Long-term metformin therapy and vitamin B12 deficiency: An association to bear in mind

To date, metformin remains the first-line oral glucose-lowering drug used for the treatment of type 2 diabetes thanks to its well-established long-term safety and efficacy profile. Indeed, metformin is the most widely used oral insulinsensitizing agent, being prescribed to more than 100 million people worldwide, including patients with prediabetes, insulin resistance, and polycystic ovary syndrome. However, over the last decades several observational studies and meta-analyses have reported a significant association between long-term metformin therapy and an increased prevalence of vitamin B12 deficiency. Of note, evidence suggests that long-term and high-dose metformin therapy impairs vitamin B12 status. Vitamin B12(also referred to as cobalamin) is a water-soluble vitamin that is mainly obtained from animal-sourced foods. At the cellular level, vitamin B12 acts as a cofactor for enzymes that play a critical role in DNA synthesis and neuroprotection. Thus, vitamin B12 deficiency can lead to a number of clinical consequences that include hematologic abnormalities(e.g., megaloblastic anemia and formation of hypersegmented neutrophils), progressive axonal demyelination and peripheral neuropathy. Nevertheless, no definite guidelines are currently available for vitamin B12 deficiency screening in patients on metformin therapy, and vitamin B12 deficiency remains frequently unrecognized in such individuals. Therefore, in this field of vision article we propose a list of criteria for a cost-effective vitamin B12 deficiency screening in metformin-treated patients, which could serve as a practical guide for identifying individuals at high risk for this condition. Moreover, we discuss additional relevant topics related to this field, including:(1) The lack of consensus about the exact definition of vitamin B12 deficiency;(2) The definition of reliable biomarkers of vitamin B12 status;(3) Causes of vitamin B12 deficiency other than metformin therapy that should be identified promptly in metformin-treated patients for a proper differential diagnosis; and(4) Potential pathophysiological mechanisms underlying metformin-induced vitamin B12 deficiency. Finally, we briefly review basic concepts related to vitamin B12 supplementation for the treatment of vitamin B12 deficiency, particularly when this condition is induced by metformin.     

Absence of luminal intrinsic factor after gastric bypass surgery for morbid obesity

Abnormally low serum cobalamin levels (<180 pg/ml) have been observed in 154 of 429 patients (36%) at an average of 22 months (range 3–64 months) after gastric bypass surgery for morbid obesity. Twenty-four patients underwent a Schilling test and retrograde endoscopy of the bypassed gastric segment to determine the presence of intrinsic factor (IF) in gastric aspirates and in mucosal biopsies at 22±4 months after surgery. Five patients had a normal cobalamin level (405±44 pg/ml), and gastric juice intrinsic factor was present in three of them (11±7 ng/ml). Nineteen patients had a low cobalamin level (113±8 pg/ml), and gastric juice IF was found in only two subjects of this group (10 ng/ml each). Basal gastric juice IF concentration of healthy control subjects was 24±5 ng/ml. Schilling test results were normal in all five patients of the first group and in only nine patients of the group with cobalamin deficiency after surgery. To assess whether IF was present within the parietal cells of subjects with absent luminal IF, we studied gastric biopsy material of 14 patients using a well-characterized indirect immunoperoxidase method. IF was identified in fundic mucosal biopsy specimens of all 14 patients with absent gastric juice IF. We conclude that cobalamin deficiency occurs in a significant number of patients after gastric bypass and is associated with absence of gastric juice IF. We propose that this abnormality might be caused by inadequate secretion of IF from the bypassed stomach.This work was presented in part at the annual meeting of the American Gastroenterological Association, San Francisco, California, May 1986, and has appeared in abstract form (Gastroenterology 90:1533, 1986).     

