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The Gpbar1 [G-protein-coupled BA (bile acid) receptor 1] is a recently identified cell-surface receptor that can bind and is activated by BAs, but its physiological role is unclear. Using targeted deletion of the Gpbar1 gene in mice, we show that the gene plays a critical role in the maintenance of bile lipid homoeostasis. Mice lacking Gpbar1 expression were viable, developed normally and did not show significant difference in the levels of cholesterol, BAs or any other bile constituents. However, they did not form cholesterol gallstones when fed a cholic acid-containing high-fat diet, and liver-specific gene expression indicated that Gpbar1-deficient mice have altered feedback regulation of BA synthesis. These results suggest that Gpbar1 plays a critical role in the formation of gallstones, possibly via a regulatory mechanism involving the cholesterol 7alpha-hydroxylase pathway.

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Although many cytokines have been implicated in the development and persistence of inflammatory immune responses, it is unknown if any of these are important in inflammatory acne. This study investigated the production of the proinflammatory cytokines interleukin-8 (IL-8), IL-1␤, and tumor necrosis factor alpha (TNF-␣) by human monocytic cell lines, ThP-1 and U937, and by freshly isolated peripheral blood mononuclear cells from acne patients. Both Propionibacterium acnes and supernatants obtained from 72-h P. acnes cultures could induce significant concentrations of IL-1␤, TNF-␣, and IL-8 by both cell lines and by peripheral blood mononuclear cells as determined by enzyme-linked immunosorbent assay. There was no significant difference between acne and non-acne subjects. Endotoxin quantification and addition of polymyxin B to assays indicated no lipopolysaccharide (LPS) contamination. P. acnes supernatant was fractionated into components with molecular weights of <3,000, <10,000, and <30,000 and assayed for the ability to induce IL-8 and TNF production in ThP-1 cells. Nearly 90% of the original activity was found in the <30,000-molecular-weight fraction, 50% was in the <10,000-molecular-weight fraction, and only 15% remained in the <3,000-molecularweight fraction. The effluent from the <3,000-molecular-weight fraction contained about 70% activity, indicating that the inducing factor was not retained in the membrane. Incubation of P. acnes supernatant with various concentrations of mutanolysin or lysozyme resulted in a loss of 60% of the original activity. The addition of jimson lectin, which binds peptidoglycan, resulted in a loss of 70% of the activity in a dose-response manner, whereas peanut lectin had little or no effect on the activity. Heating of the P. acnes supernatant to 65 C also had no effect on the activity. Blocking of CD14, a receptor for both LPS and peptidoglycan, reduced cytokine production by >50%, suggesting that the soluble stimulating factor may be a secreted form of peptidoglycan-polysaccharide.

We have identified an orphan G protein-coupled receptor, SP174, that shares a high degree of homology with the recently described ADP receptor P2Y 12 . mRNA for SP174 is abundant in the brain and in cells of the immune system. In the present study, we demonstrate that SP174 is also a receptor for ADP, which is coupled to G␣i. ADP potently stimulates SP174 with an EC 50 of 60 nM, and other related nucleotides are active as well, with a rank order of potency 2-methylthio-ADP tetrasodium ϭ adenosine 5Ј-O-2-(thio)diphosphate ϭ 2-methylthio-ATP tetrasodium Ͼ ADP Ͼ AP3A ϾATP Ͼ IDP. This pharmacological profile is similar to that for P2Y 12 . We have also identified the murine homolog of SP174, which exhibits 75% homology to the human receptor. ADP is also a potent agonist at the murine receptor, and its pharmacological profile is similar to its human counterpart, but ADP and related nucleotides are more potent at the murine receptor than the human receptor. In keeping with the general nomenclature for the purinergic receptors, we propose designating this novel receptor P2Y 13 .P2Y receptors are G protein-coupled receptors that respond to the presence of extracellular nucleotides. Both purine and pyrimidine nucleotides can modulate a variety of physiological functions by interaction with P2Y receptors (Harden et al., 1995;Burnstock, 1997;Ralevic and Burnstock, 1998). Six mammalian P2Y receptors have been cloned so far, including P2Y 1 , P2Y 2 , P2Y 4 , P2Y 6 , P2Y 11 , and more recently, P2Y 12 (Burnstock, 1997;Communi et al., 1997;Ralevic and Burnstock, 1998;Hollopeter et al., 2001;Zhang et al., 2001). P2Y 1 , P2Y 2 , and P2Y 6 couple to the activation of phospholipase C (PLC); P2Y 11 couples to the activation of both PLC and the adenylyl cyclase pathways, whereas human P2Y 4 couples to adenylyl cyclase pathways at the early stage and PLC at a later stage (Communi et al., 1996;Ralevic and Burnstock, 1998). P2Y 1 is selectively activated by ADP with ATP, being either a partial agonist or an antagonist; P2Y 2 is activated equipotently by ATP and UTP; human P2Y 11 is selectively activated by ATP; human P2Y 6 is selectively activated by UDP, whereas rat P2Y 6 is selectively activated by UTP; and human P2Y 4 is activated selectively by UTP, whereas rat and murine P2Y 4 are activated equipotently by ATP and UTP (Harden et al., 1995;Burnstock, 1997;Ralevic and Burnstock, 1998). In contrast, the P2Y 12 receptor recently described by us and Hollopeter et al. (2001) is potently activated by ADP and is coupled to the inhibition of adenylyl cyclase activity through the G␣i class of G proteins.Analysis of the expression profile of P2Y 12 receptor mRNA revealed that it is expressed at high levels in platelets, in addition to brain tissue. The data presented by Hollopeter et al. (2001) and the analysis of platelet function in P2Y 12 null mice by Foster et al. (2001) clearly indicate that P2Y 12 represents the long sought-after platelet ADP receptor. Furthermore, these studies reveal that P2Y 12 is the molecular target of th...