Which vitamin is a person over the age of 70 years most likely to absorb poorly?

Calcium

Calcium absorption is vitamin D-dependent; therefore bioavailability depends upon vitamin D intake and status. The efficiency of absorption is related to physiological requirements for calcium and is dose-dependent. Dietary inhibitors of calcium absorption include substances that form complexes in the intestine, such as phytate. Protein and sodium may also modify calcium bioavailability in that high levels increase urinary calcium excretion. Although this is accompanied by an increase in intestinal absorption, the net result may be a reduction in the proportion of dietary calcium utilized by the body, i.e., lower bioavailability. Lactose, on the other hand, promotes calcium absorption. Other potential enhancers of calcium absorption are currently under investigation with regard to the development of functional foods for the prevention of osteoporosis, such as caseinophosphopeptides and nondigestible oligosaccharides. (See CALCIUM | Physiology; CHOLECALCIFEROL | Physiology; PROTEIN | Interactions and Reactions Involved in Food Processing.)

Fractional Calcium Absorption

Calcium absorption occurs by an active, carrier-dependent process and a passive, paracellular process. The active process is vitamin-D-dependent, but the passive process is not. When calcium intakes are low, 25(OH)D is converted to 1,25(OH)2D, which up-regulates transcription of calcium transport proteins in the gut as described in Chapter 19. However, this homeostatic regulation mechanism is unable to correct for chronically low calcium intakes. If vitamin D stores are too low, conversion to 1,25(OH)2D is reduced. Vitamin D status would likely have to be very low, i.e. <10 nmol/L [97] given the substrate pool of 25(OH)D is up to 1000-fold higher than 1,25(OH)2D. Because passive absorption is not vitamin-D-dependent, high calcium intakes can mitigate reduced calcium absorption efficiency in the face of low vitamin D status. Low vitamin D status therefore becomes a risk for bone health when calcium intakes are low. The gaps in knowledge in children and adolescents involve the calcium intake needed to overcome vitamin D insufficiency and the roles of vitamin D status, among other regulators associated with reduced calcium absorption efficiency, when calcium intakes are low.

Calcium absorption efficiency in children is determined in only a few laboratories worldwide. The various approaches have been reviewed [98,99]. The best method for a double isotope tracer technique is where one calcium tracer is given by oral administration to determine absorption and adjusted for tracer dilution by a second intraveneously administered tracer. This method gives true fractional calcium absorption. Isotopic tracers are then measured in serum or urine samples. Recently, a single oral isotope tracer method has been validated in adolescents against the double isotope reference methods [100]. Another method is net calcium absorption determined by metabolic balance, which necessitates controlled diets and complete urine and fecal collections. This method does not correct for endogenous secretion, i.e., absorbed calcium that is re-excreted in the gut. Indirect methods of calcium absorption are changes in serum calcium or iPTH after a calcium challenge. Sequential blood draws following the calcium load allows calculation of area under the curve (AUC).

The relationship between vitamin D and calcium absorption efficiency has not been studied extensively. Cross-sectional studies show no relationship between serum 25(OH)D and calcium absorption efficiency in adolescents with higher 25(OH)D concentrations [68,91], but in those with serum 25(OH)D concentrations below 50–62.5 nmol/L, the relationship appears negative [91,101] (Fig. 37.5). Conversely, when expressed as total calcium absorbed, the relationship appears positive [43]. While the above data are meaningful, cross-sectional studies are unable to assign causal effects. Longer-term, supplementation studies provide greater insight with regard to the relationship between vitamin D and calcium absorption efficiency. For example, vitamin D supplementation at 1000 IU/d for 4 weeks improved vitamin D status, but resulted in decreased fractional calcium absorption despite no significant change in serum 1,25(OH)2D or PTH [102]. The unexpected decrease in calcium absorption with increased vitamin D status as shown in this study may occur if serum 25(OH)D is a limiting substrate pool for forming 1,25(OH)2D, such that at very low vitamin D status, the active hormone cannot be formed in sufficient quantities to stimulate calcium absorption. In adults, calcium absorption increases when vitamin D status falls below 10 nmol/L [97]. However, the mean presupplementation serum 25(OH)D level in the Park et al. [102] study was 48 nmol/L. Sufficiently high calcium intakes suppress iPTH release and therefore, conversion to 1,25(OH)2D. Calcium intake in the vitamin D supplementation study by Park et al. [102] was ~1000 mg/d, which may have been sufficient to suppress iPTH release, but does not explain the inverse relationship between serum 25(OH)D and calcium absorption. The inverse relationship between serum 25(OH)D and iPTH may explain the decreased calcium absorption at higher 25(OH)D concentrations through decreased conversion to 1,25(OH)2D. Indeed, serum 1,25(OH)2D has been shown to be positively correlated with calcium absorption efficiency in adolescent girls (r = 0.35, P = –0.001) [90]. Calcium absorption is influenced by an interaction of calcium intake and genetics, as well as diet. For example, the Fok1 polymorphism of the VDR gene was significantly related to calcium absorption (P = 0.008), where those with the FF genotype had the greatest calcium absorption efficiency [103]. FGF-23 has an indirect role on calcium absorption since it down-regulates renal 1α-hydroxylase and decreases 1,25(OH)2D.

