Annette Liesegang

Diet-induced pseudohypoparathyroidism: A hypocalcemia and milk fever risk factor

Subclinical hypocalcemia may affect half of all multiparous cows, and clinical hypocalcemia or milk fever affects approximately 5% of dairy cows each year. This disorder of calcium homeostasis can be induced by several dietary factors. Recent studies implicate high dietary potassium and high dietary cation-anion difference (DCAD) with increased risk of milk fever. The hypothesis tested in this study was that high-DCAD diets fed to prepartum cows reduce tissue sensitivity to parathyroid hormone (PTH), inducing a pseudohypoparathyroid state that diminishes calcium homeostatic responses. Multiparous Jersey cows were fed low- or high-DCAD diets in late gestation, creating a compensated metabolic alkalosis in the high-DCAD cows and a compensated metabolic acidosis in the low-DCAD cows. They then received synthetic PTH injections at 3-h intervals for 48 h. Parathyroid hormone is expected to cause an increase in plasma calcium by increasing renal production of 1,25-dihydroxyvitamin D and increasing bone calcium resorption. Plasma calcium concentration increased at a significantly lower rate in cows fed the high-DCAD diet. Cows fed the high-DCAD diet also produced significantly less 1,25-dihydroxyvitamin D in response to the PTH injections than cows fed the low-DCAD diet. Serum concentrations of the bone resorption marker carboxyterminal telopeptide of type I collagen were numerically lower in cows fed the high-DCAD diet but this difference was not statistically significant. These data provide direct evidence that high-DCAD diets reduce tissue sensitivity to PTH. The metabolic alkalosis associated with high-DCAD diets likely induces a state of pseudohypoparathyroidism in some dairy cows at the onset of lactation, resulting in hypocalcemia and milk fever.

Roux-en-Y gastric bypass surgery reduces bone mineral density and induces metabolic acidosis in rats

Roux-en-Y gastric bypass (RYGB) surgery leads to bone loss in humans, which may be caused by vitamin D and calcium malabsorption and subsequent secondary hyperparathyroidism. However, because these conditions occur frequently in obese people, it is unclear whether they are the primary causes of bone loss after RYGB. To determine the contribution of calcium and vitamin D malabsorption to bone loss in a rat RYGB model, adult male Wistar rats were randomized for RYGB surgery, sham-operation-ad libitum fed, or sham-operation-body weight-matched. Bone mineral density, calcium and phosphorus balance, acid-base status, and markers of bone turnover were assessed at different time points for 14 wk after surgery. Bone mineral density decreased for several weeks after RYGB. Intestinal calcium absorption was reduced early after surgery, but plasma calcium and parathyroid hormone levels were normal. 25-hydroxyvitamin D levels decreased, while levels of active 1,25-dihydroxyvitamin D increased after surgery. RYGB rats displayed metabolic acidosis due to increased plasma lactate levels and increased urinary calcium loss throughout the study. These results suggest that initial calcium malabsorption may play a key role in bone loss early after RYGB in rats, but other factors, including chronic metabolic acidosis, contribute to insufficient bone restoration after normalization of intestinal calcium absorption. Secondary hyperparathyroidism is not involved in postoperative bone loss. Upregulated vitamin D activation may compensate for any vitamin D malabsorption.

In contrast to sheep, goats adapt to dietary calcium restriction by increasing intestinal absorption of calcium.

