6,102

The Isoform Spesific Roles of Rho-Kinases in Vascular Diseases

Figen Çiçek, Murat Ayaz

Figen Çiçek, Department of Biophysics, Faculty of Medicine, Cukurova University, Adana, Turkey
Murat Ayaz, Department of Biophysics, Faculty of Medicine, Selçuk University, Konya, Turkey

Correspondence to: Murat Ayaz, Department of Biophysics, Faculty of Medicine, Selçuk University, Konya, Turkey.
Email: ayaz72@yahoo.com
Telephone: +90-332-2243863
Received: July 23, 2015
Revised: November 28, 2015
Accepted: December 2, 2015
Published online: December 10, 2015

ABSTRACT

A small GTPase RhoA and its downstream effectors Rho-associated protein kinases (ROCK) signaling pathway activation mediate smooth muscle contraction. ROCKs inhibit myosin light chain phosphatase (MLCP) dephosphorylation and therefore reduce relaxation. However, nitric oxide (NO) that is produced and released from endothelial cells has an inhibitory effect on the ROCK pathway in vasculature. Studies in which ROCK activity was inhibited by variety of pharmacological agents (HA1077 or Y-27632) have shown that it has some critical effects on systemic diseases like hypertension or diabetes mellitus. Indeed this activity may show isoform specificity (ROCK1 or ROCK2) dependent on the pathology. Therefore, in vascular pathogenesis ROCK pathway with its isoforms also need to be considered due to its direct effects on the vasoconstriction.

© 2015 ACT. All rights reserved.

Key words:Rho Kinases; ROCK1; ROCK2; Vasoconstriction; Vascular diseases

Çiçek F, Ayaz M. The Isoform Spesific Roles of Rho-Kinases in Vascular Diseases. Journal of Cardiology and Therapy 2015; 2(6): 465-468 Available from: URL: http://www.ghrnet.org/index.php/jct/article/view/1517

INTRODUCTION

Rho-associated protein kinases (ROCKs) play a critical role in smooth muscle contraction and relaxation. After activation of small GTPase RhoA its effector protein ROCK mediates contraction. Briefly after a signal arrives to cell membrane and activates membrane receptors or voltage operated Ca2+ channels (VOCC) free cytoplasmic Ca2+ concentration increases. Then Ca2+ binds to calmodulin and activates myosin light chain kinase (MLCK). MLCK phosphorylates myosin light chains (MLC), which are the regulatory subunits of the myosin heads.

MLCK phosphorylates (MLC) the subunits of the myosin heads. Phosphorylated MLCs enable the cross bridges between myosin and actin and so contraction occurs in smooth muscles[1,2]. On the other hand RhoA mediated ROCK is the other mediator of contraction with Ca2+[3]. Once activated ROCK provides continuation of contractile activity by inhibiting MLC phosphatase (MLCP) which dephosphorylate MLC and induces relaxation in smooth muscle cells. Together with Ca2+, ROCK pathways precisely control the vasoconstriction. In the arteries smooth muscle contraction is directly effects blood pressure by regulating the vessel diameter and tension[1,4]. In addition to vasoconstrictor effectors, endothelial derived Nitric Oxide (NO) is a vasodilator agent for smooth muscle cells which regulates the relaxation through cGMP pathway and also reduces ROCK activity and thereby contraction[5-8] (Figure 1). In the regulation of the vascular tonus these vasoconstrictor and vasodilator pathways mediate contraction. In recent years many studies has shown that Rho Kinase pathway should be taken into consideration in treatments of vascular diseases[9,10].

ROCK consists of two isoforms ROCK1 and ROCK2. ROCK1 enzyme is expressed in a plenty of different tissues like lung, kidney, stomach whereas ROCK2 is mostly expressed in heart, brain and skeletal muscle[11]. Cellular localization of the ROCK1 and ROCK2 also show diversity. ROCK1 is mostly localized at plasma membrane but ROCK2 at centrosomes of smooth muscle cells. At cardiomyocytes ROCK2 localized at intercalated discs, and at skeletal muscle cells Z-discs and sarcoplasmic reticulum[12].

