Loss of smooth muscle cell hypoxia inducible factor-1 alpha underlies increased vascular contractility in pulmonary hypertension
2017; 31 (2): 650-662
Pulmonary arterial hypertension (PAH) is an often fatal disease with limited treatment options. Whereas current data support the notion that, in pulmonary artery endothelial cells (PAECs), expression of transcription factor hypoxia inducible factor-1α (HIF-1α) is increased, the role of HIF-1α in pulmonary artery smooth muscle cells (PASMCs) remains controversial. This study investigates the hypothesis that, in PASMCs from patients with PAH, decreases in HIF-1α expression and activity underlie augmented pulmonary vascular contractility. PASMCs and tissues were isolated from nonhypertensive controls and patients with PAH. Compared with controls, HIF-1α and Kv1.5 protein expression were decreased in PAH smooth muscle cells (primary culture). Myosin light chain (MLC) phosphorylation and MLC kinase (MLCK) activity-major determinants of vascular tone-were increased in patients with PAH. Cofactors involved in prolyl hydroxylase domain activity were increased in PAH smooth muscle cells. Functionally, PASMC contractility was inversely correlated with HIF-1α activity. In PASMCs derived from patients with PAH, HIF-1α expression is decreased, and MLCK activity, MLC phosphorylation, and cell contraction are increased. We conclude that compromised PASMC HIF-1α expression may contribute to the increased tone that characterizes pulmonary hypertension.-Barnes, E. A., Chen, C.-H., Sedan, O., Cornfield, D. N. Loss of smooth muscle cell hypoxia inducible factor-1α underlies increased vascular contractility in pulmonary hypertension.
View details for DOI 10.1096/fj.201600557R
View details for Web of Science ID 000394235400018
Molecular characterization and functional properties of cardiomyocytes derived from human inducible pluripotent stem cells.
Journal of cellular and molecular medicine
2011; 15 (1): 38-51
In view of the therapeutic potential of cardiomyocytes derived from induced pluripotent stem (iPS) cells (iPS-derived cardiomyocytes), in the present study we investigated in iPS-derived cardiomyocytes, the functional properties related to [Ca(2+) ](i) handling and contraction, the contribution of the sarcoplasmic reticulum (SR) Ca(2+) release to contraction and the b-adrenergic inotropic responsiveness. The two iPS clones investigated here were generated through infection of human foreskin fibroblasts (HFF) with retroviruses containing the four human genes: OCT4, Sox2, Klf4 and C-Myc. Our major findings showed that iPS-derived cardiomyocytes: (i) express cardiac specific RNA and proteins; (ii) exhibit negative force-frequency relations and mild (compared to adult) post-rest potentiation; (iii) respond to ryanodine and caffeine, albeit less than adult cardiomyocytes, and express the SR-Ca(2+) handling proteins ryanodine receptor and calsequestrin. Hence, this study demonstrates that in our cardiomyocytes clones differentiated from HFF-derived iPS, the functional properties related to excitation-contraction coupling, resemble in part those of adult cardiomyocytes.
View details for DOI 10.1111/j.1582-4934.2009.00996.x
View details for PubMedID 20041972
View details for PubMedCentralID PMC3822492
Human embryonic stem cell-derived cardiomyocytes can mobilize 1,4,5-inositol trisphosphate-operated [Ca2+]i stores: the functionality of angiotensin-II/endothelin-1 signaling pathways.
Annals of the New York Academy of Sciences
2010; 1188: 68-77
Because previous findings showed that in human embryonic stem cell-derived cardiomyocytes (hESC-CM) the machinery for Ca2+-induced release of calcium is immature, we tested the hypothesis that hESC-CM contain functional 1,4,5-inositol triphosphate (IP3)-operated intracellular Ca2+ ([Ca2+]i) stores. We investigated the effects of angiotensin II (AT-II) and endothelin 1 (ET-1), which activate the 1,4,5-IP3 pathway, on [Ca2+]i transients and contractions in hESC-CM. Our major findings were that in hESC-CM, both AT-II (10(-9)-10(-7) M) and ET-1 (10(-9)-10(-7) M) exert inotropic and lusitropic effects. The involvement of 1,4,5-IP3-dependent intracellular Ca2+ release in AT-I-induced effects was supported by these findings: the effects of AT-II were blocked by 2-aminoethoxyphenyl borate (2-APB, a 1,4,5-IP3 receptor blocker) and U73122 (a phosopholipase C blocker); and hESC-CM express AT-II type 1 and IP3 type I and II receptors as determined by fluorescence immunostaining. In conclusion, hESC-CM exhibit functional AT-II and ET-1 signaling pathways, as well as 1,4,5-IP3-operated releasable Ca2+ stores.
View details for DOI 10.1111/j.1749-6632.2009.05085.x
View details for PubMedID 20201888
TVP1022 protects neonatal rat ventricular myocytes against doxorubicin-induced functional derangements.