Serum cobalamin, folate, methylmalonic acid and total homocysteine as vitamin B12 and folate tissue deficiency markers amongst elderly Swedes--a population-based study

OBJECTIVES: The possibilities of detecting tissue cobalamin and folate deficiency are under debate. In this report the levels of serum cobalamin, folate, methylmalonic acid (MMA) and total homocysteine (tHcy) and their interrelations in a representative random population sample are presented. DESIGN: Cohort study. SETTING: A general mid-Swedish population. SUBJECTS: A 20% random sample of persons 70 years or older in a defined geographical area were invited to a survey. A total of 235 (85%) persons responded, out of whom 224 had no interfering diseases. MAIN OUTCOME MEASURES: Serum cobalamin, folate, MMA and tHcy. RESULTS: The serum levels of cobalamin, folate, MMA and tHcy were all correlated to cobalamin and folic acid treatment. They were also correlated to the intake of multivitamin preparations. In addition, serum cobalamin was higher in untreated women than in men but not correlated to age. Serum folate was correlated neither to sex nor age. Serum tHcy and MMA were both directly correlated to age but MMA not to sex. MMA was inversely correlated to serum cobalamin but not to serum folate, whereas serum tHcy was inversely correlated to serum cobalamin, folate and creatinine. Neither serum cobalamin, folate, MMA nor tHcy had any significant correlation to haemoglobin, erythrocyte volume fraction (EVF) or mean red cell volume. Half of the study population had abnormal MMA or tHcy levels, suggesting a latent or overt tissue deficiency of cobalamin or folate. CONCLUSIONS: A substantial proportion of the elderly general population had signs of low tissue levels of cobalamin or folate. Amongst those who took multivitamin preparations this proportion was much lower.     

Genetic polymorphisms predisposing to hyperhomocysteinemia in cardiac transplant patients

Genetic determinants for high homocysteine (Hcy) levels are now well known. We studied several single nucleotide polymorphisms (SNP) in Hcy-regulating genes [methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C; methionine synthase (MS) A2756G; methionine synthase reductase (MTRR) A66G] in relation to total plasma Hcy levels, transplant coronary artery disease and thromboembolic episodes in 84 heart transplant patients, and we compared the incidence of these polymorphisms with those in a healthy adult controls. At least one copy of the G allele of the MTRR A66G SNP was found in a significantly greater proportion of cardiac transplant (CTX) recipients compared with controls (94.0% vs. 79.9% respectively). None of the SNP analyzed were correlated with total Hcy plasma levels or the presence of transplant coronary artery disease. However, MS A2756G was significantly associated with cobalamin levels (AA genotype: 290 +/- 122 pmol/l; AG: 381 +/- 151 pmol/l and GG: 415 +/- 100 pmol/l), as was MTRR A66G (AA: 478 +/- 219 pmol/l, AG: 306 +/- 124 pmol/l and GG: 306 +/- 123 pmol/l). MTRR A66G was also correlated with serum folate. No association was found with thromboembolic events. In conclusion, there was a significant difference in the frequency of the G allele genotype of the MTRR A66G in CTX patients versus controls. Differences in cobalamin and folate levels with the MTRR A66G and MS A2756G polymorphisms were noted. Thus, SNP in Hcy-regulating genes may be important determinants of vitamin metabolism in CTX, raising the question of increased vitamin requirements to minimize increased plasma Hcy in this high-risk group.     

Long-term effects of nitrous oxide anaesthesia on laboratory and clinical parameters in elderly Omani patients: a randomized double-blind study

