NUTRITION RESEARCH 36 (2016) 109– 116

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Review Article

Metabolic deficiency: a missed opportunity to prevent and stroke

J. David Spence⁎

Stroke Prevention & Atherosclerosis Research Centre, Robarts Research Institute, Western University, London, ON, Canada

ARTICLE INFO ABSTRACT

Article history: The purpose of this narrative review is to highlight insights into the importance and Received 17 July 2015 frequency of metabolic vitamin B12 (B12) deficiency, reasons why it is commonly missed, Revised 16 October 2015 and reasons for the widespread but mistaken belief that treatment of B12 deficiency does Accepted 19 October 2015 not prevent stroke or improve cognitive function. Metabolic B12 deficiency is common, being present in 10%-40% of the population; is frequently missed; is easily treated; and contributes importantly to cognitive decline and stroke in older people. Measuring serum Keywords: B12 alone is not sufficient for diagnosis; it is necessary to measure holotranscobalamin or Vitamin B12 functional markers of B12 adequacy such as methylmalonic acid or plasma total Cobalamin . B-vitamin therapy with cyanocobalamin reduces the risk of stroke in Homocysteine patients with normal renal function but is harmful (perhaps because of thiocyanate Stroke accumulation from cyanide in cyanocobalamin) in patients with renal impairment. Cognitive function Methylcobalamin may be preferable in renal impairment. B12 therapy slowed gray matter Cyanide atrophy and cognitive decline in the Homocysteine and in Cognitive Renal impairment Impairment Trial. Undiagnosed metabolic B12 deficiency may be an important missed opportunity for prevention of dementia and stroke; in patients with metabolic B12 deficiency, it would be prudent to offer inexpensive and nontoxic supplements of oral B12, preferably methylcobalamin or hydroxycobalamin. Future research is needed to distinguish the effects of thiocyanate from cyanocobalamin on hydrogen sulfide, and effects of treatment with methylcobalamin on cognitive function and stroke, particularly in patients with renal failure. © 2016 Elsevier Inc. All rights reserved.

1. Introduction insights into reasons why many of the randomized trials appeared to be negative are discussed. The literature There is widespread misunderstanding of the complexity of reviewed was from reference databases of the author and of interpreting clinical trial results of vitamin B12 (B12) for Profs David Smith and Helga Refsum, who advised the author prevention of stroke and dementia. In this narrative review, on early drafts. Randomized trials, although rightly regarded

Abbreviations: B12, vitamin B12; CSPPT, China Stroke Primary Prevention Trial; GFR, glomerular filtration rate; H2S, hydrogen sulfide; MMA, methylmalonic acid; NHANES, National Health and Nutrition Examination Survey; tHcy, plasma total homocysteine; VISP, Vitamin Intervention for Stroke Prevention trial. ⁎ Stroke Prevention & Atherosclerosis Research Centre, Robarts Research Institute, University of Western Ontario, 1400 Western Rd, London, Ontario, Canada, N6G 2V4. Tel.: +1 519 931 5731; fax: +1 519 931 5737. E-mail address: [email protected]. http://dx.doi.org/10.1016/j.nutres.2015.10.003 0271-5317/© 2016 Elsevier Inc. All rights reserved. 110 NUTRITION RESEARCH 36 (2016) 109– 116

Fig. 1 – Relation of serum B12 to plasma total homocysteine. Based on data from 8832 participants in the NHANES. Polynomial and inverse regression, spline, loess fits for plasma tHcy with B 12. Solid, dotted, long dashed, and short dashed lines represent loess, cubic polynomials, cubic polynomials of the inverse covariate, and smooth splines, respectively. (Reproduced by permission of Karger from Bang H, Mazumdar M, Spence D. Tutorial in biostatistics: analyzing associations between total plasma homocysteine and B vitamins using optimal categorization and segmented regression. Neuroepidemiology. 2006;27(4):188-200).

as first-tier evidence, are blunt instruments for studying Metabolic B12 deficiency is strictly defined by elevation of vascular biology; they must be interpreted with knowledge MMA levels or by elevation of Hcy in -replete individuals of the biology of the conditions being studied. [3]. The prevalence varies markedly in different populations but is typically in the range of 10%-40% [3,9]. The majority of people with metabolic deficiency of B12 have normal or low- 2. Importance of metabolic B12 deficiency normal serum B12 concentrations [3].

