An Updated Comprehensive Review on Vitamin A and Carotenoids in Breast Cancer: Mechanisms, Genetics, Assessment, Current Evidence, and Future Clinical Implications.

Vitamin A and carotenoids are fat-soluble micronutrients that play important role as powerful antioxidants modulating oxidative stress and cancer development. Breast cancer is the most common malignancy in women. As the risk of breast cancer is dependent on various lifestyle factors such as dietary modifications, there is increasing interest surrounding the anti-cancerous properties of vitamin A and carotenoids. Despite the suggested protective roles of vitamin A and carotenoids in breast cancer development, their clinical application for the prevention and treatment of breast cancer is limited. In this narrative review, we discuss the roles of vitamin A and carotenoids along with the evaluation method of vitamin A status. We also exhibit the association of genetic variations involved in metabolism of vitamin A and carotenoids with cancers and other diseases. We demonstrate the epidemiological evidence for the relationship of vitamin A and carotenoids with breast cancer risk, their effects on cancer mechanism, and the recent updates in clinical practice of vitamin A or carotenoids as a potential therapeutic agent against breast cancer. This review provides insight into the preventive and therapeutic roles of vitamin A and carotenoids in breast cancer development and progression.

Comparative assessment of immunomodulatory, proliferative, and antioxidant activities of crocin and crocetin on mesenchymal stem cells.

Saffron (Crocus sativus L) is a well-known spice with active pharmacologic components including crocin, crocetin, safranal, and picrocrocin. Similar to crocin/crocetin, mesenchymal stem cells (MSCs) have been shown to display immunomodulatory and antioxidant properties, which could be beneficial in treatment of various diseases. In the current study, we have evaluated the effects of crocin and crocetin on the functions of MSCs. We used the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay to evaluate MSCs proliferation, and flow cytometry assay to measure the percentage of apoptotic MSCs and Tregs populations. Furthermore, we used the real-time polymerase chain reaction method to quantify messenger RNA (mRNA) expression of inflammatory and anti-inflammatory cytokines. Antioxidant assay was employed to quantify antioxidant parameters including nitric oxide and malondialdehyde levels besides superoxide dismutase activity. Our findings indicated that both crocin and crocetin at low concentrations (2.5 and 5 µM) exhibited significant effects on increasing MSCs viability and on protecting them against apoptosis-induced death. Furthermore, crocin and crocetin at low concentrations (2.5 and 5 µM) displayed a better antioxidant function. Moreover, increased Treg population was observed at lower doses. In addition, crocin/crocetin at low concentrations caused an elevation in mRNA expression of anti-inflammatory cytokines (transforming growth factor-ß, interleukin-10 [IL-10], and IL-4), while at higher doses (25 and 50 µM) they led to lowering inflammatory cytokines (IL-1ß, IL-6, IL-17, and interferon gamma). Altogether, both crocin and crocetin at lower concentrations exhibited more efficacies on MSCs with a better effect toward crocin. It seems that crocin and crocetin may be considered as complementary treatments for the patients who undergo MSCs transplantation.

Combination of retinyl palmitate and UV-filters: phototoxic risk assessment based on photostability and in vitro and in vivo phototoxicity assays.

This study aimed to assess the phototoxic potential of combined UV-filters and retinyl palmitate (RP) in the presence or not of bemotrizinol (BMTZ), employing photostability and in vitro and in vivo phototoxicity assays. The formulations tested contained octocrylene (OCT), octyl methoxycinnamate (OMC), benzophenone-3 (BZP-3) and RP (photostable) or octocrylene (OCT), octyl methoxycinnamate (OMC), avobenzone (AVO) and RP (less photostable). Both formulations were supplemented with bemotrizinol. Photostability was evaluated by exposing, or not, formulations spread on a glass plate to UVA/UVB irradiation. The resulting products were quantified by HPLC analysis. In vitro phototoxicity of UV-filters and combinations were evaluated using 3T3 viable monolayer fibroblast cultures submitted, or not, to irradiation according to OECD TG 432. In vivo photoallergy and photoxicity were assessed by clinical studies (photopatch test). Photostability assays showed that UV-filter bemotrizinol was a better photostabilizer for RP/benzophenone-3 than for RP/avobenzone. The in vitro phototoxicity of the combination RP/avobenzone was reduced by bemotrizinol. Clinical studies did not indicate phototoxic or photoallergenic potentials in all formulations tested. It is concluded that the 3T3 NRU phototoxicity test may be considered a supplementary assay in formulation developments, since it can detect chemically unstable and potentially phototoxic combinations. However, extrapolation of in vitro positive results to human photopatch tests may be performed only to a limited extent.

