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. 2016 Aug 17;146(10):1953–1960. doi: 10.3945/jn.116.234310

Vitamin A Supplementation Transiently Increases Retinol Concentrations in Extrahepatic Organs of Neonatal Rats Raised under Vitamin A–Marginal Conditions1,2,3

Joanna K Hodges 1, Libo Tan 1,4, Michael H Green 1, A Catharine Ross 1,*
PMCID: PMC5037875  PMID: 27534819

Abstract

Background: Vitamin A (VA; retinol) supplementation is recommended for children aged >6 mo in countries with high rates of malnutrition, but the distribution and retention of VA in body tissues have not been extensively explored.

Objective: We sought to determine the distribution and retention of VA in tissues of neonatal rats raised under VA-marginal conditions.

Methods: Sprague-Dawley neonatal rats (n = 104; 63 males) nursed by mothers fed a VA-marginal diet (0.35 mg retinol equivalents/kg diet) were randomized and treated on postnatal day 4 with an oral dose of either VA (6 μg retinyl palmitate/g body weight) or canola oil as control. Pups (n = 4/group) were killed at 13 time points from 30 min to 24 d after dose administration. The total retinol concentration and mass were determined in all collected organs.

Results: In the control group, plasma VA was marginal (0.8 μmol/L), whereas liver VA was deficient (<70 nmol/g). Nonetheless, the liver contained most (∼76%) of the total VA mass in the body, whereas extrahepatic nondigestive organs together contained ∼13%. White adipose tissue (WAT), which was nearly absent before postnatal day 12, contained only ∼1%. In VA-supplemented neonates, the mean total retinol concentrations in all organs were significantly greater than in control pups. However, this increase lasted for only ∼1 d in most extrahepatic tissues, with the exception of WAT, in which it lasted 18 d.

Conclusions: Extrahepatic organs in neonatal rats raised under VA-marginal conditions store relatively little VA, and the scarcity of adipose tissue may predispose neonates to a low-VA status. The effect of VA supplementation on VA content in most extrahepatic organs is transient. A more frequent supplementation along with other nutritional interventions may be necessary for maintaining a steady supply of retinol to the rapidly developing extrahepatic organs.

Keywords: adipose, brain, extrahepatic, neonate, rat, retinol, skin, tissue distribution, vitamin A supplementation

Introduction

Vitamin A (VA5; retinol) is essential for the proliferation and differentiation of many different cell types and thus the development of the entire organism (1–3). Given the rapid growth that occurs during the neonatal period, it is reasonable to postulate that neonates have an increased need for VA. Studies have shown, however, that even healthy infants are born with low concentrations of VA in plasma and in the liver (<0.7 μM and < 10 μg/g, respectively), concentrations that would indicate deficiency if observed in older age groups (4, 5).

In addition to its role in cell differentiation, VA is also essential for immune response, and the problem of high infant mortality caused by infections in countries with high rates of VA deficiency has been recognized for many decades (6). In view of this problem, the WHO has initiated numerous clinical trials, several of which are still ongoing, to examine the effect of large-dose VA supplementation (50,000 IU/2.5 kg body weight) on infant survival (7). A meta-analysis of randomized clinical trials in which VA was given to children aged 6–59 mo revealed a 23–34% reduction in child mortality (8–10). On the other hand, VA supplementation administered to newborns within the first few days of life did not significantly affect mortality outcomes in infants aged <6 mo, despite several studies conducted in this age group (11, 12). The exact fate of VA administered to newborns remains, to a large extent, poorly understood.

In this study, our objective was to help fill this knowledge gap by determining the distribution of VA in tissues of neonatal rats, without and after VA supplementation at a dose equivalent—after adjusting for body weight (∼10 g)—to the dose that has been given to infants (50,000 IU/2.5 kg body weight). Specifically, we aimed to examine the concentration and mass of retinol in tissues that have been relatively unexplored from the perspective of VA metabolism, such as brain, skin, white adipose tissue (WAT), and brown adipose tissue (BAT). Previous studies have shown that neonates store a greater proportion (51% compared with 44%) of the whole-body VA mass than adult rats in tissues other than the liver, which is the main VA storage organ in VA-sufficient rodents (13). Moreover, a single dose of VA admixed with retinoic acid (VARA) stimulated the uptake of VA by extrahepatic tissues, especially the carcass and intestine, which together acquired ∼75% of the recently ingested VA (14). In view of these findings, we focused herein on extrahepatic tissues, subdividing the carcass—which in our previous experiment included several organs—into its components and determined the effect of VA supplementation, administered without retinoic acid, on total retinol concentration in WAT, BAT, skin, brain, and the remaining carcass over 24 d after dose administration. Because tissues grow rapidly during the neonatal period, organ weights and retinol mass accumulation over time were also determined. The results obtained provide a more comprehensive view of the impact of VA supplementation on neonatal tissue VA concentrations and reveal the transient nature of the effect of supplementation on extrahepatic VA storage in neonatal rats.

