204 A COMPARATIVE ANALYSIS OF CALCIUM TRANSPORT GENES EXPRESSION IN HOLSTEIN AND HANWOO (KOREAN CATTLE) IN THE DUODENUM AND KIDNEY

Abstract

Transmembranous calcium (Ca2+) channels such as Trpv5/6,Ncx1,Pmca1b are known to play an important role in maintaining homeostasis and metabolizing Ca2+ ions. Transient receptor potential cation channel subfamily V members 5 and 6 (Trpv5/6) play an important role in Ca2+ absorption. Na+/Ca2+ exchanger 1 (Ncx1) and plasma membrane Ca2+-transporting ATPase1 (Pmca1b) play a role in Ca2+ excretion. Holstein cattle are known to provide higher milk production than other cattle breeds. In this respect, a higher susceptibility to hypocalcaemia that is a risk factor for many of calcium-related diseases such as milk fever has been observed. In contrary, Hanwoo cattle are relatively resistant to the calcium-related diseases. The aim of this study was to evaluate the expression of calcium transport genes in the duodenum and kidney of Hanwoo (Korean cattle) and Holstein cattle. All samples were obtained from Holstein and Hanwoo cattle from 36 to 60 months of age at a local slaughterhouse. The collected duodenum and kidney were rapidly excised and washed in cold sterile saline (0.9% NaCl). Total RNA was prepared from duodenum and kidney tissues using TRIzol Reagent (Invitrogen, Carlsbad, CA, USA), according to the manufacturer's protocol. Protein was extracted using Pro-prep (iNtRON Bio), according to the manufacturer's protocol. Expression of Trpv5/6, Ncx1 and Pmca1b was analysed by real-time RT–PCR, Western blot analysis, and immunohistochemistry. In dairy cattle's kidney (Holstein), the level of Trpv5 mRNA was ~3-fold more than Hanwoo's kidney while Pmca1b mRNA was one-fifth less than Hanwoo's kidney. However, no difference in Trpv6 and Ncx1 mRNA expression was observed in the kidney of Holstein compared with Hanwoo. Duodenal Trpv5 and Trpv6 mRNA were scarcely expressed in both Hanwoo and Holstein cattle. In addition, the mRNA expression of Pmca1b and Ncx1 were also not different between Hanwoo and Holstein. The protein expression of calcium transport genes were similar to the mRNA levels of calcium transport genes in both Hanwoo and Holstein. Localization of calcium transporter genes were identified, the glomerulus, and proximal and distal convoluted tubules expressed TRPV5, 6, Pmca1b, and Ncx1 in the kidney. These four calcium transport genes may play an important role in the bovine duodenum and kidney. The difference between Holstein and Hanwoo animals which express different gene expression patterns may be helpful in studying diseases associated with calcium metabolism, e.g. milk fever.