Vitamina d, sus acciones "no clásicas" y su utilidad en la pandemia del covid-19 / Vitamin d, its "non-classic" actions and it utility in the covid-19 pandemic

RESUMEN El descubrimiento de que la síntesis de 1,25 vitamina D no fue solo renal, la enzima 1 alfa hidroxilasa se encuentra en numerosos tejidos del organismo, además de la evidencia de que la asociación entre el déficit de vitamina D y la presencia de enfermedades no óseas (cáncer, esclerosis múltiple, enfermedades autoinmunes, etc.) nos ofrece la posibilidad de intentar prevenir estas afecciones. Los estudios de suplementación contra placebo no han dado resultados positivos para algunas afecciones, aunque algunos de esos trials se realizaron en población "suficiente" y no "deficiente" de vitamina D. Sin embargo, otros metaanálisis han demostrado prevención en los grupos suplementados con déficit para algunas patologías (infecciones respiratorias, prediabetes). Además, existe evidencia de efecto antiviral de la misma. La acción antiinfecciosa e inmunomoduladora que ejerce y su efecto sobre el sistema renina angiotensina, estimulando la enzima convertidora de angiotensina 2 (que es el receptor virus del SARS-CoV), permiten sospechar, actualmente, que con niveles elevados podría ser más difícil, o menos grave, la infección por COVID-19. La suplementación con vitamina D es conveniente para prevenir enfermedades en sujetos con déficit, pero en medio de la grave pandemia 2020 administrarla, aún sin tener un dosaje previo en las poblaciones de mayor riesgo, podría disminuir la chance de esta enfermedad.

ABSTRACT The discovery that the synthesis of 1-25-vitamin D is not only renal and that the enzyme 1 alpha hydroxylase is found in numerous tissues of the body, together with the evidence of the association between vitamin D deficiency and the presence of non-bone diseases (cancer, multiple sclerosis, autoimmune diseases, etc.), gives us the possibility of trying to prevent these conditions. Placebo-controlled supplementation studies have not provided positive results for certain conditions, but some of these trials have been carried out on populations with "sufficient" and not "deficient" vitamin D levels. However, other meta-analyses have shown prevention of some conditions (respiratory infections, prediabetes) in groups of patients with deficiencies who were given supplements. There is also evidence of antiviral effect of vitamin D. Its anti-infective and immunomodulatory action and its effect upon the renin-angiotensin system, stimulating the angiotensin-converting enzyme 2 (the SARS-CoV virus receptor), nowadays allow us to think that, in high levels, COVID-19 infection could be less likely or serious. Vitamin D supplementation is adequate for preventing diseases in patients with deficiencies; administering vitamin D within the 2020 pandemic, even without having tested it in high-risk populations, could diminish the incidence of this disease.

Vitamin D nutritional status in the adult population in Colombia - An analytical cross-sectional study.

BACKGROUND: The key role of Vitamin D is to maintain an adequate calcium and phosphorus metabolism. Vitamin D plays an antagonistic role with the parathyroid hormone. 25 OH Vitamin D is the major circulating form and the best indicator to monitor Vitamin D levels. METHODS: A cross-sectional study was conducted in 1339 individuals ≥18 years old. The main objective was to establish the nutritional status of Vitamin D and its association with PTH and ionized calcium levels. Other objectives were to compare the levels of 25 OH Vitamin D based on sun exposure habits, and to identify the minimum cut-off point for the levels of 25 OH Vitamin D that could give rise to a concomitant increase in PTH and ionized calcium levels. RESULTS: 14.2% of participants presented Vitamin D deficiency, and 28.8% presented insufficiency; ≥89% of the participants with deficiency or insufficiency were exposed to sunlight <30 minutes per week. A value of 25 OH Vitamin D >30 ng/mL was associated with a more stable and "flat" PTH value. The median of 25 OH Vit-D associated with hypercalcemia was <10 ng/mL. CONCLUSION: In Colombia, low 25 OH Vitamin D values are highly prevalent; this may be accounted for by poor sun-exposure habits and frequent use of sunscreen. Just as in other similar trials, the lower the levels of 25 OH Vit-D, the higher the effect on PTH and ionized calcium elevation.

25-OH-vitamin D deficiency identifies poor tumor response in hepatocellular carcinoma treated with transarterial chemoembolization.

