Smooth Muscle Mitochondria Research Articles

Cаlіx[4]аrene С-956 is effective inhibitor of Н(+)-Сa(2+)-exchanger in smooth muscle mitochondria

Cаlіx[4]аrene С-956 is effective inhibitor of Н(+)-Сa(2+)-exchanger in smooth muscle mitochondria

Effect of Calix[4]arenes on Ca2+ Transport, Electric Potential and Functioning of the Electron Transport Chain in Smooth Muscle Mitochondria

Effect of Calix[4]arenes on Ca2+ Transport, Electric Potential and Functioning of the Electron Transport Chain in Smooth Muscle Mitochondria

Calix[4]arenes as Modulators of Energy- Dependent Ca2+-Accumulation and Functioning of the Electron Transport Chain in Smooth Muscle Mitochondria

Calix[4]arenes as Modulators of Energy- Dependent Ca2+-Accumulation and Functioning of the Electron Transport Chain in Smooth Muscle Mitochondria

Abstract 16983: Inhibition of Dynamin-Related Protein 1 Accelerates Vascular Calcification in apoE-Deficient Mice through AKT Activation

Background: Mitochondrial dysfunction has been implicated in the pathogenesis of cardiovascular disease. Dynamin-related protein 1 (DRP1) regulates mitochondrial function by driving membrane fission and allowing mitochondria to divide. How DRP1 acts in atherosclerosis, including the development of vascular calcification however remains unknown. Methods and Results: Aortic smooth muscle mitochondria from apoE-deficient mice (apoE-/-) exhibited a 47% increase in size compared to mitochondria from wild-type mice fed the same atherogenic diet for 18 weeks (N=103-129 mitochondria from 3 mice/group; p<0.05). In agreement with the observed changes in mitochondrial morphology, inhibitory phosphorylation of DRP1 on Ser637 was enriched in apoE-/- mouse aortas. DRP1 immunoreactivity increased around calcified areas of apoE-/- mouse aortic sections. Additionally, DRP1 mRNA was increased 1.8-fold in calcified over non-calcified areas (N=8; p<0.05). To determine the role of DRP1 inhibition on aortic lesion calcification, apoE-/- mice maintained on an atherogenic diet for 13 weeks (a point prior to calcified lesion development) were subsequently given the DRP1 inhibitor, MDIVI-1 (50mg/kg, i.p., once a week) for 5 more weeks. MDIVI-1 administration led to a 65% increase in calcified lesion area without altering lesion size (N=26 mice/group; p<0.05). A single injection of MDIVI-1 resulted in a 3-fold increase in the ratio of phospho-AKT to total pan-AKT along with a strong increase in Runx2 protein in the apoE-/- mouse aorta 6 hours post-injection. MDIVI-1 administration did not alter bone remolding as detected by micro-CT in wild type or apoE-/- mice (N=5 mice/group). Conclusion: We provide novel roles of the mitochondrial fission protein DRP1 in the vascular calcification process, and demonstrate that inhibition of DRP1 through administration of the DRP1 inhibitor MDIVI-1 in apoE-/- mice accelerates vascular calcification through the AKT/Runx2 pathway.

Vascular Bioactivation of Nitroglycerin Is Catalyzed by Cytosolic Aldehyde Dehydrogenase-2

According to general view, aldehyde dehydrogenase-2 (ALDH2) catalyzes the high-affinity pathway of vascular nitroglycerin (GTN) bioactivation in smooth muscle mitochondria. Despite having wide implications to GTN pharmacology and raising many questions that are still unresolved, mitochondrial bioactivation of GTN in blood vessels is still lacking experimental support. In the present study, we investigated whether bioactivation of GTN is affected by the subcellular localization of ALDH2 using immortalized ALDH2-deficient aortic smooth muscle cells and mouse aortas with selective overexpression of the enzyme in either cytosol or mitochondria. Quantitative Western blotting revealed that ALDH2 is mainly cytosolic in mouse aorta and human coronary arteries, with only approximately 15% (mouse) and approximately 5% (human) of the enzyme being localized in mitochondria. Infection of ALDH2-deficient aortic smooth muscle cells or isolated aortas with adenovirus containing ALDH2 cDNA with or without the mitochondrial signal peptide sequence led to selective expression of the protein in mitochondria and cytosol, respectively. Cytosolic overexpression of ALDH2 restored GTN-induced relaxation and GTN denitration to wild-type levels, whereas overexpression in mitochondria (6-fold vs wild-type) had no effect on relaxation. Overexpression of ALDH2 in the cytosol of ALDH2-deficient aortic smooth muscle cells led to a significant increase in GTN denitration and cyclic GMP accumulation, whereas mitochondrial overexpression had no effect. The data indicate that vascular bioactivation of GTN is catalyzed by cytosolic ALDH2. Mitochondrial GTN metabolism may contribute to oxidative stress-related adverse effects of nitrate therapy and the development of nitrate tolerance.