Many studies aimed at understanding calcium homeostasis in the cow use sheep or goats as ruminant models. However, the comparability of relevant homeostatic processes between ruminant species has not been assessed. Therefore, we investigated whether the mechanisms of maintaining calcium homeostasis are similar in different ruminant species. Dietary calcium of goats was restricted along with treatment with calcitriol in a similar protocol to that in a recent study with sheep. Plasma calcium and phosphate and parameters of bone metabolism were analysed. Gastrointestinal calcium transport was characterised in vitro in Ussing chambers. The expression of apical epithelial calcium channels, calbindin-D(9K), and the basolateral plasma membrane Ca(2+)-ATPase was determined by quantitative RT-PCR and Western blot analysis. In contrast to sheep, the goats were able to compensate for low dietary calcium supply by increasing active calcium absorption in the small intestine, especially in the jejunum. As in sheep, the observed ruminal calcium transport of goats was affected neither by the calcium restricted diet nor by the calcitriol treatment, thus indicating the presence of an alternative, vitamin D-independent mechanism of calcium transport in the forestomachs. These results demonstrate that mechanisms for maintaining calcium homeostasis differ significantly between ruminant species.

Effect of source and quantity of dietary vitamin D in maternal and creep diets on bone metabolism and growth in piglets.

Piglets are born with reduced plasma concentrations of 25-hydroxycholecalciferol (25-OH-D(3)) and are thus highly predisposed to vitamin D deficiency. Furthermore, sow milk contains little vitamin D, and the slow intestinal vitamin D absorption of sows limits the efficacy of dietary vitamin D supplementation. Hence, the neonate depends, to a large extent, on the vitamin D stores built up in fetal tissues from maternal sources. The current study was undertaken to evaluate whether the source and quantity of dietary vitamin D provided to the gestating and lactating sow, and also directly in the form of creep feed to the piglet, would influence the vitamin D status, growth performance, and skeletal development of piglets. A total of 39 primiparous and multiparous sows were randomly assigned to 1 of 3 dietary treatments (13 in each treatment), supplemented with either 5 or 50 μg of the commonly used cholecalciferol (vitamin D(3)) or 50 μg of 25-OH-D(3) per kilogram of feed. By wk 3 of lactation, piglets were offered a creep diet with vitamin D supplementation according to the treatment of the dam, and they were offered the same creep diets after weaning at d 35 of age until they reached a BW of approximately 20 kg. When dietary 25-OH-D(3) was provided, circulating concentrations of 25-OH-D(3) in piglet serum increased (P < 0.05) as early as d 21 and later at d 33 and 77, indicating greater body stores in those animals. Bone-breaking strength and cortical bone mineral content and density at the tibial midshaft of piglets were reduced (P < 0.05) when vitamin D(3) was supplemented at 5 μg/kg compared with the bone traits of other groups, but no differences (P > 0.05) were observed between the 2 other groups. After weaning, ADFI was greater (P < 0.05) and growth performance tended (P = 0.08) to improve when doses of 50 μg/kg were administered, regardless of the vitamin D source. In conclusion, supplementation of the diet with 50 μg/kg of either source of vitamin D was proved to be adequate in meeting the needs of gestating sows and in permitting the accumulation of vitamin D in fetal tissues, as well as for normal skeletal mineralization and growth in the offspring. Furthermore, the markedly improved vitamin D status of piglets whose mothers received 25-OH-D(3) possibly resulted from greater tissue reserves present at birth and a greater availability of vitamin D when released from those stores.

Influence of different calcium supplies and a single vitamin D injection on vitamin D receptor and calbindin D9k immunoreactivities in the gastrointestinal tract of goat kids

