- Email: [email protected]
reperfusion insult
International Journal of Cardiology 178 (2015) 200–202
Contents lists available at ScienceDirect
International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard
Letter to the Editor
Estrogen enhancement of SGK1 expression induced by urocortin contributes to its cardioprotection against ischemia/reperfusion insult☆ Binhai Cong a, Jiankui Du a, Xiaoyan Zhu a, Jianqiang Lu b,⁎, Xin Ni a,b,⁎⁎ a b
Department of Physiology, Second Military Medical University, Shanghai 200433, China School of Kinesiology, Key Laboratory of Exercise and Health Sciences of Ministry of Education, Shanghai University of Sport, Shanghai 200438, China
a r t i c l e
i n f o
Article history: Received 23 September 2014 Accepted 21 October 2014 Available online 22 October 2014 Keywords: Estrogen Serum and glucocorticoid-responsive kinase1 Cardiovascular
Serum and glucocorticoid-responsive kinase1 (SGK1) is a serine– threonine kinase, which has been reported to play a key role in cell survival in various tissues including the myocardium [1]. As estrogen has cardioprotective effects [2], we explored whether SGK1 contributes to estrogen cardioprotection. As shown in Fig. 1, ovariectomized (Ovx) rats showed deceased SGK1 mRNA and protein expression in the myocardium, which was reversed by 17β-estradiol (E2, 30 μg/kg/day, sc) replacement for 8 weeks. In order to elucidate whether estrogen modulates SGK1 expression in cardiomyocytes, we examined the effects of E2 on SGK1 expression in neonatal rat cardiomyocytes in vitro. It was found that treatment with E2 (0.1–100 nmol/L) for 24 h did not affect SGK1 mRNA and protein levels in cardiomyocytes (Fig. 1B), which indicates that E2 might regulate SGK1 expression via an indirect mechanism in the myocardium. Recently, we demonstrated that urocortin1 (UCN1) protects cardiomyocytes against hypoxia/reoxygenation (H/R) insult through upregulation of SGK1 expression [3], and E2 plays a critical role in the maintenance of corticotrophin-releasing hormone receptor 2 (CRHR2) expression in cardiomyocytes [4,5]. We therefore hypothesized that E2 might enhance UCN1-induced SGK1 expression. Primary cardiomyocytes were treated with E2 (10 nmol/L) plus UCN1 (10 nmol/L) for 24 h. Compared ☆ This work is supported by the National Natural Science Foundation of China (No. 31000516 and No. 31371197), Key Laboratory of Exercise and Health Sciences of Ministry of Education, Shanghai University of Sport and Innovation Training Program for Undergraduates, and Education Commission of Shanghai Municipality. ⁎ Correspondence to: J. Lu, School of Kinesiology, Shanghai University of Sports, 399 Changhai Road, Shanghai 200438, China. ⁎⁎ Correspondence to: X. Ni, Department of Physiology, Second Military Medical University, Shanghai 200433, China. E-mail addresses: [email protected] (J. Lu), [email protected] (X. Ni).
http://dx.doi.org/10.1016/j.ijcard.2014.10.113 0167-5273/© 2014 Elsevier Ireland Ltd. All rights reserved.
