α-Galactosylceramide stimulates splenic lymphocyte proliferation in vitro and increases antibody production in vivo in late neonatal-age mice.

The neonatal stage is characterized by weak responses to various infections and vaccines, thus the development of efficient formulas to improve vaccine effectiveness is of high priority. The glycolipid alpha galactosylceramide (αGalCer) is known as a potent immune modulator due mainly to natural killer (NK) T cell activation. Using a mouse tetanus toxoid (TT) immunization model, we observed that neonatal mice given αGalCer at the time of primary immunization on postnatal day (pnd) 17 had a significantly higher TT-specific immunoglobulin (Ig)M response as well as a memory IgG response, while αGalCer given on pnd 7 resulted in only marginal boosting. Consistently, immunostaining of the spleen sections from αGalCer-treated pnd 17 immunized neonates showed a higher number of Ki67(+) cells in the splenic germinal centre area, suggesting a stronger response after immunization. In-vitro kinetic studies revealed that spleen cells from newborn to pnd 7 neonates did not respond to αGalCer stimulation, whereas cell proliferation was increased markedly by αGalCer after pnd 7, and became dramatic around neonatal pnd 17-18, which was accompanied by increased B, T and NK T cell populations in the spleen. In addition, in pnd 17 spleen cells, αGalCer significantly stimulated the production of NK T cytokines, interleukin (IL)-4 and interferon (IFN)-γ, and promoted the proliferation of CD23(+) B cells, a subset of B cells enriched in germinal centres. These data suggest that αGalCer is an effective immune stimulus in the late neonatal stage, and thus may be useful in translational studies to test as a potential adjuvant to achieve a more efficient response to immunization.

Retinoic acid and tumor necrosis factor-α induced monocytic cell gene expression is regulated in part by induction of transcription factor MafB.

All-trans-retinoic acid (RA), the major active metabolite of vitamin A, is a regulator of gene expression with many roles in cell differentiation. In the present study, we investigated RA in the regulation of MafB, a basic leucine-zipper transcription factor with broad roles in embryonic development, hematopoiesis and monocyte-macrophage differentiation. In RA-treated THP-1 human monocytic cells, MafB mRNA and protein levels were up-regulated by RA dose and time-dependently, while, additionally, RA and tumor necrosis factor (TNF)α, also known to induce monocyte to macrophage differentiation, increased MafB expression synergistically. Screening of potential targets containing Maf recognition elements (MARE motifs) in their promoter regions identified SPOCK1, Blimp1 and CCL2 as potential targets; these genes are related to cell communication, recruitment and differentiation, respectively. Across cell treatments, SPOCK1, Blimp1 and CCL2 mRNA levels were highly correlated (P<0.001) with MafB. ChIP assays demonstrated increased MafB protein binding to MARE elements in the promoter regions of SPOCK1, Blimp1 and CCL2 in RA and TNFα-treated cells, as well as acetylation of histone-H4 in MARE-containing regions, indicative of chromatin activation. Conversely, reducing MafB protein by microRNA silencing significantly decreased the expression of SPOCK1, Blimp1 and CCL2 (P<0.01). Moreover, the reduction in MafB expression and these downstream targets correlated with decreased cell differentiation as determined by cell-surface CD11b expression and phagocytic activity. We conclude that MafB may be a key factor in mediating the ability of RA and TNFα to regulate monocytic cell communication, recruitment and differentiation through regulation of MafB target genes including SPOCK1, CCL2 and Blimp1.

Effects of in utero antiretroviral exposure on mitochondrial DNA levels, mitochondrial function and oxidative stress.

