Abstract
Introduction: Cardiovascular system is highly sensitive to LPS-induced oxidative damage. This study aimed to show the inhibitory effect of bacterial Lipase on LPS-induced cardiomyoblasts toxicity. Methods: Rat cardiomyoblasts H9C2 were classified into Control, LPS (cells received 0.1, 1 and 10 μg/mL LPS) and LPS+ Lipase groups. In LPS+Lipase group, different concentrations of lipopolysaccharide were pre-incubated with 5 mg/mL bacterial lipase at 37˚C overnight prior to cell treatment. After 72 hours, cell viability was assessed by MTT assay. The expression of key genes related to toll-like receptor signaling pathways was assessed by real-time PCR assay. Percentage of fatty acids was evaluated in each group using gas chromatography assay. The levels of NO was also measured using the Griess reaction. Results: Data showed H9C2 cells viability was decreased after exposure to LPS in a dose-dependent manner (P < 0.05). Incubation of LPS with lipase increased cell survival rate and closed to near-to-control levels (P < 0.05). Lipase had the potential to blunt the increased expression of IRAK and NF-κB in cells after exposure to the LPS. Compared to the LPS group, lipase attenuated the increased level of NO-induced by LPS (P < 0.05). Gas chromatography analysis showed the reduction of saturated fatty acids in cells from LPS group while the activity of lipase prohibited impact of LPS on cell fatty acid composition. LPS decreased the ability of cardiomyoblasts to form colonies. Incubation of LPS with lipase enhanced clonogenic capacity. Conclusion: Reduction in lipopolysaccharide-induced cytotoxicity is possibly related to lipase activity and reduction of modified lipopolysaccharide with toll-like receptor.
Highlights
Cardiovascular system is highly sensitive to LPS-induced oxidative damage
Lipase decreased the detrimental effect of LPS on rat cardiomyocytes According to data from the MTT assay, we noted that LPS influenced cell viability in a dose-dependent manner (P
The data demonstrated that the cell viability in the group given 0.1 μg/mL LPS with 93% viability reached to 69.53 ± 3.15% in cells treated with 10 μg/mL LPS compared to the control group, showing dose-dependent toxicity of LPS on rat cells
Summary
Cardiovascular system is highly sensitive to LPS-induced oxidative damage. This study aimed to show the inhibitory effect of bacterial lipase on LPS-induced cardiomyoblasts toxicity. Toll-like receptors (TLR family), which is formed of at least 10 mammalian homologs of Drosophila toll, function as pattern recognition receptors for bacterial LPS and thereby control the production of pro-inflammatory cytokines.[7,8] A member of this family named TLR-4 functions as the main receptor for LPS of gram-negative bacteria, transduces signals through effector MyD88, interleukin-1 (IL-1) receptor-associated kinase (IRAK), and TRAF6 and activates NF-ƙB and mitogenactivated protein kinase pathway.[9,10] Some of TLRs like -1, -2, -4, and -6, unlike TLR-4, constitute heterodimers, while a unique homodimer has been described for TLR4.11 a protein named MD-2 is necessary for efficient LPS signal transduction, making a complex with the extracellular domain of TLR-4.12-14 It is proposed that the attachment of soluble serum proteins such as CD14, LPS binding protein (LBP) to bacterial LPS enhances affinity of LPS to TLR-4.2 CD14 in membrane-bound form is candidate as an alternative receptor for LPS and acts
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