Fractional Calcium Absorption

Calcium absorption occurs by an active, carrier-dependent process and a passive, paracellular process. The active process is vitamin D-dependent, but the passive process is not. When calcium intakes are low, 25(OH)D is converted to 1,25(OH)2D, which upregulates transcription of calcium transport proteins in the gut as described in Chapter 30. However, this homeostatic regulation mechanism is unable to correct for chronically low calcium intakes. If vitamin D stores are too low, conversion to 1,25(OH)2D is reduced. Vitamin D status would likely have to be very low, i.e., <10 nmol/L [133] given the substrate pool of 25(OH)D is up to 1000-fold higher than 1,25(OH)2D. Because passive absorption is not vitamin D-dependent, high calcium intakes can mitigate reduced calcium absorption efficiency in the face of low vitamin D status. Low vitamin D status therefore becomes a risk for poor bone health when calcium intakes are low. The gaps in knowledge in children and adolescents involve the calcium intake needed to overcome vitamin D insufficiency and the roles of vitamin D status, among other regulators associated with reduced calcium absorption efficiency, when calcium intakes are low.

Calcium absorption efficiency in children is determined in only a few laboratories worldwide. The various approaches have been reviewed [134,135]. The best method for a double isotope tracer technique is where one calcium tracer is given by oral administration to determine absorption and adjusted for tracer dilution by a second intravenously administered tracer. This method gives true fractional calcium absorption. Isotopic tracers are then measured in serum or urine samples. A single oral isotope tracer method has been validated in adolescents against the double isotope reference methods [136]. Another method is net calcium absorption determined by metabolic balance, which necessitates controlled diets and complete urine and fecal collections. This method does not correct for endogenous secretion, i.e., absorbed calcium that is reexcreted in the gut. Indirect methods of calcium absorption are changes in serum calcium or iPTH after a calcium challenge. Conducting sequential blood draws following the calcium load allows for calculation of area under the curve.

Despite the known role of vitamin D metabolites in calcium absorption pathways, there is little evidence that vitamin D supplementation enhances calcium absorption in clinical trials unless vitamin D status is very low. Cross-sectional studies show no relationship between serum 25(OH)D and calcium absorption efficiency in adolescents with higher 25(OH)D concentrations [96,128], but in those with serum 25(OH)D concentrations below 50–62.5 nmol/L, the relationship appears negative [128,136] (Fig. 41.5) [61]. Conversely, when expressed as total calcium absorbed, the relationship appears positive [88]. Although the above data are meaningful, cross-sectional studies are unable to assign causal effects. In adolescents, vitamin D supplementation at 1000 IU/day for 4 weeks improved vitamin D status, but calcium absorption efficiency was not improved [60]. In adults, calcium absorption increases when vitamin D status falls below 10 nmol/L [133]. However, the mean presupplementation serum 25(OH)D in the Park et al. [60] study was 48 nmol/L. Sufficiently high calcium intakes suppress iPTH release and therefore conversion to 1,25(OH)2D. Calcium intake in the vitamin D supplementation study by Park et al. [60] was ∼1000 mg/d, which may have been sufficient to suppress iPTH release. The strongest study to date on the effect of vitamin D supplementation in fractional calcium absorption was the dose-response GAPI study in 323 black and white boys and girls [5]. Children 9–13 years of age were randomized to one of five doses of vitamin D3, i.e., 0, 400, 1000, 2000, or 4000 IU daily for 12 weeks. Although the changes in serum 25(OH)D were dose dependent, there was no effect of supplementation on fractional calcium absorption [5].