PURPOSE: Vitamin D is implicated linked to liver cancer and chronic liver diseases, but its association with tumor response in hepatocellular carcinoma (HCC) patients undergoing transarterial chemoembolization (TACE) remains unclear. This study aimed to determine whether vitamin D levels influence tumor response in HCC patients treated with TACE. METHODS: A total of 58 HCC patients undergoing TACE were enrolled in the study. Serum 25-hydroxyvitamin D (25-OHD) levels were determined at baseline and 1 day after TACE using electrochemiluminescence immunoassay. Response to TACE was evaluated after a 4-6 week interval. Univariate and multivariate analyses with Cox regression model were performed to determine the risk factors associated with tumor response. Receiver operating characteristic (ROC) curve analysis was performed to assess the predictive performance of baseline 25-OHD levels on tumor response in HCC patients undergoing TACE. RESULTS: 43.1% of HCC patients showed 25-OHD deficiency. Baseline 25-OHD level was associated with liver cirrhosis (P = 0.025), vascular invasion (P = 0.031), Barcelona Clinic Liver Cancer stage (P = 0.002) and an alanine aminotransferase increase after TACE (P = 0.021). Serum 25-OHD level was significantly decreased 1 day after TACE (P = 0.045). Multiple tumor numbers (P = 0.034) and low baseline 25-OHD levels (P = 0.040) were independently correlated with poor tumor response after TACE. ROC curve analysis showed that baseline 25-OHD levels present better predictive performance for OR in those patients, compared with other current clinical test pointers. CONCLUSION: Our study suggested that 25-OHD deficiency at baseline is a prognostic indicator for a poor tumor response in hepatocellular carcinoma treated with TACE.

Normal physiological functions in two animal species with highly different vitamin d status compared to that of humans / Funciones fisiológicas normales en dos especies animales con estatus de vitamina d muy diferentes del actual en seres humanos

Mole rats live in permanent darkness, in networks of underground tunnels (which extend up to 1 km in the subsoil), excavated with their incisors, in warm and semi-arid areas of South Africa. Mole rats have an unusually impoverished vitamin D3 status with undetectable and low plasma concentrations of 25- hydroxyvitamin D3 and 1α,25-dihydroxyvitamin D3, respectively. They express 25-hydroxylase in the liver and 1-hydroxylase and 24-hydroxylase in their kidneys. The presence of specific receptors (VDR) was confirmed in the intestine, kidney, Harderʼs glands and skin. In spite of their poor vitamin D3 status, the apparent fractional intestinal absorption of calcium, magnesium and phosphate was high, always greater than 90%. Oral supplementation with cholecalciferol to mole rats did not improve the efficiency of gastrointestinal absorption of these minerals. Mole ratsdo not display the typical lesion of rickets: hypertrophic and radiolucent growth cartilages. Histological studies reported normal parameters of trabecular and cortical bone quality. Marmosets (monkeys of the New World) are not hypercalcaemic, eventhough they exhibit much higher levels of 25-hydroxyvitamin D3, 1α,25-dihydroxyvitamin D3 and parathyroid hormonethan that of rhesus monkeys and humans. Fed a high vitamin D3 intake (110 IU/day/100 g of body weight), a fraction of the experimental group was found to display osteomalacic changes in their bones: distinct increases in osteoid surface, relative osteoid volume, and active osteoclastic bone resorption. These findings suggest that some marmosets appears to suffer vitamin D-dependent rickets, type II. The maximum binding capacity of the VDR or the dissociation constant of VDR1α,25(OH)2D3 complex of mole rats and New World monkeys are distinctly different of VDR isolated from human cells. Health status of those species appears to be adaptations to the mutations of their VDR. Though rare, as mutations may occur at any time in any patient, the overall message of this review to clinicians may be: recent clinical studies strongly suggests that the normality of physiological functions might be a better indicator of the health status than the serum levels of vitamin D metabolites. (AU)

