Anti-bacterial activity of Achatina CRP and its mechanism of action.

The physiological role of C-reactive protein (CRP), the classical acute-phase protein, is not well documented, despite many reports on biological effects of CRP in vitro and in model systems in vivo. It has been suggested that CRP protects mice against lethal toxicity of bacterial infections by implementing immunological responses. In Achatina fulica CRP is a constitutive multifunctional protein in haemolymph and considered responsible for their survival in the environment for millions of years. The efficacy of Achatina CRP (ACRP) was tested against both Salmonella typhimurium and Bacillus subtilis infections in mice where endogenous CRP level is negligible even after inflammatory stimulus. Further, growth curves of the bacteria revealed that ACRP (50 µg/mL) is bacteriostatic against gram negative salmonellae and bactericidal against gram positive bacilli. ACRP induced energy crises in bacterial cells, inhibited key carbohydrate metabolic enzymes such as phosphofructokinase in glycolysis, isocitrate dehydrogenase in TCA cycle, isocitrate lyase in glyoxylate cycle and fructose-1,6-bisphosphatase in gluconeogenesis. ACRP disturbed the homeostasis of cellular redox potential as well as reduced glutathione status, which is accompanied by an enhanced rate of lipid peroxidation. Annexin V-Cy3/CFDA dual staining clearly showed ACRP induced apoptosis-like death in bacterial cell population. Moreover, immunoblot analyses also indicated apoptosis-like death in ACRP treated bacterial cells, where activation of poly (ADP-ribose) polymerase-1 (PARP) and caspase-3 was noteworthy. It is concluded that metabolic impairment by ACRP in bacterial cells is primarily due to generation of reactive oxygen species and ACRP induced anti-bacterial effect is mediated by metabolic impairment leading to apoptosis-like death in bacterial cells.

Mollusc C-reactive protein crosses species barrier and reverses hepatotoxicity of lead in rodent models.

Achatina fulica C-reactive protein (ACRP) reversed the toxic effects of lead nitrate both in vivo in mice and in vitro in rat hepatocytes restoring the basal level of cell viability, lipid peroxidation, reduced glutathione and superoxides. Cytotoxicity was also significantly ameliorated in rat hepatocytes by in vitro pre-treatments with individual subunits (60, 62, 90 and 110 kDa) of ACRP. Annexin V-Cy3/CFDA dual staining showed significant reduction in the number of apoptotic hepatocytes pre-treated with ACRP. ACRP induced restoration of mitochondrial membrane potential was remarkable. ACRP pre-treatment prevented Pb-induced apoptosis mediated by caspase activation. The antagonistic effect of ACRP may be due to scavenging of reactive oxygen species which maintained the homeostasis of cellular redox potential as well as reduced glutathione status. The results suggest that ACRP crosses the species barrier and it may be utilized as a viable exogenous agent of cytoprotection against heavy metal related toxicity.

Induction of oxidative stress by non-lethal dose of mercury in rat liver:Possible relationships between apoptosis and necrosis.

Sprague Dawley strain of male rats weighing 200 ± 10.0 g, were exposed intramuscularly to non-lethal dose of mercury for short acute duration of 24 and 48 hr. Mercury treatment increased thio-barbituric acid reactive substance (TBARS) and conjugated diene (CD) content with increase in duration when compared with control. This reflects possible increase in lipid peroxidation, revealing that sufficient intoxication was generated by non-lethal dose of mercury. Furthermore, mercury treatment decreased tissue glutathione (GSH) content to 2.07 and 1.49 3g GSH mg protein-1 with concomitant decrease in glutathione-S-transferase (GST) activity by 26.06 and 36.40% after 24 and 48hr of exposure respectively. The elevations of aspartate transaminase (AST) and alanine transaminase (ALT) levels measured exhibited increase of 287.5 and 214.5% after 48 hr of exposure respectively which were found to be highly significant compared with control. Western blot analysis indicated upregulation of caspase-9 and upsurge in effector caspase-3 activity leading to apoptosis. The concluded findings of the present investigation suggests possible role of early mercury exposure in inducing oxidative stress mediated apoptosis in mammalian model systems as an indicator component of environmental toxicology.