The effect of moxifloxacin on apoptosis of airway smooth muscle cells and mitochondria membrane potential

To observe the effects of moxifloxacin at various concentrations on the expression of Caspase-3, the alteration of mitochondria membrane potential (ΔΨm) and the apoptosis of airway smooth muscle cells (ASMCs), and to explore the possible mechanisms. ASMCs were derived from rat airway tissues and cultured in vitro. The cells were randomly divided into 5 groups including a control group and 4 groups to which moxifloxacin was added at different concentrations (40, 80, 120, 200 mg/L, groups M40, M80, M120 and M200 respectively). Then the cells of different groups were incubated for 48 h. An apoptosis detection kit was used for annexin V and PI staining, and JC-1 probe was employed to measure mitochondrial depolarization in ASMCs, and the protein of Caspase-3 was measured by Western blot. The apoptosis rates of ASMCs in groups M40, M80, M120 and M200 were (2.95 ± 0.21)%, (7.39 ± 0.63)%, (13.39 ± 0.40)% and (21.20 ± 1.42)%, respectively, all of which were higher than that in the control group (0.94 ± 0.05)%, F = 399.77, P < 0.01. Furthermore, the concentration of moxifloxacin was positively related to the apoptosis rate (r = 0.974, P < 0.01). Compared to the control group (the ratio of orange-red fluorescence to green fluorescence was 10.02 ± 0.20), there was a shift from mitochondrial orange-red fluorescence to green fluorescence among groups with the concentrations of moxifloxacin increasing (6.54 ± 0.15, 4.48 ± 0.14, 2.25 ± 0.10 and 1.99 ± 0.12); the difference was significant (F = 1565.12, P < 0.01), and there was a dose-dependent response (r = -0.946, P < 0.01). The results of Western blot indicated that the expression of Caspase-3 increased with the concentrations of moxifloxacin increasing (0.45 ± 0.05, 0.59 ± 0.04, 0.69 ± 0.06 and 0.84 ± 0.04, respectively), and there was a very low expression of Caspase-3 in the control group (0.31 ± 0.03). The expression of Caspase-3 showed a positive correlation with the concentration of moxifloxacin (r = 0.979, P < 0.01). The apoptosis rate of ASMCs in the different groups had a remarkable correlation with the ΔΨm and Caspase-3 (r = -0.887, P < 0.01; r = 0.955, P < 0.01). There was also a remarkable negative correlation between ΔΨm and Caspase-3 (r = -0.951, P < 0.01). Moxifloxacin was shown to promote ASMC apoptosis by altering ΔΨm.

Contractility in vitro and mitochondrial response in small and large rabbit bowel after anastomosis.

Leakage of a large bowel anastomosis remains the most serious postoperative complication in gastrointestinal surgery. In a recent experimental study we found that surgically induced hypoxia resulted in more derangement of a variety of biochemical markers in the large bowel (LB) than in the small bowel (SB). We explored the question of whether spontaneous and agonist-induced contractility of SB and LB muscle strips was influenced by surgical procedures and how contractility was related to energetic oxidative metabolism capacity in smooth muscle mitochondria. Sixty male New Zealand rabbits were operated on under general anesthesia. Segments of ileum and colon were resected from each rabbit, and an end-to-end anastomosis was constructed. A representative subset of segments from SB (n = 14) and LB (n = 14) at time 0 was used as controls. Tracts containing an anastomosis were resected at days 2, 7, and 14 after operation. At each time point, 20 segments adjacent to the anastomosis of both SB and LB were used for tensiometric and biochemical studies. Tensiometric studies demonstrated modifications in the smooth muscle function at both the acute and chronic stages with intestinal inflammation that may contribute to surgical stress-associated abnormal motility. Biochemical data showed that the respiratory capacity of the resected LB was more impaired than that of the SB. In both SB and LB, changes in respiratory activity preceded tensiometric changes. Thus abnormalities of contractility after surgical stress are more evident in LB than SB in segments adjacent to the anastomoses. This could be the consequence of abnormal biochemical changes, as adenosine triphosphate (ATP) is required for membrane potential maintenance, calcium homeostasis, and actin-myosin interactions.