The purpose of this study was to investigate whether diets differing in Ca concentration would have an influence on vitamin D (VitD) receptor (VDR) and calbindin D9k (Calb9k) immunoreactivities in the gastrointestinal tract of growing goats. In addition, the effect of a single VitD injection was studied, to clarify whether exogenous VitD would further increase the active Ca absorption mechanisms. The hypothesis of the study was that reduced Ca intake would lead to greater active Ca absorption, and with that, to greater amounts of VDR and Calb9k immunoreactivities. The normal Ca kid group (according to age requirements) received 2.5 to 6 g of Ca/d, whereas the lesser Ca kid group (less than requirements) received 1.5 to 4 g of Ca/d from wk 6 (weaning) to 15 (slaughter). In addition, 5 and 6 goat kids, respectively, of each group (normal Ca kid group, lesser Ca kid group), were injected with VitD (0.05 mg of cholecalciferol/kg of BW) in wk 14 of life. Blood samples were taken in wk 14 and 15. Calcium and VitD (25-hydroxyvitamin D and 1,25-dihydroxyvitamin D) concentrations were determined in serum. Immediately after slaughter, the duodenum (DD) and rumen (RU) were mounted in conventional Ussing chambers. Unidirectional flux rates of Ca across gastrointestinal tissues were measured. Additionally, tissue specimens of the gastrointestinal tract were collected, and formaldehyde-fixed paraffin sections were used for VDR and Calb9k immunohistochemistry. In all kid groups, a net absorption in the RU and a net secretion of Ca in the DD were observed. Immunoreactions of VDR were greatest in the duodenal mucosa, whereas Calb9k immunoreactions were observed in the forestomach and intestinal tissues. The greatest expression was observed in the duodenal surface epithelium. Additionally, in the VitD-injected groups, an immunoreaction occurred in the jejunal superficial and basal glands and the ileal superficial epithelium. In contrast, the other groups showed no Calb9k immunoreactions at these sites. In conclusion, there is clear evidence for the RU as a main site for Ca absorption. The results of this study also indicate that VDR and Calb9k are highly expressed in the duodenal mucosa. The active absorption may not have such an important role in the DD because active transport was also evident in the RU. However, Calb9k expression seems to be stimulated by VitD administration.

Influence of Anionic Salts on Bone Metabolism in Periparturient Dairy Goats and Sheep

The purpose of the present study was to investigate the influence of diets supplemented with anionic salts on bone metabolism of dairy goats and sheep. Twelve Saanen goats and 12 Ostfrisean milk sheep (fourth lactation) were divided into 2 groups each [sheep control (SC), goat control (GC); sheep anionic salts (SA), goat anionic salts (GA)]. Each group was fed a different diet in the last 10 d of gestation. Groups SC and GC received a normal diet according to the requirements of goats and sheep in this stage of gestation. Groups SA and GA received supplemental anionic salts. The dietary cation-anion difference (DCAD) was +524 (SC) and +515 (GC) vs. -163 (SA) and -164 (GA) mEq/kg of dry matter. Blood and urine samples were collected daily until parturition. Serum Ca, P, Mg, serum crosslaps (CTX), osteocalcin, 1,25-dihydroxy-vitamin D (VITD), urinary pH, and urinary Ca concentrations were analyzed. Bone mineral density and bone mineral content were measured with peripheral quantitative computer tomography. The bone resorption marker CTX showed significant differences between the animals supplemented with anionic salts and the control animals in goats, but not in sheep. The goats receiving anionic salts had greater CTX concentrations throughout the administration of the salts. In sheep, a difference was only observed on the day of parturition. Similar observations were made in VITD concentrations, although a significant difference between the goat groups was only observed 3 d prepartum. The bone formation marker osteocalcin was lower prepartum in the animals supplemented with anionic salts. The urinary pH was lower in the SA and GA animals, whereas urinary Ca concentrations were greater. Bone mineral content and bone mineral density decreased in all groups around parturition. In conclusion, this experiment showed that the addition of anionic salts in goats led to greater bone resorption rates while on this feeding regimen. It can be concluded that the anionic salts induced a mild metabolic acidosis with all its effects on calcium metabolism. These effects were not evident in milk sheep.

Influence of benzoic acid and phytase in low-phosphorus diets on bone characteristics in growing-finishing pigs.