OBJECTIVES: HIV and antiretroviral (ART) exposure in utero may have deleterious effects on the infant, but uncertainty still exists. The objective of this study was to evaluate aspects of mitochondrial DNA (mtDNA) content, mitochondrial function and oxidative stress simultaneously in placenta, umbilical cord blood and infant blood in HIV/ART-exposed infants compared with uninfected controls. METHODS: HIV-1-infected pregnant women and HIV-1-uninfected healthy pregnant controls were enrolled in the study prospectively. Placenta and umbilical cord blood were obtained at delivery and infant blood was obtained within 48 h of delivery. mtDNA content was determined for each specimen. Nuclear [subunit IV of cytochrome c-oxidase (COX IV)]- and mitochondrial (COX II)-encoded polypeptides of the oxidative phosphorylation enzyme cytochrome c-oxidase were quantified in cord and infant blood. Placental mitochondria malondialdehyde (MDA) concentrations were measured as a marker of oxidative stress. RESULTS: Twenty HIV-positive/HIV-exposed and 26 control mother-infant pairs were enrolled in the study. All HIV-infected women and their infants received ART. Placental MDA concentration and mtDNA content in placenta and cord blood were similar between groups. The cord blood COX II:IV ratio was lower in the HIV-positive group than in the controls, whereas the infant peripheral blood mtDNA content was higher in the HIV-exposed infants, but the infant peripheral blood COX II:IV ratio was similar. No infant had clinical evidence of mitochondrial disease or acquired HIV infection. In multivariable regression analyses, the significant findings in cord and infant blood were both most associated with HIV/ART exposure. CONCLUSIONS: HIV-exposed infants showed reduced umbilical cord blood mitochondrial enzyme expression with increased infant peripheral blood mitochondrial DNA levels, the latter possibly reflecting a compensatory mechanism to overcome HIV/ART-associated mitochondrial toxicity.

Effect of fatty acid supplementation on cholesterol and retinol esterification in J774 macrophages.

J774 macrophages exposed to medium containing cholesterol-rich phospholipid dispersions accumulate cholesteryl ester. Supplementing this medium with 100 micrograms oleate/ml increased cellular cholesteryl ester contents 3-fold. Cell retinyl ester contents increased 8-fold when medium containing retinol dispersed in dimethyl sulfoxide was supplemented with oleate. These increases were not the result of increases in total lipid uptake by the cells but rather of redistribution of cholesterol and retinol into their respective ester pools. Effective oleate concentration of 15-30 micrograms/ml increased cellular retinyl and cholesteryl ester contents. The effective oleate concentration was reduced to 5 micrograms/ml when the fatty acid/albumin molar ratio was increased. The oleate-stimulated increase in cholesterol esterification was blocked by incubating cells with Sandoz 58-035, a specific inhibitor of acyl-CoA:cholesterol acyltransferase (ACAT), indicating that the effect of fatty acid exposure is mediated through changes in ACAT activity. When cholesterol or retinol was added to cells which had been exposed to oleate for 24 h to provide a triacylglycerol store, the cellular contents of cholesteryl or retinyl ester were also significantly increased compared to cells not previously exposed to oleate. The oleate-stimulated increase in the esterification of cholesterol and/or retinol was also observed in P388D1 macrophages, human (HepG2) and rat (Fu5AH) hepatomas, human fibroblasts, rabbit aortic smooth muscle cells and MCF-7 breast carcinoma cells. In addition to oleate, a number of other fatty acids increased retinol esterification in J774 macrophages; however, cellular cholesterol esterification in these cells was increased only by unsaturated fatty acids and was inhibited in the presence of saturated fatty acids. Although the cellular uptake of radiolabeled oleate and palmitate was similar, a significant difference in the distribution of these fatty acids among the lipid classes was observed. These data demonstrate that exogenous fatty acids are one factor that regulate cellular cholesteryl and retinyl ester contents in cultured cells.

Characterization of a bile salt-dependent cholesteryl ester hydrolase activity secreted from HepG2 cells.