Although, they have high genetic homology in their kinase domain (92%)[13], Yoneda et al[14] (2005) showed that they have isoform specific functions even in the same cell. According to this study ROCK1 is more active in focal adhesion and fiber formations rather than ROCK2 in primary rat embryo fibroblasts. Conversely, ROCK2 is the primer player in phagocytic activity. In another study it was shown that ROCK1 and ROCK2 have distinct roles in adhesion and differentiation in keratinocytes[15]. Also the experiments performed with the cells which derived from ROCK1 and 2 knockout animals, was shown that ROCK 1 acts in MLC2 phosphorylation and cell detachment, whereas ROCK2 in actin cytoskeleton stabilization[16]. Their mechanistic difference was indicated by Wang Y et al[17] (2009) that ROCK2 can bind directly to the myosin-binding subunit (MYPT1) of myosin phosphatase but not ROCK1. This difference reflects that ROCK1 use intermediate components for MLCP inhibition and this fringed pathway selection makes harder to understand the ROCK activity.

However lack of the isoform specific ROCK blockers (HA1077 and Y27632 are nonspecific blockers of ROCK) make difficult to distinguish the functional roles of the isoforms. But such a critical pathway that regulates constrictive mechanisms in vascular system deserves more precise evaluation. Therefore in this review we tried to we focus on these functional differences between two isoforms in vasculature from diseases perspective.

ROCK Isoforms in Arterial and Pulmonary Arterial Hypertension

Smooth muscle cells primarily regulate vascular volume and thereby blood pressure in the aorta. The role of the vasoconstrictors on the hypertension is a detailed examined in many studies concluded ROCK also play a major role in persistency of the high pressure in aorta[2,18].

It was shown in the hypertensive animal models that ROCK mediated vasoconstriction is involved arterial hypertension by blocking its function with Y-27632[19]. Other selective ROCK inhibitor HA1077 named as fasudil is believed to be a key therapeutic for human use. In one study dealing with hypertensive patients it was shown that the fasudil induce a vasodilator effect on the arterial pressure[20]. Also in a study Fukumoto et al[21] showed the effects of the fasudil on the patients with pulmonary arterial (PA) hypertension. The treatment with fasudil hydrochloride caused a slight decrease in the PA hypertension.

In both of these arterial high pressure diseases differences in the expression levels of ROCK1 and ROCK2 were observed. The immunostaining experiments ROCK2 (but not ROCK1) showed that its expression increases in arterioles of the lung sections taken from the PA hypertension patients[22]. The same study indicated that the hypoxia induced PA hypertension with vascular smooth muscle specific ROCK2 gene knockout mice, the right ventricular systolic pressure was significantly reduced versus control. Their findings indicate the importance of ROCK2 for the development of hypoxia-induced PA hypertension. Also ROCK2 gene silencing was improved erectile function on spontaneously hypertensive rats suggesting ROCK2 inhibition can be used as a specific therapeutic target for vascular dysfunctions caused by hypertension[23].

Atherosclerosis

When dealing with this very complicated inflammatory disease we see that on tunica intima, the layer surrounded with the formations such foam cells (monocytes/macrophages) that decrease vessel diameter and even make it more stiffening[24,25]. Impaired endothelium activity (endothelium dysfunction) causes dysregulation of NO release, which was thought as a major responsible factor for the initiation of atherosclerosis[26,27]. According to the study of Anju Nohria et al[28] ROCK inhibition with fasudil caused endothelial dependent vasodilation in the patients with coronary artery disease. Their measurements with brachial artery ultrasonography suggest the relation between endothelium activity and ROCK inhibition in atherosclerosis. In another study with mice ROCK inhibition with Y27632 results a protection against atherosclerosis by reducing significantly size of the atherosclerotic plaque formation significantly[29].