Methods

Animals and diet.

Pregnant Sprague-Dawley rats (n = 11) were purchased from Charles River Laboratories. Upon arrival, dams were switched to a VA-marginal AIN-93G-purified diet (15) modified to contain 0.35 mg retinol equivalents/kg diet (Research Diets) to render pups in a VA-marginal state similar to that of low-birth-weight infants in regions with a high prevalence of VA deficiency. All dams were housed individually at 22°C in a room with a 12-h light-and-dark cycle and given free access to food and water. All animal procedures were approved by the Pennsylvania State University Institutional Animal Care and Use Committee.

Dose administration.

The oral VA supplement consisted of VA in the form of all-trans-retinyl palmitate (Sigma-Aldrich) calculated to deliver 50,000 IU/2.5 kg body weight, as in human studies (7). With the use of the conversion factor 0.548 μg retinyl palmitate/IU, the dose was 6 μg body weight/g (21 nmol/g) for neonatal rats. The total amount of VA required for supplementing all pups was mixed with canola oil in proportions such that the desired VA mass would be delivered in a dose volume of 0.4 μL body weight/g + 1 μL as an allowance for retention in the pipette tip. The oil and VA doses were stored at −20°C in foil-wrapped vials to protect the VA from photodegradation.

All dams gave birth within 2 consecutive days to a total of 116 pups that were subsequently redistributed between 11 litters to avoid any effects caused by litter-size differences. Approximately 10 pups from each litter were randomly assigned to the control (n = 52; 30 males) and VA-supplemented (n = 52; 33 males) groups. On postnatal day 4, all pups were treated with an oral dose of either VA or canola oil as control. Immediately after being treated, each pup was returned to its mother and allowed to consume milk (and diet after weaning) for the remainder of the study.

Tissue collection.

At 13 time points after dose administration (0.5, 1, 4, 8, and 15 h and 1, 2, 4, 8, 11, 14, 18, and 22 d), 4 pups/group were removed from their cages, weighed, and killed with isoflurane or, if aged >14 d, CO2. Blood was collected from the vena cava into heparinized syringes, and the following tissues were dissected: liver, stomach, intestines (small and large intestine with the contents), lungs, kidneys with adrenals, brain, interscapular BAT, WAT (collected on and after postnatal day 12 from the inguinal and scapular depots), and the remaining carcass. Blood and liver were also collected from dams killed on the last day of the study. Blood samples were centrifuged at 800 × g for 15 min and stored at −20°C. Tissue samples were snap-frozen in liquid nitrogen and stored at −80°C.

Analyses of total retinol and retinyl esters.

Total retinol mass (unesterified + esterified retinol) was determined by ultra-performance liquid chromatography (UPLC) (Acquity UPLC System; Waters) with the use of an adaptation of the previously reported method (14, 16). Briefly, plasma aliquots (5–50 μL) and tissue homogenates (∼0.2 g) were incubated in 100% ethanol for 1 h for lipid extraction and then saponified with the use of potassium hydroxide. Neutral lipids were partitioned into hexanes containing 0.1% butylated hydroxytoluene. The partition step was repeated 2–3 times for lipid-rich tissues (brain, skin, BAT, WAT, and carcass) to attain ≥93% extraction efficiency, as determined in prior pilot testing for these tissues. After centrifugation, the upper-phase hexanes were removed, an internal standard (trimethylmethoxyphenyl-retinol) was added, the solvent was evaporated under nitrogen, and the residue was reconstituted immediately in 100 μL methanol for injection onto the reversed-phase column of the Acquity UPLC System. To measure retinyl esters, the saponification step was omitted, and the UPLC run time was extended from 1 to 7 min. All results were corrected for the unextracted portion of lipids.

Statistical analyses.

