Dr. Ron’s Research Review – December 10, 2014

© 2014

This week’s research review focuses on Vitamin B12 deficiency testing.

There is no "gold standard" test for B12 deficiency.

Serum B12

Serum B12 values above the cut-off point of deficiency do not necessarily indicate adequate B12 status. As B12 deficiency occurs, serum values may be maintained while tissue B12 stores become depleted.

Intrinsic Factor Antibodies

False positives in newer assays are caused by the presence of intrinsic factor antibodies and heterophilic antibodies in the test sample. (Scarpa et al., 2013) (Carmel and Agrawal, 2012)

Methylmalonic acid (MMA) and Homocysteine

Methylmalonic acid, homocysteine and holotranscobalamin testing were not significantly better than total serum B12 concentration as predictors of a hematologic response to vitamin B12 therapy. (Goringe et al., 2006) (Carmel, 2011)

Holotranscobalamin

Holotranscobalamin, the active fraction of vitamin B12, can be measured to detect vitamin B12 deficiency. Approximately 75% of serum B12 is bound to haptocorrin (formerly called transcobalamin I) whose function is unknown. Therefore, total serum B12 levels largely reflect B12 that is not bioavailable. Total transcobalamin, consisting of holoTC and apo-transcobalamin, is the major carrier protein in the plasma/serum that delivers B12 to the tissues. (Clarke et al., 2007)

Intrinsic Factor and Parietal Cell Antibodies

Without performing Schilling's test, intrinsic factor deficiency may not be proven, and intrinsic factor and parietal cell antibodies are useful surrogate markers of PA, with 73% sensitivity and 100% specificity. PA is mainly considered a disease of the elderly, but younger patients represent about 15% of patients. PA is frequently associated with autoimmune thyroid disease (40%) and other autoimmune disorders, such as diabetes mellitus (10%). (Lahner and Annibale, 2009)

Gastric Intrinsic Factor Output

A gastric intrinsic factor output under 200 U/h after pentagastrin stimulation (N > 2000 U/h) is specific for pernicious anemia. (Cattan, 2011)

Cascade Testing

A recent article recommends multiple lab tests for vitamin B12 deficiency. “Borderline low B12 level (100-250 pg/mL, normal 185-1000); suggest serum methylmalonic acid if clinically indicated.” (Berg and Shaw, 2013)

Dr. Ron


 

Articles

Laboratory evaluation for vitamin B12 deficiency: the case for cascade testing.
            (Berg and Shaw, 2013) Download
OBJECTIVE: Potential vitamin B(12) deficiency is a common clinical diagnostic problem, and many providers have a low threshold for initiating therapy. The goal of this study was to systematically evaluate current practice patterns regarding the laboratory evaluation of suspected vitamin B(12) deficiency. METHODS: This retrospective study reviewed the electronic medical records of 192 patients initiated on intramuscular vitamin B(12) injections. RESULTS: Only 12 patients had objectively documented hematologic responses: decrease of mean corpuscular volume by >/=5 fL with stable or improved hemoglobin. Another 5 patients had equivocal hematologic responses. There was one plausible neurologic response. Thus, only 18 (9.4%) of 192 patients had data supportive of a clinical response. In these 18 patients, the baseline serum B(12) level was </=107 pg/mL; only 3 patients also had a baseline serum methylmalonic acid level, which was >/=1.29 mumol/L in all 3 patients. CONCLUSIONS: Currently, only a small minority of patients initiated on intramuscular vitamin B(12) supplementation derive any meaningful clinical benefit. Furthermore, current testing recommendations for vitamin B(12) deficiency are usually not followed. Up-front ordering of a diagnostic testing cascade is recommended to improve compliance; an example is presented with decision points chosen to improve specificity for clinically evident vitamin B(12) deficiency without loss of sensitivity. Ultimately, a better understanding of vitamin B(12) physiology is needed to develop and evaluate laboratory tests that more accurately reflect true intracellular vitamin B(12) status.