The Journal of pharmacology and experimental therapeutics
2010; 332 (2): 413-20
Our recent studies demonstrated that propargylamine derivatives such as rasagiline (Azilect, Food and Drug Administration-approved anti-Parkinson drug) and its S-isomer TVP1022 protect cardiac and neuronal cell cultures against apoptotic-inducing stimuli. Studies on structure-activity relationship revealed that their neuroprotective effect is associated with the propargylamine moiety, which protects mitochondrial viability and prevents apoptosis by activating Bcl-2 and protein kinase C-epsilon and by down-regulating the proapoptotic protein Bax. Based on the established cytoprotective and neuroprotective efficacies of propargylamine derivatives, as well as on our recent study showing that TVP1022 attenuates serum starvation-induced and doxorubicin-induced apoptosis in neonatal rat ventricular myocytes (NRVMs), we tested the hypothesis that TVP1022 will also provide protection against doxorubicin-induced NRVM functional derangements. The present study demonstrates that pretreatment of NRVMs with TVP1022 (1 microM, 24 h) prevented doxorubicin (0.5 microM, 24 h)-induced elevation of diastolic [Ca(2+)](i), the slowing of [Ca(2+)](i) relaxation kinetics, and the decrease in the rates of myocyte contraction and relaxation. Furthermore, pretreatment with TVP1022 attenuated the doxorubicin-induced reduction in the protein expression of sarco/endoplasmic reticulum calcium (Ca(2+)) ATPase, Na(+)/Ca(2+) exchanger 1, and total connexin 43. Finally, TVP1022 diminished the inhibitory effect of doxorubicin on gap junctional intercellular coupling (measured by means of Lucifer yellow transfer) and on conduction velocity, the amplitude of the activation phase, and the maximal rate of activation (dv/dt(max)) measured by the Micro-Electrode-Array system. In summary, our results indicate that TVP1022 acts as a novel cardioprotective agent against anthracycline cardiotoxicity, and therefore potentially can be coadmhence, theinistered with doxorubicin in the treatment of malignancies in humans.
View details for DOI 10.1124/jpet.109.161158
View details for PubMedID 19915070
View details for PubMedCentralID PMC3202463
1,4,5-Inositol trisphosphate-operated intracellular Ca(2+) stores and angiotensin-II/endothelin-1 signaling pathway are functional in human embryonic stem cell-derived cardiomyocytes.
Stem cells (Dayton, Ohio)
2008; 26 (12): 3130-8
On the basis of previous findings suggesting that in human embryonic stem cell-derived cardiomyocytes (hESC-CM) the sarcoplasmic reticulum Ca(2+)-induced release of calcium machinery is either absent or immature, in the present study we tested the hypothesis that hESC-CM contain fully functional 1,4,5-inositol trisphosphate (1,4,5-IP(3))-operated intracellular Ca(2+) ([Ca(2+)](i)) stores that can be mobilized upon appropriate physiological stimuli. To test this hypothesis we investigated the effects of angiotensin-II (AT-II) and endothelin-1 (ET-1), which activate the 1,4,5-IP(3) pathway, on [Ca(2+)](i) transients and contractions in beating clusters of hESC-CM. Our major findings were that in paced hESC-CM both AT-II and ET-1 (10(-9) to 10(-7) M) increased the contraction amplitude and the maximal rates of contraction and relaxation. In addition, AT-II (10(-9) to 10(-7) M) increased the [Ca(2+)](i) transient amplitude. The involvement of 1,4,5-IP(3)-dependent intracellular Ca(2+) release in the inotropic effect of AT-II was supported by the findings that (a) hESC-CM express AT-II, ET-1, and 1,4,5-IP(3) receptors determined by immunofluorescence staining, and (b) the effects of AT-II were blocked by 2 microM 2-aminoethoxyphenyl borate (a 1,4,5-IP(3) receptor blocker) and U73122 (a phospholipase C blocker). In conclusion, these findings demonstrate for the first time that hESC-CM exhibit functional AT-II and ET-1 signaling pathways, as well as 1,4,5-IP(3)-operated releasable Ca(2+) stores.
View details for DOI 10.1634/stemcells.2008-0777
View details for PubMedID 18818435
Vagal stomach afferents inhibit somatic pain perception.
2005; 113 (3): 354-9
Vagal stimulation inhibits systemic pain perception in animals, probably via the nucleus tractus solitarius and its connections with descending nuclei in the brainstem which inhibit pain. Pain-inhibiting effects of such stimulation in humans, obtained from epileptic patients treated by vagal stimulation, are controversial. The aim of our study was to evaluate whether vagal stomach afferent activation inhibits pain perception in healthy humans. Pain thresholds, magnitude of tonic heat pain at 46 degrees C stimulation, pain temporal summation and laser pain evoked potentials were measured at the hand before and immediately after rapid drinking of 1500 ml water in 31 volunteers. We found an increase in heat pain threshold from 43.3+/-2.6 to 44.7+/-2.2 degrees C, P<0.0001, a decrease of peak pain magnitude to tonic heat from 56.3+/-26.2 to 43.7+/-25.8 (on 0-100 VAS), P<0.0001, a lowering of area under the curve during tonic noxious heat stimulus from 1962+/-984 to 1411+/-934, P<0.001. Additionally, we observed a decrease in the peak to peak evoked potential amplitude from 19.2 microV+/-1.2 to 15.6 microV+/-1.2 (P=0.005) together with a decrease in the estimation of mean laser induced pain from 52.28+/-18.00 to 48.14+/-20.18 (P=0.025). Mechanical pain thresholds and temporal summation did not change significantly. We conclude that vagal stomach afferents exert an inhibitory effect on somatic pain perception in humans.
View details for DOI 10.1016/j.pain.2004.11.012
View details for PubMedID 15661444