Browsing by Subject "Calpain"
Now showing items 1-3 of 3
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(2020)Growing evidence emphasizes insufficient clearance of pathological alpha-synuclein (alpha SYN) aggregates in the progression of Parkinson's disease (PD). Consequently, cellular degradation pathways represent a potential therapeutic target. Prolyl oligopeptidase (PREP) is highly expressed in the brain and has been suggested to increase alpha SYN aggregation and negatively regulate the autophagy pathway. Inhibition of PREP with a small molecule inhibitor, KYP-2407, stimulates autophagy and reduces the oligomeric species of alpha SYN aggregates in PD mouse models. However, whether PREP inhibition has any effects on intracellular alpha SYN fibrils has not been studied before. In this study, the effect of KYP2407 on alpha SYN preformed fibrils (PFFs) was tested in SH-SY5Y cells and human astmcytes. Immunostaining analysis revealed that both cell types accumulated alpha SYN PFFs intracellularly but KYP-2047 decreased intracellular alpha SYN deposits only in SH-SY5Y cells, as astrocytes did not show any PREP activity. Western blot analysis confirmed the reduction of high molecular weight alpha SYN species in SH-SY5Y cell lysates, and secretion of aSYN from SH-SY5Y cells also decreased in the presence of KYP-2407. Accumulation of alpha SYN inside the SH-SY5Y cells resulted in an increase of the auto-lysosomal proteins p62 and LC3BII, as well as calpain 1 and 2, which have been shown to be associated with PD pathology. Notably, treatment with KYP-2407 significantly reduced p62 and LC3BII levels, indicating an increased autophagic flux, and calpain 1 and 2 levels returned to normal in the presence of KYP-2407. Our findings indicate that PREP inhibition can potentially be used as therapy to reduce the insoluble intracellular alpha SYN aggregates.
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(2023)The effect of pre-rigor temperature incubation on the activity and distribution in sarcoplasmic and myofibrillar fractions of calpains, and meat quality attributes was investigated. Porcine longissimus thoracis muscles were incubated pre-rigor at 14, 22, 30 and 38 degrees C to 6 h postmortem, followed by another 2 h incubation at 14 degrees C. Thereafter, muscles were stored at 2 degrees C for 1 or 4 days. With higher pre-rigor temperature, sarcoplasmic Ca2+ concentration, purge loss and myofibril-bound calpain-1 content increased, while shear force declined. Water-holding capacity of isolated myofibrils was lower after pre-rigor incubation at 38 degrees C. Desmin and troponin T degradation, and myofibril fragmentation was greater upon incubation of isolated myofibrils with added Ca2+ in the order 800 mu M Ca2+ > 40 mu M Ca2+ > no Ca2+, suggesting that calpain-1 and calpain-2 were associated to myofibrils and proteolytically active with sufficient Ca2+. Activity of myofibril-bound calpain-1 in muscle incubated pre-rigor at 22 and 30 degrees C were higher than when incubated at 14 and 38 degrees C. These results indicate that calpains translocate from the sarcoplasm onto myofibrils with higher pre-rigor temperature to 30 degrees C and the proteolytic potential of myofibril-associated calpains is thereby increased.