Non-invasive short-term assessment of retinoids effects on human skin in vivo using multiphoton microscopy.

BACKGROUND: The occlusive patch test developed for assessing topical retinoids activity in human skin has been extended as a short-term screening protocol for anti-ageing agents. In this model, biopsies are performed at the end of the occlusion period for morphological and immuno-histochemistry analysis. Multiphoton microscopy is a recent non-invasive imaging technique that combined with image processing tools allows the in vivo quantification of human skin modifications. OBJECTIVE: To validate with gold standards of anti-ageing that are retinoids, the relevance of multiphoton microscopy for kinetic and quantitative assessment in this model. METHODS: Twenty women, aged 50-65 years, were enrolled. Retinol 0.3% (RO) and Retinoic acid 0.025% (RA) were applied to the dorsal photo-damaged side of their forearm under occlusive patches for 12 days. A patch alone was applied to a third area as control. Evaluation was performed at day D0, D12 (end of treatment), D18 and D32 using multiphoton microscopy. Epidermal thickness, normalized area of the dermal-epidermal junction (DEJ) and melanin density were estimated using 3D image processing tools. RESULTS: Main significant results are: Epidermal thickening at D12, D18 and D32 with RO and at D12, D18 with RA vs. baseline and vs. CONTROL: Increased DEJ undulation at D32 with RO and at D12 with RA vs. baseline and vs. CONTROL: Decreased melanin content with RO (at D12 and D18 vs. baseline and at D32 vs. baseline and vs. control) and with RA (at D12 vs. baseline). CONCLUSIONS: This study shows that multiphoton microscopy associated to specific 3D image processing tools allows cutaneous effects induced by topical retinoids in this in vivo model to be non-invasively detected, quantified and followed over time. This innovative approach could be applied to the evaluation of other active compounds.

Assessment of the relative dose-response test based on serum retinol-binding protein instead of serum retinol in determining low hepatic vitamin A stores.

BACKGROUND: The relative dose-response (RDR) test, which measures the percentage of change in serum retinol concentration in response to an oral vitamin A (VA) dose, is a functional reference method to assess low hepatic VA stores. However, problems due to the relative instability of retinol, which is measured in the traditional RDR test, could be circumvented if retinol-binding protein (RBP), a more stable marker of VA, could be measured instead of retinol to provide the RDR value. OBJECTIVE: The objective was to compare classification of VA status assessed by retinol-RDR with that assessed by using RBP-RDR. DESIGN: With the use of serum samples from 57 lactating women in northern Kenya collected in August-September 2006, we assessed the accuracy of RBP-RDR in predicting low hepatic VA stores through receiver operator characteristic (ROC) analysis using retinol-RDR values as the gold standard. By using regression analysis, we explored the effects of 1) body mass index (BMI) on RBP-RDR performance and 2) the oral VA dose on the retinol-RBP molar ratio. RESULTS: The classificatory accuracy of RBP-RDR was low to moderate (n = 50; ROC area: 0.56-0.72) depending on the cutoffs used. RBP-RDR systematically overestimated VA deficiency with higher BMI, although it was superior to a single measurement of serum retinol. The discrepancy between RBP-RDR and retinol-RDR appears to originate in a retinol concentration-dependent alteration of the retinol-RBP molar ratio triggered by the oral dose. CONCLUSIONS: RBP-RDR has the potential to serve as a moderately accurate surrogate measure of retinol-RDR if the variation associated with BMI is understood and adjusted. Further studies should clarify the dynamics of the retinol-RBP molar ratio and its link to RBP-RDR performance.

Effects of micronutrients on growth of children under 5 y of age: meta-analyses of single and multiple nutrient interventions.

BACKGROUND: Micronutrient interventions have received much attention as a cost-effective and promising strategy to improve child health, but their roles in improving child growth remain unclear. OBJECTIVE: Meta-analyses of randomized controlled trials were conducted to evaluate the effect of micronutrient interventions on the growth of children aged <5 y old. DESIGN: Eligible studies were identified by PubMed database searches and other methods. Weighted mean effect sizes and 95% CIs were calculated for changes in height, weight, and weight-for-height z scores (WHZ) by using random-effect models. Tests for publication bias were done by using funnel plots, heterogeneity, and stratified analyses by predefined characteristics. RESULTS: Interventions including iron (n = 27) or vitamin A (n = 17) only had no significant effects on growth. Interventions including zinc only (n = 43) had a small positive effect (effect size = 0.06; 95% CI: 0.006, 0.11) on change in WHZ but no significant effect on height or weight gain. Multiple micronutrient interventions (n = 20) improved linear growth (0.09; 95% CI: 0.008, 0.17). CONCLUSIONS: Our findings confirm earlier results of no benefits for interventions including iron and vitamin A only but differ from the earlier meta-analysis that found improvements in linear growth for zinc only interventions. This may be due to the improved nutritional status of children in the more recent studies. Multiple micronutrient interventions improve linear growth, but the benefits are small. Other strategies are needed to prevent stunting.

