Claudins are thought to be major components of tight junctions (TJs), and claudin-5 and -12 are localized at TJs of the blood-brain barrier (BBB). Claudin-5-deficient mice exhibit size-selective (<800 Da) opening of the BBB. The purpose of this study was to clarify the expression levels of claudin-5 and -12 in rat brain capillary endothelial cells, and to examine the ability of claudin-5 to form TJs in cultured rat brain capillary endothelial cells (TR-BBB). Expression of claudin-5 mRNA in rat brain capillary fraction was 751-fold greater than that of claudin-12. The level of claudin-5 mRNA in the rat brain capillary fraction (per total mRNA) was 35.6-fold greater than that in whole brain, while the level of claudin-12 mRNA was only 13.9% of that in whole brain, suggesting that expression of claudin-12 mRNA is not restricted to brain capillaries. Transfection of TR-BBB cells with the claudin-5 gene afforded TR-BBB/CLD5 cells, which showed no change in expression of claudin-12 or ZO-1, while the expressed claudin-5 was detected at the cell-cell boundaries. The permeability surface product of [ 14 C]inulin at a TR-BBB/CLD5 cell monolayer was significantly smaller (P < 0.01) than that for the parental TR-BBB cells, and the values of the permeability coefficient (Pe) were 1.14 Â 10 À3 and 11.6 Â 10 À3 cm/ min, respectively. These results indicate that claudin-5, but not claudin-12, is predominantly expressed in brain capillaries, and plays a key role in the appearance of barrier properties of brain capillary endothelial cells. J. Cell. Physiol. 210: 81-86, 2007. Tight junctions (TJs) between brain capillary endothelial cells play a key role in the formation of the blood-brain barrier (BBB) because TJs block paracellular diffusion between endothelial cells (Pardridge, 2005). To date, three distinct types of integral membrane proteins have been reported to be localized at TJs: occludin (Furuse et al., 1993), junctional adhesion molecule (JAM) (Martin-Padura et al., 1998), and claudins (Furuse et al., 1998a). Among these TJ proteins, claudins are thought to be the major constituents of TJs (Furuse et al., 1998b). Claudins are a multigene family consisting of more than 20 members (Tsukita et al., 2001). They interact homogeneously and heterogeneously with each other to form TJs, and the properties of the resulting TJs depend on the claudin subtypes involved . Therefore, clarifying the contributions of claudin subtypes to TJ formation in the BBB is important in order to understand the development of the BBB and changes in the BBB that may occur in disease, as well as for developing novel techniques to deliver drugs to the brain.Claudin-5 is localized at the cell-cell boundaries of brain capillary endothelial cells (Morita et al., 1999). Nitta et al. (2003) reported size-selective opening of the BBB in claudin-5-deficient mice. In these mice, TJs with a normal appearance were present at the brain capillary endothelial cell-cell contact regions, but leakage of small molecules (<800 Da), though not larger molecules, was obs...

Members of the ATP binding cassette (ABC) transporter family transport various substrates coupled with hydrolysis of ATP. The human ABC transporter superfamily consists of 49 subtypes, some of which function as drug efflux transporters (ABCB1, ABCC subfamily and ABCG2) or sterol transporters (ABCA1, ABCA7, ABCG1, ABCG5 and ABCG8).1,2) However, the role of the ABCA5-like transporters remains largely unknown.The ABCA family forms the second largest gene family, consisting of 12 subtypes, after the ABCC family. Evolutional analysis has revealed a gene cluster encoding ABCA5, ABCA6, ABCA8, ABCA9 and ABCA10, which are known as the ABCA5-like transporters.3) Interestingly, although these five ABCA transporters form a unique cluster on human Chr17q24, most other ABC transporter genes are dispersed in the mammalian genome.3) Among the ABCA5-like transporters, ABCA8 expressed in oocytes was shown to exhibit ATP-dependent transport of estradiol-b-glucuronide. 4)In the testis, ABCA5 is localized in Leydig cells, which form the blood-testis barrier, and ABCA5 knockout mice develop an enlarged heart, injured liver and decreased plasma levels of thyroid hormones. 5,6) These observations imply the physiological importance of ABCA5-like transporters.Since ABCA8 has been reported to act as a transporter, 4) it is possible that ABCA5-like transporters have functions related to multi-drug resistance in tumor cells. The purpose of this study is, therefore, to investigate mRNA expression of ABCA5-like transporters in human tissues and tumors, in order to establish whether there is a relationship between ABCA5-like transporters and tumor development. MATERIALS AND METHODScDNA Samples Human tissue normalized first-strand cDNAs were purchased as MTC Multiple Tissue cDNA Panel I and II from Clontech (Palo Alto, CA, U.S.A.). Human tumor normalized first-strand cDNAs were purchased as MTC Multiple Tumor cDNA Panel from Clontech. The first-strand cDNA was reverse-transcribed from poly-A RNA purified from each tissue or tumor. MTC Panel I contains cDNA preparations from heart, brain, placenta, lung, liver, skeletal muscle, kidney and pancreas pooled from 2-15 male/female Caucasians. MTC Panel II contains cDNA preparations from spleen, thymus, prostate, testis, ovary, small intestine without mucosal lining, and colon pooled from 7-45 male/female Caucasians, and peripheral blood leukocytes pooled from male/female Caucasians negative for HIV-1, HIV-2, hepatitis B and syphilis.MTC Tumor Panel contains cDNA preparations from breast carcinoma (GI-101), lung carcinoma (LX-1), colon adenocarcinoma (CX-1), lung carcinoma (GI-117), prostate adenocarcinoma (PC3), colon adenocarcinoma (GI-112), ovarian carcinoma (GI-102) and pancreatic adenocarcinoma (GI-103). GI-101 is a poorly differentiated mammary carcinoma isolated from recurrent ductal carcinoma. 7) LX-1 is a poorly differentiated carcinomal surgical explant from a metastasis in a 48-year-old male.8) CX-1 is a moderately well-differentiated adenocarcinoma consistent with gastrointestinal origin, ...

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