Identification of key regulatory kinases in the intestinal epithelium are useful to understand the molecular mechanisms that underlie proliferation and differentiation in cells found in this compartment. We used the polymerase chain reaction (PCR) to amplify the catalytic kinase domain of serine-threonine kinases by employing degenerate primers and then screened an intestinal crypt cDNA library to clone and sequence the open reading frame of a novel serine-threonine kinase. This was then further characterized by Northern blot analysis and RNA in situ hybridization. This kinase, designated intestinal cell kinase, harbors a dual phosphorylation site found in mitogen-activating protein (MAP) kinases that is important for kinase activity.
We continuously measured hepatic absorbance of in-and exclusive biliary excretion with neither intradocyanine green (ICG) using near-infrared (NIR) spec-hepatic conjugation nor enterohepatic circulation.1,2 troscopy after intravenous bolus injection in rabbits. He-Conventionally, ICG has been used as a test substance patic ICG concentration was obtained by subtracting for the estimation of hepatic blood flow, as measured out the absorbance of hemoglobin and other pigments by plasma clearance determined from the initial disapwithin the liver. Two exponential rate constants, the pearance pattern in plasma concentration decay. then used in liver-injured rabbits inflicted with hemor-liver-bile interaction. 13 Thus, plasma decay fails to prorhagic shock and ischemia-reperfusion, to show that the vide us with detailed information about ICG disappearfirst rate constant is primarily affected by hepatic micro-ance from the liver. Direct assessment of the transicirculatory condition, and the second refers closely to tional dye concentration within the liver would be parenchymal liver damage. In another series of partial much more effective, considering the drawbacks of conliver ischemia-reperfusions, it was possible to simultane-ventional methods. hemoglobin has an absorption peak at 760 nm. Recently, we have developed a new method of quantitative Hepatic clearance of indocyanine green (ICG) occurs evaluation of hemoglobin oxygenation in the liver siby two major processes: by uptake across the sinusoidal nusoid with NIR spectroscopy. In this method, the plasma membrane with a high extraction ratio and by deformed absorption spectrum caused by photon scatremoval from the hepatocytes by cytoplasmic transport tering in the living tissue was corrected, and a multicomponent least square curve-fitting analysis was perAbbreviations: ICG, indocyanine green; NIR, near-infrared; ATP, adenosine formed to avoid the influence of the spectra of triphosphate.cytochromes and other pigments in the liver on the Otsuka Electronics, Osaka, Japan.tion. 16,17 ICG has an absorbance peak at 805 nm within
The Epstein-Barr (EBV) virus induces a lytic state after infecting epithelial cells. Subsequently, there is infection of B lymphocytes with two types of cycles, latent and lytic. Apart from linkage of the EBV latent membrane protein-1 (LMP-1) with benign and malignant conditions of squamous epithelial cells, little is known about other EBV gene products that may be important in these processes as well as cellular transcriptional factors that regulate EBV gene expression in these epithelial cells. The EBV ED-L2 promoter, an early lytic cycle promoter, is located upstream of a transcription start site for a short open reading frame designated BNLF2 and just downstream of the BNLF1 (LMP-1) open reading frame. We have previously used the EBV ED-L2 promoter to target oncogenes in transgenic mice, resulting in tissue-specific expression in the tongue, esophagus, forestomach, and skin, all sharing stratifying squamous epithelia, alternatively called keratinocytes. In the present study, we have functionally dissected the ED-L2 promoter by making deletion constructs fused to the luciferase reporter gene with transient transfections into squamous and nonsquamous epithelial cell lines as well as B lymphocytes. A CACCC box-like cisregulatory element has been identified that is located between ؊218 and ؊187 base pairs of the ED-L2 promoter that confers significant promoter activity only in squamous epithelial cells. This cis-regulatory element is active in a heterologous minimal herpes simplex virus thymidine kinase promoter reporter gene construct when transfected into squamous epithelial cells but not in nonsquamous epithelial cells. DNA gel mobility shift assays have led to the identification of DNA-protein complexes that bind the CACCC box-like element. One of these proteins is a novel transcriptional factor that is uniquely active in stratified squamous epithelial cells, designated as keratinocyte specific factor (KSF). KSF may be related to Sp1 but appears to be distinct from Sp1. In addition, KSF may interact with related or identical cis-regulatory elements found in human papillomavirus-11 E6 and cytokeratin K3 promoters that are active in keratinocytes. In aggregate, KSF may be important in the transcriptional regulation of viral and eukaryotic genes in keratinocytes.
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