In 2 simultaneous experiments (Exp. 1 and Exp. 2), the effects of benzoic acid (BA) and phytase (Phy) in low-P diets on bone metabolism, bone composition, and bone stability in growing and growing-finishing pigs were examined. Experiment 1 was conducted with 16 crossbred gilts in the BW range of 25 to 66 kg of BW, whereas in Exp. 2, 32 crossbred gilts (25 to 108 kg of BW) were used. All pigs were individually housed in pens and restrictively fed 1 of 4 diets throughout the experiment. Total P content of the wheat-soybean diets was 4 g/kg (all values on an as-fed basis). The experimental diets were 1) unsupplemented control diet; 2) control diet with 0.5% BA; 3) Phy diet with 750 Phy units (FTU) of Phy/kg and no BA; and 4) PhyBA, control diet with 750 FTU of Phy/kg and 0.5% BA. Blood samples were taken at the beginning of the experiment, wk 3 (only for pigs in Exp. 1), wk 6, and before slaughter to determine P and Ca in serum and concentrations of total alkaline phosphatase, serum crosslaps (marker for bone resorption), and osteocalcin (marker for bone formation). Ash, P, and Ca contents of bones and bone stability were examined using the left metatarsal bones and tibia of the pigs after slaughter. Benzoic acid did not influence any of the blood variables (P > 0.09). The addition of Phy increased (P < or =0.03) P concentration in serum from 2.71 +/- 0.08 to 3.03 +/- 0.07 mmol/L at wk 3 and content of serum crosslaps from 0.39 +/- 0.02 to 0.45 +/- 0.02 ng/mL at wk 6 and decreased (P < 0.05) osteocalcin at wk 6 by 160 ng/mL. No long-term effect of diets on serum mineral concentrations, alkaline phosphatase, and bone markers in serum could be detected. Benzoic acid negatively affected (P < or = 0.03) Ca content in bones and distal bone mineral density, especially in the younger pigs. In the control diet with 0.5% BA and the control diet with 750 FTU of Phy/kg and 0.5% BA, the CA content in bones and distal bone mineral density were reduced by 6 and 11%, respectively. Throughout the whole growing and finishing period, Phy increased (P < or =0.02) ash, P, and Ca contents in bones by 29.4, 4.8, and 11.6 g/kg of DM, respectively. Bone mineral density and bone mineral content were greater in diets with Phy (P < or = 0.03), as well as breaking strength of tibia (+22%) and metatarsal bones (+27%; P < 0.01). The results of this study indicate that for a healthy skeleton, BA should not be used in low-P diets without the addition of Phy.

Organic matter and macromineral digestibility in domestic rabbits (Oryctolagus cuniculus) as compared to other hindgut fermenters

It is generally assumed that animal species differ in physiological characteristics of their digestive tract. Because investigating digestive processes is often labour-intensive, requiring lengthy adaptation and collection periods, comparisons are often made on the basis of data collated from different studies. We added a new data set on dietary crude fibre (CF) and macromineral (Na, K, Ca, P, Mg) composition and apparent digestibility (aD) of organic matter (OM) and macrominerals in rabbits (Oryctolagus cuniculus; a total of 180 measurements with 12 individuals and 10 different diets) to a literature data collection for rodents and domestic horses (Equus caballus). Significant negative relationships between dietary CF and aD OM were obtained, but the significant difference in digestive efficiency between rabbits and guinea pigs (Cavia porcellus) reported from studies where both species had been investigated under identical conditions were not detectable in the larger data collection. However, the 95% confidence interval of the negative slope of the CF-aD OM relationship did not overlap between domestic horses on the one hand, and rabbits and guinea pigs on the other hand, indicating a less depressing effect of CF in horses. Akaikes information criterion indicated that aD of a macromineral did not only depend on its concentration in the diet, but also on the individual and various other diet characteristics and the presence of other macrominerals, indicating complex interactions. The data indicate similar mechanisms in the absorption of macrominerals amongst rabbits, rodents and horses, with the exception of Na and K in guinea pigs and Ca in chinchillas (Chinchilla laniger) that should be further corroborated. In particular, the occurrence of high aD Ca in hindgut fermenters, regardless of whether they employ a digestive strategy of coprophagy or whether they have fossorial habits, suggests that this peculiarity might represent an adaptation to hindgut fermentation in general.

