SummaryInterleukin (IL)-36a, IL-36b and IL-36g are expressed highly in skin and are involved in the pathogenesis of psoriasis, while the antagonists IL-36Ra or IL-38, another potential IL-36 inhibitor, limit uncontrolled inflammation. The expression and role of IL-36 cytokines in rheumatoid arthritis (RA) and Crohn's disease (CD) is currently debated. Here, we observed that during imiquimod-induced mouse skin inflammation and in human psoriasis, expression of IL-36a, g and IL-36Ra, but not IL-36b and IL-38 mRNA, was induced and correlated with IL-1b and T helper type 17 (Th17) cytokines (IL-17A, IL-22, IL-23, CCL20). In mice with collageninduced arthritis and in the synovium of patients with RA, IL-36a, b, g, IL36Ra and IL-38 were all elevated and correlated with IL-1b, CCL3, CCL4 and macrophage colony-stimulating factor (M-CSF), but not with Th17 cytokines. In the colon of mice with dextran sulphate sodium-induced colitis and in patients with CD, only IL-36a, g and IL-38 were induced at relatively low levels and correlated with IL-1b and IL-17A. We suggest that only a minor subgroup of patients with RA (17-29%) or CD (25%) had an elevated IL-36 agonists/antagonists ratio, versus 93% of patients with psoriasis. By immunohistochemistry, IL-36 cytokines were produced by various cell types in skin, synovium and colonic mucosa such as keratinocytes, CD68 1 macrophages, dendritic/Langerhans cells and CD79a 1 plasma cells. In primary cultures of monocytes or inflammatory macrophages (M1), IL-36b and IL-36Ra were produced constitutively, but IL-36a, g and IL-38 were produced after lipopolysaccharide stimulation. These distinct expression profiles may help to explain why only subgroups of RA and CD patients have a potentially elevated IL-36 agonists/ antagonists ratio.

Hypertension is one of the most frequent pathologies in the industrialized world. Although recognized to be dependent on a combination of genetic and environmental factors, its molecular basis remains elusive. Increased activity of the monomeric G protein RhoA in arteries is a common feature of hypertension. However, how RhoA is activated and whether it has a causative role in hypertension remains unclear. Here we provide evidence that Arhgef1 is the RhoA guanine exchange factor specifically responsible for angiotensin II-induced activation of RhoA signaling in arterial smooth muscle cells. We found that angiotensin II activates Arhgef1 through a previously undescribed mechanism in which Jak2 phosphorylates Tyr738 of Arhgef1. Arhgef1 inactivation in smooth muscle induced resistance to angiotensin II-dependent hypertension in mice, but did not affect normal blood pressure regulation. Our results show that control of RhoA signaling through Arhgef1 is central to the development of angiotensin II-dependent hypertension and identify Arhgef1 as a potential target for the treatment of hypertension.

The monolayer of columnar epithelial cells lining the gastrointestinal tract--the intestinal epithelial barrier (IEB)--is the largest exchange surface between the body and the external environment. The permeability of the IEB has a central role in the regulation of fluid and nutrient intake as well as in the control of the passage of pathogens. The functions of the IEB are highly regulated by luminal as well as internal components, such as bacteria or immune cells, respectively. Evidence indicates that two cell types of the enteric nervous system (ENS), namely enteric neurons and enteric glial cells, are potent modulators of IEB functions, giving rise to the novel concept of a digestive 'neuronal-glial-epithelial unit' akin to the neuronal-glial-endothelial unit in the brain. In this Review, we summarize findings demonstrating that the ENS is a key regulator of IEB function and is actively involved in pathologies associated with altered barrier function.

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Small G proteins of the Rho family function as tightly regulated molecular switches that govern a wide range of cell functions (1). A large body of evidence has now been obtained regarding the important functions of Rho proteins in the vasculature, and RhoA has been shown to play a major role in vascular processes such as smooth muscle cell contraction, proliferation, and differentiation; endothelial permeability; platelet activation; and leukocyte migration (2-4). The activity of Rho is under the direct control of a large set of other regulatory proteins (1). In the inactive GDP-bound form, RhoA is locked in the cytosol by guanine dissociation inhibitors. The guanine nucleotide exchange factors catalyze the exchange of GDP for GTP to activate RhoA (5). Activation is then turned off by GTPase-activating proteins that hydrolyze GTP to GDP. Therefore, both the relative expression of these proteins (in particular, that of RhoA) and the fraction of active GTP-bound RhoA are key determinants of RhoA protein activity.Data are now accumulating regarding the regulation of the amount of active GTP-bound RhoA. In vascular smooth muscle cells, several agonists of G protein-coupled receptors, including thrombin, thromboxane A 2 , endothelin, carbachol, angiotensin, ␣-adrenergic agonists, sphingolipids, and extracellular nucleotides, stimulate RhoA activity through the activation of guanine nucleotide exchange factors and increases in the fraction of GTP-bound RhoA. This RhoA activation is accompanied by the membrane translocation of GTP-bound RhoA (5-8). On the other hand, the NO, cGMP, and cGMP-dependent kinase (PKG) 1 signaling pathway exerts inhibitory action on RhoA functions in cells stimulated by these G protein-coupled receptor agonists. We have previously demonstrated that PKG phosphorylates RhoA at Ser 188 in vitro and that the effects of PKG activation on actin cytoskeleton are lost in cells expressing the non-phosphorylatable RhoA A188 mutant, suggesting that inhibitory effects of PKG on RhoA-mediated contraction and actin organization are due to phosphorylation of RhoA at Ser 188 (9). This effect involves inhibition of membrane translocation of GTP-bound RhoA. Several additional reports have now confirmed the inhibitory effect of the NO/cGMP/PKG signaling pathway on the RhoA-dependent component of agonist-induced contraction (10 -13). Recently, it has also been shown that PKG inhibits RhoA-mediated serum response element (SRE)-dependent transcription (14). PKG inhibits SRE-dependent transcription induced by serum, constitutively active G␣ 12 or G␣ 13 , constitutively active Rho exchange factor p115 RhoGEF , or constitutively active RhoA L63 . This inhibition is associated with a decrease in the amount of active GTP-bound RhoA in cells stimulated with serum or constitutively active G␣ 12 or G␣ 13 , but not in p115 RhoGEF -or RhoA L63 -expressing cells, suggesting that PKG can act both upstream and downstream of RhoA. The effect on steps downstream of RhoA has been confirmed by showing that SRE-dependent transcrip...