HepG2 cells and medium were assayed for cholesteryl ester hydrolase (CEH) activity in the presence and absence of sodium cholate. Although bile salt-dependent CEH activity was measured in the medium at 6 to 96 h (up to 4500 pmol/h per mg cell protein), there was very little activity detected in the corresponding cell homogenates (less than 70 pmol/h per mg cell protein). Activity in the medium was expressed only in the presence of trihydroxy bile salts and was maximal at 40 mM cholate and pH 7.5. Incubation of HepG2 cells with brefeldin A resulted in an 80 to 90% inhibition of secretion of the bile salt-dependent CEH activity, while only inhibiting total protein secretion by 42%. Bile salt-dependent CEH activity could also be detected in rat liver perfusates. Although there was measurable activity in all of 14 livers analyzed (47 +/- 10 and 53 +/- 17 nmol/h per g liver per h perfusion during two 5-min collections after 15 and 30 min of perfusion, respectively), it did not correlate with the activity found in corresponding liver homogenates, as only four livers had detectable bile salt-dependent CEH activity. These results provide evidence for the secretion of a bile salt-dependent CEH activity, from both a hepatic cell line and the intact liver, that has similar properties to the enzyme previously isolated from rat liver homogenates and rat pancreas.

Potentiation of interferon-gamma-stimulated nitric oxide production by retinoic acid in RAW 264.7 cells.

Nitric oxide (NO) production is essential for normal immunity. We have examined the capacity of retinoic acid (RA), a pleiotropic hormone necessary for normal immunity, to modulate NO production in RAW 264.7 cells. NO production induced by suboptimal concentrations of interferon-gamma (IFN-gamma) was significantly greater in cells cultured in low-retinoid medium and treated with all-trans-RA (10(-10) - 10(-6) M, P <0.05), as well as with 9-cis-RA and several retinoids selective for the RA receptor subfamily of nuclear retinoid receptors. Similar results were obtained with lipopolysaccharide and monophosphoryl lipid A as stimuli. The RA-potentiated production of NO was positively correlated with inducible NO synthase (iNOS) protein (r =0.94, P <0.002), although the expression of iNOS mRNA was not altered. We hypothesize that modulation of the macrophage response to suboptimal immune stimuli by physiological concentrations of RA, as observed in these studies, may be important in establishing an optimal balance between T helper (Th) 1- and Th2-mediated immunity.

Regulation of thyroid-stimulating hormone beta-subunit and growth hormone messenger ribonucleic acid levels in the rat: effect of vitamin A status.

T3 inhibits transcription of the rat TSH beta gene, and two T3 response elements have been identified that bind T3 receptors and that share sequence homology with the consensus sequence that is also recognized by retinoic acid receptors (RARs). We, therefore, asked whether RA was a regulator of TSH beta gene expression in vivo. Using RNase protection analysis, we found that vitamin A deficiency led to a 2-fold increase in rat pituitary TSH beta messenger RNA (mRNA) levels, which returned to normal 18 h after treatment with RA (20 micrograms/rat). Vitamin A deficiency had no effect on TSH beta mRNA levels in hypothyroid rats. Coadministration of RA and T3 (10 micrograms/100g body wt) to either vitamin A-deficient or vitamin A-deficient, hypothyroid animals caused decreases in TSH beta mRNA content that were indistinguishable from those seen with T3 alone. Surprisingly, vitamin A deficiency had no significant effect on GH mRNA levels in euthyroid or hypothyroid rats. Furthermore, treatment of vitamin A-deficient, hypothyroid animals with RA for either 18 or 72 h had no effect on GH mRNA levels, whereas T3 caused 11-fold and 18-fold increases in GH mRNA, respectively, at these times. We also used transient transfection to test for direct, retinoid receptor-mediated regulation of TSH beta gene transcription by RA. A plasmid TSH beta luciferase, containing 0.8 kilobases of rat TSH beta gene 5'-flanking sequences, exon 1, and 150 base pairs of intron 1, was transfected into CV-1 cells. Cotransfection with RAR alpha and retinoid X receptor-beta induced TSH beta expression by 3.5-fold, and treatment with RA suppressed this induction by 46%. These results show that vitamin A levels play a significant role in regulating the expression of the TSH beta gene, but not the GH gene, in vivo and suggest that RA may suppress TSH beta gene transcription directly by an RAR-retinoid X receptor heterodimer-mediated mechanism.