Biomarkers of cobalamin (vitamin B-12) status in the epidemiologic setting: a critical overview of context, applications, and performance characteristics of cobalamin, methylmalonic acid, and holotranscobalamin II.
            (Carmel, 2011) Download
Cobalamin deficiency is relatively common, but the great majority of cases in epidemiologic surveys have subclinical cobalamin deficiency (SCCD), not classical clinical deficiency. Because SCCD has no known clinical expression, its diagnosis depends solely on biochemical biomarkers, whose optimal application becomes crucial yet remains unsettled. This review critically examines the current diagnostic concepts, tools, and interpretations. Their exploration begins with understanding that SCCD differs from clinical deficiency not just in degree of deficiency but in fundamental pathophysiology, causes, likelihood and rate of progression, and known health risks (the causation of which by SCCD awaits proof by randomized clinical trials). Conclusions from SCCD data, therefore, often may not apply to clinical deficiency and vice versa. Although many investigators view cobalamin testing as unreliable, cobalamin, like all diagnostic biomarkers, performs satisfactorily in clinical deficiency but less well in SCCD. The lack of a diagnostic gold standard limits the ability to weigh the performance characteristics of metabolic biomarkers such as methylmalonic acid (MMA) and holotranscobalamin II, whose specificities remain incompletely defined outside their relations to each other. Variable cutoff selections affect diagnostic conclusions heavily and need to be much better rationalized. The maximization of reliability and specificity of diagnosis is far more important today than the identification of ever-earlier stages of SCCD. The limitations of all current biomarkers make the combination of >/=2 test result abnormalities, such as cobalamin and MMA, the most reliable approach to diagnosing deficiency in the research setting; reliance on one test alone courts frequent misdiagnosis. Much work remains to be done.

Failures of cobalamin assays in pernicious anemia.
            (Carmel and Agrawal, 2012) Download

Pernicious anemia: what are the actual diagnosis criteria?
            (Cattan, 2011) Download
A gastric intrinsic factor output under 200 U/h after pentagastrin stimulation (N > 2000 U/h) is specific for pernicious anemia. The other findings are either variable or non specific. Serum intrinsic factor antibodies, considered as specific in general practice, are present only in half of the patients with pernicious anemia. In their absence, since the disappearance of the Schilling tests, the gastric tubage currently used for the study of gastric acid secretion, is obligatory for the simultaneous study of intrinsic factor output. This study is important to eliminate another disease much more frequent than pernicious anemia, the protein bound to cobalamin malabsorption was observed in achlorhydric simple atrophic gastritis in the presence of intrinsic factor secretion.

Detection of vitamin B12 deficiency in older people by measuring vitamin B12 or the active fraction of vitamin B12, holotranscobalamin.
            (Clarke et al., 2007) Download
BACKGROUND: Impaired vitamin B(12) function and decreased vitamin B(12) status have been associated with neurological and cognitive impairment. Current assays analyze total vitamin B(12) concentration, only a small percentage of which is metabolically active. Concentrations of this active component, carried on holotranscobalamin (holoTC), may be of greater relevance than total vitamin B(12). METHODS: We compared the utility of serum holoTC with conventional vitamin B(12) for detection of vitamin B(12) deficiency in a population-based study of older people, using increased methylmalonic acid (MMA) concentrations as a marker of metabolic vitamin B(12) deficiency in the overall population (n = 2403) and in subsets with normal (n = 1651) and abnormal (n = 752) renal function. RESULTS: Among all participants, 6% had definite (MMA >0.75 micromol/L) and 16% had probable (MMA >0.45 micromol/L) metabolic vitamin B(12) deficiency. In receiver operating characteristic curves for detection of definite vitamin B(12) deficiency, holoTC had a greater area under the curve (AUC) compared with vitamin B(12) in all participants (0.85 vs 0.76; P <0.001) and in subsets with normal (AUC: 0.87 vs 0.79; P <0.001) and abnormal (AUC: 0.85 vs 0.74; P = 0.002) renal function. Similar findings were observed for detection of moderate vitamin B(12) deficiency. Whereas the positive predictive value for both holoTC and vitamin B(12) was greater for detection of probable than definite vitamin B(12) deficiency, both tests were associated with more false-positive than true-positive test results. CONCLUSIONS: HoloTC has a modestly superior diagnostic accuracy compared with conventional vitamin B(12) for the detection of vitamin B(12) deficiency, but neither test can be recommended to screen asymptomatic populations.