Calcium absorption is influenced by an interaction of calcium intake and genetics, as well as diet. For example, the Fok1 polymorphism of the VDR gene was significantly related to calcium absorption (P = .008), where those with the FF genotype had the greatest calcium absorption efficiency [137]. FGF-23 has an indirect role on calcium absorption because it downregulates renal 1α-hydroxylase and decreases 1,25(OH)2D.

A. Intestinal Calcium Absorption and Osteoporosis

Calcium absorption by the transcellular pathway is an active process requiring influx from the lumen of the gut into the enterocyte by epithelial calcium channels (especially TRPV6 and to a lesser extent TRPV5), followed by transcellular CaBP9k-mediated transport and extrusion into the bloodstream by PMCA pumps. The vitamin D endocrine system regulates all three steps, but the most regulated and essential step is the epithelial calcium influx [5–7]. The paracellular calcium transport is functional only during a high calcium intake and is probably only partially vitamin D dependent.

Intestinal absorption of calcium declines with age, particularly after the age of 70 years [8, 9]. This has been suggested to contribute to the decline in bone density with age that occurs in both sexes. Several groups have compared calcium absorption in osteoporotic patients with that [10] in age-matched normal subjects. Most [9, 10], but not all [11], studies have found that calcium absorption is lower in osteoporotic subjects. The reason for impaired intestinal calcium absorption with aging in general and in osteoporotic patients is not clear. VDR and calbindin 9K levels do not decrease with age [12]. Serum 1,25-(OH)2D decreases with age [13] and is sometimes but not consistently lower in osteoporotic patients than in age-matched controls [9, 10]. Dual calcium isotope studies, however, reveal a good correlation between active calcium absorption and serum 1,25-(OH)2D in adolescents [14], but such well-validated studies are not available or less convincing [15] for a large group of elderly subjects with or without osteoporosis. Heaney reported a positive relation between serum 25(OH)D and intestinal calcium absorption with lower absorption when 25(OH)D levels fall below 80 nmol/L. As 25(OH)D is only a precursor for the active hormone, local 1-hydroxylase activity is then supposed to be the mediator of active calcium absorption in the intestine [7].

2.2 Intestinal Absorption of Calcium

Intestinal calcium absorption more than doubles during pregnancy and this appears to be the main mechanism that enables the fetal calcium demand to be met.2 Placental phosphorus and magnesium absorption are similarly increased.2 Studies using stable isotopes of calcium and other methods have revealed that intestinal calcium absorption is upregulated by midpregnancy in rodents and 12 weeks of gestation in humans; the most marked increase is late in pregnancy in both species, corresponding to the interval of greatest fetal demand for mineral.2 A positive calcium balance results by midpregnancy in rodents and humans, and skeletal mineral content may also increase.20,49 Isotope studies in rats have revealed that 92% of fetal skeletal mineral content derives from the maternal diet during pregnancy.50

As total calcitriol levels double or triple during pregnancy, it has been assumed that calcitriol regulates the increased intestinal calcium absorption. However, studies in rodents indicate that a pregnancy-related increase in intestinal calcium absorption precedes the rise in calcitriol and occurs despite severe vitamin D deficiency,51,52 absence of the vitamin D receptor,22 or maternal parathyroidectomy.53 This upregulated calcium absorption is physiologically significant because vitamin D-deficient rats and Vdr null mice both achieved a significant increase in skeletal mineral content during pregnancy, which reached 155% of baseline in Vdr nulls.2,22 In humans intestinal calcium absorption doubles from the first trimester and may precede an increase in free calcitriol.2 These data suggest that factors other than calcitriol must also stimulate intestinal calcium absorption during pregnancy.

Additional animal studies suggest that the Vdr null mouse compensates by upregulating intestinal expression of the calcium channel TRPV6,22 and that PRL and placental lactogen may stimulate intestinal calcium absorption independently of calcitriol, possibly by stimulating TRPV6.54–58 Although compelling animal data indicate that calcitriol or its receptor are not required to upregulate intestinal calcium absorption during pregnancy, no studies have examined the effects of vitamin D deficiency on intestinal calcium absorption during human pregnancy. A clinical study found that hyperprolactinemic men and women had calcitriol levels and rates of intestinal calcium absorption that were similar to normoprolactinemic controls59; however, the hormonal milieu of hyperprolactinemia differs from that of pregnancy, especially with respect to the relative concentrations of sex steroids.