The individual roles of ROCK1 and ROCK2 in atherosclerosis tried to be explained in several studies. ROCK1 knockout was decreased atherosclerotic lesion formations in aortas from the bone marrow (BM) derived macrophage transplanted LDLr knockout mice[30]. While the experiments with ROCK2 lacking in the cultured BM differentiated macrophages was shown the importance of ROCK2 in the foam cell formations[31].

FUTURE DEVELOPMENT AND RESEARCH

mHealth in CHD prevention remains an emerging area of research. To try to estimate how much evidence can be built in the next few years, we conducted a search on Clinicaltrials.gov database using the search terms mHealth, mobile health, mobile phone, cell phone OR smartphone AND cardiovascular in June 2015. We found another five ongoing RCTs evaluating either text-messaging or smartphone apps for either primary or secondary prevention of CHD. The results of ongoing studies will add to the mHealth body of evidence.

It is important to highlight that there is a need for large, high-quality RCTs with hard outcomes to provide more robust evidence of effectiveness. Alongside evaluation of effectiveness, future studies should have a process evaluation to better understand why an intervention works or not and which components or features of the intervention are essential and contribute most to the success of the intervention. Furthermore, future research should also include cost-effectiveness analysis as the development of interventions using technology can be expensive and time-consuming, therefore increasing the costs of the intervention. Another important aspect to be considered when developing future mHealth interventions is having mechanisms to ensure privacy and security in the data transmission and storage. Finally, future research should also investigate mHealth effectiveness in different age groups, socio-economic status and cultural contexts.

Diabetes

Type independently, diabetes mellitus (DM) patients frequently suffer from the complications of circulatory system diseases such as cardiovascular or other vascular diseases. These complications may accompany with hypertension, atherosclerosis and thereby some ischemic diseases or systemic dysfunctions (peripheral, pulmonary, renin-angiotensin)[32-35]. It was shown that Rho kinase has a promoter effect on Ca2+ sensitive vasocontraction with PKC in STZ induced DM model studies[36]. Also in the study by Sandu OA et al[37] (2001) the interaction of insulin with Rho kinase from phosphatidylinositol 3-kinase (PI3-kinase) and iNOS activated NO-cGMP pathway was specified in vascular smooth muscle cells (VSMC). According to them insulin receptor activation inhibits ROCK activity by the NO pathway and a defectiveness in this pathway in diabetes and hypertension may lead an impaired relaxation with increased ROCK activity and resulting vasoconstriction. Also from the Rho-kinase activity experiments it was observed that arteries from Zucker diabetic fatty (ZDF) rats or incubated with high glucose concentrations, ROCK activity increase parallel with the glucose concentration[38]. Rikiteka et al (2005) also has shown the correlation between vascular endothelial cells (HSVECs) and ROCK activity that increases in high glucose[39]. In the same study the high levels of Plasminogen activator inhibitor-1 (PAI-1) protein expression induced with hyperglycemia decreased in ROCK1 knockout (ROCK I+/−) murine lung endothelial cells. While PAI-1 is a risk factor in many vascular diseases[40], the effect of ROCK on the expression of this protein in hyperglycemia will also show the key role of ROCK activity in vascular dysfunctions.

In DM induced circulatory system diseases endothelial dysfunction which led impaired NO bioavailability causes impaired vasodilation. Many study show the effect of the ROCK pathway on the endothelial dysfunction and which then leads to impaired relaxation. In DM induced vascular endothelial dysfunction (VED) Rho kinase inhibition with fasudil improved eNOS/NO dependent vasodilation is stimulated by acetylcholine[41]. Also in diabetic retinopathy, a microvascular endothelial dysfunction, it was found that high glucose concentration has increased ROCK activity in retinal endothelial cell line, RF/6A cells[42].

There are also ROCK1 and ROCK2 isoform specific studies in DM. Yao L (2013) by partly deletion both isoforms showed that ROCK1 is more effective in diabetic mice aorta according to vasorelaxive response to acetylcholine[43]. However in endothelial cells of rat thoracic aorta ROCK2 protein expression was found higher in DM with respect to the control group[44]. This difference may reflect different functional properties of ROCKs in the regulation of vascular smooth muscle contractions in DM.

