Calcium transport in male reproduction is possibly influenced by vitamin D and CaSR

Vitamin D is important for gonadal function in rodents, and improvement of vitamin D status in men with low sperm counts increases live birth rate. Vitamin D is a regulator of transcellular calcium transport in the intestine and kidney and may influence the dramatic changes in the luminal calcium concentration in epididymis. Here, we show spatial expression in the male reproductive tract of vitamin D receptor (VDR)-regulated factors involved in calcium transport: Transient receptor potential vanilloid 5/6 (TRPV5/6), sodium/calcium exchanger 1 (NCX1), plasma membrane calcium ATPase 1 (PMCA1), calbindin D9k, calcium-sensing receptor (CaSR), and parathyroid hormone-related peptide (PTHrP) in mouse and human testis and epididymis. Testicular Casr expression was lower in Vdr ablated mice compared with controls. Moreover, expression levels of Casr and Pthrp were strongly correlated in both testis and epididymis and Pthrp was suppressed by 1,25(OH)2D3 in a spermatogonial cell line. The expression of CaSR in epididymis may be of greater importance than in the gonad in mice as germ cell-specific Casr deficient mice had no major reproductive phenotype, and coincubation with a CaSR-agonist had no effect on human sperm-oocyte binding. In humans, seminal calcium concentration between 5-10 mM was associated with a higher fraction of motile and morphologically normal sperm cells and the seminal calcium concentration was not associated with serum calcium levels. In conclusion, VDR regulates CaSR and PTHrP, and both factors may be involved in the regulation of calcium transport in the male reproductive tract with possible implications for sperm function and storage.

Abstract P401: The Anti-arrhythmic Effects Of VDAC Isoforms Are Mediated By Glutamate 73 Through Regulation Of Mitochondrial Calcium Transport

Mitochondria critically regulate cellular processes such as bioenergetics, metabolism, calcium homeostasis and apoptosis. VDAC proteins are abundant proteins that control the passage of ions and metabolites across the outer mitochondrial membrane. We have previously shown that activation of VDAC2, is able to buffer excess calcium and thereby suppress calcium overload induced arrhythmogenic events in vitro and in vivo. However, the mechanism by which VDAC2 regulates calcium transport and cardiac contractions remained unclear. It is also unclear whether all three VDAC isoforms (VDAC1,2 and 3) possess similar cardioprotective activity. The zebrafish tremblor/ncx1 mutant lacks functional NCX1 in cardiomyocytes leading to calcium overload, and the manifestation of fibrillation-like phenotypes. Using the tremblor/ncx1 mutant as a model, we observed isoform-specific differences between the VDAC family members. VDAC1 and VDAC2 enhanced mitochondrial calcium trafficking and restore rhythmic contraction in tremblor mutants, whereas, VDAC3 did not. We found that the differing rescue capabilities of VDAC proteins were dependent upon residues in their N-terminal halves. Phylogenetic analysis further revealed the presence of an evolutionarily conserved glutamate at position 73 (E73) within VDAC1 and VDAC2, but a glutamine (Q73) in VDAC3. Excitingly, we showed that replacing VDAC2 E73 with Q73 ablated its calcium transporting activity. Conversely, substituting the Q73 with E73 allows VDAC3 to gain calcium trafficking and cardioprotective abilities. Overall, our study demonstrates an essential role for the evolutionarily conserved glutamate-73 in determining the anti-arrhythmic effect of VDAC isoforms through their regulation of mitochondrial calcium uptake.