AIMS: This study examined the long-term effects of nitrous oxide anaesthesia on serum levels of cobalamin and folate, red cell folate levels and haematological parameters, and neurological status in elderly Omani patients. METHODS: Sixty-nine consecutive patients undergoing ophthalmic surgery were randomly and double-blind assigned to nitrous oxide or propofol anaesthesia. They met the following entry criteria: age 55 years or above, no major organ failure, no clinical signs or symptoms of cobalamin or folate deficiency, mean cell volume (MCV) cobalamin and/or folate substitution therapy during the preceding months. Serum levels of cobalamin and folate, red cell folate levels, and haematological parameters were measured prior to anaesthesia and 3-5 weeks later. At that time, the patients also underwent thorough neurological examination. RESULTS: Data of 51 patients were complete and considered for analysis. In both nitrous oxide and propofol group, the range of exposure time was comparable (+/-1 h). In the nitrous oxide group, a slight but significant decrease in haemoglobin, Hct, and red blood cell count (RBC) (P < 0.001) was observed, whereas there was a mild increase in mean cell haemoglobin (MCH) and mean cell volume (P < 0.05). In addition, there was a significant decrease in serum folate levels (P < 0.05). Hct and RBC decreased slightly in the propofol group (P < 0. 05), whereas there was a small increase in MCH. There was no difference between the two anaesthetics with regard to serum cobalamin and red cell folate levels, but there was a significant decrease in serum folate levels in the nitrous oxide group compared to those in the propofol group. Three patients with pre-existing low red cell folate levels, who were randomized to nitrous oxide anaesthesia, developed clinical symptoms of folate deficiency. CONCLUSION: This study showed that short-term (40-80 min) nitrous oxide anaesthesia did not affect cobalamin levels but reduced serum folate levels in this elderly population. Although this reduction was clinically irrelevant, some patients with pre-existing asymptomatic folate deficiency developed nitrous oxide-induced folate deficiency.     

Diversity in rat tissue accumulation of vitamin B12 supports a distinct role for the kidney in vitamin B12 homeostasis.

BACKGROUND: Vitamin B(12) in plasma is complexed to the carrier proteins transcobalamin (TC) and haptocorrin. The TC-B(12) complex is filtered in the glomeruli and reabsorbed in the renal tubules by receptor-mediated endocytosis, providing a route for a significant renal accumulation of vitamin B(12). The present study investigates the role of the rodent kidney in B(12) homeostasis by examining the distribution of vitamin B(12) in rats during vitamin B(12) depletion or B(12) load, and compares kidney accumulation with the vitamin distribution in other tissues including brain, liver, testes, intestine, spleen and plasma. METHODS: Fifteen rats were fed on a diet containing different concentrations of B(12) supplemented with s.c. injections of B(12). Twenty four hours prior to sacrifice, all animals were injected with [(57)Co]B(12). The vitamin contents of kidneys, liver, spleen, brain, testis, intestine, skeletal muscle, serum and urine were analysed. Both total tissue vitamin B(12) accumulation and [(57)Co]B(12) were determined to compare steady-state B(12) and the distribution of an acutely injected dose. In the kidney, free and protein-bound B(12) was determined by gel filtration. RESULTS: The rat kidneys accumulated more B(12) during normal and loaded conditions than any other tissue. A 110-fold increase in vitamin content was observed from the deficient to the loaded conditions in the kidney compared with a 3.5-fold increase in the liver. In contrast to all other organs, significantly smaller amounts of acutely injected B(12) accumulated in the kidneys in the vitamin-deprived state compared with both the normal and the vitamin-loaded condition. CONCLUSIONS: The present study suggests a significant role for the rodent kidney in vitamin B(12) metabolism. We propose a model for rat tissue uptake consistent with the presence of two different TC-B(12) receptors and renal uptake following filtration of TC-B(12) in the glomeruli. The presented model allows for the reduced renal uptake and accumulation in vitamin-deprived conditions, thus reserving the vitamin for other tissues, including nerve tissue and bone marrow, which are more sensitive to vitamin B(12) deficiency.     

Serum transferrin receptor in the megaloblastic anemia of cobalamin deficiency.

In order to further study the relation between transferrin receptor and erythropoiesis we examined serum receptor levels in megaloblastic anemia, which is the classic example of ineffective erythropoiesis. We studied 33 patients with unequivocal cobalamin deficiency, only 22 of whom were anemic. High serum transferrin receptor levels were found in 12 patients, all of whom were anemic and had high lactate dehydrogenase (LDH) levels; in contrast, only 10 of the 21 patients with normal receptor levels were anemic. Receptor correlated most strongly with LDH (r = 0.573, p < 0.001) and, inversely, with hemoglobin values (r = -0.560, p < 0.001); it also correlated with ferritin and total bilirubin levels, but not with cobalamin, MCV or erythropoietin. No association was found with the hemolytic component of megaloblastic anemia, represented indirectly by haptoglobin levels. Changes induced by cobalamin therapy were also examined in 13 patients. Transferrin receptors rose in all 6 patients who initially had high levels and in 2 of 3 patients who had borderline levels, but not in the 4 patients with initially normal levels. The receptor levels began to rise within 1-3 days, peaked at about 2 weeks and returned to normal at about the 5th wk. The findings indicate that serum transferrin receptor levels reflect the severity of the megaloblastic anemia. The elevated receptor levels rise further with cobalamin therapy, however, as effective erythropoiesis replaces ineffective erythropoiesis, and these persist until the increased erythropoiesis returns to normal.     

