**14. miRNAs in histone deacetylase (HDAC) inhibitor arrested VSMC proliferation**

Butyrate, a dietary-derived epigenetic histone modifier and a histone deacetylase (HDAC) inhibitor, is a strong inhibitor of VSMC proliferation [72-75]. Butyrate elicits many cytoprotec‐ tive, chemopreventive and chemotherapeutic activities mainly through inhibition of cell proliferation,inductionof celldeathor stimulationof celldifferentiationbyselectivelymodulat‐ ing gene expression via epigenetic changes [72-75]. Incidentally, the cellular effects that are stimulated by butyrate are also regulated by miRNAs and expression of some of these miR‐ NAs is regulated by epigenetic mechanisms including DNA methylation and histone modifica‐ tion [76, 77]. Because butyrate is an established epigenetic histone modifier it is possible that butyrate may alter expression of some of the miRNAs in butyrate arrested VSMC proliferation. To explore this possibility, we recently examined expression profile of 650 miRNAs in butyrate inhibitedratVSMCproliferationbyqRT-PCRarrayplatform.Ourpreliminaryunpublisheddata indicates differential expression of about 60 miRNAs. Among these, members of the miR‐ NA-17-92 cluster are some of the miRNAs that are downregulated by butyrate in VSMC suggesting that antiproliferation action of butyrate is linked to downregulation of miRNAs of miRNA-17-92 cluster (Table 2). Studies have shown that the miRNAs of this cluster are not only involvedinnormaldevelopmentofheart,lungandimmunesystembuttheyalsoexhibitessential role in tumor formation by promoting cell proliferation and suppressing apoptosis [78].


\*Values represent fold changes relative to untreated rat VSMC.

**Table 2.** Changes in miRNA-17-92 cluster mature miRNAs levels in butyrate treated rat VSMC

The miRNA-17-92 cluster is a polycistronic miRNA gene, which is titled as oncomir-1 in humans because of their oncogenic properties and overexpression in different cancers [79]. The miRNA-17-92 primary transcript encodes six mature miRNAs: miRNA-17,-18a, 19a, 20a, 19b-1, and 92a-1 that are tightly grouped within an 800 base-pair region of human chromosome 13 [80]. For some of these members corresponding target genes have been identified, which include cell cycle inhibitor CDKN1A (p21Cip1) and pro- apoptotic PTEN and BCL2L11 (Bim). Furthermore, transcription of miRNA-17-92 has been shown to be activated by c-myc tran‐ scription factor [78]. In our earlier studies butyrate has been shown to downregulate c-myc [81] and upregulate CDKN1A (p21Cip1) [72-75] in proliferation inhibited VSMC. Based on these observations, it appears by downregulating c-myc expression potentially via epigenetic modification, butyrate inhibits expression of miRNA-17-92 cluster with a corresponding increase in miRNA-17-92 target genes such as CDKN1A (p21Cip1). Taken together, our preliminary miRNA expression data emphasizes role of miRNAs in antiproliferative and chemoprotective effects of butyrate in VSMC. Further studies are under investigation to confirm the role of miRNA-17-92 cluster in the regulation of VSMC proliferation by investi‐ gating the effects of miRNA mimics of miRNA-17-92 cluster in reversing the effect of butyrate on VSMC proliferation and on decreasing the levels of their target proteins. Utilization of this information is beneficial in targeting miRNAs aimed to decrease the level of pathogenic/ aberrantly expressed miRNAs or to increase miRNAs with valuable functions in the interven‐ tion of occlusive vascular proliferative diseases.

VSMC. Taken together, these studies indicate significant role of miRNA-143/miRNA-145 in

**14. miRNAs in histone deacetylase (HDAC) inhibitor arrested VSMC**

**mature miRNAs Fold change \* rno-miR-17-1-3p** -2.77 **rno-miR-17-2-3p** -2.65 **rno-miR-17-5p** -2.15 **rno-miR-18a\*** -1.80 **rno-miR-19a** -2.26 **rno-miR-19a\*** -2.31 **rno-miR-19b** -2.32 **rno-miR-19b-1\*** -2.84 **rno-miR-20a\*** -2.40 **rno-miR-92a** -2.45 **rno-miR-92a-1\*** -8.62

**Table 2.** Changes in miRNA-17-92 cluster mature miRNAs levels in butyrate treated rat VSMC

\*Values represent fold changes relative to untreated rat VSMC.