Abstract MP43: Knockout Of ACE-N Facilitates Improved Cardiac Function After Myocardial Infarction

Angiotensin-converting enzyme (ACE) hydrolyzes N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) into inactive fragments through its N-terminal domain (ACE-N). We reported that cardioprotective effects of ACE inhibitor in angiotensin II-induced hypertension were partially mediated through increased Ac-SDKP bioavailability. Whether increased endogenous Ac-SDKP by knocking out ACE-N leads to improved cardiac function in myocardial infarction (MI) is unknown. Wild-type (WT) and ACE-N knockout (ACE-N -/- ) mice were subjected to MI induced by ligating the left anterior descending artery and treated with either vehicle or Ac-SDKP (1.6 mg/kg/day, s.c.) for 5 weeks. Echocardiography was performed on awake mice at the end of experiment and left ventricles (LV) were harvested for histology and molecular biology studies. ACE-N-/- mice showed increased plasma Ac-SDKP level in Sham or MI group compared to WT. Exogenous Ac-SDKP further increased Ac-SDKP level in both WT and ACE-N-/-. SF and EF were significantly decreased in both WT and ACE-N-/- mice post-MI, . Exogenous Ac-SDKP further increased EF and SF post-MI only in WT, but not in ACE-N-/- mice. Sarcoendoplasmic reticulum calcium transport ATPase 2 (SERCA2), a marker of cardiac calcium homeostatsis, significantly decreased in WT post-MI was rescued with Ac-SDKP, whereas ACE-N-/- mice displayed less reduction in SERCA2 expression. These results demonstrate that gene deletion of ACE-N improves cardiac function in mice post-MI, which was associated with increased Ac-SDKP level and minimally reduced expression of SERCA2.Therefore,this study illustrates that endogenous Ac-SDKP results in cardioprotective role in MI.

Promoting the Calcium-Uptake Bioactivity of Casein Phosphopeptides in vitro and in vivo

Casein phosphopeptides have been studied widely for their ability to chelate calcium. However, systematic studies on the effects of casein phosphopeptides (CPP) on calcium absorption in vitro and in vivo are scarce. The purities of two commercially available products, CPP1 and CPP2, are 18.37 and 25.12%, respectively. Here, the in vitro calcium binding capacity of CPP2 was 142.56 ± 7.39 mg/g, which was higher than that of CPP1 (107.15 ± 6.27 mg/g). The calcium transport results in a Caco-2 monolayer model indicated that, relative to controls, CPP1 and CPP2 increased calcium transport by 21.78 and 53.68%, respectively. Subsequent animal experiments showed that the CPP2-Ca-H group (1% Ca, 0.4% CPP2) had significant increases in the femur index, serum Ca2+ and serum osteocalcin levels, and femoral Ca content. The CPP2-Ca-H animal also had decreased serum alkaline phosphatase levels, parathyroid hormone content, and urinary pyridinoline content. Overall, our results demonstrated that CPP2 had stronger effects on promoting calcium uptake than CPP1.

Calcium Transport along the Axial Canal in Acropora

In Acropora, the complex canals in a coral colony connect all polyps to a holistic network, enabling them to collaborate in performing biological processes. There are various types of canals, including calice, axial canals, and other internal canals, with structures that are dynamically altered during different coral growth states due to internal calcium transport. In this study, we investigated the morphological changes in the corallite of six Acropora muricata samples by high resolution micro-computed tomography, observing the patterns of calcium carbonate deposition within axial corallite during processes of new branch formation and truncated tip repair. We visualized the formation of a new branch from a calice and the calcium carbonate deposition in the axial canal. Furthermore, the diameter and volume changes of the axial canal in truncated branches during rebuilding processes were calculated, revealing that the volume ratio of calcareous deposits in the axial canal exhibit significant increases within the first three weeks, returning to levels in the initial state in the following week. This work demonstrates that calcium carbonate can be stored temporarily and then remobilized as needed for rapid growth. The results of this study shed light on the control of calcium carbonate deposition and growth of the axial corallite in Acropora.