Calcium Homeostasis

Calcium absorption across the intestinal membrane occurs via both a vitamin D–dependent, saturable pathway and a vitamin D–independent, nonsaturable pathway. The duodenum is the major source of calcium absorption, although the remainder of the small intestine and the colon also contribute. In the kidney, approximately 60% to 70% of calcium is reabsorbed passively in the proximal tubule, driven by a transepithelial electrochemical gradient generated by sodium and water reabsorption. A further 10% is absorbed in the thick ascending limb by paracellular transport. The regulation of reabsorption is via transcellular pathways that occur in the distal convoluted tubule, the connecting tubule, and the initial portion of the cortical collecting duct.

Proximal Straight Tubule

Because the proximal straight tubule (pars rectae) does not extend to the surface of the kidney and is not amenable to conventional micropuncture techniques, calcium transport by this segment has been less thoroughly studied than in proximal convoluted tubules. Indirect estimates of calcium reabsorption derived from the difference between the calcium concentration of fluid samples obtained from late proximal convoluted tubules and those from the tip of the loop of Henle suggested that 10% of the filtered calcium is recovered by proximal straight tubules (116, 231, 260). This approach necessitates obtaining proximal samples from a superficial nephron, while the samples from the loop of Henle are taken from the tip of a juxtamedullary nephron. Nonetheless, direct in vitro microperfusion studies of proximal straight tubules substantiated the conclusion that proximal straight tubules absorb 10% of the filtered calcium (44, 400).

MECHANISMS OF PROXIMAL TUBULE CALCIUM TRANSPORT

Calcium absorption by proximal tubules may be mediated by a combination of passive and active transport mechanisms. The transepithelial absorption of calcium can be conceptually described by the following relation: calcium absorption = passive transport + active transport where passive absorption is the sum of diffusion and solvent drag. These relations can be expressed formally as (468):

(2)JCa2+=PCa2+(ΔC Ca2++ziRTC¯ Ca2+Δψ)diffision+ (1−σCa2+)C¯Ca2+ +JVsolventdrag+ JCa2+activeactive transport

where PCa2+ is the apparent calcium permeability, ΔCCa2+ is the transepithelial calcium concentration difference ([lumen-to-bath] – [bath-to-lumen]), zi is the valence, R the gas constant, T the absolute temperature,

Ca2+ is the average transmural calcium concentration across the tubule ([lumen-to-bath + bath-to-lumen]/2), Δψ is the transepithelial voltage, σCa2+ is the reflection coefficient for calcium, Jv is the net fluid absorption, and JCa2+active is the metabolically active, transcellular calcium transport.

Most evidence suggests that proximal tubule calcium transport proceeds primarily by passive diffusion and solvent drag mechanisms. These mechanisms would dictate a paracellular route for absorption. Active transport is a two-step process, wherein calcium enters the cell across apical plasma membranes and is then extruded across basolateral plasma membranes. Evidence for passive calcium absorption will be discussed first and that for active transport second. A general schematic representation of these processes in proximal tubules is shown in Fig. 5.

FIGURE 5. Model of proximal tubular calcium absorption. The inset shows the portion of the nephron referred to in the cell model. In proximal tubules, the majority of calcium is absorbed by passive mechanisms through the paracellular pathway. Evidence supports the presence of a small component of active, transcellular calcium transport.

The finding that the (TF/GF)Ca2+ ratio in proximal convoluted tubules is virtually unity (Table 2) suggests that the permeability to calcium is high. Direct assessments of calcium permeability support this inference. The apparent calcium permeability of rat proximal convoluted tubule is 15–21 × 10−5 cm sec−1 (Table 3), whereas that of rabbit proximal convoluted tubules is approximately 10 × 10−5 cm sec−1. The divergence between these values is likely to be attributable to the generally higher permeability coefficients for all ionic species in the rat compared with the rabbit (202). However, methodological differences may also contribute to the disparity since the measurements on the rat were performed in vivo, where the rigorous conditions required to measure permeability are more difficult to achieve than with isolated tubules studied in vitro. Irrespective of the origin of these differences, the relatively high apparent permeability coefficients are consistent with a mechanism of transport that proceeds primarily by passive diffusion. Thus, in the latter portions of S1 proximal convoluted tubules, where the (TF/GF)Ca2+ ratio is greater than unity, the calcium concentration gradient, combined with the high permeability, constitute a favorable driving force for passive calcium absorption. Furthermore, tubular fluid traversing late proximal convoluted tubules contains more chloride than bicarbonate. Since chloride permeability is greater than bicarbonate permeability in proximal convoluted tubules, the concentration differences for these anions result in a diffusion voltage that is oriented as electropositive in the lumen with respect to the peritubular fluid (17). This positive voltage also serves as a driving force for passive calcium absorption. In proximal straight tubules, the development of an electropositive transepithelial voltage, together with the high permeability, would likewise represent a significant driving force for passive calcium absorption. In both cases such passive calcium absorption proceeds through the paracellular pathway between adjacent cells (Fig. 5).