Values were expressed as means ± SEMs. Group means for plasma and tissue retinol mass and concentration at individual times were compared with the use of Student’s t test with Bonferroni correction for multiple comparisons (GraphPad Prism version 5.0). Changes over time were assessed with the use of simple linear regression analysis, with time as the independent variable and retinol concentration/mass as the dependent variable. A significant nonzero slope (β) indicated an increase or decrease over time. P < 0.05 was considered significant. To compare organs in terms of VA storage capacity, the organ retinol mass was averaged between postnatal days 4 and 8, when the relative organ weights were nearly constant (Supplemental Table 1). This period in rats corresponds to ∼1–2.5 y of age in humans (17). The means were then compared with the use of Wilcoxon matched-pairs signed rank test. Plasma total volume was ∼3.5% of body weight based on previously published data (18).

Results

Animal and relative organ growth.

Animals appeared to be healthy and grew rapidly (from ∼10 to 80 g) without differences caused by VA supplementation (Figure 1). The organ:body weight ratio was highest for the skin (0.2 ± 0.0) and carcass (0.4 ± 0.0). During the study, the organ:body weight ratio increased for the liver, intestine, and carcass and decreased for the lungs and brain. For the skin, this ratio increased before day 11 of the study and declined afterward (Supplemental Table 1).

FIGURE 1.

FIGURE 1

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Neonatal rat body weights between postnatal days 4 and 28. Inset shows the first 24 h after dose administration. Each point represents the mean ± SEM of 4 rats. VA, vitamin A.

Digestive organs.

The concentration of retinol in the stomach was elevated from 30 min to 1 h (P < 0.01) after VA supplementation and declined to the control group concentration thereafter (Figure 2A). Total retinol mass in the stomach followed a similar pattern (Figure 2B). In the intestine, the concentration of retinol peaked at 1 h after dose administration and remained significantly elevated for 24 h (P < 0.001) (Figure 2C). Total retinol mass in the intestine remained relatively constant despite a ∼900% increase in organ weight during the study (Figure 2D, inset).

FIGURE 2.

FIGURE 2

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Stomach (A, B) and intestine (C, D) total (unesterified + esterified) retinol concentration and mass in control and VA-supplemented neonatal rats between postnatal days 4 and 28. Insets show the first 24 h after dose administration (A, C) and organ weights (B, D). Each point represents the mean ± SEM of 4 rats. *P < 0.05. VA, vitamin A.

Plasma.

In the control group, the mean concentration of retinol in plasma (0.8 ± 0.1 μmol/L) was within the marginal status of 0.7–1 μM (19). In the VA-supplemented neonates, plasma retinol concentration peaked at 1 h after dose administration (3.6 ± 1.7 μmol/L) and returned to baseline 15 h later (Figure 3A). This increase was mainly caused by retinyl esters (Figure 3B), which are presumably present in postprandial chylomicrons. After 15 h, the plasma retinol concentration in the VA-supplemented group did not differ from that in the VA-marginal control group. Total retinol mass in plasma was also no different, except on day 14, when there was a ∼30% decrease in the VA-supplemented neonates (P < 0.001). After day 14, the mass of retinol in plasma increased more rapidly over time than did plasma volume (Figure 3C, inset).

FIGURE 3.

FIGURE 3

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Plasma total (unesterified + esterified) retinol concentration (A), retinyl ester concentration (B), and retinol mass (C) in control and VA-supplemented neonatal rats between postnatal days 4 and 28. Insets show the first 24 h after dose administration (A) and plasma volume (B). Each point represents the mean ± SEM of 4 rats (A, C) or a pooled sample of 4 rats killed at 1 sampling time (B). *P < 0.05. The dotted line represents the boundary between adequate and marginal plasma retinol concentrations (17). VA, vitamin A.

Liver, lungs, and kidneys.

In the liver of control neonates, the mean retinol concentration (59.4 ± 1.8 nmol/g) was deficient (<70 nmol tissue/g) (20) and decreased steadily over time (β = −5.9 ± 1.2; P < 0.001). VA supplementation increased the total retinol concentration by 302% from the control group value at 24 h after dose administration, and the concentration remained elevated at most time points until day 11 (P < 0.001) (Figure 4A). Total retinol mass in the liver increased in both groups until day 14 and then declined and remained steady until the end of the study (Figure 4B) despite continuous liver growth (Figure 4B, inset).

FIGURE 4.

FIGURE 4

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Liver (A, B), lung (C, D), and kidney (E, F) total (unesterified + esterified) retinol concentration and mass, respectively, in control and VA-supplemented neonatal rats between postnatal days 4 and 28. Insets show the first 24 h after dose administration (A, C, E) and organ weights (B, D, F). Each point represents the mean ± SEM of 4 rats. *P < 0.05. The dotted line represents the boundary between sufficient and deficient liver retinol concentrations (18). VA, vitamin A.