Las ratas topo viven en la oscuridad permanente, en redes de túneles subterráneos excavadas con sus incisivos (que se extienden hasta 1 km en el subsuelo), en áreas cálidas y semiáridas de Sudáfrica. Las ratas topo tienen un estatus de vitamina D3 inusualmente empobrecido con concentraciones plasmáticas indetectables de 25-hidroxivitamina D3 y bajas de 1α, 25-dihidroxivitamina D3. Poseen 25-hidroxilasa en el hígado y 1-hidroxilasa y 24-hidroxilasa en sus riñones. La presencia de receptores específicos (VDR) ha sido confirmada en el intestino, el riñón, las glándulas de Harder y la piel. A pesar de su pobre estatus de vitamina D3,la absorción fraccional intestinal aparente de calcio, magnesio y fosfato fue alta, siempre superior al 90%. La suplementación oral con colecalciferol a las ratas topo no mejoró la eficacia de la absorción gastrointestinal de estos minerales. No muestran la lesión típica del raquitismo: cartílagos de crecimiento hipertróficos y radiolúcidos. Varios estudios histológicos confirman los hallazgos radiológicos y se informan parámetros normales de la calidad ósea trabecular y cortical. Los titíes (monos del Nuevo Mundo) exhiben calcemias normales con niveles más elevados de 25-hidroxivitamina D3, 1α,25-dihidroxivitamina D3 y hormona paratiroidea que los monos rhesus y los seres humanos. Un tercio de un grupo de titíes alimentados con una alta ingesta de vitamina D3 (110 I/día/100 g de peso corporal) exhibió cambios osteomalácicos en sus huesos: aumento en la superficie osteoide, volumen osteoide y activa reabsorción osteoclástica. Estos hallazgos sugieren que una fracción de la población de titíes padece raquitismo dependiente de vitamina D, tipo II. Debido a mutaciones ocurridas hace millones de años, las máximas capacidades de ligamiento del VDR o los valores de la constante de disociación del complejo VDR-1α,25(OH)2D3 de las ratas topo o monos del Nuevo Mundo son muy diferentes de los verificables en receptores aislados de células humanas actuales. El mensaje de esta revisión a los médicos clínicos podría ser: varios estudios clínicos recientes indican que la normalidad de las funciones fisiológicas de un paciente es un mejor indicador de su salud que los niveles séricos de los metabolitos de la vitamina D. (AU)

Effect of Season on 25-OH Vitamin D in Serum of Rhesus Monkeys Housed in Puerto Rico.

OBJECTIVE: Vitamin D blood levels have been shown to be partially dependent upon season in temperate climates, however, this same evaluation has not yet been reported in fully tropical climates. Herein, we assessed the vitamin D levels in the blood of Rhesus monkeys housed at the Puerto Rico Caribbean Primate Research Center collected in the island's "summer"(May-October) and "winter" (November-April) months. MATERIALS AND METHODS: In 2006 through 2014, repeated measurements of blood samples were collected from 5 Rhesus monkeys (IACUC-approved) during "summer" and "winter" months to assess 25-OH vitamin D, determined via HPLC. UV-B and UV-A (KJ/m2/day) were measured using a ground based radiometer for these time periods. A paired t-test and a multilevel mixed- effect model approach was performed for data analysis. RESULTS: The difference of the mean serum values of 25-OH vitamin D between seasons showed lower levels during "winter" than "summer" months. About 23% of the variance in levels can be attributed to difference between the monkeys. The means of UV-B and UV-A, as a proxy for sunlight intensity, were greater (over the entire study interval) during the "summer" as opposed to "winter" months (p < 0.001). CONCLUSION: Vitamin D levels were substantially higher in the "summer" rather than the "winter" months. This observation implies that even in fully tropical regions, such as Puerto Rico, time of year can have an influence on vitamin D status. While comparable studies have not been undertaken in humans, it would not be unreasonable to suggest that similar results would be obtained should such a study be done.

Association between Vitamin D Deficiency and Single Nucleotide Polymorphisms in the Vitamin D Receptor and GC Genes and Analysis of Their Distribution in Mexican Postmenopausal Women.