Both stroke and dementia increase steeply with age. With the aging of the population, the burden of dementia and stroke, in 3. Level of serum B12 that would define both human and economic terms, is expected to increase adequacy of functional B12 markedly. A policy statement from the American Heart Association [1] predicted that stroke in the United States will Vogiatzoglou et al [10] reported in the Hordaland study that more than double by 2030, with total annual costs of stroke of levels of MMA and tHcy begin to rise already at a serum B12 of ~$240 billion. In view of recent calls for integrated approaches 350-400 pmol/L, with a much steeper rise in MMA in the to prevention of dementia [2], it is timely to address a elderly. That level coincides with the inflection point of serum potentially treatable contributory factor for both dementia B12, 400 pmol/L, below which tHcy levels began to rise in the and stroke: inadequate B12 status. Underdiagnosis of this National Health and Nutrition Examination Survey (NHANES) condition results largely from failure to understand that a [11] (Fig. 1). A more complex relationship between serum B12 normal serum B12 may not reflect an adequate functional B12 and MMA has recently been described in the NHANES cohort, status [3,4]. Only ~25% of serum total B12 is in the active form with 33% of those studied having serum B12 between 126 and of holotranscobalamin [5]. Thus, total serum B12 determina- 287 pmol/L and so considered intermediate between deficien- tion is not sensitive for B12 deficiency. A potentially more cy and sufficiency [12]. Both these large cohorts indicate that accurate way to diagnose functional adequacy of B12 is to B12 sufficiency is first obtained with at least 300-350 pmol/L in measure holotranscobalamin [5–7] or to use functional or serum B12. In the zone below 300-400 pmol/L but within the metabolic markers of B12 such as plasma methylmalonic acid normal range of serum B12, metabolic B12 deficiency is (MMA) or total homocysteine (tHcy) [3,8]. common but is seldom accompanied by the classical markers NUTRITION RESEARCH 36 (2016) 109– 116 111

present in 9% of the patients. Another 44% belonged in the borderline zone (B12 of 160-299 pmol/L) where metabolic testing with MMA, tHcy, and/or holotranscobalamin would be useful. Only 26% had B12 greater than 400 pmol/L, that is, levels that are associated with optimally low levels of tHcy and MMA.

5. Causes of metabolic B12 deficiency

There are many ways for B12 status to become inadequate besides deficiency of intrinsic factor and gastric acid. The acquired causes increase with age, and this probably accounts for the finding that higher doses of oral B12 are needed to achieve adequate absorption in the elderly [14]. Among elderly patients with low serum B12 levels (≤221 pmol/L) and elevated MMA, peroral doses of up to 1000 μgofcyanocobalaminwererequired to achieve normal MMA levels [14]. An unexpected potential cause of inadequacy of B12 is high circulating concentrations of folate, which have been associated with increased levels of tHcy and MMA and lower levels of holotranscobalamin in individuals Fig. 2 – Serum B12 by age in patients at the Stroke Prevention with poor B12 status [15,16]. Several reports indicate that this Clinic at University Hospital, London, Ontario, Canada. imbalance may lead to an increased risk of anemia and/or Among 3025 patients referred for secondary stroke cognitive impairment [17–19]. prevention, 281 (9%) had a serum B12 <160 pmol/L (the lower limit of the reference range for the hospital laboratory), and 1317 (44%) had a B12 level in the range of 160-299 pmol/L that 6. Metabolic B12 deficiency and cognitive decline requires further testing such as MMA or tHcy to assess metabolic adequacy of B12. Only 797 (26%) had clearly Metabolic B12 deficiency is also important in cognitive decline. adequate levels of serum B12 >400 pmol/L associated with Approximately half of patients presenting with megaloblastic optimal function of B12. anemia will have cognitive impairment [20,21],butthis association is not limited to those with frank B12 deficiency. In community-dwelling elderly, the risk of cognitive impair- ment extends across the normal range of holotranscobalamin of neuropathy or megaloblastic anemia [3,4]. A definition of [22,23], and many prospective studies show that raised tHcy is a metabolic B12 deficiency cannot be precise because the cutoff strong risk factor for dementia [24]. Patients with serum B12 in values for tHcy and for MMA vary according to the population the lower third of the normal range or with tHcy in the upper being investigated, most notably with age. Furthermore, third are >4 times more likely to have confirmed Alzheimer the levels of these markers are often elevated in renal disease. disease [25]. Serum B12 across the normal range, or its markers In adults, MMA values greater than 260-360 nmol/L, and MMA and tHcy are strongly related to brain atrophy [26,27] and values for tHcy greater than 12-15 μmol/L in folate-replete to white matter lesions on magnetic resonance imaging [28].A individuals, are indicative of metabolic deficiency. Renal recent meta-analysis claiming no apparent benefit on cognitive impairment would interfere with interpretation of these aging with B-vitamin treatment [29] was based on studies in cutoffs. The higher cutoff relates to older people. The which participants were not selected for inadequate B12 status limitations of published cutoffs are discussed in detail in a and so highlights the importance of identifying patients with recent review [8]. metabolic B12 deficiency. The Homocysteine and B Vitamins in Cognitive Impair- ment Trial randomized elderly participants with mild cogni- 4. Frequency of metabolic B12 deficiency tive impairment to B-vitamin therapy vs placebo. Higher among vascular patients levels of tHcy at baseline were associated with faster gray matter atrophy and cognitive decline in the placebo group, In vascular patients, evidence of metabolic B12 deficiency is but these effects were greatly reduced by B-vitamin treatment common and increases with age. For instance, in a Canadian [30,31]. In particular, the beneficial effect of B vitamins was secondary stroke prevention clinic population, metabolic B12 confined to participants with high tHcy (greater than the deficiency was present in 12.5% of those younger than 71 median, 11 μmol/L). In those participants, a Bayesian network years and in 30% of those older than 71 years [13]. Fig. 2 shows analysis revealed a causal pathway linking B12 administra- the frequency of serum B12 levels by age in the clinic tion to lowering homocysteine, which in turn decreases gray population. Levels less than 160 pmol/L (the lower reference matter atrophy, thereby slowing cognitive decline. It is limit of for that laboratory), that is, B12 deficiency, were notable that the administration of B12 almost abolished the 112 NUTRITION RESEARCH 36 (2016) 109– 116

