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Pulsatile shear stress increased mitochondrial membrane potential: implication of Mn-SOD.

ABSTRACT: Mitochondrial dysfunction is intimately involved in cardiovascular diseases. Mitochondrial membrane potential (DeltaPsi(m)) is coupled with oxidative phosphorylation to drive ATP synthesis. In this study, we examined the effect of physiological pulsatile shear stress (PSS) on DeltaPsi(m) and the role of Mn-SOD expression on DeltaPsi(m). Confluent human aortic endothelial cells (HAEC) were exposed to PSS, and DeltaPsi(m) was monitored using tetramethylrhodamine methyl ester (TMRM(+)), a mitochondrial membrane potential probe. PSS significantly increased DeltaPsi(m) and the change in DeltaPsi(m) was a dynamic process. DeltaPsi(m) returned to baseline level after PSS for 2h followed by static state for 4h. Mitochondrial Mn-SOD expression and activities were also significantly up-regulated in response to PSS. Silencing Mn-SOD attenuated PSS-mediated DeltaPsi(m) increase while adding Mn-SOD mimetic, MnTMPyP, increased DeltaPsi(m) to the similar extent as induced by PSS. Our findings suggest that PSS-increased mitochondrial DeltaPsi(m), in part, via Mn-SOD up-regulation.

SUBMITTER: Li R

PROVIDER: S-EPMC2746082 | biostudies-literature | 2009 Oct

REPOSITORIES: biostudies-literature

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Pulsatile shear stress increased mitochondrial membrane potential: implication of Mn-SOD.

Li Rongsong R Beebe Tyler T Cui Jeffrey J Rouhanizadeh Mahsa M Ai Lisong L Wang Pin P Gundersen Martin M Takabe Wakako W Hsiai Tzung K TK

Biochemical and biophysical research communications 20090808 2

Mitochondrial dysfunction is intimately involved in cardiovascular diseases. Mitochondrial membrane potential (DeltaPsi(m)) is coupled with oxidative phosphorylation to drive ATP synthesis. In this study, we examined the effect of physiological pulsatile shear stress (PSS) on DeltaPsi(m) and the role of Mn-SOD expression on DeltaPsi(m). Confluent human aortic endothelial cells (HAEC) were exposed to PSS, and DeltaPsi(m) was monitored using tetramethylrhodamine methyl ester (TMRM(+)), a mitochond ...[more]

PMID: 19666009

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Project description:Insects have developed a complex network of enzymatic antioxidant systems for handling reactive oxygen species (ROS) generated during stress. Superoxide dismutases (SODs) play a determinant role in balancing ROS in insect. However, studies devoted to SODs functions in insects under cold stress are limited. In the present study, we attempted to identify and characterize a mitochondrial manganese SOD (mMn-SOD) from the desert beetle Micordera punctipennis (denoted as MpmMn-SOD) and explore its protective effects on bacteria cells under cold stress. MpmMn-SOD is composed of 202 amino acids with conserved domains required for metal ions binding and enzyme activity. RT-qPCR experiments revealed that the expression of MpmMn-SOD was ubiquitous but tissue-specific and was induced by cold stress. An E. coli (BL21) system was applied to study the function of MpmMn-SOD. The MpmMn-SOD gene was cloned into the prokaryotic expression vector pET-32a to generate a recombinant plasmid pET-32a(MpmMn-SOD). After transformation of the plasmid into E. coli BL21, the fusion protein Trx-His-MpmMn-SOD was overexpressed and identified by SDS-PAGE and Western blotting. Antioxidant activity assay showed that the death zones of the transformed bacteria BL21 (pET32a-mMn-SOD) were smaller in diameter than the control bacteria BL21 (pET32a). Survival curves under -4 °C showed that BL21 (pET32a-mMn-SOD) had significant enhanced cold resistance compared to BL21 (pET32a). Its SOD activity under -4 °C had a significant negative correlation (r = - 0.995) with superoxide anion O2 •- content. Accordingly, under cold stress BL21 (pET32a-mMn-SOD) had lower electric conductivity and malondialdehyde (MDA) content than BL21 (pET32a). Taken together, our results showed that cold stress stimulated the expression of MpmMn-SOD in M. punctipennis. The E. coli cells that overexpress MpmMn-SOD increase their resistance to cold stress by scavenging ROS, and mitigate potential cell damage caused by ROS under cold conditions.

Project description:The objective of this study was to advance the understanding of how in vivo arterial shear forces affect vascular endothelial gene expression. Complicated blood flow patterns at arterial branches create small regions that experience fluctuations in shear stress at frequencies higher than the heart rate. To assess whether such temporal variations in shear stress can affect endothelial gene expression, a series of in vitro microarray experiments was performed. The effects of three sinusoidal waveforms (1, 2, and 3 Hz) and one physiological waveform were compared to the expression profiles under steady flow. At each frequency, three levels of mean shear stress (0, 7.5, and 15 dyn/cm2) were used. Porcine aortic endothelial cells were exposed for 24 hours to each combination, replicated four times. Following shear exposure, phase contrast images of the cells were acquired, and RNA was extracted for microarray analysis against about 10,000 porcine oligonucleotides. Cell alignment with the flow was positively correlated with mean shear (p < 0.001) and independent of frequency. A two-way ANOVA identified 232 genes that were differentially regulated by frequency. The frequency sensitive genes were clustered to identify groups of genes exhibiting similar frequency responses. The largest response was seen at 2 Hz. At this frequency, several inflammatory molecules were upregulated, including VCAM, CTGF, TGF-beta2, c-jun, and IL-8, indicating a potential endothelial atherosusceptibility at this frequency. Mean shear significantly affected the expression of ~3,000 genes. Purely oscillatory flow (zero mean shear) enhanced the expression of several growth factors and adhesion molecules (E-selectin, VCAM, MCP-1, IL-8, c-jun), relative to non-reversing flow (15 dyn/cm2 mean shear). The 2 Hz upregulation of certain atherogenic molecules such as VCAM, c-jun, and IL-8 was enhanced as the mean shear was reduced. Thus, the inflammatory response evoked at certain frequencies appears to be exacerbated by low, oscillatory shear. Keywords: Shear stress response

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Pulsatile shear stress increased mitochondrial membrane potential: implication of Mn-SOD.

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Pulsatile shear stress increased mitochondrial membrane potential: implication of Mn-SOD.

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