In the lungs of control pups, retinol concentration increased by ∼400% from days 0 to 18 (β = 0.3 ± 0.0; P < 0.001). After VA supplementation, lung retinol concentration was greater than in the control group, but only at 1 h (P < 0.01) and 24 h (P < 0.001) after dose administration. This was most likely caused by the large variation between pups within groups. After day 1, lung retinol concentration tracked similarly in both groups, with a gradual increase and then a rapid decline during the last 6 d of the study (Figure 4C). The lungs also showed a progressive accumulation of retinol relative to lung weight, which lasted until days 18 and 24 in the control and VA-supplemented neonates, respectively (Figure 4D).

The concentration of retinol in the kidneys of control neonates decreased by ∼80% from days 0 to 8 (β = −0.2 ± 0.0; P < 0.001). VA supplementation resulted in a significantly higher kidney retinol concentration than that of the control group that lasted until day 2. There were no differences thereafter that resulted from treatment (Figure 4E). Total retinol mass in kidneys remained steady (1.8 ± 0.1 nmol) in both groups, although gradual accumulation was observed during the last 10 d of the study (Figure 4F).

Other organs and the remaining carcass.

In the brain, VA supplementation increased retinol concentration by ∼300% from the control group value at 8 h after dosing (P < 0.001), with the difference lasting until day 1 (P < 0.05 for all) (Figure 5A). Retinol mass in the brain increased slightly over time (Figure 5B), but the increase was insufficient to match the rapid increase in brain weight (Figure 5B, inset). In BAT, VA supplementation increased retinol concentration by ∼400% from the control group value at 4 h (P < 0.05), and the concentration remained elevated until day 4 (P < 0.01 for all) (Figure 5C). Retinol did not accumulate in BAT despite continuous tissue growth (Figure 5D). In WAT, the concentration of retinol was still significantly elevated from that of the control group at day 18 after dose administration (P < 0.01) and declined gradually over the last 10 d of the study (Figure 5E). Retinol did not accumulate in WAT over time, and WAT weight did not significantly increase (Figure 5F). In the skin, VA supplementation increased total retinol concentration from that of the control group for ≤15 h after dose administration (P < 0.001) (Figure 5G). Retinol mass in the skin increased over time by ∼500% in both groups (βcontrol = 0.5 ± 0.0; βVA = 0.4 ± 0.1; P < 0.001 for both) (Figure 5H) in parallel to the increase in skin weight (Figure 5H, inset).

FIGURE 5.

FIGURE 5

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Brain (A, B), WAT (C, D), BAT (E, F), skin (G, H), and carcass (I, J) total (unesterified + esterified) retinol concentration and mass, respectively, in control and VA-supplemented neonatal rats between postnatal days 4 and 28. Insets show the first 24 h after dose administration (A, C, E, G, I) and organ weights (B, D, F, H, J). Each point represents the mean ± SEM of 4 rats. *P < 0.05. BAT, brown adipose tissue; VA, vitamin A; WAT, white adipose tissue.

In the carcass, the concentration of retinol was low (0.1 ± 0.0 nmol/g) and declined over time (except for day 24 in the VA-supplemented neonates), with no significant difference by treatment at any time during the study (Figure 6A). Retinol mass in the carcass was also low (0.7 ± 0.1 nmol) and declined over time (except for day 24 in the VA-supplemented group) despite a ∼900% increase in carcass weight (Figure 6B, inset).

FIGURE 6.

FIGURE 6

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Carcass total (unesterified + esterified) retinol concentration (A) and retinol mass (B) in control and VA-supplemented neonatal rats between postnatal days 4 and 28. Insets show the first 24 h after dose administration (A) and carcass weight (B). Each point represents the mean ± SEM of 4 rats. VA, vitamin A.

Discussion

This study provides comprehensive and, to our knowledge, novel information on the distribution of VA in tissues of neonatal rats raised under VA-marginal conditions. Previous studies examined the kinetics of VA administered orally to neonatal rats on postnatal day 4 and showed that ∼51% of the whole-body VA mass resided in the extrahepatic tissues (compared with ∼44% in adult rats) (14). In these experiments, however, total retinol mass in extrahepatic tissues was not measured directly but was predicted using mathematical modeling based on tracer kinetics in plasma and tissues. Furthermore, several extrahepatic tissues, including skin, brain, and adipose tissue, were lumped together with the carcass. We analyzed these tissues individually herein, leaving only the skeleton, muscle, and some connective tissue as the remaining carcass.