Biomarker and animal models for assessment of retinoid efficacy in cancer chemoprevention.

Vitamin A is essential for normal growth and development. Epidemiology and laboratory studies suggest that decreased vitamin A levels and defective metabolism/ action may contribute to the genesis of certain cancers. Based on this information, natural and synthetic derivatives of vitamin A (retinoids) have been used for chemoprevention of cancer. Retinoids have had some success in the chemoprevention of leukoplakia and in the decreased incidence of second primaries in head and neck cancer. There is little information on biomarkers that can be used to assess the efficacy of the chemopreventive activity of retinoids. The ability of retinoids to induce RARb has been consistently shown to correlate with the response of cells and tissues to retinoic acid, but few other biomarkers have been certified as indicators of retinoid activity. In light of the failure of the ATBC and CARET clinical intervention trials for chemoprevention of lung cancer, greater use of animal models for chemoprevention studies is necessary. The potential combination of phytochemicals that inhibit DNA methyltransferase activity with retinoids holds promise for more effective chemoprevention of retinoid-unresponsive premalignant lesions.

Assessment of antioxidant effect of 2,5-dihydroxybenzoic acid and vitamin a in brains of rats with induced hyperoxia.

The aim of this study was to evaluate the effect of 2,5-dihydroxybenzoic acid, a salicylate derived from Acetyl salicylic acid (ASA) and vitamin A (vit A) on Na(+), K(+) ATPase enzyme and GSH levels in brain of rats exposed to hyperoxia (Hyp) as oxidant protocol. Rats were treated as follow: group I (control), group II (Hyp), group III (Hyp, ASA), group IV (vit A), group V (Hyp, vit A), group VI (Hyp, vit A, ASA). Vit A was given 5 days before and during Hyp, aspirin at the end of Hyp. Na(+),K(+) ATPase and total ATPase activity was significantly increased in group V. Levels of GSH showed a significant increase in group III, besides, levels of 2,5-dihydroxybenzoic acid as salicylate in plasma were significantly increased in group II. These results elucidate differences in the biochemical response of animal towards intake of various types of antioxidant substances, with increased GSH and salicylate in hyperoxia.

A critical assessment of the upper intake levels for infants and children.

The dietary reference intake (DRI) committees of the Food and Nutrition Board of the Institute of Medicine were charged with establishing DRIs for all of the essential nutrients, including fat and energy. These reference values are quantitative estimates of the nutrient intakes to be used to plan and assess the diets of healthy people. The reference values include both recommended intakes and tolerable upper intake levels (ULs). The ULs are defined as "the highest level of daily nutrient intake that is likely to pose no risk of adverse health effects to almost all individuals in the general population." As intake increases above the UL, the risk of adverse effects increases. The term "tolerable intake" was chosen to avoid the implication of a possible beneficial effect. Instead, the term is intended to connote a level of intake that can, with a high probability, be tolerated biologically. Yet, with intakes above the ULs, the risk of adverse health effects increases. Thus, there is no obvious benefit for healthy individuals to consume quantities of nutrients above the UL.

Assessment of antioxidative activity of lipid- and water-soluble vitamins in human whole blood. Comparative analysis between a biological test and chemical methods.

Assessment of the antioxidant activity of vitamins and other compounds is of interest in the understanding of their in vivo effects. In this study, we have investigated the activity of several lipid and water-soluble vitamins in human whole blood. Measurements were carried out using a biological test that enables the evaluation of both red blood cells and plasma resistance against free radical activity induced by 2,2'-azobis (2-amidinopropane)hydrochloride (AAPH). Antioxidant activity of vitamins has been determined by using the biological test versus chemical methods (chemiluminescence, DMPD radical). We have observed strong anti-oxidant potentials for vitamins B6 and B9 with biological tests, but not with chemical methods. At 10 microM, the vitamin B9 efficiency in inhibiting radical-induced red blood cell hemolysis was almost three times higher than vitamin C efficiency and two times higher than alpha-tocopherol efficiency. Antioxidant activity was not observed for vitamins B1 or B2, nor for retinol. The weak activity of beta-carotene still remains to be investigated particularly in relation to oxygen pressure. Our study demonstrated that the biological test is more useful than the chemical methods employed in this instance, for the evaluation of antioxidant capacity of lipophilic and putatively biologically active compounds.

