Alteration in endogenous proteolytic pathways caused by Bothrops Jararacussu venom in skeletal muscle / Alterações nas vias proteoliticas endogenas causados pela peçonha de Bothrops Jararacussu em musculo esqueletico

AUTOR(ES)
DATA DE PUBLICAÇÃO

2008

RESUMO

Bites by Bothrops snakes produce intense local hemorrhage and myonecrosis, often with extensive tissue degradation. In this work, we investigated the activities of the proteasome (pathway for selective protein degradation), cathepsins (lysosomal proteinases) and calpains (neutral, calcium-dependent proteinases) in envenoming by Bothrops jararacussu. We also examined the ability of MG-132, a proteasome inhibitor, to attenuate the venom-induced effects. Mice were injected with venom (25 µg or 75 µg) in the left gastrocnemius muscle and then killed 1, 3, 6, 12, 24, 48, 72 h and 7, 14, 21 and 28 days post-venom. The venom-injected and contralateral muscles were removed and processed for histological analysis or enzymatic assays using fluorimetric or colorimetric substrates. Blood was also collected for the quantification of plasma creatine kinase (CK, an indicator of myonecrosis). Bothrops jararacussu venom caused hemorrhage and necrosis (phase 1, up to 6-12 h post-venom), an inflammatory cell infiltrate and it generated the formation of myotubes and myoblasts (phase 2, 12-72 h post-venom), and the appearance of regenerative cells with increase of collagen deposition around these cells (phase 3, 7-28 days post-venom). The alterations were generally more marked with the higher dose of venom. The early tissue damage was confirmed by an increase in plasma CK levels 1-6 h post-venom, with a peak at 3 h. The muscle content of free amino acids decreased during the first 24 h, followed by a return to normal levels. Proteasomal activity was significantly inhibited for up to 48 h post-venom, followed by recuperation and a significant increase during muscle regeneration. During regeneration, there was also an increase in the expression of the 20Sα and 11S proteasomal subunits, mainly with the highest dose of venom. The proteasomal inhibitor reduced the chymotrypsin activity of the proteasome and the number of regenerating cells, but these effects were also seen in mice that received vehicle (DMSO) alone. The highest dose of venom caused an increase in calpain activity in the first 6 h whereas both of the venom doses significantly increased the activities of cathepsins B and H during the regeneration phase (48 h–28 days post-venom). These results indicate that B. jararacussu venom differentially affects the proteolytic activities studied. Calpain may be involved in phase 1 of tissue damage and cathepsin activity may be related to the presence of an inflammatory infiltrate (phase 2) and/or regeneration (phase 3). The lack of proteasomal activation in the early stages of envenoming suggests that this proteolytic pathway is not involved in early venom-induced muscle damage. However, the involvement of proteasomal activity during muscle regeneration remains to be established.

ASSUNTO(S)

proteossomo proteolysis muscle damage bothrops proteolise lesão muscular proteosome necrose necrosis

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