Identification and characterization of aquaporin-9 (AQP9) in porcine hepatic tissue and hepatocytes in monolayer culture

Abstract

Aquaporins (AQPs) are members of a large family of integral membrane proteins involved in the rapid movement of water and neutral solutes across cell membranes. In this study, we have prepared an affinity-purified porcine-specific polyclonal antiserum to AQP9 and have investigated the distribution and expression of AQP9 in pig liver tissue and in hepatocytes in primary culture. Immunocytochemical analysis showed that AQP9 was primarily localized in the membrane structures of hepatocytes and was not associated with intrahepatic bile ducts or blood vessels. Western blot analysis indicated that AQP9 ranged in apparent molecular mass between 27 and 38 kD in whole liver and hepatocyte membrane fractions; minor components were also observed at approximately 34 kD in the cytosol compartment of hepatocytes, bile duct and gall bladder. A prominent immunoreactive band at 44 kD was shown to be an artifact of Western blot analysis. In primary cultures of porcine hepatocytes, glucagon enhanced absolute levels of AQP9 protein, while gene expression was enhanced by T3 and glucagon. Insulin alone had no discernable influence on AQP9 gene expression or its cellular protein levels. These data ଝ Mention of trade name, proprietary product or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture or imply its approval to the exclusion of other products or vendors that also may be suitable.* This article is a U.S. government work, and is not subject to copyright in the United States. 274T.J. Caperna et al. / Domestic Animal Endocrinology 32 (2007) [273][274][275][276][277][278][279][280][281][282][283][284][285][286] suggest that AQP9 is a major AQP in porcine hepatic tissue and appears to be primarily responsive to glucagon induction.

Triiodothyronine modulates the expression of aquaporin-8 in rat liver mitochondria

“…This idea agrees with the functional and regulatory distinctions known to exist between AQP8 and AQP9 in rat liver (Garcia et al 2001, Kuriyama et al 2002. The absence of T3 regulation of AQP9 in rat liver does not confirm the increase in the mRNA levels of AQP9 seen by Caperna et al (2007) who looked at the expression and distribution of AQP9 in pig liver tissues after stimulation with hormones, such as glucagon, insulin, and T3. Because the T3-induced enhancement of AQP9 transcript seen by Caperna et al (2007) was not accompanied by a parallel increase in the same AQP at a protein level, it is conceivable to speculate that the T3 modulation of AQP9 in porcine liver is not of major physiological relevance.…”

“…These data suggest a negative effect exerted by T3 on the expression of the AQP8 gene. Unlike AQP8 in this study and AQP9 in a recent work using porcine liver (Caperna et al 2007), no significant changes were found at the mRNA level for AQP9, the other major AQP expressed in the rat liver, throughout the various thyroid states (data not shown).…”

“…The absence of T3 regulation of AQP9 in rat liver does not confirm the increase in the mRNA levels of AQP9 seen by Caperna et al (2007) who looked at the expression and distribution of AQP9 in pig liver tissues after stimulation with hormones, such as glucagon, insulin, and T3. Because the T3-induced enhancement of AQP9 transcript seen by Caperna et al (2007) was not accompanied by a parallel increase in the same AQP at a protein level, it is conceivable to speculate that the T3 modulation of AQP9 in porcine liver is not of major physiological relevance. As also concluded by the same authors who found that glucagon enhanced the absolute levels of AQP9 protein (in addition to the mRNA ones), in pig liver AQP9 may be primarily responsive to glucagon induction.…”

“…Aquaporins (AQP9 and AQP1) have already been reported to be directly modulated by T3 at a transcriptional level in porcine liver (Caperna et al 2007) and rabbit developing kidney (Mulder et al 2003) respectively. However, no information is yet available about possible effects exerted by the different thyroid states on the expression of AQP8 in mitochondria.…”

A gel-based reference map of the porcine hepatocyte proteome

“…There is growing interest in porcine hepatocyte models for use in the investigation of swine metabolic processes [1][2][3][4], as well as, in human clinical applications for transplantation/artificial liver support [5] or toxicological analyses [6]. In all cases, whether fresh ଝ Mention of trade name, proprietary product or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture or imply its approval to the exclusion of other products or vendors that may also be suitable.…”