TABLE 3. Proximal Tubule Calcium Permeability Coefficients and Absorption

Segment	Pca2+ cm sec−1× 10−5	Jca2+ mol min−1 mm−1 × 10−12	Species	Reference
PCT	15a	4.4	Rat	31
	17a	3.4b		469
	21a,c	6.9		329
	19a	10.2		467
		5.2		379
	7.0d	0.9e	Rabbit	336
	7.1a,c	3.3		157
PST S2	4.0	5.6	Rabbit	400
	2.4	0.65		44
	1.1	0.56		380
PST S3	1.2	0.61	Rabbit	380

PCT, proximal convoluted tubule; PST S2, superficial S2 proximal straight tubule; PST S3, juxtamedullary S3 proximal straight tubule.

aAssuming luminal diameter of 30 μm.bCalculated from average perfused tubule length of 1.95 mm.cEstimated from net flux and average difference between perfused and collected calcium concentrations.dMeasured as bath-to-lumen influx coefficient.eNet transport estimated from the difference between the lumen-to-bath and bath-to-lumen fluxes.

Solvent drag may represent a second passive means of calcium absorption in proximal convoluted tubules. According to this view, calcium is entrained during fluid absorption. This is theoretically possible if the reflection coefficient, σCa, is less than unity. The magnitude of solvent drag is a function not only of σCa but also of the rate of fluid absorption, Jv, and the average transmural calcium concentration across the tubule,

Ca2+. Bomsztyk and Wright estimated the apparent σCa to be 0.72 (32). They microperfused rat proximal tubules in situ and simultaneously manipulated net fluid absorption by adding mannitol, an impermeant solute, at various concentrations to the luminal perfusion fluid. Reduction or reversal of fluid absorption was simultaneously accompanied by changes of calcium absorption. Thus, in these experiments a small component of proximal calcium absorption may be mediated by solvent drag or by solute polarization in an unstirred layer adjacent to the membrane. A change in Jv of 1.15 nl min−1 mm−1 was associated with a change of calcium transport of about 1 pmol min−1 mm−1. Since control rates of Jv in the rat proximal tubule measured under the same conditions were 2.34 nl min−1 mm−1, solvent drag accounts for less than 15% of the total calcium absorption (30, 32) (Table 3). Ng et al. (336) reported a similar value for σCa of 0.89 in single isolated perfused rabbit proximal S2 convoluted tubules. However, calculations of the calcium absorption attributable to solvent drag suggest that it is effectively negligible (0.06 × 10−12 mol min−1 mm−1). Moreover, no evidence for active calcium transport was adduced (336).

Factoring (TF/GF)Ca2+ (or [TF/UF]Ca2+) by the extent of fluid absorption ([TF/P]inulin) provides a measure of the delivery of calcium to that point in the nephron, while (1–[TF/UF]Ca2+/[TF/P]inulin) is the fractional calcium absorption. Maximal calcium absorption by proximal convoluted tubules reaches some 70% of that filtered. It is generally accepted that most calcium absorption by proximal tubules is passive and proceeds through the paracellular pathway, which provides an aqueous conduit between adjacent cells (31, 44, 336). These comments notwithstanding, careful examination of calcium absorption under conditions where driving forces for passive calcium movement and fluid absorption were eliminated does provide convincing evidence that a finite component of calcium absorption proceeds by an active transport pathway in proximal convoluted tubules (31, 400, 469). Ullrich et al. (469) estimated that one-third of the net transport in rat proximal tubules occurs by an active mechanism and that passive mechanisms account for the remaining two-thirds. Bomsztyk and colleagues (30, 31) confirmed the presence of active calcium absorption in microperfusion experiments. In these studies, the electrochemical driving force for calcium across the proximal tubule epithelium was determined by measuring transepithelial voltage and ionized Ca2+ activity under conditions of zero net fluid flux. Under these conditions, calcium absorption varied with the magnitude of the electrochemical driving forces. When the transepithelial electrochemical gradient was abolished, some residual net calcium absorption remained, confirming the presence of a small component of active transport.