Fig. 3 – B-vitamin therapy reduces loss of gray matter (GM) in Alzheimer disease–related regions of the brain in elderly individuals with mild cognitive impairment. B-vitamin treatment was more effective in participants with higher tHcy levels (P <.05FWE corrected). (A) Brain regions in green demonstrate where B-vitamin treatment significantly reduces GM loss in participants with high tHcy levels (>11.06 μmol/L) at baseline. (B) Percentage of GM loss for each of the 77 participants with high tHcy level: the placebo group (n = 35) showed an average loss of 5.2% (±3.4) of GM volume over 2 years, whereas the B-vitamin group (n =42) showed an average loss of 0.6% (±2.1). (Reproduced by permission of the National Academy of Sciences of the United States of America, from: Douaud G, Refsum H, de Jager CA, Jacoby R, Nichols TE, Smith SM et al. Preventing Alzheimer's disease–related gray matter atrophy by B-vitamin treatment. Proc Natl Acad Sci U S A. 2013;110:9523-8).

atrophy of brain regions specifically linked to the Alzheimer contributor to stroke risk [35,36]. B12 deficiency raises levels of disease process [32] (Fig. 3). tHcy, increasing carotid plaque [37]; the risk of deep vein thrombosis, retinal vein thrombosis, and cerebral vein throm- bosis; as well as stroke risk in patients with atrial fibrillation [36]. High levels of tHcy quadruple the risk of stroke in patients 7. Metabolic B12 deficiency and vascular disease with atrial fibrillation [38]. In addition to elevation of tHcy, another possible role of B12 An important aspect of preventing dementia is the prevention deficiency in vascular disease has recently come to light. of stroke [33,34], and B12 deficiency is an easily treated Deficiency of B12 has been shown to impair bone metabolism NUTRITION RESEARCH 36 (2016) 109– 116 113 through reduced levels of taurine [39]; taurine is thought to have important effects in reducing atherosclerosis [40] and 9. How could B vitamins cause harm in cardiovascular disease [41,42] and has neuroprotective effects persons with renal failure? in animal models of stroke [43]. Possible mechanisms for toxicity of high-dose B vitamins in patients with renal failure were discussed by Spence and 8. B12 deficiency, elevated tHcy, and stroke Stampfer [61]. One possibility relates to unmetabolized folic acid that has not been converted to tetrahydrofolate [17]; this In the Framingham study, the proportion of strokes due to may interact with metabolic B12 deficiency via the “ ” atrial fibrillation increased steeply with age, from 1.5% at age methylfolate trap, in which B12 deficiency prevents metab- 50 years to 23% at age 80-89 years [44]. Among patients olism of 5-methyltetrahydrofolate [62,63]. Another relates to attending the Stroke Prevention Clinic at University Hospital cyanide from cyanocobalamin. Koyama et al [64] showed that in London, Ontario, stroke subtypes changed markedly with dialysis patients accumulate thiocyanate from the cyanide in improved treatment of dyslipidemia: the proportion of cyanocobalamin; they also showed in dialysis patients [65] strokes identified as cardioembolic increased from 26% in that methylcobalamin lowers levels of both tHcy and asym- 2002 to 56% in 2012 [45]. In that same population, the metric dimethylarginine. In contrast, Løland et al [66] found prevalence of tHcy >14 increased from 20% at age 35 years to that cyanocobalamin did not lower levels of asymmetric 40% above age 80 years [46]. Plasma levels of tHcy are dimethylarginine in the Western Norway B Vitamin Interven- significantly higher among patients with paradoxical embo- tion Trial. (Similarly, hydroxycobalamin, but not cyanocobal- lism [47] and among those patients with asymptomatic amin, is effective in alcohol/tobacco amblyopia, a condition in carotid stenosis who have microemboli on transcranial which cyanide plays a key role [67]). Therefore, in patients Doppler [48,49]. with renal failure, methylcobalamin may be superior to Although it is commonly claimed that B-vitamin therapy cyanocobalamin [68]. A possible connection between cyanide to lower plasma levels of tHcy does not prevent stroke, the and vascular disease is that cyanide consumes hydrogen issue is more complicated than originally thought. The sulfide in its elimination as thiocyanate, and hydrogen sulfide recent large China Stroke Primary Prevention Trial (CSPPT) is a recently recognized endothelium-derived relaxing factor, showed significant reduction of stroke with folic acid [50]; analogous to nitric oxide [69]. Thiocyanate also catalyzes Stampfer and