with UCN1 treatment alone, E2 did increase SGK1 mRNA and protein levels in the presence of UCN1 (Fig. 1C). As UCN1 has been shown to be released from cardiomyocytes under ischemia [6], we examined whether E2 increases SGK1 expression under H/R condition. Treatment of cardiomyocytes with E2 (10 nmol/L) under the simulated H/R condition resulted in an increase in SGK1 expression, which was abolished by CRHR2 antagonist (Fig. 1D). These data indicate that E2 induces SGK1 expression under H/R, which might be through enhancement of the UCN1 effect. To examine whether SGK1 is involved in the cardioprotective effects of E2, we knocked down SGK1 expression by specific SGK1 siRNA, and then the cardioprotective effects of E2 upon H/R in cardiomyocytes were examined. As shown in Fig. 2, the E2 induced increasing cell viability, decreasing LDH release and inhibiting cleaved caspase 3 level were all attenuated by SGK1 siRNA (Fig. 2A–C). These data indicate that SGK1 involves in E2 cardioprotection against H/R injury. In summary, our data demonstrated that E2 regulates SGK1 expression via enhancing UCN1 action in the myocardium, which contributes to cardioprotective effects of E2 against H/R. These findings provide new evidences that UCN1-CRHR2 system plays important roles in the cardiac effect of E2 [4,5]. However, our findings that SGK1 is involved in E2 cardioprotection were on the basis of experiments in vitro. Further studies are required to confirm the role of SGK1 in estrogen cardioprotection. The authors of this manuscript have certified that they comply with the Principles of Ethical Publishing in the International Journal of Cardiology. References [1] T. Aoyama, T. Matsui, M. Novikov, J. Park, B. Hemmings, A. Rosenzweig, Serum and glucocorticoid-responsive kinase-1 regulates cardiomyocyte survival and hypertrophic response, Circulation 111 (13) (2005) 1652–1659. [2] E. Murphy, Estrogen signaling and cardiovascular disease, Circ. Res. 109 (6) (2011) 687–696. [3] B. Cong, L. Wang, X. Zhu, X. Li, B. Liu, X. Ni, SGK1 is involved in cardioprotection of urocortin-1 against hypoxia/reoxygenation in cardiomyocytes, Can. J. Cardiol. 30 (6) (2014) 687–695. [4] B. Cong, X. Zhu, B. Cao, J. Xiao, Z. Wang, X. Ni, Estrogens protect myocardium against ischemia/reperfusion insult by up-regulation of CRH receptor type 2 in female rats, Int. J. Cardiol. 168 (5) (2013) 4755–4760. [5] B. Cong, Y. Xu, H. Sheng, X. Zhu, L. Wang, W. Zhao, Z. Tang, J. Lu, X. Ni, Cardioprotection of 17β-estradiol against hypoxia/reoxygenation in cardiomyocytes is partly through up-regulation of CRH receptor type 2, Mol. Cell. Endocrinol. 382 (1) (2014) 17–25. [6] R.A. Knight, C. Chen-Scarabelli, Z. Yuan, et al., Cardiac release of UCN precedes the occurrence of irreversible myocardial damage in the rat heart exposed to I/R injury, FEBS Lett. 582 (6) (2008) 984–990.
B. Cong et al. / International Journal of Cardiology 178 (2015) 200–202
201
Fig. 1. The effect of estrogen on SGK1 mRNA and protein expression in the myocardium and cardiomyocytes. A. Effect of Ovx and E2 replacement (Ovx-E2) on SGK1 expression in rat myocardium. E2 was administrated (30 μg/kg/day, sc) for 8 weeks. The mRNA and protein level of SGK1 in the myocardium was determined by real-time RT-PCR and Western blot, respectively. n = 10 in each group. *P b 0.05, **P b 0.01 compared with sham; B–D. Effect of E2 on SGK1 expression in cardiomyocytes in vitro. B. Neonatal rat cardiomyocytes were treated with E2 (0.1–100 nmol/L) for 24 h. C. Cells were treated with E2 (10 nmol/L) plus UCN1 (10 nmol/L) for 24 h. D. Cells were treated with E2 (10 nmol/L) plus CRHR2 antagonist astrissin2B (Ast2B) (10 nmol/L) under H/R for 24 h. Four cultures (n = 4) were performed in triplicate. *P b 0.05, **P b 0.01 compared with control; #P b 0.05. Relative SGK1 was normalized to β-actin and presented as mean ± SD. Representative Western blot bands are presented on the top of the corresponding graphs.
202
B. Cong et al. / International Journal of Cardiology 178 (2015) 200–202
Fig. 2. SGK1 involves in the protective effects of E2 upon H/R injury in cardiomyocytes. Cardiomyocytes were transfected with control siRNA or SGK1 siRNA for 24 h, and then treated with E2 (10 nmol/L) under H/R for 24 h. A. Overall cell viability assessed by MTT method; B. cell damage determined by supernatant LDH concentration; C. apoptosis assessed by cleaved caspase3 level in cardiomyocytes; relative cleaved caspase3 level was normalized to β-actin level and the representative Western blotting bands are on the top of corresponding graphs. All data are presented as mean ± SD for four cultures (n = 4) performed in triplicate. *P b 0.05, **P b 0.01.