Effect of low-dosage vitamin A and riboflavin on iron-folate supplementation in anaemic pregnant women.

A double-blind, placebo, controlled trial was conducted in Banyudono subdistrict, Boyolali regency, Central Java province, Indonesia. The aim of the study was to determine whether adding low-dosage vitamin A and riboflavin can enhance the effect of iron-folate supplementation in anaemic pregnant women. From July to November 2000, 202 pregnant women were screened for anaemia (haemoglobin < 11.0 g/dL). One hundred and three pregnant women (51%) were found to be anaemic and were then allocated alternately into four groups. Over a period of 60 days, group IF (n = 29) received iron-folate tablets (200 mg FeSO4 and 250 microg folic acid) + 5 mg glucose: group IFR (n = 22) received iron-folate tablets + 5 mg riboflavin; group IFA (n = 29) received iron-folate tablets + 2.75 mg retinyl palmitate (equal to 5000 IU vitamin A); and group IFRA (n = 23) received iron-folate tablets + 5 mg riboflavin + 2.75 mg retinyl palmitate. At the end of the study 19 pregnant women (18.4%) were excluded from the analysis because of various reasons. Statistical analysis was based on 84 women (81.5%): group IF, n = 25; group IFR, n = 22; group IFA, n = 18; and group IFRA, n = 19. Haemoglobin measurements were carried out using the Technicon H1* (cyanmethaemoglobin method). All groups showed a significant increase in haemoglobin concentration (P < 0.05), except group IFA (P > 0.05), with the highest increment being in group FR. Multiple comparisons only showed significant differences between group IFR and group IFA (P < 0.05). It can be concluded that iron-folate supplementation can increase haemoglobin concentrations in anaemic pregnant women. Adding riboflavin tends to enhance the effect of iron-folate supplementation, but this is not the case with adding vitamin A.

Assessment of vitamin A status in chronic obstructive pulmonary disease patients and healthy smokers.

The relation between vitamin A status and the degree of lung airway obstruction was examined in a cross-sectional study of 36 male subjects aged 43-74 y who were assigned to five groups as follows: healthy nonsmokers (n = 7), healthy smokers (n = 7), mild chronic obstructive pulmonary disease (COPD-mild) patients (n = 9), COPD-moderate-severe patients (n = 7), and COPD-moderate-severe patients with exacerbation (+ex; n = 6). Smoking habits, pulmonary function tests, energy-protein status were assessed; serum concentrations of retinyl esters, retinol, retinol binding protein, and transthyretin and relative dose responses were measured. In addition, 12 male smokers aged 45-61 y with mild COPD were randomly assigned to two groups for a longitudinal study: six subjects consumed vitamin A (1000 RE/d; COPD-vitamin A) and six subjects received placebo for 30 d. Lowered serum retinol concentrations were found in the COPD-moderate-severe and COPD-moderate-severe+ex groups. Measurements of vitamin A status in healthy smokers and in COPD-mild patients were not different from those in healthy nonsmokers. The improvement of pulmonary function test results after vitamin A supplementation [mean increase for 1-s forced expiratory volume (FEV1) = 22.9% in the COPD-vitamin A group] may support the assumption of a local (respiratory) vitamin A deficiency in patients with this disease.

Separation of specific stages of spermatids from vitamin A-synchronized rat testes for assessment of nucleoprotein changes during spermiogenesis.

Studies of the precise steps of spermiogenesis at which dramatic changes occur in the nuclear proteins have been limited by the inability to obtain sufficient quantities of these cells in narrowly defined developmental stages, especially those between steps 1 and 12. This limitation can now be overcome by vitamin A-induced synchronization of rat testes into a few stages of the seminiferous epithelial cycle. Cell suspensions from stage-synchronized rat testes were separated by centrifugal elutriation, and selected fractions were further purified on Percoll gradients. Fractions enriched in spermatids in steps 1-3, 7-8, 9-10, and 11-12 were obtained. Analysis of the basic nucleoproteins from these cells by PAGE revealed the following changes. Between steps 3 and 7, histone (H) 2A variants, H2A.1, H2A.2, and TH2A, became post-translationally modified; and during steps 9-11, H1t became modified. H4, which was monoacetylated in steps 1-3, showed maximal levels of hyperacetylation in steps 11-12. The histones were the major basic nuclear proteins in spermatids through step 12. The low levels of transition proteins 1 and 2 observed in a fraction enriched in steps 11-12 could be largely accounted for by contamination from step 13-15 spermatids. All results were consistent with those obtained from normal, unsynchronized rats. The technique of vitamin A synchronization is therefore useful in more precisely defining biochemical changes during spermiogenesis.