Intact smooth muscle metabolism: its responses to cyanide poisoning and pyruvate stimulation.

Smooth muscle mitochondria are unique in their compartmentalization of metabolism with the contractile proteins and putative role in cell fate choice. In this study we examine the relative and quantitative differences that smooth muscle mitochondria have with regard to cyanide inhibition. The effect of cyanide poisoning in the mitochondria of smooth muscle was examined in the intact porcine carotid artery and intact guinea pig stomach. The respiratory responses of these tissues were monitored in the presence of cyanide and following the addition of various metabolites. In HEPES buffer with 10 mM glucose as the substrate, it was found that the EC50 for cyanide inhibition was 0.11+/-0.02 and 0.14+/-0.02 mM in the pig carotid and guinea stomach respectively. We also found that the signaling metabolite, pyruvate could partially reverse this inhibition. With pyruvate (10 mM) as the substrate, the EC50 increased significantly to 6.52+/-0.11 (carotid artery) and 1.95+/-0. 30 (stomach) mM as well as being significantly different between the tissues. This apparent resistance to cyanide inhibition caused by pyruvate was lost when sodium bicarbonate buffer was used (EC50 in the presence of pyruvate of 0.13+/-0.04 and 0.25+/-0.04 for carotid and stomach respectively). HEPES buffer permitted pyruvate's protection from cyanide poisoning whereas bicarbonate buffer did not. The rate of respiration caused by pyruvate re-stimulation was not significantly different than control in the stomach (2.63+/-0.77 and 2.69+/-0.3??Mol O2/min/g dw respectively) but was significantly greater in the carotid artery. Therefore, smooth muscle with 10 mM pyruvate and 1 mM cyanide had a rate or respiration significantly greater than with 10 mM glucose alone (0.90+/-0.2 and 0.59+/-0. 06??Mol O2/min/g dw respectively). Using 31P NMR spectroscopy, we observed a complete normalisation of high energy phosphates and pH in the guinea pig stomach smooth muscle caused by pyruvate after cyanide poisoning. These results suggest that cyanide's toxicity of smooth muscle oxidative metabolism is affected by the buffer (HEPES versus Bicarbonate) and metabolic signalling molecules or substrates (pyruvate versus glucose) in which the tissues are exposed to as well as tissue to tissue variations.

Brain SPECT Imaging with Tc-99m HMPAO in Ophthalmoplegia Plus

In 14 patients with ophthalmoplegia plus Tc-99m hexamethylpropylene amineoxime (HMPAO) brain SPECT imaging was performed. Three patients showed bilateral parieto-occipital hypoperfusions, and two patients had left parietal defects (one with an additional right temporal lesion). Two patients presented with occipital hypoperfusions and one with a distinct hypoperfusion in the medial part of both frontal lobes. Hypoperfusion of the visual cortex might result from permanently decreased input from the degenerated retina. The hypoperfusion in the visual cortex and in other regions alternatively might reflect direct central nervous system (CNS) involvement due to neuronal degeneration or to impaired vascular function caused by morphologically changed mitochondria in smooth muscle and endothelial cells. The data suggest CNS involvement in ophthalmoplegia plus with abnormalities of cerebral blood flow similar to those found in other mitochondrial encephalomyopathies.

Quantitative electron microscopic study on mitochondria of smooth muscle in piqixuzheng rat

Johnson stated that man has always been interested in the structure of things in the world around him. In this regard, the electron microscopy, a modern technique, is used for research in traditional Chinese medicine. The terms and theories of TCM are quite different from those of Western medicine. The theory of TCM indicates that pi (spleen in TCM) has important roles in human health. According to the basic theory of TCM, one such function of the pi is nourishing the muscles. A significant decrease in the number of mitochondria with normal structure and a significant increase in the number of mitochondria with abnormal structure in skeletal muscle of piqixuzheng (spleen energy deficiency in TCM) rats have been reported. The present work is a quantitative electron microscopic study on mitochondria of smooth muscle in piqixuzheng rats.Male Wistar rats were used in this study. The piqixuzheng rats were produced by treatment with houpo (Magnolia officinalis), Zhishi(Fructus aurantii immaturus), dahuang (Radix et rhizoma) and fasting day other day for six weeks.