Enteric glial cells (EGCs) are in many respects similar to astrocytes of the central nervous system and express similar proteins including glial fibrillary acidic protein (GFAP). Changes in GFAP expression and/or phosphorylation have been reported during brain damage or central nervous system degeneration. As in Parkinson's disease (PD) the enteric neurons accumulate a-synuclein, and thus are showing PDspecific pathological features, we undertook the present survey to study whether the enteric glia in PD become reactive by assessing the expression and phosphorylation levels of GFAP in colonic biopsies. Twenty-four PD, six progressive supranuclear palsy (PSP), six multiple system atrophy (MSA) patients, and 21 age-matched healthy controls were included. The expression levels and the phosphorylation state of GFAP were analyzed in colonic biopsies by western blot. Additional experiments were performed using real-time PCR for a more precise analysis of the GFAP isoforms expressed by EGCs. We showed that GFAPj was the main isoform expressed in EGCs. As compared to control subjects, patients with PD, but not PSP and MSA, had significant higher GFAP expression levels in their colonic biopsies. The phosphorylation level of GFAP at serine 13 was significantly lower in PD patients compared to control subjects. By contrast, no change in GFAP phosphorylation was observed between PSP, MSA and controls. Our findings provide evidence that enteric glial reaction occurs in PD and further reinforce the role of the enteric nervous system in the initiation and/or the progression of the disease.

The small G proteins of the Rho family are identified as key signaling molecules in the vasculature. RhoA and its downstream effector Rho kinase have been shown to play a major role in vascular processes such as smooth muscle cell contraction, proliferation and differentiation, endothelial permeability, platelet activation, and leukocyte migration (1, 2). Abnormal activation of the RhoA/Rho kinase pathway has been observed in major cardiovascular disorders such as atherosclerosis, restenosis, hypertension, pulmonary hypertension, and cardiac hypertrophy. The activity of RhoA is under the direct control of a large set of regulatory proteins (3). In the inactive GDP-bound form, RhoA is locked in the cytosol by guanine dissociation inhibitors (4). The guanine nucleotide exchange factors catalyze the exchange of GDP for GTP to activate RhoA (5). In the active GTP-bound form, RhoA translocates to plasma membrane, where it interacts with effectors to transduce the signal downstream. GTPase-activating proteins that hydrolyzed GTP to GDP then turn off activation. In addition to this activation/ inactivation cycle, recent reports have proposed that regulation of Rho protein degradation could participate in the regulation of Rho protein functions. Phosphorylation of RhoA by cGMPdependent protein kinase protects RhoA from ubiquitin/proteasome-mediated degradation in vascular smooth muscle cells (6). In contrast, constitutive activation of Rho proteins by the bacterial toxin cytotoxic necrotizing factor 1 enhances their ubiquitin/proteasome-mediated degradation (7). To date, the relevance of this mechanism of Rho protein depletion in a physiological/pathophysiological context, in response to membrane receptor agonist stimulation, has not been addressed.In vascular physiology, Akt signaling has been shown to mediate survival signals of many angiogenic factors and plays central roles in the regulation of vascular homeostasis and angiogenesis (8). Recent reports also reveal that Akt signaling plays important roles in vascular smooth muscle cells. Akt signaling mediates cell survival, proliferation, and migration of vascular smooth muscle cells induced by angiotensin II, serotonin 5-hydroxytryptamine (5-HT), 4 and several growth factors (9 -12). Akt signaling pathway is suggested to play critical roles in pathological accumulation of vascular smooth muscle cells observed in various types of vascular lesions (11,13). Recently, it has been shown that Akt is negatively regulated by the RhoA/Rho kinase pathway in endothelial cells (14). Accord-* This work was supported by grants from the INSERM, the Agence Nationale de la Recherche, and the Fondation pour la Recherche Mé dicale. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C.

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