Contents lists available at ScienceDirect
International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard
Letter to the Editor
Estrogen enhancement of SGK1 expression induced by urocortin contributes to its cardioprotection against ischemia/reperfusion insult☆ Binhai Cong a, Jiankui Du a, Xiaoyan Zhu a, Jianqiang Lu b,⁎, Xin Ni a,b,⁎⁎ a b
Department of Physiology, Second Military Medical University, Shanghai 200433, China School of Kinesiology, Key Laboratory of Exercise and Health Sciences of Ministry of Education, Shanghai University of Sport, Shanghai 200438, China
a r t i c l e
i n f o
Article history: Received 23 September 2014 Accepted 21 October 2014 Available online 22 October 2014 Keywords: Estrogen Serum and glucocorticoid-responsive kinase1 Cardiovascular
Serum and glucocorticoid-responsive kinase1 (SGK1) is a serine– threonine kinase, which has been reported to play a key role in cell survival in various tissues including the myocardium [1]. As estrogen has cardioprotective effects [2], we explored whether SGK1 contributes to estrogen cardioprotection. As shown in Fig. 1, ovariectomized (Ovx) rats showed deceased SGK1 mRNA and protein expression in the myocardium, which was reversed by 17β-estradiol (E2, 30 μg/kg/day, sc) replacement for 8 weeks. In order to elucidate whether estrogen modulates SGK1 expression in cardiomyocytes, we examined the effects of E2 on SGK1 expression in neonatal rat cardiomyocytes in vitro. It was found that treatment with E2 (0.1–100 nmol/L) for 24 h did not affect SGK1 mRNA and protein levels in cardiomyocytes (Fig. 1B), which indicates that E2 might regulate SGK1 expression via an indirect mechanism in the myocardium. Recently, we demonstrated that urocortin1 (UCN1) protects cardiomyocytes against hypoxia/reoxygenation (H/R) insult through upregulation of SGK1 expression [3], and E2 plays a critical role in the maintenance of corticotrophin-releasing hormone receptor 2 (CRHR2) expression in cardiomyocytes [4,5]. We therefore hypothesized that E2 might enhance UCN1-induced SGK1 expression. Primary cardiomyocytes were treated with E2 (10 nmol/L) plus UCN1 (10 nmol/L) for 24 h. Compared ☆ This work is supported by the National Natural Science Foundation of China (No. 31000516 and No. 31371197), Key Laboratory of Exercise and Health Sciences of Ministry of Education, Shanghai University of Sport and Innovation Training Program for Undergraduates, and Education Commission of Shanghai Municipality. ⁎ Correspondence to: J. Lu, School of Kinesiology, Shanghai University of Sports, 399 Changhai Road, Shanghai 200438, China. ⁎⁎ Correspondence to: X. Ni, Department of Physiology, Second Military Medical University, Shanghai 200433, China. E-mail addresses: [email protected] (J. Lu), [email protected] (X. Ni).
http://dx.doi.org/10.1016/j.ijcard.2014.10.113 0167-5273/© 2014 Elsevier Ireland Ltd. All rights reserved.