Expression of renal and intestinal Na/Pi cotransporters in the absence of GABARAP

We have recently shown that the abundance of the renal sodium (Na)/inorganic phosphate (Pi) cotransporter NaPi-IIa is increased in the absence of the GABAA receptor-associated protein (GABARAP). Accordingly, GABARAP-deficient mice have a reduced urinary excretion of Pi. However, their circulating levels of Pi do not differ from wild-type animals, suggesting the presence of a compensatory mechanism responsible for keeping serum Pi values constant. Here, we aimed first to identify the molecular basis of this compensation by analyzing the expression of Na/Pi cotransporters known to be expressed in the kidney and intestine. We found that, in the kidney, the upregulation of NaPi-IIa is not accompanied by changes on the expression of either NaPi-IIc or PiT2, the other cotransporters known to participate in renal Pi reabsorption. In contrast, the intestinal expression of NaPi-IIb is downregulated in mutant animals, suggesting that a reduced intestinal absorption of Pi could contribute to maintain a normophosphatemic status despite the increased renal retention. The second goal of this work was to study whether the alterations on the expression of NaPi-IIa induced by chronic dietary Pi are impaired in the absence of GABARAP. Our data indicate that, in response to high Pi diets, GABARAP-deficient mice downregulate the expression of NaPi-IIa to levels comparable to those seen in wild-type animals. However, in response to low Pi diets, the upregulation of NaPi-IIa is greater in the mutant mice. Thus, both the basal expression and the dietary-induced upregulation of NaPi-IIa are increased in the absence of GABARAP.

Effect of different potassium levels in hay on acid–base status and mineral balance in periparturient dairy cows

Forages commonly used in dry cow rations contain high K concentrations. This results in a high dietary cation-anion difference (DCAD), which can compromise the calcium homeostasis of periparturient cows. The aim of this study was to determine the effect of 2 types of hay, fed during the prepartum period and differing in their K concentrations, on the peripartum acid-base status and mineral balance of dairy cows. During the prepartum period, the cows of group K(33) (n = 6) received a diet based on hay with a high K concentration (33 g/kg of DM), whereas the cows of group K(13) (n = 6) received a diet based on hay with a low K concentration (13 g/kg of DM). Both experimental diets were formulated to be isoenergetic and isonitrogenous. After calving, all cows received the same diet based on hay K(33). Blood and urine samples were taken on d 14, 7, and 3 before parturition, at parturition, and then daily during the first 8 d after calving. Concentrations of minerals were analyzed in both blood and urine. Creatinine was also measured in urine for the calculation of the mineral:creatinine ratio. The acid-base parameters in blood (pH and HCO(3)(-) concentration) and urine (pH, net acid-base excretion, and base-acid quotient) were determined on d 14, 7, and 3 before parturition, at parturition, and on d 1 after parturition. The use of hay K(13) reduced the DCAD value of the prepartum diet by half (195 vs. 514 mEq/kg of DM). No significant differences between the 2 groups were observed for blood acid-base indicators or plasma minerals except for the Mg plasma concentration, which tended to be higher in group K(13) from d 3 prepartum to d 2 after calving. In group K(13), urinary Ca excretion tended to be higher from d 3 prepartum to d 1 after parturition than that in group K(33). On d 3 before parturition, urinary pH and net acid-base excretion were significantly lower in group K(13) than in group K(33). On d 14, 7, and 3 before parturition, base-acid quotient was significantly lower in group K(13) than in group K(33). In group K(13), daily feed intake and hence daily intake of Ca, P, and Mg during d 3 and 4 after parturition were higher than in group K(33). The decrease of the DCAD in positive ranges by feeding a low-K hay before parturition induced a reduction of the metabolic alkalotic charge, as observed in acid-base parameters in urine, and increased the availability of Ca and P as a result of higher feed intake at the onset of lactation.