It should be noted that, although small by comparison with paracellular calcium absorption, active cellular absorption by proximal tubules amounts to some 20 μmol/min (469), which, in fact, is approximately twice that of the distal nephron, where calcium absorption is entirely cellular.

The mechanism of calcium transport by the S2 segment of the proximal straight tubule resembles that of the proximal convoluted tubule. Studies using isolated perfused rabbit S2 proximal tubules, under experimental conditions designed to minimize net fluid movement and the electrochemical gradient for Ca2+, generally are consistent with the idea that passive driving forces are the major determinant of calcium absorption (44, 336). However, evidence for a significant amount of active calcium transport has been reported (400). Sacks and Bourdeau (403) showed that when passive driving forces across isolated S2 segments of rabbit proximal straight tubules were experimentally manipulated, the direction and rate of net calcium flux were predicted by the magnitude of the imposed electrochemical gradient. Thus, passive diffusion appears to be the major mechanism of transport in proximal straight tubules.

Intestinal Absorption of Calcium

Intestinal calcium absorption is normal in lactating women [9,36,79–81,241–243], and this change appears to coincide with calcitriol falling to normal (nonpregnant) levels. Adding 1 g of calcium or placebo daily did not alter the fractional absorption of calcium in lactating women [242]. Intestinal calcium absorption is somewhat higher in women whose menses have resumed while they are still lactating [242].

Lactating women evidently do not need increased efficiency of intestinal calcium absorption to meet the calcium requirements of milk production. This is supported by evidence from randomized trials and observational studies. Increased dietary intake of calcium does not reduce skeletal resorption during lactation, nor does it alter breast milk calcium content [201,244–248].

During the postweaning interval, intestinal calcium absorption increased 20% in one study [242] but did not change significantly in another study [36].

Intestinal Calcium Absorption

Calcium absorption is moderately efficient in humans, with 35% of dietary intake typically absorbed. The transfer of calcium across the intestinal barrier occurs through both saturable (presumably transcellular) and nonsaturable (presumably paracellular diffusion) pathways. Calcium absorption is saturable in the duodenum (proximal segment of small intestine) and to a lesser extent in the jejunum (midportion of the small intestine). Animal studies suggest that saturable calcium transport may also occur in the large intestine. The saturable pathway is energy dependent. Calcium moves from the mucosal to serosal side of the intestine, even against a concentration gradient. This pathway is under nutritional and physiologic regulation. For the standard recommended calcium intake (i.e., 400–500 mg per meal), the saturable transport accounts for more than 60% of total calcium absorption in the small intestine, thus demonstrating the importance of the saturable calcium absorption pathway under normal dietary loads.

In contrast, passive transport, involving claudins-2, -12, and -15, occurs throughout the entire intestine. Passive transport is a nonsaturable, linear function, dependent on the calcium concentration found in a given segment of the intestinal lumen.

When calcium intake is adequate to high, the proportion of calcium transported in any given intestinal segment is determined by the following: (1) the presence of saturable and nonsaturable pathways, (2) the transit time through the intestinal tract, and (3) the solubility of calcium within the intestinal segment. As a result, even though calcium solubility is low and the saturable pathway is absent or downregulated in the ileum (the final segment of the small intestine), the total amount of calcium absorbed is actually greatest in the ileum because transit time through this segment is 10 or more times longer than through the more proximal intestinal segments.

Habitual consumption of a low-calcium diet stimulates processes to increase small intestine calcium absorption efficiency. This effect is mediated in part through changes in the plasma concentrations of the most active vitamin D metabolite, 1,25(OH)2D. There are four different models for regulated calcium transport (Fig. 13.3).

In the facilitated diffusion model, calcium enters epithelial cells through the apical membrane calcium channel TRPV6. TRPV6 delivers calcium to calbindin-D9K, a low molecular weight, cytosolic, calcium-binding protein proposed to facilitate transcellular calcium movement. Calcium is then actively extruded across the basolateral membrane. This is primarily mediated by PMCA1b moving calcium against a concentration gradient, though the NCX1 sodium-calcium exchanger also contributes. Each of these proteins are transcriptionally regulated by 1,25(OH)2D.

