Abstract-We have recently shown that all-trans retinoic acid (tRA) modulates arterial smooth muscle cell (SMC) morphologic features and biochemical composition in vitro. It has been proposed that different SMC phenotypes coexist in arteries, which may be retrieved in culture: hence, a differential action of tRA on distinct SMC subsets is conceivable.We have examined the effect of tRA on SMC proliferation, migration, plasminogen activator activity, and ␣-smooth muscle actin expression in 2 phenotypically different rat SMC populations, cultured respectively from the normal aortic media and from the intimal thickening (IT) after endothelial injury. tRA inhibited proliferation and increased migration and tissue-type plasminogen activator activity in both SMC populations, but decreased ␣-smooth muscle actin only in SMC cultured from the IT. The action of tRA is mediated by 2 families of nuclear receptors, RAR and RXR, each containing 3 isoforms, ␣, ␤, and ␥. RAR and RAR-␣ agonists, but not RXR agonists, inhibited SMC proliferation in both cell populations and ␣-smooth muscle actin expression only in IT SMC. When administered intraperitoneally to balloon-injured rats, tRA and RAR-␣ agonists reduced the intimal hyperplasia in the carotid artery. Our results show that tRA and synthetic retinoids can affect the proliferation, migration, and differentiation of SMC in vitro. Key Words: smooth muscle cell heterogeneity Ⅲ smooth muscle cell motility Ⅲ smooth muscle cell differentiation Ⅲ intimal thickening S mooth muscle cell (SMC) differentiation, migration, and proliferation are key factors in the development of atherosclerosis and restenosis after angioplasty. 1,2 These phenomena involve an SMC phenotypic modulation characterized by modification of gene expression. 3 It has been suggested that SMC subsets present in the normal media are particularly prone to undergoing phenotypic modulation, and the concept of SMC heterogeneity is gaining wider acceptance (for review see Reference 4). For instance, SMC populations cultured from the rat aortic media and from the intima 15 days after endothelial injury are characterized by a spindle-shaped and an epithelioid morphology, respectively (for discussion of this point see Reference 5). These 2 phenotypes are also obtained when SMC are cloned from the aortic media or the intima, albeit in different proportions. 6 The regulation of SMC phenotype is thought to be exerted by cytokines, growth factors, and agents regulating differentiation. 7 Within the last category, some agents are known to govern SMC differentiation during embryogenesis, and there is a growing body of evidence suggesting that the genetic programs used during embryogenesis may act also during arterial disease processes. 8,9 Vitamin A plays a crucial role in the regulation of cell growth and differentiation, and its active form, retinoic acid, is involved in signal transduction pathways regulating embryonic development. 10 These effects are mediated by 2 families of nuclear receptors that are ligand-dependent trans...

Since nitric oxide (NO) has been implicated in the pathogenesis of various hyperproliferative and inflammatory diseases, the mRNA expression of constitutive nitric oxide synthase (cNOS) and inducible nitric oxide synthase (iNOS) were investigated in psoriatic skin by reverse transcriptase coupled to the polymerase chain reaction (PCR). The study showed that the mRNA expression of brain nitric oxide synthase (bNOS), one of two isoforms of cNOS, was weak in both psoriatic plaques lesions and uninvolved skin, while mRNA transcripts for the second isoform, endothelial nitric oxide synthase (eNOS), were not detectable using the present method. In contrast, the mRNA expression of iNOS was markedly increased in lesional skin as compared to uninvolved skin. Cultured human keratinocytes exposed to a combination of interleukin-1 beta (IL-1 beta) and tumour necrosis factor-alpha (TNF-alpha) for 4 h, showed strong gene expression of iNOS, while in 24 h, the expression had returned to baseline expression. In summary, the study demonstrates that mRNA for the inducible form of NOS is over-expressed in psoriatic lesions. The cause of this may be the local presence of inflammatory cytokines. These findings imply that iNOS may play an important part in local regulation of NO synthesis in psoriasis and other inflammatory dermatoses.

To evaluate a newly developed high-resolution laser-Doppler perfusion imager (HR-LDPI) for analysis of local tissue perfusion, blood flow measurements in the rabbit tenuissimus muscle were carried out in combination with intravital microscopic observation. The principle of the LDPI method is based on a low-power laser beam scan of the exposed tissue from which a two-dimensional color-coded perfusion map is created through computerized signal analysis. The perfusion of the tenuissimus muscle prepared for microscopic observation was analyzed in a 5 mm x 8 mm area as the muscle was exposed to atmospheric oxygen tension (Po2; 20 kPa), a low Po2 (approximately 3 kPa), and after vasodilatation induced by topical application of prostaglandin E2 (PGE2). In selected areas free from larger vessels, a significantly lower perfusion average reading was demonstrated under high Po2 conditions compared with low Po2 conditions (P < 0.05, n = 5 animals), and application of PGE2 gave rise to an average reading significantly higher than that at low Po2 (P < 0.01, n = 6 animals). The results were in good agreement with the flow changes observed microscopically, and the architecture of the microvascular network, as depicted by in vivo micrographs, was clearly recognizable in the perfusion images. In conclusion, blood flow changes in the rabbit tenuissimus muscle induced by various stimuli were quantitated with the HR-LDPI method and could be spatially resolved in great detail, illustrating the potential of using HR-LDPI for analysis of local blood flow and to reveal spatial perfusion heterogeneity in tissues.

Aim: The cytochrome P450 enzymes of the CYP26 family are involved in the catabolism of the biologically active retinoid all-trans-retinoic acid (atRA). Since it is possible that an increased local CYP26 activity would reduce the effects of retinoids in vascular injury, we investigated the role of CYP26 in the regulation of atRA levels in human aortic smooth muscle cells (AOSMCs). Methods: The expression of CYP26 was investigated in cultured AOSMCs using real-time PCR. The metabolism of atRA was analyzed by high-performance liquid chromatography, and the inhibitor R115866 or small interfering RNA (siRNA) was used to suppress CYP26 activity/expression. Results: AOSMCs expressed CYP26B1 constitutively and atRA exposure augmented CYP26B1 mRNA levels. Silencing of the CYP26B1 gene expression or reduction of CYP26B1 enzymatic activity by using siRNA or the inhibitor R115866, respectively, increased atRA-mediated signaling and resulted in decreased cell proliferation. The CYP26 inhibitor also induced expression of atRA-responsive genes. Therefore, atRA-induced CYP26 expression accelerated atRA inactivation in AOSMCs, giving rise to an atRA-CYP26 feedback loop. Inhibition of this loop with a CYP26 inhibitor increased retinoid signaling. Conclusion: The results suggest that CYP26 inhibitors may be a therapeutic alternative to exogenous retinoid administration.