Cell-type specific interactions between retinoic acid and thyroid hormone in the regulation of expression of the gene encoding ornithine aminotransferase.

The purposes of this study were to determine whether expression of the gene encoding ornithine aminotransferase (OAT) in the rat liver and kidney is regulated by retinoic acid (RA) and to characterize further the role of thyroid hormone in regulating the expression of this gene. The level of OAT messenger RNA (mRNA) was reduced 70% in the liver of animals fed a vitamin A-deficient diet relative to that in animals fed a vitamin A-sufficient diet. RA, administered at a dose of 20 micrograms/rat to A-deficient rats for 1 or 3 days, restored OAT mRNA to near the level observed in animals fed the A-sufficient diet. Retinol was also effective in this regard. T3, when injected alone at a dose of 10 micrograms/100 g BW, had no effect on the level of OAT mRNA in the liver. However, when injected concurrently with RA, T3 blocked the ability of RA to induce OAT mRNA in the liver of rats fed the vitamin A-deficient diet. Animals made both vitamin A deficient and hypothyroid responded to RA in a manner similar to vitamin A-deficient animals. The vitamin A-deficient, hypothyroid rats responded somewhat differently to T3, however. T3 was unable to block the induction of OAT mRNA in the liver of vitamin A-deficient, hypothyroid rats when injected concurrently with RA for 1 day, but did block the induction of OAT mRNA by RA when these two hormones were injected concurrently for 3 days. These data indicate that RA and T3 exert opposing effects on the level of OAT mRNA in the liver. The effects of RA and T3 on OAT mRNA were markedly different in the kidney. Neither vitamin A deficiency nor RA had any apparent affect on the level of OAT mRNA in the kidney. T3, however, increased the level of OAT mRNA in the kidney of vitamin A-deficient rats. In the kidney of vitamin A-deficient, hypothyroid rats, T3 was unable to increase OAT mRNA when injected for 1 day, but did increase this mRNA when injected for 3 days. Together, these data indicate cell-type specific effects of both RA and T3 on the OAT gene.

Antibody response against tetanus toxoid is enhanced by lipopolysaccharide or tumor necrosis factor-alpha in vitamin A-sufficient and deficient rats.

Reduced antibody response to tetanus toxoid (TT) was previously demonstrated during vitamin A deficiency but the response to bacterial lipopolysaccharide (LPS) was normal. We addressed whether anti-TT IgG responses are enhanced in vitamin A-sufficient and deficient rats by immunization with LPS plus TT. Antibody responses in vitamin A-sufficient and deficient rats increased significantly after coimmunization (LPS + TT) compared with the response of rats immunized with TT alone. In additional studies, recombinant tumor necrosis factor-alpha (TNF-alpha) also significantly increased the anti-TT IgG concentrations. Because pretreatment with anti-TNF before coimmunization or immunization with TT alone markedly reduced the anti-TT IgG responses, we infer TNF to be a mediator of both the adjuvanticity of LPS and the unstimulated response to TT. In conclusion, vitamin A-deficient rats can be stimulated to respond to TT by coimmunization with LPS or by treatment with TNF.

Immunologic memory is established in nursling rats immunized with tetanus toxoid, but is not affected by concurrent supplementation with vitamin A.

Experiments were conducted to determine whether nursling rats immunized with tetanus toxoid (TT) are able to produce a specific antibody response and whether oral treatment with retinyl palmitate, concurrent with immunization, affects the magnitude of the anti-TT response. When rats aged 8-15 d and nursed by vitamin A-sufficient dams were immunized with TT, no primary anti-TT immunoglobulin (Ig) M or IgG response was detected. However, nursling rats formed immunologic memory to TT because, when they were reimmunized at 40 d of age, their secondary anti-TT IgG response exceeded the primary response of 40-d-old vitamin A-sufficient rats (P < 0.02). Provision of retinyl palmitate (equal to 37.5 or 150 micrograms retinol equivalents) by mouth with early primary immunization did not change the magnitude of the secondary anti-TT IgG response. However, the age of nursling rats at first immunization significantly affected the magnitude of their secondary anti-TT IgG response, because rats first immunized at 15 d of age and reimmunized at 40 d of age produced a secondary response that was nearly fivefold greater than that of rats immunized at 8 and 40 d of age. In conclusion, nursling rats immunized with TT formed immunologic memory, which was affected significantly by the timing of the primary immunization. However, the administration of retinyl palmitate concurrent with early primary immunization did not significantly affect the development of memory to TT.