Genome-wide association studies in people with European ancestry suggest that polymorphisms in genes involved in vitamin D (VD) metabolism have an effect on serum concentrations of 25-hydroxyvitamin D. However, nothing is known about these polymorphisms in populations with Amerindian ancestry. Our aim was to evaluate the association between genetic variants on the vitamin D receptor (VDR) and the vitamin D binding protein (GC) genes, involved in the VD pathway, and VD deficiency in 689 unrelated Mexican postmenopausal women. We also described the frequencies of these variants in 355 postmenopausal women from different ethnic groups. Based on our preliminary results of 400 unrelated Mexican postmenopausal women, three single nucleotide polymorphisms (SNPs) were selected for genotyping. The SNPs rs4516035 in VDR and rs2282679 in GC were associated with VD deficiency. Additionally, women who carried three risk alleles had a 3.67 times higher risk of suffering VD deficiency, compared to women with no risk alleles (p = 0.002). The rs4516035-C allele frequency in the Amerindian population was enriched in the South East region of Mexico. In contrast, the highest frequency of the rs2298850-C allele, a proxy for the tag SNP rs2282679, was observed in the South region. Our results indicate that genetic variants in VDR and GC genes are associated with VD deficiency in Mexican postmenopausal women. Moreover, an association was observed for the variants rs3794060 and rs4944957 of the DHCR7/NADSYN1 gene with osteopenia/osteoporosis.

Effect of Housing Type on 25 OH Vitamin D in Serum of Rhesus Monkeys.

OBJECTIVE: Vitamin D status is primarily dependent upon sun exposure and dietary sources, however genetic, cultural, and environmental factors can have a modulating role in the measured amount. One under-reported factor is the effect of regular living quarters on the degree of sun exposure. Herein, we assess vitamin D status in the blood of Rhesus monkeys (Macaca mulatta) housed in high amounts of sunlight (corn-cribs), medium sunlight (corrals with shaded areas), and minimal sunlight (quarantine cages). METHODS: Fifty-five male Rhesus monkeys, aged 1 to 31 years were housed in varying amounts of sun exposure at the Caribbean Primate Research Center. Serum was collected and analyzed for 25 OH Vitamin D which is the preferred metabolite for determination of Vitamin D using High Performance Liquid Chromatography (HPLC). RESULTS: 25 OH Vitamin D levels in blood were significantly greater in corn-cribhoused monkeys than in corral or quarantine-housed animals (p > 0.01 and p > 0.001 respectively). Significant differences of serum levels were not found when ages of animals housed in the same environment were compared. CONCLUSION: Monkeys housed in a tropical environment with the greatest amount of exposure to sunlight maintain the highest serum levels of 25 OH vitamin D independent of age. These findings emphasize the importance of documenting the environment in which subjects typically spend their time when Vitamin D results are interpreted.

Avaliação dos níveis séricos de vitamina D em crianças e adolescentes com diabetes mellitus tipo I / Evaluation of vitamin D serum levels in children and adolescents with diabetes mellitus type 1

Introdução: A deficiência de vitamina D é considerada um problema de saúde pública no mundo todo em razão de suas implicações no desenvolvimento de diversas doenças, incluindo a Diabetes Mellitus tipo I (DMI). Os objetivos deste estudo são avaliar os níveis séricos de 25 OH Vitamina D, Cálcio, Fosfatase alcalina, fósforo e hemoglobina glicada em crianças diabéticas tipo I e correlacionar os níveis de vitamina D com a hemoglobina glicada e com a presença de Tireoidite de Hashimoto, microalbuminúria e Doença celíaca. Métodos: Estudo do tipo quantitativo, observacional transversal obtido pela analise de exames laboratoriais de 48 pacientes, maiores de 3 anos, com diagnóstico de Diabetes Mellitus tipo I. Os dados foram obtidos mediante análise dos prontuários. Resultados: Dezenove pacientes (59,4%) apresentaram vitamina D deficiente/insuficiente. A média dos níveis de cálcio, fósforo, fosfatase alcalina e HbA1c foram de 9,24, 281,17, 4,69 e 9,34, respectivamente. Em 31 pacientes (91,17%) o controle da DMI foi considerado ruim ou regular, sendo este avaliado pelo valor da hemoglobina glicosilada. Dez pacientes (29,41%) apresentaram uma ou mais das comorbidades associadas à DMI. Conclusão: Em 59,4 % dos pacientes o nível de vitamina D foi deficiente ou insuficiente. A média dos níveis de cálcio, fósforo e fosfatase alcalina dos pacientes estavam dentro dos valores normais. Em 31 pacientes (91,17%) o controle da DMI, avaliado pela HbA1c, foi considerado ruim ou regular. Não houve associação significativa entre a presença de Tireoidite de Hashimoto, doença celíaca ou microalbuminúria e os níveis de vitamina D.

