RESUMEN
Objective To observe the enzymatic changes of myocardial sarcoplasmic reticulum in rats after injecting endotoxin (LPS), and provide basic research results for the future study of myocardial sarcoplasmic reticulum dysfunction caused by LPS in rats. Methods Ten SD rats were randomly divided into blank control group and LPS injection group with 5 rats in each group. In the LPS injection group, endotoxin was injected into the tail vessels of the rats. Results The heart rate (HR) of the LPS injection group increased and was faster than that in the blank control group [(204±18) beat per min vs. (139±10) beat per min on the first day, and (199±22) beat per min vs. (143±17) beat per min on the next day, both P values were less than 0.05]. The mean arterial pressure (MAP) was lower than that of the blank control group on the first day [(87±12) mmHg vs. (102±7) mmHg, P<0.05]. Under light and electron microscope, the myocardial cells of rats with LPS injection were loosely arranged, with intercellular infiltration with inflammatory cells, muscle fibers broken, and difficult to identify the morphology of mitochondria and sarcoplasmic reticulum. Quantitative PCR results showed that after endotoxin injection, troponin (CASQ1), sodium-calcium exchanger (NCX), calmodulin phosphatase 1 (ppplCa), phospholipid protein (PLN), sarcoplasmic reticulum Ca2+-ATPase (SERCA2) increased significantly (P<0.05). Conclusion Endotoxin can inhibit cardiomyocyte function by affecting the activity of sarcoplasmic reticulum calcium regulatory protein-related enzymes through various mechanisms.
RESUMEN
Background: The pathogenesis of Graves’ ophthalmopathy (GO) and the mechanism for its link to thyroid autoimmunity is poorly understood. Our present research focuses on the role of the skeletal muscle calcium binding protein calsequestrin (CASQ1). Earlier studies showed that the CASQ1 gene was up-regulated in thyroid tissue from patients with GO compared to those with Graves’ hyperthyroidism (GH) without eye signs, however, the protein levels remained the same in both groups, raising the possibility that the orbital autoimmune reaction begins in the thyroid gland. Here, we measured the concentration of the CASQ1 protein in normal and Graves’ thyroid tissue, correlating levels with parameters of the eye signs, CASQ1 antibody levels and the CASQ1 gene polymorphism rs3838216, shown previously to be a risk factor for ophthalmopathy. Methods: The CASQ1 protein was measured by quantitative Western blotting. Following electrophoresis, samples were transblotted to polyvinyl difluoride (PVDF) membranes incubated with a 1:1000 dilution of a rabbit anti-CASQ1 antibody and incubated with an horseradish peroxidase (HRP)-conjugated goat anti-rabbit antibody, or anti-mouse antibodies for Glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The protein concentrations were determined from density quantification using the Quantity One 4.4.0 ChemiDoc program and expressed as pmol/mg total protein by reference to CASQ1 standards. Results: Western blot analysis showed the presence of two forms of CASQ1 in the thyroid, of 50 and 60 kDa molecular weight respectively. In thyroid tissues, the mean (± SD) (GO 54.9±86.8 pmol/mg) concentration of the CASQ1 protein (GH 37.1±51.8 pmol/mg) was significantly reduced in patients with Graves’ disease , with or without ophthalmopathy, compared to normal thyroid tissues from control subjects with multinodular goitre or thyroid cancer (144.3±162.5 pmol/mg). The difference between GO and GH was not significant. The decreased CASQ1 protein levels in Graves’ thyroid tissues correlated with the homozygous genotype of the rs3838216 CASQ1 polymorphism. A two-fold increase in Levels of CASQ1 protein in toxic nodules compared to Graves’ hyperthyroidism was markedly significant. Conclusions: Decreased CASQ1 in the thyroid tissues of patients with Graves’ disease compared to normal thyroid tissues from control subjects may reflect consumption of the protein in the course of an autoimmune reaction against CASQ1 in the thyroid. Comparing CASQ1 protein levels in thyroid tissue from five patients with toxic nodular goitre (74.5±52.8) to controls showed no significance. The relative two fold increase in CASQ1 levels in toxic nodules compared to Graves’ disease suggests that this is due to the autoimmune reaction rather than the hyperthyroidism.
RESUMEN
We characterized and compared the characteristics of Ca2+ movements through the sarcoplasmic reticulum of inferior oblique muscles in the various conditions including primary inferior oblique overaction (IOOA), secondary IOOA, and controls, so as to further understand the pathogenesis of primary IOOA. Of 15 specimens obtained through inferior oblique myectomy, six were from primary IOOA, 6 from secondary IOOA, and the remaining 3 were controls from enucleated eyes. Ryanodine binding assays were performed, and Ca2+ uptake rates, calsequestrins and SERCA levels were determined. Ryanodine bindings and sarcoplasmic reticulum Ca2+ uptake rates were significantly decreased in primary IOOA (p < 0.05). Western blot analysis conducted to quantify calsequestrins and SERCA, found no significant difference between primary IOOA, secondary IOOA, and the controls. Increased intracellular Ca2+ concentration due to reduced sarcoplasmic reticulum Ca2+ uptake may play a role in primary IOOA.