Based on previous findings, we hypothesized that the extrahepatic tissues in neonates would contain a relatively higher proportion of the whole-body VA than that observed in adults and that VA supplementation would increase the proportion present in the liver. Our findings did not support these hypotheses. Instead, the results demonstrated that, despite a liver retinol concentration that would be categorized as deficient, most VA reserves were still hepatic. VA supplementation had a transient effect on retinol concentrations in all extrahepatic tissues examined except WAT, but the supplementation did not affect tissue retinol distribution.

VA body distribution in the control neonates.

The liver contained ∼76% of the whole-body VA mass in control pups aged 4–8 d, a proportion similar to the 86% of hepatic retinol found in adult rats and humans in the VA-adequate state (21, 22). This relatively high proportion of hepatic retinol was maintained despite a liver retinol concentration that was deficient and significantly lower than that measured in adult mother rats fed the VA-marginal diet (Figure 7), a finding consistent with the low neonatal liver retinol concentration found in piglets fed a VA-free diet (∼25 nmol/g) and in newborn humans (5, 23).

FIGURE 7.

FIGURE 7

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Mean organ total (unesterified + esterified) retinol concentration (A) and mass (B) in control and VA-supplemented rats between postnatal days 4 and 8 (0–4 d after VA supplementation). Each bar represents the mean ± SEM of 32 rats, P < 0.05. Adult liver retinol concentration in A represents the mean ± SEM of 4 mother rats used in the study. *P < 0.05. BAT, brown adipose tissue; VA, vitamin A.

The lungs and kidneys together stored ∼5% of the whole-body VA and showed a relatively high retinol concentration consistent with the previously demonstrated role of these organs in VA storage and metabolism in rodents. Interestingly, our study showed that rats in the control group accumulated retinol in the lungs for the 22 d of this study despite the VA-marginal conditions. This trend may reflect the need for retinol in the lungs to support the process of secondary alveolar septation, which in rats occurs from postnatal days 1 to 14, unlike in humans, in whom this process is completed during gestation (24).

The remaining extrahepatic tissues each contributed <1% to the total VA mass in the body, except the skin, digestive organs, and carcass. The skin contained ∼6% of the whole-body VA mass, mostly because of its large weight (∼20% of the whole-body weight in pups aged 4–8 d) rather than high retinol concentration (∼1 nmol/g). Given the large surface:body weight ratio in newborns (25) and the importance of VA for the differentiation of cells in epithelial layers (26), it is possible that neonates have a greater requirement for VA in supporting normal skin development. The skin may also act as a reservoir of VA in neonates, in which WAT is nearly absent. The stomach and intestine each contained ∼5% of the whole-body VA mass. These organs, however, were analyzed together with their contents, which contributed to the measurement. The carcass, which constituted 40% of the whole-body weight, contained only 1.2% of the whole-body retinol mass and, together with the brain, had the lowest VA concentration (∼0.1 nmol/g).

Collectively, our results show that the extrahepatic nondigestive organs in control neonates contained ∼13% of the whole-body VA mass, a proportion similar to that observed under VA-adequate conditions. In adult VA-sufficient rats, 14% of the whole-body VA mass was extrahepatic (compared with 44% in VA-marginal and 93% in VA-deficient adult rats) (27–29). In humans, the corresponding percentage under VA-adequate conditions was ∼10% (30). The fact that neonates in our study stored only ∼13% of retinol outside of the liver suggests that the nonhepatic tissues may not be sufficiently developed to store VA at the adult capacity. Although the retinoid receptors, cellular retinoid-binding proteins, and enzymes involved in retinol storage are expressed early in life (13), they may not be rate-limiting.

The low VA content in extrahepatic tissues may be attributed to the specific characteristics of the neonatal body, particularly the scarcity of WAT. WAT, which normally constitutes 5–10% of the adult rat’s body weight (31), was nearly absent before postnatal day 12 in neonatal rats and thereafter contained only ∼1% of the whole-body VA mass, compared with 10–20% in adult rats (32, 33). BAT was present from birth, but its contribution was also low (∼0.5%), as was the concentration of retinol in the WAT and BAT of neonates (∼2 nmol/g compared with 21–25 nmol/g in adult rats) (34). Such low VA storage capacity of adipose tissue may be caused by the lower lipid and higher water content of neonatal tissues in general (25).

The effect of VA supplementation.

