Biomedical subjects

Ronnie Blazev

Publications and source records attributed to Ronnie Blazev.

4 recordsLinked to original sources

PubMed2026

Quantitative interactome mapping of skeletal muscle insulin resistance.

Protein-protein interactions (PPIs) are dynamic and critical to adaptive homeostasis. While there have been massive efforts to catalogue proteome-wide PPIs, global quantification of changes remains a challenge. Here, we integrate dynamic protein correlation profiling - mass spectrometry (PCP-MS) and quantitative cross linking-mass spectrometry (qXL-MS) using multiplexed stable isotope labelling to characterise global PPI remodelling following the development of chronic skeletal muscle insulin resistance (IR) with or without acute insulin stimulation. We quantify >7,000 unique PPIs amongst 5,346 proteins and show changes in the interactome network dominate the proteome response. Our data show the dysregulation of protein processing in the endoplasmic/sarcoplasmic reticulum involving changes in PPIs with protein chaperones and disulfide isomerases is a major hallmark of skeletal muscle IR. Mechanistically, we show the dysregulation of PPIs with Protein-Disulfide Isomerase 6 (PDIA6) regulates cysteine oxidation and insulin sensitivity. Taken together, we show in vivo quantitative interactome mapping is a powerful approach to understand disease mechanisms and provide new insights into protein network re-organisations with IR.

Insulin Resistance↗

PubMed2005

Electrophoretic and functional identification of two troponin C isoforms in toad skeletal muscle fibers.

The differential sensitivity of frog twitch and slow-tonic fibers to Ca(2+) and Sr(2+) suggests that these two fiber types express different troponin C (TnC) isoforms. To date, only one TnC isoform from anurans (resembling the mammalian fast-twitch isoform) has been isolated and characterized. In this study, we examined the possibility that anuran striated muscle contains more than one TnC isoform. Toward this end, we determined the TnC isoform composition of 198 single fibers from the rectus abdominis of the cane toad (a mixed slow-tonic and twitch muscle) and of toad cardiac muscle using a method that enables the identification of TnC isoforms on the basis of the effect of Ca(2+) on their electrophoretic mobility. The fibers were typed according to their myosin heavy chain (MHC) isoform composition. The data indicate that striated muscle of the cane toad contains two TnC isoforms, one of which (TnC-t) is present in all fibers displaying only twitch MHC isoforms and the other of which (TnC-T/c) is present in fibers displaying the tonic MHC isoform and in cardiac muscle. For a subpopulation of 15 fibers, the TnC isoform composition was also compared with Ca(2+) and Sr(2+) activation characteristics. Fibers containing the TnC-T/c isoform were approximately 3-fold more sensitive to Ca(2+), approximately 40-fold more sensitive to Sr(2+), and responded to a approximately 4.6-fold broader range of [Ca(2+)] than did fibers containing the TnC-t isoform. The Ca(2+) activation properties of toad fibers containing the TnC-T/c isoform appear to be consistent with the previously reported physiological characteristics of amphibian slow-tonic muscle fibers.

Animals↗

PubMed2005

Glycogen content and contractile responsiveness to T-system depolarization in skinned muscle fibres of the rat.

1. Glycogen content (determined microfluorometrically), response capacity to transverse tubular (T) system depolarization and the relationship between these two parameters were examined in single, mechanically skinned fibres from rat extensor digitorum longus (EDL) muscle in the presence of high and constant concentrations of ATP and creatine phosphate. 2. The mean total glycogen content (tGlyc) in freshly dissected fibres was 58.1 +/- 4.2 mmol glucosyl units/L fibre (n = 53). 3. A large proportion of tGlyc was retained in the skinned fibres (SFGlyc) after 2 and 30 min exposure to an aqueous relaxing solution (73.1 +/- 2.8 and 64 +/- 12.3%, respectively). 4. When fibres were incubated for 30 min in a high (30 micromol/L)-Ca2+ solution, the proportion of SFGlyc was markedly lower (approximately 28%), which suggests that rat skinned fibres contain a Ca2+-sensitive glycogenolytic system. 5. In rat skinned fibres, T-system depolarization-induced Ca2+ release was not accompanied by a detectable loss of fibre glycogen and there was no correlation between response capacity and initial SFGlyc, indicating that other factors, unrelated to glycogen depletion, ultimately limited the capacity of rat skinned fibres to respond to T-system depolarization. 6. It is concluded that rat mechanically skinned fibre preparations are well suited for studies of glycogenolysis at a cellular level and that, with further refinement of the depolarization protocol, they may be suitable for studies of the non-metabolic role of glycogen in mammalian skeletal muscle contractility.

Animals↗

PubMed2004

Troponin C isoform composition determines differences in Sr(2+)-activation characteristics between rat diaphragm fibers.

Single fibers of rat diaphragm containing different naturally occurring combinations of myofibrillar protein isoforms were used to evaluate the contribution of troponin C (TnC) isoforms to fiber type-related differences with respect to sensitivity to Sr(2+) of the contractile system. Mechanically skinned fibers were studied for their isometric force vs. Sr(2+) concentration ([Sr(2+)]) relationships and then analyzed electrophoretically for myofibrillar protein isoform composition. Our data demonstrate that fiber-type differences in Sr(2+) dependence of contractile activation processes are primarily determined by the TnC isoform composition, with the slow isoform conferring on average a sevenfold greater sensitivity to Sr(2+) than the fast isoform. Moreover, the ratio of TnC isoforms determined functionally from the force-pSr (-log(10) [Sr(2+)]) curves is tightly (r(2) = 0.97) positively correlated with that estimated electrophoretically. Together, these results validate the use of Sr(2+) activation characteristics to distinguish fibers containing different proportions of fast and slow TnC isoforms and to study the mechanisms by which divalent cations activate the contractile apparatus. We also found that the functionally and electrophoretically determined ratios of TnC isoforms present in a fiber display similar sigmoidal relationships with the ratio of myosin heavy chain (MHC) isoform types expressed. These relationships 1) offer further insight in the functional and molecular expression of TnC in relation to the molecular expression of MHC isoform types and 2) may provide the basis for predicting sensitivity to Sr(2+), TnC, and MHC isoforms in pure and hybrid skeletal muscle fibers.

Animals↗