Overall ROCK is a key player of many cellular functions. In recent years growing studies elicited its role in regulation of blood pressure in the vessels and therefore should be considered along with other contraction parameters. The isoforms ROCK1 and ROCK2 show branched functions, and regulate many diverse cellular activities on the circulatory system cells. Therefore, particularly in the treatment of cardiovascular diseases ROCKs with their isoforms should be taken into consideration because of their direct interventions on vasoconstriction

CONFLICT OF INTERESTS

There are no conflicts of interest with regard to the present study.

REFERENCES

1.Webb, R. C. (2003) Smooth muscle contraction and relaxation. Advances in physiology education 27, 201-206

2.Lee, D. L., Webb, R. C., and Jin, L. (2004) Hypertension and RhoA/Rho-kinase signaling in the vasculature: highlights from the recent literature. Hypertension 44, 796-799

3.Berridge, M. J. (2008) Smooth muscle cell calcium activation mechanisms. The Journal of physiology 586, 5047-5061

4.Zicha, J., Behuliak, M., Pinterova, M., Bencze, M., Kunes, J., and Vaneckova, I. (2014) The interaction of calcium entry and calcium sensitization in the control of vascular tone and blood pressure of normotensive and hypertensive rats. Physiological research / Academia Scientiarum Bohemoslovaca 63 Suppl 1, S19-27

5.Michel, T., and Vanhoutte, P. M. (2010) Cellular signaling and NO production. Pflugers Archiv: European journal of physiology 459, 807-816

6.Ignarro, L. J. (1989) Endothelium-derived nitric oxide: actions and properties. FASEB journal: official publication of the Federation of American Societies for Experimental Biology 3, 31-36

7.Chitaley, K., and Webb, R. C. (2002) Nitric oxide induces dilation of rat aorta via inhibition of rho-kinase signaling. Hypertension 39, 438-442

8.Wirth, A. (2010) Rho kinase and hypertension. Biochimica et biophysica acta 1802, 1276-1284

9.Nunes, K. P., Rigsby, C. S., and Webb, R. C. (2010) RhoA/Rho-kinase and vascular diseases: what is the link? Cellular and molecular life sciences: CMLS 67, 3823-3836

10.Kataoka, C., Egashira, K., Inoue, S., Takemoto, M., Ni, W., Koyanagi, M., Kitamoto, S., Usui, M., Kaibuchi, K., Shimokawa, H., and Takeshita, A. (2002) Important role of Rho-kinase in the pathogenesis of cardiovascular inflammation and remodeling induced by long-term blockade of nitric oxide synthesis in rats. Hypertension 39, 245-250

11.Nakagawa, O., Fujisawa, K., Ishizaki, T., Saito, Y., Nakao, K., and Narumiya, S. (1996) ROCK-I and ROCK-II, two isoforms of Rho-associated coiled-coil forming protein serine/threonine kinase in mice. FEBS letters 392, 189-193

12.Iizuka, M., Kimura, K., Wang, S., Kato, K., Amano, M., Kaibuchi, K., and Mizoguchi, A. (2012) Distinct distribution and localization of Rho-kinase in mouse epithelial, muscle and neural tissues. Cell structure and function 37, 155-175

13.Noma, K., Oyama, N., and Liao, J. K. (2006) Physiological role of ROCKs in the cardiovascular system. American journal of physiology. Cell physiology 290, C661-668

14.Yoneda, A., Multhaupt, H. A., and Couchman, J. R. (2005) The Rho kinases I and II regulate different aspects of myosin II activity. The Journal of cell biology 170, 443-453

15.Lock, F. E., and Hotchin, N. A. (2009) Distinct roles for ROCK1 and ROCK2 in the regulation of keratinocyte differentiation. PloS one 4, e8190

16.Shi, J., Wu, X., Surma, M., Vemula, S., Zhang, L., Yang, Y., Kapur, R., and Wei, L. (2013) Distinct roles for ROCK1 and ROCK2 in the regulation of cell detachment. Cell death & disease 4, e483

