Cytokine patterns in experimental schistosomiasis mansoni infected mice treated with silymarin.

The purpose of this study was to determine cytokine patterns in experimental schistosomiasis mansoni infected mice treated with silymarin. The study was conducted upon 100 mice that were divided into five groups; 20 each: uninfected control group, Schistosoma mansoni infected untreated mice (infected control), infected mice treated with praziquantel (PZQ), infected mice treated with silymarin and infected mice treated with both praziquantel and silymarin. 10 mice from each group were sacrificed at 10th and 18th weeks post infection respectively. Histopathological investigations were performed. Liver sections were stained with hematoxylin-eosin and Masson's trichrome stain to evaluate changes of granuloma sizes and numbers. Serum levels of the cytokines (TNF-α, IFN-γ, IL-4 and TGF-ß1) were assessed in the sera of all groups by immunoassay. The measured levels of cytokines (IFN-γ, IL-4, TNF-α, TGF-ß1) were found to be significantly increased in infected mice compared to normal control. At the same time, treated groups with silymarin alone or combined with PZQ showed significant decrease in IL-4, TNF-α and TGF-ß1 levels compared to infected control. On the other hand, there was a significant increase in IFN-γ level observed in all treated groups compared to infected control. In addition, the histopathological examination of the liver in the group treated with PZQ showed a reduction in the number of livers eggs granuloma at all periods of sacrification compared with the infected untreated group. However, there was more decrease in granulomas diameter in both silymarin treated group or combined with PZQ at all periods of sacrification when compared to infected untreated group. In conclusion; treatment with silymarin combined with PZQ in murine schistosomiasis could reduce hepatic fibrosis by their action on the production of pro-inflammatory cytokines.

Studies on the role of tumor necrosis factor- alpha (TNF-α) in hepatocytes induced apoptosis in vaccinated, Schistosoma mansoni-challenged mice.

Tumour Necrosis Factor-alpha (TNF-α) plays a complex role in pathophysiological changes caused by schistosomiasis in the liver cells as induced apoptosis. So, The highlighted experimentally the role of TNF-α in hepatocytes apoptosis, using that as an assessment of the efficacy of antischistosomal vaccination by mixed crude antigens preparations [Cercarial antigen preparation (CAP) + soluble worm antigen preparation (SWAP) + soluble egg antigen(SEA)] by parasitological, histo-pathological and histochemical studies using Feulgen stain of hepatoytes DNA, a serological study also of serum TNF-α level by ELISA. Fifty two laboratory bred Albino male mice, were used in this study. They were classified into four groups (13 mice in each group), G1: normal control, G2 as infected control while G3 supported by Freund's Adjuvant (F. Adj) then infected and G4 vaccinated with combined antigens (CAP, SWAP and SEA) + F. Adj, then infected. Mice were sacrificed by cervical dislocation 9 weeks post infection, parasitological (Kato-Katz thick smear for egg count), histopathologial {haematoxylin and eosin (H&E) staining of hepatic sections}, histochemical (feulgen staining of hepatocytes DNA) and ELISA to estimate serum TNF-α level were performed. The data showed that vaccination with combined antigens showed protective effect on vaccinated then Schistosoma challenged mice, hepatocytes induced apoptosis was directly proportional with the TNF-α serum level, and the protection degree of potential combined vaccine was inversely proportional with serum TNF-α level and induced apoptosis.

In vitro effect of Mirazid on Schistosoma mansoni worms.

Measuring Mirazid ability for contracting the worm muscle and its effect on the worm surface ultra-structure can be used to monitor the in vitro effect of any drug. This study aims at investigating the actual effect of Mirazid (a new schistosomicide; purified oleo-resin extract of Myrrh, derived from Commiphora molmol plant) on S. mansoni worms by detecting its in vitro effect. Three groups of white albino mice (5 mice in each group) were infected by 100 cercariae for each mouse. The 3rd group served as a control group. Seven weeks post-infection the mice were sacrificed, perfused and worms were collected. Muscle tension of the worms collected from the first group of mice, was assayed in response to Mirazid in rising concentrations of 100, 200, 300 and 400 nM. The in vitro effect of Mirazid on the muscle tension of single S. mansoni worm was determined using a special device to determine percentage of change in worm length (% shortening). The drug elicited somatic muscle contraction and reached the highest response with 400 nM Mirazid. The maximal increase in the muscle tension (48% shortening) was induced by the highest concentration (400 nM) of the drug. The worms collected from the second group of mice were scanned by electron microscopy. The worms were exposed each to 10 ul of Mirazid in concentration of 10(-6) and collected after 10, 20 and 30 minutes of exposure. Ten minutes exposure caused disruption of the tegument and collapse of tubercles. After 20 minutes, the tegument appeared edematous with more disruption and more shrinkage of tubercles. After 30 minutes, the tegument appeared markedly edematous with extensive disruption of the inter-papillary areas and sensory bulbs. The spines covering the tubercles appeared to be lost.

Parasitological and immunological changes in murine hepatic schistosomiasis before and after praziquantel treatment.

Hepatic fibrosis was assessed by estimating hydroxyproline content in liver tissues at the course of the disease and after PZQ treatment. Parasitological investigations included egg count in faeces and tissues (liver and intestine) and oogram pattern. The results showed that treatment of infected mice with PZQ, modulated the course of schistosomiasis as evaluated by highly significant decline in hepatic hydroxyproline content as well as egg count in stool and tissues. In addition, the oogram pattern showed that PZQ had a lethal effect on mature eggs in tissues. PZQ treatment of S. mansoni infected mice, is effective in reducing the severity of the disease and in attenuating hepatic fibrosis, particularly when the treatment starts early with a suitable dose.

Resistance to praziquantel, effect of drug pressure and stability test.

Various isolates of S. mansoni, originally showed marked diminished susceptibility to PZQ, were used in this work. These isolates were taken from patients not cured after two or three doses of the drug. They were passed in experimental mice and treated with sub-curative doses of PZQ to determine the effect of drug pressure on the offspring of these isolates. Upon treatment of the second generation of these isolates with curative doses of PZQ, they showed significant less response to the drug in terms of both the drug efficacy (percent of worm reduction), and the ED50 (the effective dose that kills 50% of worms). Also, stability test was performed on some S. mansoni isolates. This means repeated treatment of unsusceptible isolates that have been passed for several passages in the lab. and results compared to that of a control susceptible isolate (originally taken from a patient cured after a single dose of PZQ). The results showed that repeated passage of S. mansoni isolates in the lab. does not render them more susceptible to PZQ. Indeed, these resistant isolates showed less susceptibility to the drug than before, or at least they retain their original level of insusceptibility to PZQ.