Effects of timing and dose of vitamin A on tissue retinol concentrations and antibody production in the previously vitamin A-depleted rats.

The effect of retinol repletion in previously vitamin A-depleted Lewis rats on antibody production to pneumococcal polysaccharide (SSS-III) was studied. When vitamin A-depleted rats were given either 0.35 mumol (0.1 mg) or 5.2 mumol (1.5 mg) retinol, plasma retinol became normal within 8 h. Liver and lymphoid-organ retinol concentrations were normalized by 1 d after repletion with 5.2 mumol but not 0.35 mumol retinol. Antibody production to SSS-III was compared after administering 5.2 mumol retinol either as a divided dose (half given 4 d before and half given on the day of immunization) or as a single dose concurrent with immunization. Vitamin A-depleted rats produced very little SSS-III-specific antibody. The divided dose of retinol consistently restored anti-SSS-III production whereas the single concurrent dose was less effective despite equal effects on tissue retinol concentrations. Interestingly, normalization of plasma retinol was not always a good predictor of the immune response to pneumococcal polysaccharide.

Vitamin A depletion and repletion: effects on antibody response to the capsular polysaccharide of Streptococcus pneumoniae, type III (SSS-III).

To determine the ability of young rats to produce antibody against pneumococcal polysaccharide (SSS-III), weaning rats were fed a semipurified diet containing no retinol (A-) or 4 micrograms retinol/g diet (A+). Splenic antibody response specific to SSS-III was 17% (p less than or equal to 0.05) of control for A- Sprague-Dawley rats; similarly, the response of retinol-depleted Lewis rats was 22% (p less than or equal to 0.05) of pair-fed controls. No kinetic differences were observed in the antibody response between A- and control Lewis rats. Retinol depletion more markedly reduced the antibody response of male rats than female rats despite equally low tissue retinol concentrations. For both strains, retinol repletion near the time of immunization normalized antibody production. When male Lewis rats were fed the A- diet longer, the antibody response of A- rats was only 3% of pair-fed controls; repletion again normalized antibody production. Thus, retinol supplementation near the time of immunization can restore the immune response in previously compromised A- rats.

Relation of serum retinol to acute phase proteins and malarial morbidity in Papua New Guinea children.

BACKGROUND: Acute phase proteins (APPs) are associated with malaria-induced hyporetinemia (serum retinol <0.70 micromol/L); however, the degree of the association is not well documented. OBJECTIVE: The association between malaria-induced hyporetinemia and APPs was assessed. DESIGN: In a cross-sectional study, 90 children with serum retinol concentrations from <0.35 to >1.05 micromol/L were selected from children in a clinical trial of vitamin A supplementation. Serum was collected before treatment allocation. Retinol binding protein (RBP) concentrations were determined by radioimmunoassays, and transthyretin, alpha(1)-acid glycoprotein (AGP), alpha(1)-antichymotrypsin, C-reactive protein (CRP), haptoglobin, and albumin concentrations by radial immunodiffusion assays. RESULTS: Children in the subsample had high rates of splenomegaly and Plasmodium-positive blood-smear slides (P < 0.01); AGP (Pearson's r = -0.40, P < 0.001) and CRP (r = -0.21, P = 0.04) were inversely correlated with retinol. The negative APPs RBP, transthyretin, and albumin were positively and significantly associated with retinol. All APPs, except alpha(1)-antichymotrypsin, were significantly correlated with splenomegaly. Of the positive APPs, AGP correlated with CRP (r = 0.37, P < 0.001), indicating chronic inflammation. In a stepwise regression analysis, 73% of retinol's variability was explained by RBP and transthyretin. The model predicted that a 1-SD increase in RBP or transthyretin increases retinol by approximately 0.38 or 0.47 micromol/L, respectively, whereas an equivalent increase in AGP decreases retinol by 0.12 micromol/L. CONCLUSIONS: The RBP-transthyretin transport complex of retinol is not altered by inflammation. Positive APPs are useful markers of type and severity of inflammation; however, except for AGP, it is unlikely that they can correct for malaria-induced hyporetinemia.