Introdução: Vitamin D deficiency is considered a public health problem worldwide because of its implications in the development of various diseases, including diabetes mellitus type I (DMI). The aims of this study are to evaluate the serum levels of 25 OH Vitamin D, calcium, alkaline phosphatase, phosphorus and glycated hemoglobin in diabetic children type I and correlate vitamin D levels with glycated hemoglobin and presence of Hashimoto's thyroiditis, microalbuminuria and celiac disease. Methods: A quantitative, observational cross-sectional study performed by analysis of laboratory tests from 48 patients over three years of age and diagnosed with diabetes mellitus type I. The data were obtained by analyzing medical charts. Results: Nineteen patients (59.4%) were vitamin D deficient/insufficient. Calcium, phosphorus, alkaline phosphatase and HbA1c mean levels were 9.24, 281.17, 4.69, and 9.34, respectively. In 31 patients (91.17%), control of DMI was considered poor or fair, as measured by the value of glycosylated hemoglobin. Ten patients (29.41%) had one or more comorbidities associated with DMI. Conclusion: In 59.4% of the patients vitamin D level was deficient or insufficient. Patients’ mean levels of calcium, phosphorus and alkaline phosphatase were within the normal range. In 31 patients (91.17%) the control of DMI, assessed by HbA1c, was considered poor or fair. There was no significant association between presence of Hashimoto's thyroiditis, celiac disease or microalbuminuria and vitamin D levels.

Hipovitaminosis D en pacientes hemodiálizados (HD): factores relacionados e influencia sobre la fuerza muscular / Hypovitaminosis D in patients on hemodialysis (HD): related factors and influence on muscle strength

Introducción: La deficiencia de 25 (OH) vitamina D es una alteración prevalente en los pacientes con enfermedad renal crónica (ERC) , sin embargo en nuestro medio no es medida de manera rutinaria y por ende no suele hacerse reposición vitamínica. Nuestro objetivo fue determinar la prevalencia y los factores relacionados a deficiencia de 25 (OH) D en pacientes con ERC en hemodiálisis (HD), particularmente la relación con la función y masa muscular. Métodos: Efectuamos un estudio prospectivo, multicéntrico, en pacientes adultos en HD crónica que no estuvieran recibiendo ningún derivado de la vitamina D. Se midieron en sangre los niveles de 25(OH) D, Hemoglobina, PCR, Albúmina, Ca, P, FAL, PTHi. Se realizó la medición de la fuerza del puño con dinamómetro, y la prueba de sentado-parado. Se aplicó el índice de Karnofsky para clasificar el estado funcional., Se realizó una bioimpedanciometría (BCM; Frese nius Medical Care) en aquellos pacientes sin, contraindicación. Resultados: Se incluyeron 138 pacientes. La 25(OH) vitamina fue de 20.43 ± 10.5 ng/ml, la prevalencia de insuficiencia /defi ciencia 87% (37% con menos de 15 ng/ml). Las concentraciones de vitamina D/deficiencia mostraron correlación/relación significativa con la edad, la presencia de diabetes, los niveles de hemoglobina y albúmina, la fuerza y la masa muscular y la clase funcional (p<0.05) . Conclusión: Alta prevalencia de hipovitaminosis D en pacientes hemodializados particularmente gerontes y diabéticos. Esto estaría relacionado con la desnutrición, anemia, clase funcional y la fuerza/masa muscular de los pacientes, estos últimos dos factores no reportados hasta ahora. Todos estos factores deben ser considerados al momento de la sustitución vitamínica y en la evaluación de la efectividad de la misma.(AU)

Background: 25 (OH) vitamin D deficiency is a prevailing alteration in patients with chronic kidney disease (CKD); however, in our environment, it is not routinely measured and, therefore, vitamin replacement is unusual. Our purpose was assessing the prevalence of and the factors related to 25 (OH) vitamin D deficiency in patientswith CKD in hemodialysis (HD), especially the relation to function and muscle mass. Methods: We conducted a prospective, multicenter study in adult patients on chronic HD who were not receiving any vitamin D derivative. Blood levels of 25 (OH) D, Hemoglobin, CRP, Albumin, Ca,P, ALP and PTHi were measured. The handgrip strength was measured with a dynamometer and the sitting-rising test was carried out. A bioimpedance analysis (BCM; Fresenius Medical Care) was conducted in the patients who had no contraindications. Results: 138 patients were included. The levels of 25 (OH) vitamin D were 20.43±10.5 ng/ml; the insufficiency/deficiency had 87% prevalence (and 37% prevalence with less than 15 ng/ml). Vitamin D concentrations/ deficiency showed a significant correlation with/ relation to age, diabetes, hemoglobin and albumin levels, muscle strength and mass, and functional class (p<0.05). Conclusion: High prevalence of hypovitaminosis D in patients on hemodialysis, particularly in the elderly and in patients with diabetes. This should be related to undernutrition, anemia, the functional class and the muscle strength/mass of patients, the latter two being unreported factors until now. All these factors should be considered when vitamin replacement is conducted and when its effectiveness is assessed.(AU)