The validity of the facilitated diffusion model has been challenged. Animals lacking TRPV6 or calbindin-D9K still increase the efficiency of intestinal calcium absorption in response to dietary calcium restriction, and 1,25(OH)2D still increases calcium absorption in TRPV6 knockout mice. Furthermore, mice without calbindin-D9K have normal calcium absorption, both at baseline and in response to 1,25(OH)2D. However, mice with the combined knockouts of TRPV6 and calbindin-D9K have a limited response to 1,25(OH)2D. These studies suggest that other mechanisms in addition to facilitated diffusion contribute to the process of active calcium transport across enterocytes.

The vesicular transport model predicts that calcium absorption required cycling of calcium-containing lysosomes in enterocytes (as in Fig. 13.3). In enterocytes, 1,25(OH)2D increases both lysosomal number and calcium content. Lysosomal calcium uptake models occurring after TRPV6 transport and endocytosis into cells have been proposed. Transient receptor potential cation channel subfamily V member 5 (TRPV5) and TRPV6 may be present in some vesicular structures and facilitate calcium transport. Enterocyte vesicle calbindin-D28K is also reported in chicks. However, it is not clear whether calcium accumulation in vesicles is specific to mammalian transcellular calcium transport regulation.

Transcaltachia refers to a rapid, 1,25(OH)2D-stimulated increase in calcium transport (Fig. 13.3). In contrast to the facilitated diffusion model, transcaltachia does not require gene transcription, although, like in the vesicular transport model, transportation across the cell may still involve vesicles. In ex vivo perfused chick intestine, exposure to 1,25(OH)2D for 14 min dramatically increases calcium transport across enterocytes.

Transcaltachia appears to be mediated by a basolateral membrane receptor: either by a unique role for the vitamin D3 receptor (VDR) at the basolateral surface, by a novel membrane vitamin D receptor called the membrane-associated, rapid response steroid-binding protein (MARRS), or by the PTH-PTHrP receptor. However, MARRS knockout mice do not have disrupted transcellular calcium absorption or whole body calcium metabolism.

Regulated paracellular transport also contributes to calcium absorption. 1,25(OH)2D increases production of the tight junction proteins claudin-2 and claudin-12 in the jejunum and ileum facilitating passive transport. However, vitamin D regulates active calcium absorption in the proximal duodenum and jejunum, as opposed to the ileum, where claudin-2 and claudin-12 expression is highest. Prolactin upregulates claudin-15 during pregnancy and lactation, contributing to paracellular calcium absorption, as well as TRPV5, TRPV6, and calbindin-D9K also upregulating transcellular calcium transport. In addition, the voltage-dependent L-type calcium channel subunit alpha-1D (also known as voltage-gated calcium channel subunit alpha Cav1.3) may also contribute to intestinal calcium absorption. However, this protein’s gene is not vitamin D regulated and Cav1.3 knockout mice do not have strong disruptions in either calcium or bone metabolism.

In sum, no single model is able to explain fully intestinal calcium transport mechanisms and the timing of responses to 1,25(OH)2D. Data support both transcriptional and more rapid responses of calcium transport to 1,25(OH)2D; more rapid increases in calcium transport suggest that mechanisms such as vesicular transport may be important. Redundancy in this system likely enables greater control and efficiency of calcium absorption, given that relative dietary deficiency is common and excess calcium consumption can also occur. Further studies are necessary to delineate the relative contributions of these various models and their mechanisms.

Intestinal Calcium Absorption

Calcium absorption is moderately efficient in humans, with 35% of dietary intake typically being absorbed. The transfer of calcium across the intestinal barrier occurs through both saturable (presumably transcellular) and nonsaturable (presumably paracellular diffusion) pathways. Calcium absorption is saturable in the duodenum (proximal segment of small intestine) and to a lesser extent in the jejunum (midportion of the small intestine). Animal studies suggest that saturable calcium transport may also occur in the large intestine. The saturable pathway is energy dependent, with calcium movement from the mucosal to serosal side of the intestine, even against a concentration gradient. This pathway is under nutritional and physiologic regulation. For the standard, recommended amounts of calcium intake (i.e. 400–500 mg per meal), the saturable component of calcium absorption will account for more than 60% of total calcium absorption in the small intestine, thus demonstrating the importance of the saturable calcium absorption pathway under normal dietary loads.

In contrast, passive transport occurs throughout the length of the intestine and is a nonsaturable, linear function, dependent upon the calcium concentration found in a given segment of the intestinal lumen. Under adequate to high calcium intakes, the proportion of calcium transported in any given intestinal segment is determined by the presence of the saturable and nonsaturable pathways, the transit time through the intestinal tract, and the solubility of calcium within the intestinal segment. As a result, even though calcium solubility is low and the saturable pathway is absent or downregulated in the ileum (the final segment of the small intestine), the total amount of calcium absorbed is actually greatest in the ileum since transit time through this segment is 10 or more times greater than through the more proximal intestinal segments.