An analysis of breast cancer surgery after free transverse rectus abdominis myocutaneous (TRAM) flap reconstruction.

BACKGROUND: Breast reconstruction is currently offered on a more routine basis to patients after mastectomy for breast cancer. This paper analyzes the outcomes of breast cancer surgery, and the results and effects of breast reconstruction using free TRAM flaps. METHODS: A retrospective review of 75 consecutive patients who had free transverse rectus abdominis myocutaneous (TRAM) flap breast reconstruction after breast cancer surgery was performed. A total of 92 free TRAM flaps were performed on 75 patients in Victoria, British Columbia, from January 1992 to May 1999. Thirty-three patients (44%) underwent primary breast cancer surgery and an immediate reconstruction (7 bilateral and 27 unilateral) and 42 patients (56%) had delayed reconstruction (10 bilateral and 32 unilateral). RESULTS: Twenty- one patients (28%) had stage 0 disease, 20 (26.7%) had stage I disease, 17 (22.7%) had stage IIA disease, 12 (15%) had stage IIB disease, and 4 (5.3%) had stage IIIA disease. In 1 patient the stage of disease was unknown. The mean patient age was 49.4 years (range 33 to 73). Of the patients undergoing immediate reconstruction 3 had postoperative chemotherapy and 1 had postoperative radiotherapy. Three patients had combined chemoradiotherapy. In none of these cases was the adjuvant therapy delayed by the reconstructive surgery. Overall mean follow-up time from cancer diagnosis was 56.8 months and from the time of TRAM flap reconstruction, 36.7 months. To date, 5 recurrences have been detected (6.6%). Mean time between reconstruction and detection of recurrence was 22.8 months. Detection of recurrence was achieved clinically and was not impaired in any of the cases by the presence of the free flap. Patient satisfaction was assessed via a telephone survey, with 93% of patients pleased with the cosmetic results of their surgery. CONCLUSIONS: For those patients with breast cancer requiring mastectomy, free TRAM flap reconstruction is a safe, cosmetically acceptable surgical alternative that impairs neither effective breast cancer surgery nor detection of recurrent disease.

Vitamin A as a hormone: recent advances in understanding the actions of retinol, retinoic acid, and beta carotene.

Within the past few years, much has been learned about the metabolism and actions of vitamin A and the carotenoids. This article reviews the biochemical and cellular events in retinoid metabolism that lead to production of retinoic acid, an active metabolite of vitamin A. Retinoic acid functions in a hormone-like manner to regulate the expression of a number of genes. Beta carotene is now under study as an anticancer agent and for its possible beneficial effects in a number of chronic diseases. Current recommendations for carotene intake exceed the usual daily intake nearly fourfold.

Scoliosis in juvenile chronic arthritis.

Structural scoliosis occurs more commonly in patients with juvenile chronic arthritis than in the normal population. We have reviewed 32 patients with both juvenile arthritis and a scoliosis and suggest that structural curves may arise from postural curves associated with asymmetrical involvement of lower limb joints.

Endoprosthetic replacement of the humerus and elbow joint.