Hipovitaminosis D en pacientes hemodiálizados (HD): factores relacionados e influencia sobre la fuerza muscular / Hypovitaminosis D in patients on hemodialysis (HD): related factors and influence on muscle strength

Introducción: La deficiencia de 25 (OH) vitamina D es una alteración prevalente en los pacientes con enfermedad renal crónica (ERC) , sin embargo en nuestro medio no es medida de manera rutinaria y por ende no suele hacerse reposición vitamínica. Nuestro objetivo fue determinar la prevalencia y los factores relacionados a deficiencia de 25 (OH) D en pacientes con ERC en hemodiálisis (HD), particularmente la relación con la función y masa muscular. Métodos: Efectuamos un estudio prospectivo, multicéntrico, en pacientes adultos en HD crónica que no estuvieran recibiendo ningún derivado de la vitamina D. Se midieron en sangre los niveles de 25(OH) D, Hemoglobina, PCR, Albúmina, Ca, P, FAL, PTHi. Se realizó la medición de la fuerza del puño con dinamómetro, y la prueba de sentado-parado. Se aplicó el índice de Karnofsky para clasificar el estado funcional., Se realizó una bioimpedanciometría (BCM; Frese nius Medical Care) en aquellos pacientes sin, contraindicación. Resultados: Se incluyeron 138 pacientes. La 25(OH) vitamina fue de 20.43 ± 10.5 ng/ml, la prevalencia de insuficiencia /defi ciencia 87% (37% con menos de 15 ng/ml). Las concentraciones de vitamina D/deficiencia mostraron correlación/relación significativa con la edad, la presencia de diabetes, los niveles de hemoglobina y albúmina, la fuerza y la masa muscular y la clase funcional (p<0.05) . Conclusión: Alta prevalencia de hipovitaminosis D en pacientes hemodializados particularmente gerontes y diabéticos. Esto estaría relacionado con la desnutrición, anemia, clase funcional y la fuerza/masa muscular de los pacientes, estos últimos dos factores no reportados hasta ahora. Todos estos factores deben ser considerados al momento de la sustitución vitamínica y en la evaluación de la efectividad de la misma.

Background: 25 (OH) vitamin D deficiency is a prevailing alteration in patients with chronic kidney disease (CKD); however, in our environment, it is not routinely measured and, therefore, vitamin replacement is unusual. Our purpose was assessing the prevalence of and the factors related to 25 (OH) vitamin D deficiency in patientswith CKD in hemodialysis (HD), especially the relation to function and muscle mass. Methods: We conducted a prospective, multicenter study in adult patients on chronic HD who were not receiving any vitamin D derivative. Blood levels of 25 (OH) D, Hemoglobin, CRP, Albumin, Ca,P, ALP and PTHi were measured. The handgrip strength was measured with a dynamometer and the sitting-rising test was carried out. A bioimpedance analysis (BCM; Fresenius Medical Care) was conducted in the patients who had no contraindications. Results: 138 patients were included. The levels of 25 (OH) vitamin D were 20.43±10.5 ng/ml; the insufficiency/deficiency had 87% prevalence (and 37% prevalence with less than 15 ng/ml). Vitamin D concentrations/ deficiency showed a significant correlation with/ relation to age, diabetes, hemoglobin and albumin levels, muscle strength and mass, and functional class (p<0.05). Conclusion: High prevalence of hypovitaminosis D in patients on hemodialysis, particularly in the elderly and in patients with diabetes. This should be related to undernutrition, anemia, the functional class and the muscle strength/mass of patients, the latter two being unreported factors until now. All these factors should be considered when vitamin replacement is conducted and when its effectiveness is assessed.