Alpha-Chymotrypsin Protects Against Acute Lung, Kidney, and Liver Injuries and Increases Survival in CLP-Induced Sepsis in Rats Through Inhibition of TLR4/NF-κB Pathway

Abstract

Inflammation and oxidative stress play a major role in the development of sepsis and its associated complications, leading to multiple organ failure and death. The lungs, liver, and kidneys are among the early affected organs correlated with mortality in sepsis. Alpha-chymotrypsin (α-ch) is a serine protease that exerts anti-inflammatory, anti-edematous, and anti-oxidant properties. Purpose This study was undertaken to elucidate if the anti-inflammatory and anti-oxidant effects of α-ch observed in previous studies can alleviate lung, liver, and kidney injuries in a cecal ligation and puncture (CLP)-induced sepsis model, and thus decrease mortality. Materials and Methods Septic animals were given α-ch 2 h post CLP procedure. Sepsis outcomes were assessed in the lungs, liver, and kidneys. Separate animal groups were investigated for a survival study. Results CLP resulted in 0% survival, while α-chymotrypsin post-treatment led to 50% survival at the end of the study. Administration of α-chymotrypsin resulted in a significant attenuation of sepsis-induced elevated malonaldehyde (MDA) and total nitrite/nitrate (NOx) levels. In addition, there was a significant increase in reduced glutathione (GSH) content and superoxide dismutase (SOD) activity in the lungs, liver, and kidneys. Administration of α-ch reduced elevated tissue expression of toll-like receptor-4 (TLR4), nuclear factor kappa-B (NF-κB), myeloperoxidase (MPO), and inducible nitric oxide synthase (iNOS). Alpha-chymotrypsin resulted in a significant reduction in serum levels of tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6). Alpha-chymotrypsin attenuated the rise in serum creatinine, cystatin C, blood urea nitrogen (BUN), alanine aminotransferase (ALT), and aspartate aminotransferase (AST) levels that was observed in the septic group. In addition, α-ch significantly reduced the lung wet/dry weight ratio, total protein content, and leukocytic counts in bronchoalveolar lavage fluid (BALF). Histopathological examination of the lungs, liver, and kidneys confirmed the protective effects of α-ch on those organs. Conclusion α-ch has protective potential against sepsis through lowering tissue expression of TLR4, NF-κB, MPO, and iNOS leading to decreased oxidative stress and inflammatory signals induced by sepsis. This effect appeared to alleviate the damage to the lungs, liver, and kidneys and increase survival in rats subjected to sepsis.

Research Progress of Macromolecules in the Prevention and Treatment of Sepsis

“…By inhibiting the TLR4/NF-κB pathway, α-ch, a serine protease, reduces the levels of inflammatory cytokines IL-1β, IL-6, and TNF-α, and inhibits the expression levels of myeloperoxidase (MPO) and inducible nitric oxide synthase (iNOS) in a cecum ligation and puncture (CLP)-induced sepsis mouse model. After α-ch treatment, the survival rate of CLP mice increased by 50% [ 25 ].…”

Novel Peptide-Based Fluorescent Probe for Simultaneous Sensing of Chymotrypsin and Hydrogen Peroxide

“…While some serine proteases such as, e.g., trypsin or neutrophil elastase, are cytotoxic and proven to increase ROS levels including H 2 O 2 in cells, 12 others, such as α-chymotrypsin, were found to be capable of suppressing oxidative stress and inflammation processes, consequently exhibiting protective potential against sepsis, and alleviating the damage to the kidneys, liver, and lungs. 13 During the last couple of decades, numerous systems for the sole detection of ROS including hydrogen peroxide 14−18 or various enzymes 19−21 have been reported, among which fluorescence-based sensors are heavily represented. Because of their sensitivity, prompt responsiveness, and accuracy, they have been found to be indispensable in bioimaging 22 and realtime sensing of numerous chemical, 23 biochemical, 24 and biological 25 species.…”

“…Although the scientists may not always uniformly agree on, e.g., which initiator caspases are responsible for the activation of executioner caspases, there is no doubt that H 2 O 2 -induced apoptosis proceeds through a caspase activation pathway. , Apart from the cysteine aspartic proteases, a direct connection between the serine proteases and hydrogen peroxide has been established as well. While some serine proteases such as, e.g., trypsin or neutrophil elastase, are cytotoxic and proven to increase ROS levels including H 2 O 2 in cells, others, such as α-chymotrypsin, were found to be capable of suppressing oxidative stress and inflammation processes, consequently exhibiting protective potential against sepsis, and alleviating the damage to the kidneys, liver, and lungs …”

“…While some serine proteases such as, e.g., trypsin or neutrophil elastase, are cytotoxic and proven to increase ROS levels including H 2 O 2 in cells, 12 others, such as α-chymotrypsin, were found to be capable of suppressing oxidative stress and inflammation processes, consequently exhibiting protective potential against sepsis, and alleviating the damage to the kidneys, liver, and lungs. 13 …”

Regulatory effects of mangiferin on LPS‐induced inflammatory responses and intestinal flora imbalance during sepsis

“…Further, the deletion of TLR4 significantly attenuated LPS‐induced acute liver injury in mice (Chen, Tan, et al., 2021 ). Thus, it is a promising strategy to modulate TLR4 to treat sepsis (Kumar, 2020 ; Senousy et al., 2022 ).…”