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An endogenous cannabimimetic molecule, 2-arachidonoylglycerol, induces a rapid, transient increase in intracellular free Ca 2؉ concentrations in NG108 -15 cells through a cannabinoid CB1 receptor-dependent mechanism. We examined the activities of 24 relevant compounds (2-arachidonoylglycerol, its structural analogues, and several synthetic cannabinoids). We found that 2-arachidonoylglycerol is the most potent compound examined so far: its activity was detectable from as low as 0.3 nM, and the maximal response induced by 2-arachidonoylglycerol exceeded the responses induced by others. Activities of HU-210 and CP55940, potent cannabinoid receptor agonists, were also detectable from as low as 0.3 nM, whereas the maximal responses induced by these compounds were low compared with 2-arachidonoylglycerol. Anandamide was also found to act as a partial agonist in this assay system. We confirmed that free arachidonic acid failed to elicit a response. Furthermore, we found that a metabolically stable ether-linked analogue of 2-arachidonoylglycerol possesses appreciable agonistic activity, although its activity was apparently lower than that of 2-arachidonoylglycerol. We also confirmed that pretreating cells with various cannabinoid receptor agonists nullified the response induced by 2-arachidonoylglycerol, whereas pretreating cells with other neurotransmitters or neuromodulators did not affect the response. These results strongly suggested that the cannabinoid CB1 receptor is originally a 2-arachidonoylglycerol receptor, and 2-arachidonoylglycerol is the intrinsic physiological ligand for the cannabinoid CB1 receptor.It is well known that ⌬ 9 -tetrahydrocannabinol (⌬ 9 -THC), 1 a psychoactive ingredient of marijuana, possesses a variety of pharmacological activities in vitro and in vivo (1), although, until recently, the mechanism of the action of ⌬ 9 -THC had long been unelucidated. In 1988, Devane et al. (2) provided evidence that a specific binding site(s) for cannabinoids is present in the brain. Soon after, Matsuda et al. (3) cloned a cDNA encoding a cannabinoid receptor (CB1) from a rat brain cDNA library. These findings raised the possibility that at least part of the action of ⌬ 9 -THC is mediated through such a specific receptor and prompted the search for endogenous cannabinoid receptor ligands in mammalian tissues.In 1992, Devane et al. (4) isolated N-arachidonoylethanolamine (anandamide) from porcine brain as the first endogenous cannabinoid receptor ligand. They demonstrated that anandamide exhibits several cannabimimetic activities in vitro and in vivo (4, 5). So far, a number of studies have been carried out on anandamide, and it has been assumed that anandamide is one of the important lipid mediators in the nervous system as well as in other systems (5). However, we (6, 7) and others (8 -11) have found that the levels of anandamide are very low in several mammalian tissues. In addition, the biosynthetic pathways for anandamide, either the N-acylphosphatidylethanolamine pathway (6,7,(11)(12)(13)(14) or t...

In most autoimmune diseases the serologic hallmarks of disease precede clinical pathology by years. Therefore the use of animal models in defining early disease events becomes critical. Herein we have taken advantage of a “designer” mouse with dysregulation of interferon gamma (IFNγ) characterized by prolonged and chronic expression of IFNγ through deletion of the IFNγ 3′ UTR AU-rich element. These mice develop primary biliary cholangitis (PBC) with a female predominance that mimics human disease and is characterized by upregulation of total bile acids, spontaneous production of AMA, and portal duct inflammation. Transfer of CD4 T cells from ARE-Del−/− to B6/Rag1−/− mice induced moderate portal inflammation, and parenchymal inflammation, RNA-sequencing of liver gene expression revealed that upregulated genes potentially define early stages of cholangitis. Interestingly, upregulated genes specifically overlap with the gene expression signature of biliary epithelial cells in PBC, implying that IFNγ may play a pathogenic role in biliary epithelial cells (BEC) in the initiation stage of PBC. Moreover, differentially expressed genes in female mice have stronger Type I and II interferon signaling and lymphocyte-mediated immune responses and thus may drive the female bias of the disease. In conclusion, changes in IFNγ expression are critical for the pathogenesis of PBC.

SUMMARY:Although stromal cell-derived factor-1 (SDF-1) plays an important role in hematopoiesis in the fetal liver, the role after birth remains to be clarified. We investigated the role of SDF-1 and its receptor, CXCR4, in 75 patients; this included controls and patients with viral hepatitis, liver cirrhosis, primary biliary cirrhosis, primary sclerosing cholangitis, and autoimmune hepatitis. Interestingly, SDF-1 appeared up-regulated in biliary epithelial cells (BEC) of inflammatory liver disease. Furthermore, in inflammatory liver diseases, SDF-1 was expressed by BEC of interlobular and septal bile ducts and by proliferated bile ductules. The message expression of SDF-1 in BEC was confirmed at a single-cell level by RT-PCR and laser capture microdissection. The plasma levels of SDF-1 were significantly higher in patients with liver diseases than in normal controls. Flow cytometric analysis of the surface expression of CXCR4 showed that most liver-infiltrating lymphocytes express CXCR4 and the intensity was up-regulated more significantly in liver-infiltrating lymphocytes than in peripheral blood lymphocytes. These results suggest that increased SDF-1 production by BEC may play an important role in the recruitment of CXCR4-positive inflammatory cells into the diseased livers. These data are significant because modulation of the SDF-1/CXCR4 interaction has therapeutic implications for inflammatory liver diseases. (Lab Invest 2003, 83:665-672).

To establish a sensitive and specific antibody assay, potent antigenic proteins encoded by human herpesvirus 8 (HHV8) were studied. Fifteen recombinant HHV8-encoded proteins were produced as glutathione S-transferase fusion proteins. The sera from AIDS-associated Kaposi's sarcoma (