Supplementation with VA resulted in a pronounced 1–4-fold increase in retinol concentrations in nondigestive organs between 4 and 15 h after dose administration. These elevated concentrations returned to baseline within 24 h after dose administration in plasma and all organs except the kidney, liver, WAT, and BAT (Table 1). In kidneys, the significant elevation lasted for 2 d, most likely because of an increased rate of retinol disposal after supplementation. A 4- and 11-d long elevation was observed in BAT and in the liver, respectively. WAT was the only organ in which the supplementation effect lasted for longer than in the liver (18 d), indicating that it may serve as a long-term VA storage depot. This finding further suggests that the scarcity of WAT in neonates may predispose them to a low VA status.

TABLE 1.

Effect of supplementation on organ VA concentration in neonatal rats administered 50,000 IU VA on postnatal day 41

Organ Time of peak VA concentration, h Increase from control group concentration at peak, % P Supplementation effect duration,2 h
Plasma 1 417 0.128 0
Carcass 1 350 0.144 0
Stomach 0.5 4551 <0.001 1
Skin 4 295 <0.01 15
Intestine 1 944 <0.05 24
Lungs 15 171 <0.05 24
Brain 8 327 <0.001 24
Kidneys 15 133 <0.01 48
BAT 4 429 <0.05 96
Liver 24 302 <0.001 264
WAT3 — — — 432
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1

BAT, brown adipose tissue; VA, vitamin A; WAT, white adipose tissue.

2

Duration of significant elevation (P < 0.05) in organ VA concentration in the VA-supplemented compared with the control group as determined by Student’s t test with Bonferroni correction for multiple comparisons.

3

Data for WAT were available only after postnatal day 12.

The return of plasma VA concentration to a marginal level in the supplemented pups despite a simultaneous increase in liver VA concentration suggests that neonates may have a lower “set point” for plasma retinol, which does not necessarily indicate deficiency. The rapid (within <24 h) decline of VA concentrations in all other organs suggests that they have a relatively low VA retention capacity and use it locally or release it back to plasma for transfer to the liver. This finding agrees with the early and transient (∼24 h) peak in retinol concentration in the lungs, spleen, and adrenal gland of neonatal piglets aged 28 d administered 50,000 IU VA (35). From these results, Riabroy and Tanumihardjo (35) concluded that extrahepatic tissues in neonates rely on recently ingested chylomicron retinyl esters as the main source of VA, making a constant supply of VA in the mother’s milk or diet necessary for maintaining a steady extrahepatic VA concentration.

VA supplementation did not affect the rank order of tissues in terms of their retinol mass or concentration, except for the higher position of stomach and intestine relative to skin (retinol mass) and kidneys (retinol concentration), as is expected from the presence of the VA dose in these organs (Figure 7). This observation was similar to findings in neonatal piglets, in which the liver ranked as the highest in terms of retinol concentration, followed by the kidneys and lungs, irrespective of VA treatment (23). However, kinetic studies in rats supplemented with VARA have shown a dramatic stimulatory effect of supplementation on the uptake and retention of retinyl esters by the lungs and intestine (14, 36), consistent with the previously demonstrated role of retinoic acid in upregulating genes responsible for VA storage (37–39). Based on these results, we speculate that VA alone may not be as effective as VARA in promoting extrahepatic retinol deposition, although future kinetic analyses of our data are needed to confirm this hypothesis.

In conclusion, our study demonstrated that male and female Sprague-Dawley neonatal rats raised under VA-marginal conditions stored most of their VA in the liver despite its deficient retinol concentration, an effect that may be explained by the low VA storage capacity of extrahepatic tissues. We also showed that supplementation with VA in a dose equivalent to that given to human newborns caused a transient increase in retinol concentration in all extrahepatic organs except WAT. This may indicate that the scarcity of subcutaneous fat in neonates predisposes them to VA deficiency. These findings also suggest that a more frequent VA supplementation, along with an improved dietary intake, may be necessary to meet the needs of rapidly developing neonatal tissues. More research is needed, however, to examine the link between VA storage in adipose tissue and other extrahepatic organs and the health benefits of VA supplementation.

Acknowledgments

JKH conducted the research, analyzed the data, and wrote the manuscript; LT designed and conducted the research; MHG designed the research; and ACR designed and conducted the research and had primary responsibility for the final content. All authors read and approved the final manuscript.

Footnotes

5

Abbreviations used: BAT, brown adipose tissue; UPLC, ultra-performance liquid chromatography; VA, vitamin A; VARA, VA admixed with retinoic acid; WAT, white adipose tissue.

