Effect of sex hormones on the expression of chloride/formate exchanger (Cfex, Slc26a6) in rat organs

Abstract

Izmjenjivač klora i mravlje kiseline (Cfex/Slc26a6) je transmembranski protein koji prenosi ione klora, bikarbonata, oksalata, mravlje kiseline i hidroksilne ione u sisavaca. Dosadašnja istraživanja pretpostavljaju ulogu tog proteina u nastanku oksalatnih kamenaca u urinarnom traktu, s većom učestalošću u muškaraca. Zbog slabe istraženosti u štakora, kao učestalo rabljenih pokusnih životinja, njegova lokalizacija i moguća spolno-ovisna ekspresija istražena je u različitim tkivima/organima štakora. Specifičnost anti-CFEX protutijela utvrđena je heterolognim ekspresijskim sustavom u kojemu su stanice HEK293 transfecirane cjelovitom cDNA za rCfex. Metodom RT-PCR, ekspresija mRNA za rCfex utvrđena je u tankom crijevu, bubrezima, jetri i gušterači. Imunofluorescencijskom analizom protein rCfex je lokaliziran u četkastim membranama enterocita i proksimalnih kanalića bubrega, kanalikularnim membranama hepatocita i izvodnim kanalićima gušterače. U bubrezima je kastracija smanjivala, a testosteron povećavao njegovu ekspresiju. Dakle, mRNA/protein rCfex je prisutan u različitim tkivima/organima štakora, dok je u bubrezima njegova ekspresija dominantna u mužjaka uslijed stimulacijskog učinka androgena nakon puberteta.The chloride/formate exchanger (Cfex/Slc26a6) is a transmembrane protein mediating chloride, bicarbonate, oxalate, formate and hydroxyl ions transport in mammals. Previous findings support its possible role in the oxalate urinary tract stones formation, with higher incidence in men. Due to lack of data in rats, as commonly used experimental animals, its cell localization and possible sex-related expression was investigated in various rat organs/tissues. Specificity of anti-CFEX antibody was verified by heterologous expression systems using a full-length rCfex cDNA-transfected HEK293 cells. By RT-PCR, rCfex mRNA was detected in the kidneys, liver, pancreas and small intestine. By immunofluorescence, protein rCfex was localized in the apical membrane of enterocytes and renal proximal tubules, hepatocyte canalicular membranes, and luminal domain of pancreatic ducts. Its renal expression was downregulated by castration, and upregulated by testosteron. Therefore, the rCfex mRNA/protein is expressed in various rat organs/tissues, whereas its renal expression is maledominant due to post-pubertal androgen stimulation