Differential role of prolyl hydroxylases on mitochondrial function of HL-1 cells in a model of iron deficiency.

Iron deficiency is observed in nearly half of the heart failure patients whilst closely correlated with mitochondrial dysfunction. Besides the structural roles in mitochondria, iron is also the cofactor of the hypoxia inducible factor (Hif) degradating enzyme, prolylhydroxylase, thereby Hif accumulation and its related metabolic effects commonly involve in iron deficiency. In this study, we used atrium derived HL-1 cells to investigate the effects of iron depletion on mitochondrial function under in vitro conditions. We aimed to discriminate the Hif dependent effects of iron deprivation on mitochondrial function to reveal the mechanisms leading to cardiac failure. For this purpose, HL-1 cells were either directly incubated with the iron chelating agent deferoxamine (DFO) or with dimethyloxalylglycine (DMOG, inhibitor of prolylhydroxylase). Mitochondrial function was evaluated by measuring cellular ATP content and mitochondrial potential (Ψ). According to our results, 48 h of DFO incubation affected cell viability and ATP production through further mechanisms additional to Hif-1α accumulation. Unlike DMOG group, DFO incubation did not disturb mitochondrial function probably due to its low permeability. Whether or not, prolyl hydroxylase inhibition without iron depletion may negatively affect mitochondrial function through Hif dependent mechanisms.

Enhancement of S1P-induced contractile response in detrusor smooth muscle of rats having cystitis.

Interstitial cystitis is a chronic disease characterized by lower abdominal pain and some nonspecific symptoms including an increase in urinary frequency and urgency. Sphingosine 1-phosphate (S1P) is a bioactive sphingolipid that controls smooth muscle tone via G-protein coupled receptors (S1P1-3 receptors). S1P production is known to take place both in physiological states and some pathological situations, such as in overactive bladder syndrome. The intracellular mechanism of S1P-induced contractile response was investigated in ß-escin permeabilized detrusor smooth muscle of rats having cyclophosphamide-induced cystitis. The bladder was isolated from rats and detrusor smooth muscle strips were permeabilized with ß-escin. S1P (50µM)-induced contraction and calcium sensitization response were significantly increased in cystitis. S1P-induced augmented contractile response was inhibited by S1P2 receptor antagonist JTE-013 and S1P3 receptor antagonist suramin. S1P2 receptor protein expressions were increased in cystitis, where no change was observed in S1P3 expressions between control and cystitis groups. S1P-induced contraction was reduced by Rho kinase (ROCK) inhibitor Y-27632 and protein kinase C (PKC) inhibitor GF-109203X in both control and cystitis group. S1P-induced increased calcium sensitization response was decreased by ROCK inhibitor and PKC inhibitor in cystitis. Our findings provide the first evidence that interstitial cystitis triggers S1P-induced increase in intracellular calcium in permeabilized detrusor smooth muscle of female rats. Both S1P2 and S1P3 receptors are involved in S1P mediated enhanced contractile response. The augmentation in S1P-induced contraction in interstitial cystitis involves both PKC and ROCK pathways of calcium sensitization.

Tumor necrosis factor-alpha antagonist administration recovers skeletal muscle dysfunction in ovariectomized rats.

Skeletal muscles deteriorate after ovariectomy. Molecular pathway of this deterioration has not been defined. Tumor necrosis factor (TNF)-alpha activation is assumed to trigger muscle atrophy and administration of its antagonist is hypothesized to recover this atrophy in rats. Slow-twitch soleus and fast-twitch extensor digitorum longus muscle functions were investigated in intact, ovariectomized (OVX), and OVX plus 10 µg/g/week TNF-alpha antagonist administered female rats. Maximum isometric twitch and tetanic contraction responses were lower in the OVX groups. Maximum isometric twitch amplitudes recovered in the extensor digitorum longus but not in the soleus muscles after TNF-alpha antagonist administration. The decrease in responses to tetanic stimulations recovered in the OVX-TNF group at frequencies higher than 20 Hz in both muscle types. OVX animals body weight was 21% higher than intact animals. Muscle weight to body weight ratios of the OVX groups were higher than the control group which recovered after TNF-alpha antagonist administration. Findings suggest that the functional loss in OVX rat muscles is TNF-alpha pathway dependent. Skeletal muscle atrophy and function after OVX recovered by TNF-alpha antagonist administration.