17.Wang, Y., Zheng, X. R., Riddick, N., Bryden, M., Baur, W., Zhang, X., and Surks, H. K. (2009) ROCK isoform regulation of myosin phosphatase and contractility in vascular smooth muscle cells. Circulation research 104, 531-540

18.Jalil, J., Lavandero, S., Chiong, M., and Ocaranza, M. P. (2005) [Rho/Rho kinase signal transduction pathway in cardiovascular disease and cardiovascular remodeling]. Revista espanola de cardiologia 58, 951-961

19.Uehata, M., Ishizaki, T., Satoh, H., Ono, T., Kawahara, T., Morishita, T., Tamakawa, H., Yamagami, K., Inui, J., Maekawa, M., and Narumiya, S. (1997) Calcium sensitization of smooth muscle mediated by a Rho-associated protein kinase in hypertension. Nature 389, 990-994

20.Masumoto, A., Hirooka, Y., Shimokawa, H., Hironaga, K., Setoguchi, S., and Takeshita, A. (2001) Possible involvement of Rho-kinase in the pathogenesis of hypertension in humans. Hypertension 38, 1307-1310

21.Fukumoto, Y., Matoba, T., Ito, A., Tanaka, H., Kishi, T., Hayashidani, S., Abe, K., Takeshita, A., and Shimokawa, H. (2005) Acute vasodilator effects of a Rho-kinase inhibitor, fasudil, in patients with severe pulmonary hypertension. Heart 91, 391-392

22.Shimizu, T., Fukumoto, Y., Tanaka, S., Satoh, K., Ikeda, S., and Shimokawa, H. (2013) Crucial role of ROCK2 in vascular smooth muscle cells for hypoxia-induced pulmonary hypertension in mice. Arteriosclerosis, thrombosis, and vascular biology 33, 2780-2791

23.Zhu, X., Lin, H., Jiang, R., Wang, R., Jiang, J., Peng, Q., and Fan, Z. (2014) Improving erectile function of spontaneously hypertensive rats by silencing ROCK2. Urology 84, 983 e911-988

24.Bentzon, J. F., Otsuka, F., Virmani, R., and Falk, E. (2014) Mechanisms of plaque formation and rupture. Circulation research 114, 1852-1866

25.Zieman, S. J., Melenovsky, V., and Kass, D. A. (2005) Mechanisms, pathophysiology, and therapy of arterial stiffness. Arteriosclerosis, thrombosis, and vascular biology 25, 932-943

26.Davignon, J., and Ganz, P. (2004) Role of endothelial dysfunction in atherosclerosis. Circulation 109, III27-32

27.Kratzer, A., Giral, H., and Landmesser, U. (2014) High-density lipoproteins as modulators of endothelial cell functions: alterations in patients with coronary artery disease. Cardiovascular research 103, 350-361

28.Nohria, A., Grunert, M. E., Rikitake, Y., Noma, K., Prsic, A., Ganz, P., Liao, J. K., and Creager, M. A. (2006) Rho kinase inhibition improves endothelial function in human subjects with coronary artery disease. Circulation research 99, 1426-1432

29.Mallat, Z., Gojova, A., Sauzeau, V., Brun, V., Silvestre, J. S., Esposito, B., Merval, R., Groux, H., Loirand, G., and Tedgui, A. (2003) Rho-associated protein kinase contributes to early atherosclerotic lesion formation in mice. Circulation research 93, 884-888

30.Wang, H. W., Liu, P. Y., Oyama, N., Rikitake, Y., Kitamoto, S., Gitlin, J., Liao, J. K., and Boisvert, W. A. (2008) Deficiency of ROCK1 in bone marrow-derived cells protects against atherosclerosis in LDLR-/- mice. FASEB journal: official publication of the Federation of American Societies for Experimental Biology 22, 3561-3570