References

  • 1.Pfahl M, Chytil F. Regulation of metabolism by retinoic acid and its nuclear receptors. Annu Rev Nutr 1996;16:257–83. [DOI] [PubMed] [Google Scholar]
  • 2.Ross AC. Vitamin A In: Ross AC, Caballero B, Cousins RJ, Tucker KL, Ziegler TR, editors. Modern nutrition in health and disease. 11th ed Baltimore (MD): Lippincott Williams & Wilkins; 2014. p. 260–277. [Google Scholar]
  • 3.Cunningham TJ, Duester G. Mechanisms of retinoic acid signalling and its roles in organ and limb development. Nat Rev Mol Cell Biol 2015;16:110–23. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Dahro M, Gunning D, Olson JA. Variations in liver concentrations of iron and vitamin A as a function of age in young american-children dying of the sudden infant death syndrome as well as of other causes. Int J Vitam Nutr Res 1983;53:13–8. [PubMed] [Google Scholar]
  • 5.Berggren Söderlund M, Fex GA, Nilsson-Ehle P. Concentrations of retinoids in early pregnancy and in newborns and their mothers. Am J Clin Nutr 2005;81:633–6. [DOI] [PubMed] [Google Scholar]
  • 6.Sommer A. Vitamin A deficiency and clinical disease: an historical overview. J Nutr 2008;138:1835–9. [DOI] [PubMed] [Google Scholar]
  • 7.WHO. Neonatal vitamin A supplementation. Geneva (Switzerland): WHO; 2011. [Google Scholar]
  • 8.WHO. Technical consultation on neonatal vitamin A supplementation research priorities: meeting report. Geneva (Switzerland): WHO; 2009. [Google Scholar]
  • 9.Fawzi WW, Chalmers TC, Herrera MG, Mosteller F. Vitamin A supplementation and child mortality. A meta-analysis. JAMA 1993;269:898–903. [PubMed] [Google Scholar]
  • 10.Bhutta ZA, Ahmed T, Black RE, Cousens S, Dewey K, Giugliani E, Haider BA, Kirkwood B, Morris SS, Sachdev HP, et al. What works? Interventions for maternal and child undernutrition and survival. Lancet 2008;371:417–40. [DOI] [PubMed] [Google Scholar]
  • 11.Gogia S, Sachdev HS. Neonatal vitamin A supplementation for prevention of mortality and morbidity in infancy: systematic review of randomised controlled trials. BMJ 2009;338:b919. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Imdad A, Yakoob MY, Sudfeld C, Haider BA, Black RE, Bhutta ZA. Impact of vitamin A supplementation on infant and childhood mortality. BMC Public Health 2011;11(Suppl 3):S20. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Tan L, Green MH, Ross AC. Vitamin A kinetics in neonatal rats vs. adult rats: comparisons from model-based compartmental analysis. J Nutr 2015;145:403–10. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Tan L, Wray AE, Green MH, Ross AC. Compartmental modeling of whole-body vitamin A kinetics in unsupplemented and vitamin A-retinoic acid-supplemented neonatal rats. J Lipid Res 2014;55:1738–49. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 15.Reeves PG, Nielsen FH, Fahey GC Jr. AIN-93 purified diets for laboratory rodents: final report of the American Institute of Nutrition ad hoc writing committee on the reformulation of the AIN-76A rodent diet. J Nutr 1993;123:1939–51. [DOI] [PubMed] [Google Scholar]
  • 16.Ross AC. Separation and quantitation of retinyl esters and retinol by high-performance liquid chromatography. Methods Enzymol 1986;123:68–74. [DOI] [PubMed] [Google Scholar]
  • 17.Sengupta P. The laboratory rat: relating its age with human’s. Int J Prev Med 2013;4:624–30. [PMC free article] [PubMed] [Google Scholar]
  • 18.Masters AM, Leslie AJ, Kaldor I. Red cell and plasma volume development in newborn rats measured with double label. Am J Physiol 1972;222:49–54. [DOI] [PubMed] [Google Scholar]
  • 19.Ross AC, Harrison EH. Vitamin A: nutritional aspects of retinoids and carotenoids. In: Zempleni J, Rucker RB, McCormick DB, Suttie JW, editors. Handbook of vitamins. 4th ed Boca Raton (FL): Taylor & Francis; 2007. p. 1–39. [Google Scholar]
  • 20.Underwood BA. Methods for assessment of vitamin A status. J Nutr 1990;120(Suppl 11):1459–1463. [DOI] [PubMed] [Google Scholar]
