Expression of TRPV6 and CaBP-D28k in the egg shell gland (uterus) during the oviposition cycle of the laying hen

Abstract

1. The aim of this study was to investigate the localisation of the transient receptor potential vanilloid channel type 6 (TRPV6) in egg shell gland (ESG) and examine the dynamic expression of TRPV6 and Calbindin-d28k (CaBP-D28k), as well as the changes in concentration of total calcium (Ca), total inorganic phosphorus (P), alkaline phosphatase (ALP), parathyroid hormone (PTH) and calcitonin (CT) in plasma during the oviposition cycle. 2. The plasma ALP activity was notably increased at 8 h. In addition, plasma CT was highest at 0 h and significantly lower at 8 h. The change of plasma PTH concentration increased slightly post-oviposition and reached a maximum at 16 h. 3. Immunohistochemical analysis indicated that TRPV6 was strongly localised to the apical luminal epithelium of the mucosa. The mRNA levels of TRPV6 and CaBP-D28k in the ESG remained very low from 0 to 4.5 h, but were significantly increased at 16 h. Furthermore, Western blotting analysis showed that the expression of TRPV6 and CaBP-D28k also reached a maximum at 16 h and was different from the concentration of CaBP-D28k. 4. In conclusion, the epithelial Ca(2+) channel TRPV6 is strongly expressed in the epithelial cells of the eggshell gland, and the increase of TRPV6 and CaBP-D28k mRNA and protein expression during eggshell formation suggests that active Ca(2+) transcellular transport exerts significant effects in delivering active calcium in the ESG.

Avian eggshell thinning caused by transovarian exposure to <i>o,p</i>′-DDT: changes in histology and calcium-binding protein production in the oviduct uterus

“…These two proteins have been observed in the oviduct of birds, but their functional properties are poorly understood. CALB1 is present in the cytosol of gland cells of the uterus at high concentration (Jande et al, 1981), and strong TRPV6 immunostaining is found specifically in the apical regions of the luminal epithelium of the uterus (Yang et al, 2013). These findings generally agreed with our results, but we also observed CALB1 in the normal uterine epithelium and TRPV6 throughout the cytoplasm of the gland cells.…”

“…Because Japanese quail are not commonly used as experimental animals, we could not obtain species-specific target proteins and could not examine the reactivities of these primary antibodies in quail. However, the cross-reactivity of the CALB1 antibody against its target protein in chickens has been confirmed by its manufacturer, and immunostaining of chicken tissues with TRPV6 antibody has been reported previously (Yang et al, 2013). We found here that the CALB1, SPP1, and TRPV6 antibodies immunostained specific regions of the quail uterus; as described in the DISCUSSION, the respective proteins are located similarly in chickens.…”

“…SPP1 and its mRNA have been reported to be exclusively located in the epithelial cell layer of the uterus (Pines et al, 1995), unlike our finding that SPP1 was present both throughout the normal gland cell cytoplasm and in the epithelium. Because production of CALB1, SPP1, and TRPV6, or expression of their genes, or both, has been observed or found to be elevated during eggshell calcification, when the egg is present within the uterus (Jonchère et al, 2012;Pines et al, 1995;Yang et al, 2013), there can be little doubt that these proteins play key roles in eggshell calcification. Hypoplasia of the uterus caused by o,p′-DDT or DES leads to depletion of molecules involved in eggshell calcification, such as the calcium-binding proteins examined here, and could thus impair eggshell-forming ability.…”

Vitamin D: calcium and bone homeostasis during evolution

“…CT may, however, have a greater role than presently accepted in situations of calcium stress such as during pregnancy or lactation in mammals 100 or during egg shell calcification in birds. 102 Parathyroid glands first appeared during evolution in amphibians but PTH genes are widely expressed in most tissues of fish. 90 PTH receptors are older and were already used for signaling of PTHrP, which functions as a major paracrine factor 103 and can also act as a hypercalcemic factor in early vertebrates.…”

Exogenous estradiol improves shell strength in laying hens at the end of the laying period

“…Evidence is accumulating that the surface epithelium provides the bulk of the Ca 2+ needed for shell formation [37,38]. The latest finding supporting this is the localization of transient receptor potential vanilloid channel type 6 (TRPV6) to the apical surface of these cells and that its mRNA increases in the shell gland during shell formation [39]. Since estrogen increases TRPV6 in human uterus [12] this is a very interesting candidate for future studies to elucidate the effects of estrogen on shell gland function.…”