31.Zhou, Q., Mei, Y., Shoji, T., Han, X., Kaminski, K., Oh, G. T., Ongusaha, P. P., Zhang, K., Schmitt, H., Moser, M., Bode, C., and Liao, J. K. (2012) Rho-associated coiled-coil-containing kinase 2 deficiency in bone marrow-derived cells leads to increased cholesterol efflux and decreased atherosclerosis. Circulation 126, 2236-2247

32.Adler, A. I., Stratton, I. M., Neil, H. A., Yudkin, J. S., Matthews, D. R., Cull, C. A., Wright, A. D., Turner, R. C., and Holman, R. R. (2000) Association of systolic blood pressure with macrovascular and microvascular complications of type 2 diabetes (UKPDS 36): prospective observational study. Bmj 321, 412-419

33.Moral-Sanz, J., Moreno, L., Cogolludo, A., and Perez-Vizcaino, F. (2014) Pulmonary vascular function in insulin resistance and diabetes. Current vascular pharmacology 12, 473-482

34.Bugger, H., and Abel, E. D. (2014) Molecular mechanisms of diabetic cardiomyopathy. Diabetologia 57, 660-671

35.Jandeleit-Dahm, K., and Cooper, M. E. (2006) Hypertension and diabetes: role of the renin-angiotensin system. Endocrinology and metabolism clinics of North America 35, 469-490, vii

36.Kizub, I. V., Pavlova, O. O., Johnson, C. D., Soloviev, A. I., and Zholos, A. V. (2010) Rho kinase and protein kinase C involvement in vascular smooth muscle myofilament calcium sensitization in arteries from diabetic rats. British journal of pharmacology 159, 1724-1731

37.Sandu, O. A., Ito, M., and Begum, N. (2001) Selected contribution: insulin utilizes NO/cGMP pathway to activate myosin phosphatase via Rho inhibition in vascular smooth muscle. Journal of applied physiology 91, 1475-1482

38.Bagi, Z., Feher, A., Cassuto, J., Akula, K., Labinskyy, N., Kaley, G., and Koller, A. (2011) Increased availability of angiotensin AT 1 receptors leads to sustained arterial constriction to angiotensin II in diabetes - role for Rho-kinase activation. British journal of pharmacology 163, 1059-1068

39.Rikitake, Y., and Liao, J. K. (2005) Rho-kinase mediates hyperglycemia-induced plasminogen activator inhibitor-1 expression in vascular endothelial cells. Circulation 111, 3261-3268

40.Binder, B. R., Christ, G., Gruber, F., Grubic, N., Hufnagl, P., Krebs, M., Mihaly, J., and Prager, G. W. (2002) Plasminogen activator inhibitor 1: physiological and pathophysiological roles. News in physiological sciences: an international journal of physiology produced jointly by the International Union of Physiological Sciences and the American Physiological Society 17, 56-61

41.Shah, D. I., and Singh, M. (2006) Involvement of Rho-kinase in experimental vascular endothelial dysfunction. Molecular and cellular biochemistry 283, 191-199

42.Lu, Q. Y., Chen, W., Lu, L., Zheng, Z., and Xu, X. (2014) Involvement of RhoA/ROCK1 signaling pathway in hyperglycemia-induced microvascular endothelial dysfunction in diabetic retinopathy. International journal of clinical and experimental pathology 7, 7268-7277

43.Yao, L., Chandra, S., Toque, H. A., Bhatta, A., Rojas, M., Caldwell, R. B., and Caldwell, R. W. (2013) Prevention of diabetes-induced arginase activation and vascular dysfunction by Rho kinase (ROCK) knockout. Cardiovascular research 97, 509-519

44.Cicek, F. A., Kandilci, H. B., and Turan, B. (2013) Role of ROCK upregulation in endothelial and smooth muscle vascular functions in diabetic rat aorta. Cardiovascular diabetology 12, 51

Peer reviewer:Hua He, Department of Cardiovascular Internal Medicine, Beijing Anzhen Hospital, Capital Medical University, Anzhen Road Second, Chaoyang District, Beijing, China.

Refbacks

  • There are currently no refbacks.


Creative Commons License
This work is licensed under a Creative Commons Attribution 3.0 License.