The limited value of methylmalonic acid, homocysteine and holotranscobalamin in the diagnosis of early B12 deficiency.
            (Goringe et al., 2006) Download
Treatment of B12 deficiency is important to prevent progressive neurological and/or hematologic disease but requires a secure diagnosis. The aim of this study was to evaluate second line tests of B12 status as prognostic indicators of a hematologic response to vitamin B12 therapy. Forty-nine patients referred with low, serum vitamin B12 concentrations were treated with intramuscular B12 and re-assessed after 3 months. Methylmalonic acid, homocysteine, holotranscobalamin and neutrophil hypersegmentation index were measured before and after treatment. Before treatment 27/49 patients were anemic or macrocytic of whom 15 had a clear hematologic response. All the tests had a similar prognostic accuracy. Symptomatic improvement did not correlate with hematologic response. Supplementary tests of vitamin B12 status were not significantly better than total serum B12 concentration as predictors of a hematologic response to vitamin B12 therapy.

Pernicious anemia: new insights from a gastroenterological point of view.
            (Lahner and Annibale, 2009) Download
Pernicious anemia (PA) is a macrocytic anemia that is caused by vitamin B(12) deficiency, as a result of intrinsic factor deficiency. PA is associated with atrophic body gastritis (ABG), whose diagnosis is based on histological confirmation of gastric body atrophy. Serological markers that suggest oxyntic mucosa damage are increased fasting gastrin and decreased pepsinogen I. Without performing Schilling's test, intrinsic factor deficiency may not be proven, and intrinsic factor and parietal cell antibodies are useful surrogate markers of PA, with 73% sensitivity and 100% specificity. PA is mainly considered a disease of the elderly, but younger patients represent about 15% of patients. PA patients may seek medical advice due to symptoms related to anemia, such as weakness and asthenia. Less commonly, the disease is suspected to be caused by dyspepsia. PA is frequently associated with autoimmune thyroid disease (40%) and other autoimmune disorders, such as diabetes mellitus (10%), as part of the autoimmune polyendocrine syndrome. PA is the end-stage of ABG. Long-standing Helicobacter pylori infection probably plays a role in many patients with PA, in whom the active infectious process has been gradually replaced by an autoimmune disease that terminates in a burned-out infection and the irreversible destruction of the gastric body mucosa. Human leucocyte antigen-DR genotypes suggest a role for genetic susceptibility in PA. PA patients should be managed by cobalamin replacement treatment and monitoring for onset of iron deficiency. Moreover, they should be advised about possible gastrointestinal long-term consequences, such as gastric cancer and carcinoids.

Undetected vitamin B12 deficiency due to false normal assay results.
            (Scarpa et al., 2013) Download

 


 

References

Carmel, R. and Agrawal, Y. P. (2012), ‘Failures of cobalamin assays in pernicious anemia.’, N Engl J Med, 367(4) (4), 385-86. PubMedID: 22830482
Berg, RL and GR Shaw (2013), ‘Laboratory evaluation for vitamin B12 deficiency: the case for cascade testing.’, Clin Med Res, 11 (1), 7-15. PubMedID: 23262189
Carmel, R (2011), ‘Biomarkers of cobalamin (vitamin B-12) status in the epidemiologic setting: a critical overview of context, applications, and performance characteristics of cobalamin, methylmalonic acid, and holotranscobalamin II.’, Am J Clin Nutr, 94 (1), 348S-58S. PubMedID: 21593511
Cattan, D (2011), ‘Pernicious anemia: what are the actual diagnosis criteria?’, World J Gastroenterol, 17 (4), 543-44. PubMedID: 21274387
Clarke, R, et al. (2007), ‘Detection of vitamin B12 deficiency in older people by measuring vitamin B12 or the active fraction of vitamin B12, holotranscobalamin.’, Clin Chem, 53 (5), 963-70. PubMedID: 17363419
Goringe, A, et al. (2006), ‘The limited value of methylmalonic acid, homocysteine and holotranscobalamin in the diagnosis of early B12 deficiency.’, Haematologica, 91 (2), 231-34. PubMedID: 16461308
Lahner, E and B Annibale (2009), ‘Pernicious anemia: new insights from a gastroenterological point of view.’, World J Gastroenterol, 15 (41), 5121-28. PubMedID: 19891010
Scarpa, E, et al. (2013), ‘Undetected vitamin B12 deficiency due to false normal assay results.’, Blood Transfus, 11 (4), 627-29. PubMedID: 23356970