Between 1969 and 1985 26 patients with destructive lesions of the distal humerus were treated by endoprosthetic replacement; each implant was custom-made and incorporated part of the distal humerus or the entire bone as well as a hinged total elbow replacement. Recurrence occurred in three of the patients with tumours, and three prostheses were removed because of deep infection in patients with previously compound injuries of the elbow. Another three loosened without infection, but none needed revision or removal and no amputations resulted. Other complications included nerve palsies, but the only deaths were from metastases. A useful range of elbow movement, with a stable arm and good hand function, was achieved in every patient.

Endoprosthetic replacement of the proximal humerus.

Endoprosthetic replacement of the proximal humerus has been performed in our unit on 25 occasions between 1950 and 1982. The indication for surgery was destruction of the proximal half of the humerus so extensive that the only alternatives were reconstruction or amputation. Of the patients with tumours two died from metastases, and three from unrelated causes; local recurrence necessitated amputation in two patients. Minor complications were frequent, but there were no deep infections and, after 1964, no prosthesis became loose. Active shoulder movement after operation was considerably limited, but passive movement was good and function of the elbow and hand were preserved.

Chronopharmaceutical drug delivery from a pulsatile capsule device based on programmable erosion.

We report the development of a chronopharmaceutical capsule drug delivery system capable of releasing drug after pre-determined time delays. The drug formulation is sealed inside the insoluble capsule body by an erodible tablet (ET). The release time is determined by ET erosion rate and increases as the content of an insoluble excipient (dibasic calcium phosphate) and of gel-forming excipient (hydroxypropylmethylcellulose; HPMC) increases. The time-delayed release of a model drug (propranolol HCI) was investigated by dissolution testing (USP XXIII paddle method). Both composition and weight of ET influence the time of drug release. Moreover it was found that drug release was controlled by the quantity of HPMC, irrespective of lactose content within the tablet weight range 80-160 mg, when above a threshold concentration of 20% HPMC. Programmable pulsatile release has been achieved from a capsule device over a 2-12-h period, consistent with the demands of chronotherapeutic drug delivery. The time of drug release can be controlled by manipulation of tablet formulation.

The function of vitamin A in cellular growth and differentiation, and its roles during pregnancy and lactation.

Recent advances in the molecular biology of the retinoids have provided a mechanistic explanation for the observations, first made several decades ago, that vitamin A profoundly influences the differentiation of tissues throughout the body. A central concept has recently emerged, namely that retinoids seldom exist "free" in solution but, rather, are nearly always associated with specific retinoid-binding proteins. In plasma, these include RBP and the chylomicron whereas, in cells two distinct classes of retinoid-binding proteins exist: the cellular (cytoplasmic) proteins (CRBPs and CRABPs) and the nuclear receptors proteins (RARs and RXRs). Whereas the cellular retinoid-binding proteins serve as buffers and as chaperones during metabolism (Ross, 1993b), the nuclear receptors are now recognized to be the direct mediators of retinoid actions on the genome. Both the cytoplasmic and nuclear classes of retinoid-binding proteins are expressed early in development and are proposed to control the concentration of retinoic acid and the transcription of retinoid-responsive genes, respectively. Given the profound effects of retinoic deficiency or excess on the developing fetus, it is not surprising that mechanisms have evolved to control the placental transfer of vitamin A. Transfer is nearly uniform over a rather wide range of maternal dietary vitamin A intake. The importance of RBP in transporting retinol to tissues is suggested by the observations that the visceral yolk sac and the liver of the fetus transcribe and translate RBP. In comparison to pregnancy, vitamin A transport during lactation is much more responsive to variations in maternal vitamin A intake. The young of mothers with good vitamin A nutriture may thus accumulate significant retinol reserves during the suckling period. Conversely, young nursed by mothers with poor vitamin A status and low intake during lactation may fail to develop adequate stores and be vulnerable to vitamin A deficiency if the post-weaning diet is also poor in vitamin A. In populations with low vitamin A status, the lactation period provides an excellent window of opportunity for supplementing mothers and, indirectly, their offspring, with vitamin A to replenish the mother's vitamin A reserves and assure that the infant's growth and development are not limited by an inadequate quantity of this essential nutrient.