Calcium Transport in Specialized Dental Epithelia and Its Modulation by Fluoride

Most cells use calcium (Ca2+) as a second messenger to convey signals that affect a multitude of biological processes. The ability of Ca2+ to bind to proteins to alter their charge and conformation is essential to achieve its signaling role. Cytosolic Ca2+ (cCa2+) concentration is maintained low at ~100 nM so that the impact of elevations in cCa2+ is readily sensed and transduced by cells. However, such elevations in cCa2+ must be transient to prevent detrimental effects. Cells have developed a variety of systems to rapidly clear the excess of cCa2+ including Ca2+ pumps, exchangers and sequestering Ca2+ within intracellular organelles. This Ca2+ signaling toolkit is evolutionarily adapted so that each cell, tissue, and organ can fulfill its biological function optimally. One of the most specialized cells in mammals are the enamel forming cells, the ameloblasts, which also handle large quantities of Ca2+. The end goal of ameloblasts is to synthesize, secrete and mineralize a unique proteinaceous matrix without the benefit of remodeling or repair mechanisms. Ca2+ uptake into ameloblasts is mainly regulated by the store operated Ca2+ entry (SOCE) before it is transported across the polarized ameloblasts to reach the insulated enamel space. Here we review the ameloblasts Ca2+ signaling toolkit and address how the common electronegative non-metal fluoride can alter its function, potentially addressing the biology of dental fluorosis.

Calcium Transport along the Axial Canal in Acropora

In Acropora, the complex canals in a coral colony connect all polyps into a holistic network to collaborate in performing biological processes. There are various types of canals, including calice, axial canals, and other internal canals, with structures that are dynamically altered during different coral growth states due to internal calcium transport. However, few studies have considered the regulation of calcium transport in Acropora. In this study, we investigated the morphological changes of the axial canal in six Acropora muricata samples by high resolution micro-computed tomography, observing the patterns of the axial canal during the processes of new branch formation and truncated branch rebuilding. We visualized the formation of a new branch from a calice and deposition of the iconic hexactin skeletons in the axial canal. Furthermore, the diameter and volume changes of the axial canal in truncated branches during rebuilding processes were calculated, revealing that the volume ratio of calcareous deposits in the axial canal exhibit significant increases within the first three weeks, returning to levels in the initial state in the following week. This work indicates that the axial canal can transport calcium to form hexactin skeletons in a new branch and rebuild the tip of a truncated branch. The calcium transport along canal network regulates various growth processes, including budding, branching, skeleton forming, and self-rebuilding of an Acropora colony. Understanding the changes in canal function under normal and extreme conditions will provide theoretical guidance for restoration and protection of coral reefs.

Calcium isotope fractionation by osteoblasts and osteoclasts, across endothelial and epithelial cell barriers and with binding to proteins

Timely and accurate diagnosis of osteoporosis is essential for adequate therapy. Calcium isotope ratio (δ44/42Ca) determination has been suggested as a sensitive, non-invasive and radiation-free biomarker for the diagnosis of osteoporosis, reflecting bone calcium balance. The quantitative diagnostic is based on the calculation of the δ44/42Ca difference between blood, urine and bone. The underlying cellular processes, however, have not been studied systematically. We quantified calcium transport and δ44/42Ca fractionation during in-vitro bone formation and resorption by osteoblasts and osteoclasts and across renal proximal tubular epithelial cells (HK-2), endothelial cells (HUVEC) and enterocytes (Caco-2) in transwell systems, and determined transepithelial electrical resistance characteristics. δ44/42Ca fractionation was furthermore quantified with calcium binding to albumin and collagen. Calcified matrix formed by osteoblasts was isotopically lighter than culture medium by -0.27 ± 0.03‰ within 5 days, while a consistent effect of activated osteoclasts on δ44/42Ca could not be demonstrated. A transient increase in δ44/42Ca in the apical compartment by 0.26‰ occured across HK-2 cells, while δ44/42Ca fractionation was small across the HUVEC barrier, and absent with Caco-2 enterocytes, and with binding of calcium to albumin and collagen. In conclusion, δ44/42Ca fractionation follows similar universal principles as during inorganic mineral precipitation; osteoblast activity results in δ44/42Ca fractionation. δ44/42Ca fractionation also occurs across the proximal tubular cell barrier and needs to be considered for in-vivo bone mineralization modelling. In contrast, the effect of calcium transport across endothelial and enterocyte barriers on blood δ44/42Ca should be low and is absent with physiochemical binding of calcium to proteins.