  • 21.Green MH, Green JB, Lewis KC. Model-based compartmental analysis of retinol kinetics in organs of rats at different levels of vitamin A status. In: Livrea MA, Packer L, editors. Retinoids: progress in research and clinical applications. 1st ed New York: Marcel Dekker; 1993. p. 1535–1548. [Google Scholar]
  • 22.Reinersdorff DV, Bush E, Liberato DJ. Plasma kinetics of vitamin A in humans after a single oral dose of [8,9,19–13C]retinyl palmitate. J Lipid Res 1996;37:1875–85. [PubMed] [Google Scholar]
  • 23.Sun T, Surles RL, Tanumihardjo SA. Vitamin A concentrations in piglet extrahepatic tissues respond differently ten days after vitamin A treatment. J Nutr 2008;138:1101–6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Biesalski HK, Nohr D. Importance of vitamin-A for lungs function and development. Mol Aspects Med 2003;24:431–40. [DOI] [PubMed] [Google Scholar]
  • 25.WHO. Principles for evaluation health risks in children associated with exposure to chemicals. Geneva (Switzerland): WHO; 2006. [Google Scholar]
  • 26.De Luca LM, Roop D, Huang FL. Vitamin A: a key nutrient for the maintenance of epithelial differentiation. Acta Vitaminol Enzymol 1985;7(Suppl):13–20. [PubMed] [Google Scholar]
  • 27.Green MH, Uhl L, Green JB. A multicompartmental model of vitamin A kinetics in rats with marginal liver vitamin A stores. J Lipid Res 1985;26:806–18. [PubMed] [Google Scholar]
  • 28.Lewis KC, Green MH, Green JB, Zech LA. Retinol metabolism in rats with low vitamin A status: a compartmental model. J Lipid Res 1990;31:1535–48. [PubMed] [Google Scholar]
  • 29.Green MH, Green JB. Vitamin A intake and status influence retinol balance, utilization and dynamics in rats. J Nutr 1994;124:2477–85. [DOI] [PubMed] [Google Scholar]
  • 30.Cifelli CJ, Gree JB, Wang Z, Yin S, Russell RM, Tang G, Green MH. Kinetic analysis shows that vitamin A disposal rate in humans is positively correlated with vitamin A stores. J Nutr 2008;138:971–7. [DOI] [PubMed] [Google Scholar]
  • 31.Wronska A, Sledzinski T, Goyke E, Lawniczak A, Wierzbicki P, Kmiec Z. Short-term calorie restriction and refeeding differently affect lipogenic enzymes in major white adipose tissue depots of young and old rats. J Physiol Pharmacol 2014;65:117–26. [PubMed] [Google Scholar]
  • 32.Ross R, Léger L, Guardo R, De Guise J, Pike BG. Adipose tissue volume measured by magnetic resonance imaging and computerized tomography in rats. J Appl Physiol 1991;70:2164–72. [DOI] [PubMed] [Google Scholar]
  • 33.D’Ambrosio DN, Clugston RD, Blaner WS. Vitamin A metabolism: an update. Nutrients 2011;3:63–103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 34.Tsutsumi C, Okuno M, Tannous L, Piantedosi R, Allan M, Goodman DS, Blaner WS. Retinoids and retinoid-binding protein expression in rat adipocytes. J Biol Chem 1992;267:1805–10. [PubMed] [Google Scholar]
  • 35.Riabroy N, Tanumihardjo SA. Oral doses of alpha-retinyl ester track chylomicron uptake and distribution of vitamin A in a male piglet model for newborn infants. J Nutr 2014;144:1188–95. [DOI] [PubMed] [Google Scholar]
  • 36.Ross AC, Ambalavanan N, Zolfaghari R, Li NQ. Vitamin A combined with retinoic acid increases retinol uptake and lung retinyl ester formation in a synergistic manner in neonatal rats. J Lipid Res 2006;47:1844–51. [DOI] [PubMed] [Google Scholar]
  • 37.Senoo H, Yoshikawa K, Morii M, Miura M, Imai K, Mezaki Y. Hepatic stellate cell (vitamin A-storing cell) and its relative - past, present and future. Cell Biol Int 2010;34:1247–72. [DOI] [PubMed] [Google Scholar]
  • 38.Ross AC. Retinoid production and catabolism: role of diet in regulating retinol esterification and retinoic acid oxidation. J Nutr 2003;133:291S–6S. [DOI] [PubMed] [Google Scholar]
  • 39.Dirami G, Massaro GD, Clerch LB, Ryan US, Reczek PR, Massaro D. Lung retinol storing cells synthesize and secrete retinoic acid, an inducer of alveolus formation. Am J Physiol Lung Cell Mol Physiol 2004;286:L249–56. [DOI] [PubMed] [Google Scholar]

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