ; Tetradecanoylphorbol Acetate ; pharmacology ; Zinc Finger Protein GLI1 ; genetics ; immunology ; Zygote ; chemistry