Willett [51] pointed out that folate supple- oxidation of low-density lipoprotein [70]. ments were particularly effective in participants with low The hypothesis that it was not folate but rather the baseline serum folate. Importantly, folate fortification does cyanocobalamin that accounted for the adverse outcomes in not exist in China. The Vitamin Intervention for Stroke the Diabetic Intervention with Vitamins in Nephropathy trial Prevention trial (VISP) trial [52] was initiated at the same time and negated the benefit of folate/B6/cyanocobalamin in the as folate fortification in North America and used only 400 μg VISP trial is supported by the recent observation that, in the daily of cyanocobalamin, a dose that is not sufficient for CSPPT, folate was not harmful in participants with estimated elderly persons with low serum B12 [14]. In folate-replete GFR less than 60. To the contrary, folate slowed the decline of persons, the key nutritional determinant of elevated tHcy is renal function in the CSPPT [71]. B12 [53,54]; in North America and other jurisdictions where folate fortification pertains, B12 is now the main dietary determinant of tHcy [37]. There are 3 particular factors that 10. Beyond dementia and stroke may interfere with the potential beneficial effect of homo- cysteine-lowering therapy: pretreatment B12 status, the Besides increasing the risk of stroke and dementia, function- concomitant use of antiplatelet drugs, and renal failure. al B12 deficiency causes neuropathy and myelopathy; it is Each of these factors is discussed below. likely also that loss of position sense from myelopathy Vitamin therapy to lower homocysteine reduced the risk contributes to the risk of falls in the elderly. B12 deficiency of stroke in the Heart Outcomes Prevention Evaluation trial is also associated with orthostatic hypotension (possibly due [55], in the SUFOLOM3 study in France [56], in participants to autonomic neuropathy), another cause of falls in the not taking antiplatelet agents in the VITATOPS study [57], elderly [72]. Raised tHcy, for which the main nutritional cause and in a subgroup of the VISP study [58].Thatsubgroup in countries with folic acid fortification is B12 deficiency [37], excluded patients with renal impairment (a glomerular also increases the risk of deep vein thrombosis, pulmonary filtration rate [GFR] <47 mL/min; the lowest decile) and embolism, retinal vein thrombosis, and thrombosis of the patients who received injections of B12 (regardless of the cerebral venous sinuses [36]. treatment arm to which they were assigned). The signifi- cance of poor renal function became evident in the Diabetic Intervention with Vitamins in Nephropathy trial; among patients with diabetic nephropathy, high doses of B vita- 11. Treatment of B12 deficiency mins (folic acid 2.5mg, pyridoxine 25 mg, and cyanocobal- amin 1000 μg daily) accelerated the decline of GFR and It should be noted that B12 supplementation for the doubled the risk of vascular events [59].Inthatstudy,allthe population should not be implemented until there is evi- vascular events occurred in patients with a GFR <50 mL/min/ dence from trials of benefit but no harm; however, in 1.73 m2 [60]. individual patients, it is important to improve detection and 114 NUTRITION RESEARCH 36 (2016) 109– 116 treatment of metabolic B12 deficiency, which should not be regarded as “subclinical.” Disclosures Most B12 deficiency is due to malabsorption of B12, but adequate absorption of B12 can be achieved with high doses of None relevant to the topic of this article. There was no oral B12 in most patients, even in the absence of intrinsic factor funding. and the other factors required for B12 absorption [73].Van Walraven et al [74] calculated that switching from injections to oral B12 could substantially reduce the cost of B12 therapy. Acknowledgment For elderly patients with low serum B12 levels, high doses of oral B12, as much as 1000 μg daily, may be needed [75]. This will The author is indebted to A. David Smith, University of probably depend on the cause of the B12 deficiency, that is, Oxford, and Helga Refsum, Oxford University and University whether it is related to pernicious anemia (lack of intrinsic factor) of Norway, for important revisions of the manuscript. or, more commonly, failure to release protein-bound B12 due to lack of gastric acid. The former may require very high peroral doses or injection therapy; the latter can usually suffice with REFERENCES lower doses provided as supplement. 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