Assessment of vitamin A status in pregnant women as reflected by in vitro destruction of vitamin A by hemolysates and urinary ammonium nitrogen to creatinine ratio.

To investigate the significance of the lowered plasma vitamin A during the last few weeks of pregnancy, the capacity of erythrocyte lysates to destroy vitamin A in vitro and the alteration in ammonium nitrogen to creatinine ratio (Am N/Cr) in random urine samples was followed up at different gestational ages in women from low income groups. One group received no supplementation and the other received supplements of 1800 retinol equivalents (RE) and at most points the number of observations were not less than 12. In the unsupplemented women there was a significant increase in the capacity of erythrocytes to destroy vitamin A in vitro, which reflects an increased oxidative stress as compared to the supplemented group. While differences in mean Am N/Cr did not follow any consistent pattern, there was a strong association between lowered plasma vitamin A (< 35 micrograms/dl) and increased Am N/Cr ratio (> 0.5). This was true only when combined data of both groups were considered till 26 weeks of gestation and not beyond. As expected in supplemented women there was no significant fall in the plasma vitamin A during term. These data reveal that the lowered plasma vitamin A levels are suggestive of greater risk of hypovitaminosis A during pregnancy as indicated by the oxidative stress in erythrocytes.

Assessment of the oral administration of a high dose of retinol on vitamin E status of sheep.

The effect of dietary excess of retinol on vitamin E turnover in sheep's body was studied. Five of 10 adult Canadian Arcott sheep were given daily for 30 d an oral dose of 250 mg vitamin A as retinol palmitate, while the remaining sheep were used as control. Then a single oral dose of D-alpha(5-Me-3H) tocopherol (0.2 microCi/kg vitamin B) was administered to each sheep in both groups. The kinetic data were modelled as a 2-compartment model which provided a simple kinetic description of vitamin E absorption. The analysis showed no difference between the 2 groups of sheep regarding their various pharmacokinetic values. It is suggested that oversupply of dietary retinol does not antagonize vitamin E turnover in the sheep.

Effect of supplementation with vitamin A or plant carotenes on plasma retinol levels among young Egyptian males.

Thirteen young Egyptian males with initial low plasma retinol level were divided into three groups and received single oral vitamin A dose of 200,000 IU, or carotene supplement in the form of boiled spinach leaves, providing roughly 617 RE/day or carrots providing 394 RE/day. The three supplements were equally effective, when the plasma retinol level was measured 40 days later. On a price basis, the vitamin A capsules and the carrots were much cheaper than the spinach supplement.

Methodological proposal in behavioural teratogenicity testing: assessment of propoxyphene, chlorpromazine, and vitamin A as positive controls.

Pregnant Sprague-Dawley rats received either 80 mg/kg d-propoxyphene HCl or 20 mg/kg chlorpromazine HCl or 80,000 and 160,000 IU/kg vitamin A palmitate daily between the 6th and 20th days of gestation. Vehicle control groups were similarly treated with saline or corn oil and considered as negative controls. Offspring were examined for physical landmarks, neuromotor development, and behaviour using righting reflex, swimming, negative geotaxis, open field, rotarod, water maze, and nocturnal activity. This test battery included biochemical measurements. No reduction in parental weight and physical offspring development was observed. All these treatments produced long-term changes in more than one test. Vitamin A palmitate (160,000 IU/kg) was judged as the best positive control with this test battery for future investigation of the behavioural teratology of drugs.

Assessment of the ability of mice fed on vitamin A supplemented diet to respond to a variety of potential contact sensitizers.

The contact sensitizing potential of various compounds was tested in mice maintained on a diet supplemented with vitamin A acetate. Substances were applied epicutaneously. After challenge, increases in ear thickness were measured and the differences in response between control and treated groups determined by the Mann Whitney test. A number of sensitizers, including Dowicil 200, cinnamaldehyde, hydroxycitronellal and Kathon CG gave positive sensitivity responses.