Butyrate, a dietary-derived epigenetic histone modifier and a histone deacetylase (HDAC) inhibitor, is a strong inhibitor of VSMC proliferation [72-75]. Butyrate elicits many cytoprotec‐ tive, chemopreventive and chemotherapeutic activities mainly through inhibition of cell proliferation,inductionof celldeathor stimulationof celldifferentiationbyselectivelymodulat‐ ing gene expression via epigenetic changes [72-75]. Incidentally, the cellular effects that are stimulated by butyrate are also regulated by miRNAs and expression of some of these miR‐ NAs is regulated by epigenetic mechanisms including DNA methylation and histone modifica‐ tion [76, 77]. Because butyrate is an established epigenetic histone modifier it is possible that butyrate may alter expression of some of the miRNAs in butyrate arrested VSMC proliferation. To explore this possibility, we recently examined expression profile of 650 miRNAs in butyrate inhibitedratVSMCproliferationbyqRT-PCRarrayplatform.Ourpreliminaryunpublisheddata indicates differential expression of about 60 miRNAs. Among these, members of the miR‐ NA-17-92 cluster are some of the miRNAs that are downregulated by butyrate in VSMC suggesting that antiproliferation action of butyrate is linked to downregulation of miRNAs of miRNA-17-92 cluster (Table 2). Studies have shown that the miRNAs of this cluster are not only involvedinnormaldevelopmentofheart,lungandimmunesystembuttheyalsoexhibitessential role in tumor formation by promoting cell proliferation and suppressing apoptosis [78].

VSMC differentiation and vascular disease.

**proliferation**

156 Current Trends in Atherogenesis

**miRNA-17-92 cluster**

## **15. miRNAs as new therapeutic targets for vascular proliferative diseases**

Despite the substantial progress in understanding the etiology and clinical management of vascular proliferative diseases, they are still life threatening diseases responsible for the global burden of cardiovascular diseases. Clinically, medications and surgical procedures are the only methods of treatment for patients with atherosclerotic disease. Atherosclerotic patients are generally treated by angioplasty with stent replacement but it commonly leads to restenosis in significant number of angioplasty patients. Phenotypic modification of VSMC from contractile differentiated state to proliferative dedifferentiation state is the primary pathophy‐ siological mechanism in the development of atherosclerosis and in different clinical patholo‐ gies such as postangioplasty restenosis, in-stent restenosis, vein bypass graft failure and transplant vasculopathy [33,34]. Therefore, understanding the molecular mechanisms of VSMC proliferation may offer novel insights into disease pathogenesis leading to targeted therapies. Vascular phenotypic modulation is a multifactorial process involving multiple pathways and multiple genes. Based on the current understanding of the roles of miRNAs in the normal development and in disease pathogenesis, it appears miRNA-based therapy has a potential in vascular proliferative diseases, particularly because one endogenous miRNA can target its multiple target genes. Moreover, demonstration of changes in expression of certain miRNAs that is specifically associated with particular VSMC phenotype in different models of studies, as depicted in this article, clearly suggests that expression analysis of miRNA will provide insights into vascular proliferative disease mechanisms and possibly identifies novel targets for future vascular therapy. This information is important in targeting miRNAs aimed to decrease the level of abnormally expressed miRNAs and/or to increase miRNAs with valuable functions in the intervention of occlusive vascular proliferative diseases.

potential, identification of circulating miRNAs released from injured tissues or highly expressed in patients with cardiovascular diseases suggest miRNAs can also be useful as

MicroRNAome of Vascular Smooth Muscle Cells: Potential for MicroRNA-Based Vascular Therapies

http://dx.doi.org/10.5772/54636

159

Our preliminary data presented in this article was made possible in part by research infra‐ structure support from grant numbers RR03045-21 and CO6 RR012537 from the National Center for Research Resources (NCRR), a component of the National Institutes of Health (NIH)

Department of Pharmaceutical Sciences, College of Pharmacy, Texas Southern University,

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**Acknowledgements**

**Author details**

Houston, Texas, USA

(12): 861-874.

vember 10

**References**

The recent demonstration that changes in expression of certain miRNAs in neointimal lesions, particularly upregulation of miRNA-21 and miRNA-221/-222 and downregulation of miRNA-145 support proliferative phenotype of VSMC suggests targeting miRNAs may represent a new form of therapy for vascular proliferative diseases [62, 64]. Furthermore, silencing of miRNA-21 and miRNA-221/222 by the local delivery of chemically engi‐ neered oligonucleotide-based miRNA inhibitors referred as "antigomirs" are efficient and specific silencers targeted for miRNA-21 and miRNA-221/222 was shown to reduce neointima formation [62, 64]. Similarly, use of an antagomir against miRNA -122, specifical‐ ly silenced miRNA-122 expression in the liver, lung, intestine, heart, skin and bone marrow for more than a week after one intravenous injection [82]. In another method, silencing of mir-145 or miRNA- 143 was achieved by adenovirus-mediated delivery of these miRNAs to vascular lesion, which appears to restore miRNA profile of vascular lesion that resem‐ bles normal tissue [66, 71]. Although these studies suggest that targeting miRNAs may represent a new therapy for vascular proliferative diseases, the miRNA-based technology is still long way from being translated to clinical therapy.