with UCN1 treatment alone, E2 did increase SGK1 mRNA and protein levels in the presence of UCN1 (Fig. 1C). As UCN1 has been shown to be released from cardiomyocytes under ischemia [6], we examined whether E2 increases SGK1 expression under H/R condition. Treatment of cardiomyocytes with E2 (10 nmol/L) under the simulated H/R condition resulted in an increase in SGK1 expression, which was abolished by CRHR2 antagonist (Fig. 1D). These data indicate that E2 induces SGK1 expression under H/R, which might be through enhancement of the UCN1 effect. To examine whether SGK1 is involved in the cardioprotective effects of E2, we knocked down SGK1 expression by specific SGK1 siRNA, and then the cardioprotective effects of E2 upon H/R in cardiomyocytes were examined. As shown in Fig. 2, the E2 induced increasing cell viability, decreasing LDH release and inhibiting cleaved caspase 3 level were all attenuated by SGK1 siRNA (Fig. 2A–C). These data indicate that SGK1 involves in E2 cardioprotection against H/R injury. In summary, our data demonstrated that E2 regulates SGK1 expression via enhancing UCN1 action in the myocardium, which contributes to cardioprotective effects of E2 against H/R. These findings provide new evidences that UCN1-CRHR2 system plays important roles in the cardiac effect of E2 [4,5]. However, our findings that SGK1 is involved in E2 cardioprotection were on the basis of experiments in vitro. Further studies are required to confirm the role of SGK1 in estrogen cardioprotection. The authors of this manuscript have certified that they comply with the Principles of Ethical Publishing in the International Journal of Cardiology. References [1] T. Aoyama, T. Matsui, M. Novikov, J. Park, B. Hemmings, A. Rosenzweig, Serum and glucocorticoid-responsive kinase-1 regulates cardiomyocyte survival and hypertrophic response, Circulation 111 (13) (2005) 1652–1659. [2] E. Murphy, Estrogen signaling and cardiovascular disease, Circ. Res. 109 (6) (2011) 687–696. [3] B. Cong, L. Wang, X. Zhu, X. Li, B. Liu, X. Ni, SGK1 is involved in cardioprotection of urocortin-1 against hypoxia/reoxygenation in cardiomyocytes, Can. J. Cardiol. 30 (6) (2014) 687–695. [4] B. Cong, X. Zhu, B. Cao, J. Xiao, Z. Wang, X. Ni, Estrogens protect myocardium against ischemia/reperfusion insult by up-regulation of CRH receptor type 2 in female rats, Int. J. Cardiol. 168 (5) (2013) 4755–4760. [5] B. Cong, Y. Xu, H. Sheng, X. Zhu, L. Wang, W. Zhao, Z. Tang, J. Lu, X. Ni, Cardioprotection of 17β-estradiol against hypoxia/reoxygenation in cardiomyocytes is partly through up-regulation of CRH receptor type 2, Mol. Cell. Endocrinol. 382 (1) (2014) 17–25. [6] R.A. Knight, C. Chen-Scarabelli, Z. Yuan, et al., Cardiac release of UCN precedes the occurrence of irreversible myocardial damage in the rat heart exposed to I/R injury, FEBS Lett. 582 (6) (2008) 984–990.
B. Cong et al. / International Journal of Cardiology 178 (2015) 200–202
201
Fig. 1. The effect of estrogen on SGK1 mRNA and protein expression in the myocardium and cardiomyocytes. A. Effect of Ovx and E2 replacement (Ovx-E2) on SGK1 expression in rat myocardium. E2 was administrated (30 μg/kg/day, sc) for 8 weeks. The mRNA and protein level of SGK1 in the myocardium was determined by real-time RT-PCR and Western blot, respectively. n = 10 in each group. *P b 0.05, **P b 0.01 compared with sham; B–D. Effect of E2 on SGK1 expression in cardiomyocytes in vitro. B. Neonatal rat cardiomyocytes were treated with E2 (0.1–100 nmol/L) for 24 h. C. Cells were treated with E2 (10 nmol/L) plus UCN1 (10 nmol/L) for 24 h. D. Cells were treated with E2 (10 nmol/L) plus CRHR2 antagonist astrissin2B (Ast2B) (10 nmol/L) under H/R for 24 h. Four cultures (n = 4) were performed in triplicate. *P b 0.05, **P b 0.01 compared with control; #P b 0.05. Relative SGK1 was normalized to β-actin and presented as mean ± SD. Representative Western blot bands are presented on the top of the corresponding graphs.
202
B. Cong et al. / International Journal of Cardiology 178 (2015) 200–202
Fig. 2. SGK1 involves in the protective effects of E2 upon H/R injury in cardiomyocytes. Cardiomyocytes were transfected with control siRNA or SGK1 siRNA for 24 h, and then treated with E2 (10 nmol/L) under H/R for 24 h. A. Overall cell viability assessed by MTT method; B. cell damage determined by supernatant LDH concentration; C. apoptosis assessed by cleaved caspase3 level in cardiomyocytes; relative cleaved caspase3 level was normalized to β-actin level and the representative Western blotting bands are on the top of corresponding graphs. All data are presented as mean ± SD for four cultures (n = 4) performed in triplicate. *P b 0.05, **P b 0.01.