Cobalamin binding proteins in patients with HIV infection

Abstract: P-Cobalamins have been reported to be decreased in patients with HIV infection. Because of this, we found it of interest to examine both cobalamin-saturated binding proteins (holo-transcobalamin, holo-TC and holo-haptocorrin, holo-HC) and cobalamin unsaturated binding proteins (apo-transcobalamin, apo-TC and apo-haptocorrin, apo-HC). The results are given as range and (median). Eighteen male HIV-infected patients with plasma cobalamins below 200 pmol/1 were studied. We found low concentrations of holo-TC (37–88 (47.5) pmol/1) and holo-HC (64–184 (135.5) pmol/1). The concentration of apo-TC and apo-HC was increased (480–1730 (1025) pmol/1; 70–800 (235) pmol/1). It is concluded that, in HIV-infected patients, low plasma cobalamin does not reflect a low concentration of transcobalamin or haptocorrin. In 20 HIV-infected patients and 31 patients with malignant haematological diseases, the TC isopeptide patterns were determined. In the HIV group, an increased frequency of TC isopeptide X was found and the overall distribution of TC isopeptides was significantly different from the reference population (p < 0.05). There was no difference between the group of patients with malignant haematological diseases and the reference group.     

Increased Concentration of Transcobalamin I in a Patient with Metastatic Carcinoma of the Breast

A patient with metastatic carcinoma of the breast and increased plasma cobalamin binding capacity (about 50 nmol/1) is described. The binding protein was identified as transcobalamin I (TCI) by DEAE cellulose ion-exchange chromatography, Sephadex G200 gel filtration and agar gel electrophoresis. Although the total plasma cobalamin concentration (about 20 nmol/1) was elevated, the patient complained of neurological symptoms in accordance with a functional vitamin B₁₂ deficiency. Hence, an inactivation of the coenzyme is suggested by the demonstration of considerable amounts of 5′-deoxyadenosylcobalamin bound to the plasma TCI. Both urinary excretion of FIGLU and methylmalonic acid were within the reference ranges. Reported cases of increased cobalamin binding in patients with nonhaematological malignancy are reviewed. Further investigations to characterize the function of the cobalamin dependent metabolic pathways are necessary to determine the importance of the increased transcobalamin binding in these patients.     

Vitamin B12 Metabolism in Myelomatosis

In 38 patients with myelomatosis the serum cobalamin varied from 34 pmol/l to 404 pmol/l, median 181.5 pmol/l, which is significantly lower than the levels in 22 control persons with range 173–535 pmol/l, median 265 pmol/l. In spite of low serum cobalamin no symptoms of vitamin B₁₂ deficiency could be demonstrated in any of the patients, except for the one patient who had a serum cobalamin of 34 pmol/l. Mean values for Hb, MCV, PCV, serum lactate-dehydrogenase, adjusted red cell folate and nucleated neutrophil count were similar in a group of patients with a serum cobalamin below 160 pmol/l and a group of patients with higher serum cobalamin values. The decrease in serum cobalamin is due in part to a reduction in the major cobalamin binder (TC-I) in serum. Measuring serum cobalamin in relationship to gastric acid secretion, we found a significantly higher frequency of hypo- and achlorhydria in patients with serum cobalamin below 160 pmol/l although the intestinal absorption of vitamin B₁₂ was normal by a Schilling test. Although our finding of low saturation of TC-I in serum seems to demonstrate decreased vitamin B₁₂ content in the body in myelomatosis, the lack of evidence for a functional vitamin B₁₂ deficiency speaks against giving a supplement to patients with myelomatosis.