Specific, selective, and complete staining of neurons of the myenteric plexus, using Cuprolinic blue

Wholemounts consisting of both muscular layers of the rat's intestine have been stained with Cuprolinic blue and magnesium chloride. The procedure, which is specific for RNA, gives selective coloration of the somata of the neurons of the myenteric plexus. The neuronal nucleoli and Nissl substance are the only blue structures in such preparations. For descriptive and quantitative work, Cuprolinic blue is more selective than other cationic dyes, and it colors more neurons than can be shown by staining for mitochondrial NADH diaphorase. The absence of background color enables neurons to be identified more confidently and counted more quickly than is possible with other techniques, in which nuclei or mitochondria in smooth muscle and neuroglial cells are also stained. Numbers of myenteric neurons determined using the new method are higher than those obtained by other means.

Biochemical basis of the spasmolytic effect of isoquinoline and phenothiazine derivatives

Experiments with isolated ring-like strips of pig heart veins and arteries and those with rat small intestinal strips made in the presence of hyperpotassium contracture have demonstrated that spasmolytic activity of papaverine, nospa, nonachlazine, and ethmozine is a consequence of the reduced respiratory intensity and phosphorylation in smooth muscle mitochondria. No changes in the anaerobic glycolysis rate have been found after drug administrations.

Differences between esophageal body and lower esophageal sphincter in mitochondria of smooth muscle in opossum

In opossum, contractions of smooth muscle of the circular layer of the esophageal body resist oxygen deprivation, while tone of the lower esophageal sphincter requires oxygen. Thus, morphologic differences in mitochondria could distinguish the two types of muscle. Transverse strips, cut at regular intervals along the smooth-muscle segment of the esophageal body, were physiologically identified as muscle of body or sphincter. Transverse sections of the circular muscle layer were examined by transmission electron microscopy. Muscle cell profiles located between the ends of the nucleus and the tips of the cells were used to evaluate the number of mitochondrial profiles per muscle cell profile, mitochondrial mass (relative total area of mitochondrial profiles in each muscle cell profile), mitochondrial position (distance from the center of each mitochondrial profile to the nearest point on the cell membrane relative to mean radius of that muscle cell profile), and mass of smooth endoplasmic reticulum (relative total area of profiles in each muscle cell profile). In the nonnucleated part of the muscle cells, sphincter muscle had a greater ratio of mitochondrial profile area to cell profile area (p < 0.05), mitochondrial profiles in sphincter muscle were more central in location than in esophageal body muscle (p < 0.05), and sphincter muscle contained more endoplasmic reticulum than body muscle (p < 0.05). Sphincter muscle did not differ from body muscle in mean muscle cell profile area and number of mitochondrial profiles per muscle cell profile in this region of the muscle cell (p > 0.05).

Electron microscopy and electron probe analysis of mitochondrial cation accumulation in smooth muscle.

The contractile responses to barium and the ultrastructure and ionic composition of mitochondria were studied in vascular smooth muscle. In normal rabbit portal anterior mesenteric vein (PAMV) and main pulmonary artery (MPA) smooth muscle mitochondria were frequently associated with the surface vesicles. The average distance between the outer mitochondrial and inner surface vesicle membrane was 4-5 nm. Ba contractures of MPA were tonic and of PAMV were phasic. Incubation of MPA and PAMV with Ba resulted in the accumulation of mitochondrial granules, followed in the MPA by massive mitochondrial swelling. Oligomycin and anoxia inhibited the appearance of mitochondrial electron-opaque granules and prevented the Ba-induced mitochondrial swelling in the MPA. Electron probe analysis of mitochondria in PAMV incubated with Ba and containing granules showed characteristic Ba signals over the mitochondria. Electron probe X-ray microanalysis also showed a highly significant (P < 0.001) correlation of P with mitochondrial Ba, in an estimated elemental ratio of approximately 3 Ba/4 P. Mitochondrial granules were still prominent after block staining of the osmium-fixed, Ba-loaded PAMV, but electron probe microanalysis showed no Ba, but only U, emissions. Tissues incubated with strontium had electron-opaque mitochondrial granules and deposits in the sarcoplasmic reticulum. X-ray microanalysis of mitochondria containing granules showed the presence of characteristic Sr and Ca emissions. The presence of Sr was similarly verified in the sarcoplasmic reticulum. These findings indicate the energy dependent uptake of divalent cations, in association with phosphate, by mitochondria in vascular smooth muscle in situ and the possibility that mitochondria may contribute to the regulation of intracellular divalent cation levels in smooth muscle.