Isola 2017 MiPschool Obergurgl
Event: MiPschool Obergurgl 2017
Diabetic cardiomyopathy is one of the most common outcomes of Type I diabetes. Animal models of diabetes showed that, early in the development of diabetes, cardiac mitochondria bear defective complex I and II functions and it was hypothesized that this could lead to cardiomyocytes damage [1,2]. We wished to assess whether a prolonged diabetic condition (19-20 weeks) kept the same mitochondrial alterations, and if these could be associated to changes in mitochondrial morphology.
Diabetes was induced by a single injection of 65mg/kg of streptozotocin (STZ) i.p. on female Wistar Han rats, controls were injected with vehicle. One week after injection, and then monthly, blood level of glucose and ketone bodies was checked. After 19-20 weeks rats were euthanized, hearts were excised and 3 left and right ventricles (LV or RV, respectively) were pooled together, in order to isolate interfibrillar (IFM) or subsarcolemmal (SSM) LV or RV mitochondria . Mitochondrial oxidative phosphorylation (OXPHOS) was assessed by measuring, with a Clark-type electrode, on isolated mitochondria, oxygen consumption after the addition of different substrates. In addition to substrates directed to the specific OXPHOS complexes, we evaluated fatty acid oxidation by testing OXPHOS driven by PalCoA plus carnitine and malate or Palcarnitine plus malate. Moreover, global activity of CPT1 and CPT2 (carnitine palmitoyl-transferase one and two) was assessed by a spectrophotometric method. Finally, for ultrastructural studies we fixed both isolated mitochondria and myocardial tissue (for in situ mitochondria) by conventional transmission electron microscopy procedure. Samples were photographed with a JEOL 100S transmission electron microscope. Rats developed a diabetic syndrome as soon as one week after STZ injection, as they displayed higher blood glucose and ketone bodies than matched controls. At the end of the experimental period, diabetics maintained altered blood values and, at variance with controls, displayed low body weight, together with dyspepsia and corneal opacity.
Both in SSM and IFM, enzymatic activity of fatty acid import enzymes (CPT1+CPT2) was statistically significantly increased in STZ-treated rats compared to controls.
OXPHOS data showed that diabetes lightly impaired only complex II of RV mitochondria, as both IFM and SSM showed lower oxygen rates in state IV after succinate addiction. LV mitochondria, paradoxically, had a better oxygen consumption rate than controls for state III after glutamate, but this datum wasn’t paralleled by adding pyruvate as substrate, so it can’t be ascribed to a better efficiency of complex I, but, rather, to an improvement in glutamate uptake. LV diabetic mitochondria showed higher RCR for complex I, II and III, than controls. This could possibly suggest increased electron leakage in diabetic LV OXPHOS. Diabetic LV mitochondria also showed increased oxygen consumption rates in complex IV stimulated by high ADP concentration.
• Bioblast editor: Kandolf G
Labels: Pathology: Diabetes
Preparation: Isolated mitochondria
Coupling state: OXPHOS Pathway: F, N
Event: B2, Oral MiPschool Obergurgl 2017
Mitochondrial morphologies varied for isolated mitochondria compared to in situ ones, as it was already observed by Riva et al., 2005. In isolated mitochondria, we describe 5 main morphologies: lamellar, tubular, coiled tubular, mixed, unorganized. After diabetes, in isolated mitochondria cristae morphologies changed only in LV SSM and IFM. When we observed in situ mitochondria, we haven’t observed coiled tubular cristae, and instead of unorganized cristae, sometimes rarified cristae were evident. In diabetics, we haven’t observed a loss of cristae as it was described as secondary to diabetes, but instead, we found an increasing appearance of rarified cristae in every class checked. Rarified cristae were more similar to the shape that cristae developed in the akita mice model of type-I diabetes (4) than to STZ- induced cristae destruction as usually claimed . Rarified cristae number was almost null before diabetes, thus statistics was impossible to take. The only significantly statistic differences were in LV SSM with a decrease in lamellar and mixed cristae.
Taken together these data showed that after about 20 weeks from developing diabetes, rats have clearly developed a diabetic metabolic syndrome, but their heart mitochondrial impairment is less severe than it was described elsewhere [1-2]. Changes in morphologies were statistically significant only in left ventricle isolated mitochondria, and associating this with the maintenance of mitochondrial function would be tempting. On the other hand, in situ data showed, in diabetics, a shift from lamellar and mixed morphology to the rarified morphology, that has been shown in genetically diabetic akita mice with mild uncoupling mitochondria. Whether this mild impairment is due to gender differences or to other causes, such as morphological changes due to mitofusin expression, will be pursued next.
- Isola R(1), Casti A(1), Broccia F(2), Loy F(1), Isola M(1), Vargiu R(2)
- Div Cytomorphol
- Div Physiol
- Dept Biomedical Sciences, Univ Cagliari, Cittadella Univ Monserrato.– email@example.com
- This work was supported by the Sardinia Region Government (RAS), L.R. 7/2007, Bando ricerca di Base 2012 (Grant CRP 60052).
- Vazquez EJ, Berthiaume JM, Kamath V, Achike O, Buchanan E, Montano M, Chandler MP, Miyagi M, Rosca MG (2015) Mitochondrial complex I defect increased fatty acid oxidation enhance protein lysine acetylation in the diabetic heart. Cardiovascular Research 107:453-65.
- Shen X, Zheng S, Thongboonkerd V, Xu M, Pierce WM Jr, Klein JB, Epstein PN (2004) Cardiac mitochondrial damage and biogenesis in a chronic model of type 1 diabetes. Am J Physiol - Endocrinol Metabol 287:E896-905.
- Rosca MG, Vazquez EJ, Kerner J, Parland W, Chandler MP, Stanley W, Sabbah HN, Hoppel CL (2008) Cardiac mitochondria in heart failure: decrease in respirasomes and oxidative phosphorylation. Cardiovascular Research 80:30–9.
- Bugger H, Boudina S, Hu X X, Tuinei J, Zaha VG, Theobald HA, Yun UJ, McQueen AP, Wayment B, Litwin SE, Abel ED (2008) Type 1 diabetic akita mouse hearts are insulin sensitive but manifest structurally abnormal mitochondria that remain coupled despite increased uncoupling protein 3. 2008 Diabetes 57:2924–32.