Habitual consumption of a low-calcium diet stimulates increased small intestine calcium absorption efficiency. This effect is mediated in part through changes in the serum concentrations of the most active vitamin D metabolite, 1,25(OH)2D. Over 40 years of research on the mechanism of calcium absorption has resulted in the development of the facilitated diffusion model (Fig. 13.3). In this model calcium enters the epithelial cell through the apical membrane calcium channel TRPV6 which delivers calcium to calbindin-D9K, a low molecular weight, cytosolic, calcium-binding protein proposed to facilitate transcellular calcium movement. Active extrusion of calcium across the basolateral membrane is mediated by PMCA1b moving calcium against a concentration gradient. 1,25(OH)2D regulates the gene transcription of each of these proteins.

Several studies have challenged the validity of the facilitated diffusion model. For example, in animals lacking TRPV6 or calbindin-D9K, dietary calcium restriction can still increase the efficiency of intestinal calcium absorption, and 1,25(OH)2D is still able to increase calcium absorption in TRPV6 knockout mice. Furthermore, mice without calbindin-D9K demonstrate normal calcium absorption at baseline, and in response to 1,25(OH)2D. However mice with the combined knockouts of TRPV6 and calbindin-D9K do have a limited response to 1,25(OH)2D.

These studies suggest that other mechanisms also contribute to the process of active calcium transport in the enterocytes. Three additional alternative models have been proposed but each has weaknesses that limit their acceptance. The vesicular transport model predicts that cycling of calcium-containing lysosomes in the intestine is necessary for calcium absorption (as in Fig. 13.3). 1,25(OH)2D increases lysosomal calcium in enterocytes, as well as increasing the number of lysosomes. Models of uptake into lysosomes after transport into the cell by TRPV6, and also of endocytosis, have been proposed. Transient receptor potential cation channel subfamily V member 5 (TRPV5) and TRPV6 may be present in some vesicular structures, which may facilitate transport, and the presence of calbindin-D28K in chick enterocyte vesicles is reported. However, it is not yet clear whether this accumulation of calcium in vesicles is specific to regulation of transcellular calcium transport in mammals.

An alternative model is known as transcaltachia (Fig. 13.3). This is a model for a rapid, 1,25(OH)2D-stimulated increase in calcium transport that does not require gene transcription. During transcaltachia, the process of transportation across the cell may still involve vesicular transport, while the effects of 1,25(OH)2D on molecules involved in the facilitated diffusion model require transcription. In ex vivo perfused chick intestine, exposure to 1,25(OH)2D for 14 min dramatically increases calcium transport across enterocytes, detectable as increased calcium in the perfusate. Transcaltachia appears to be mediated by a basolateral membrane receptor: either by a unique role for the vitamin D3 receptor (VDR) at the basolateral surface, by a novel membrane vitamin D receptor called the membrane-associated, rapid response steroid-binding protein (MARRS), or by PTH-PTHrP. However, MARRS knockout mice do not have disrupted transcellular calcium absorption or whole body calcium metabolism.

A third alternative model involves regulated paracellular movement of calcium through tight junctions. Production of the tight junction proteins claudin-2 and claudin-12 is increased in response to 1,25(OH)2D and decreased in Vdr-null mice; thus, this may provide vitamin D-dependent regulation of paracellular calcium transport. However, vitamin D regulates active calcium absorption in the proximal small intestine, as opposed to the ileum, where claudin-2 and claudin-12 expression is highest. Finally, the voltage-dependent L-type calcium channel subunit alpha-1D (also known as voltage-gated calcium channel subunit alpha Cav1.3) may also contribute to intestinal calcium absorption. However, the gene for this protein is not regulated by vitamin D and neither calcium nor bone metabolism is strongly disrupted in Cav1.3 knockout mice.

Currently, no single model fully explains observations of intestinal calcium transport, and the timing of responses to 1,25(OH)2D. Data supports both transcriptional and more rapid responses of calcium transport to 1,25(OH)2D; these more rapid increases in calcium transport suggest that mechanisms such as vesicular transport may be important. Redundancy in this system may enable greater control and efficiency of calcium absorption, given that relative dietary deficiency is common. Further studies are necessary to delineate the relative contributions of these various mechanisms.