Association Between Homocysteine and Vitamin Levels in Demented Patients

Abstract

Background:

Although it is known that the nutritional status among elderly persons and, in particular, patients with dementia, is compromised, malnutrition that results in insufficient uptake of several vitamins is often not diagnosed.

Objective:

An elevated homocysteine level is a known strong risk factor for vascular dementia (VaD) and Alzheimer’s disease (AD). Several B vitamins are involved in the metabolism of homocysteine. Therefore, we investigated the serum levels of vitamin B1, vitamin B6, folate, and vitamin B12 in 97 patients with mild cognitive impairment (MCI) or different forms of dementia and 54 elderly control persons without dementia.

Results:

Compared to aged non-demented people, vitamins B1, B6, B12, and folate were decreased in serum of patients with AD, and patients with Lewy body dementia had reduced vitamin B12 level. Vitamin B6 was diminished in VaD. Patients with frontotemporal dementia showed no alterations in vitamin levels. Age was identified as an important factor contributing to the concentrations of vitamin B1 and B6 in serum, but not vitamin B12 and folate. Increased levels of total homocysteine were detected especially in MCI and AD. Homocysteine correlated negatively with levels of vitamins B6, B12, and folate and positively with Q Albumin.

Conclusion:

Our data suggest that despite increased homocysteine already present in MCI, vitamin levels are decreased only in dementia. We propose to determine the vitamin levels in patients with cognitive decline, but also elderly people in general, and recommend supplementing these nutrients if needed.

Keywords

B vitamins dementia homocysteine

INTRODUCTION

Alzheimer’s disorder (AD) is the most common form of dementia. The pathophysiological hallmarks of this disorder are changes in amyloid-β protein precursor (AβPP) cleavage and production of the AβPP fragment amyloid-β (Aβ), resulting in formation of plaques, along with hyperphosphorylated tau protein aggregation, leading to the formation of neurofibrillary tangles [1]. This results in cognitive impairment seen in this disorder [2]. A cerebral microangiopathy or a cerebral infarction can often lead to vascular dementia (VaD). Frontotemporal dementia (FTD) is characterized by progressive alterations in behavior, affective symptoms dysfunction, and language impairment [3]. Although this disorder is genetically heterogenous, in some cases a common pathophysiological characteristic between FTD and AD is described: the intraneuronal deposition of the protein tau. Patients with Lewy body dementia (LBD) present visual hallucinations, extrapyramidal motor symptoms, and a fluctuating cognitive impairment. Histopathological hallmarks of LBD are the Lewy bodies and Lewy neuritis composed of alpha-synuclein [4].

It is still under debate whether the metabolic syndrome influences the risk for the development of dementia or the conversion from mild cognitive impairment (MCI) to dementia [5 –7]. Although each factor of the metabolic syndrome including abdominal obesity [8, 9], hypertension [10], dyslipidemia [11, 12], and hyperglycemia [13], had been individually associated with dementia, it was shown that a metabolic syndrome in elderly patients was associated with a lower risk of AD [14]. Moreover, abdominal obesity in older people was associated with a lower risk of overall dementia [15]. On the other hand, the nutritional status among older adults might be compromised, with an adequate uptake of daily calories but a decreased uptake of essential vitamins, or in general with a decreased uptake of energy and vitamins, mounting in the risk or even the presence of malnutrition [16]. Malnutrition is associated with cognitive impairment among elderly people [17, 18]. The cause of malnutrition is usually multifactorial the result of loss of appetite, poor food intake, pain, and acute gastrointestinal symptoms [19]. Consequently, the intake of essential vitamins is decreased and low serum concentrations of B vitamins and vitamin C and accumulation of homocysteine [20] were associated with an increased risk of AD [21 –23]. The B vitamins folate and vitamin B12 play an important role in cognitive function. Insufficient intake of folate contributes to neurological deficits including cognitive impairment and is associated with poor cognitive function [24], and also vitamin B12 deficiency is associated with cognitive decline [22, 25]. Vitamin B1 (thiamine) deficiency can cause neurological deficits similar to AD, since thiamine dependent enzymes decline as they are actually important for the glucose metabolism in brain [26].

Homocysteine is a product of the methionine metabolism. High homocysteine levels correlate with low levels of folate, B12, B6 levels [27]. The B vitamins folate, B12, and B6 directly participate in the metabolism of homocysteine. Elevated homocysteine levels are a risk factor for disorders like AD, MCI, and VaD [28, 29]. Interestingly, homocysteine is an antagonist of the N-methyl-D-aspartate receptor (NMDA-R) which is relevant for the memory function of patients with AD.

Despite these circumstances, the supplementation of specific vitamins is neither clinically established nor routinely practiced in the treatment of patients with dementia. In our study, we wanted to determine the serum levels of vitamins B1, B6, B12, and folate that are involved in metabolism of homocysteine in patients recently diagnosed with AD, VaD, FTD, or LBD, but also patients with MCI.

MATERIALS AND METHODS

Study population

The study was performed in accordance with German laws, the Declaration of Helsinki and the guidelines of the local institutional review board. We included patients that were admitted at the Department of Psychiatry and Psychotherapy, University of Magdeburg, between 2012 and 2015. Diagnosis of dementia was performed by psychiatrists, neurologists and psychologists on the gerontopsychiatric ward and gerontopsychiatric ambulance based on international guidelines [30 –32]. Dementia diagnosis was performed according to the DSM-IV criteria and based on the results obtained by magnetic resonance imaging (MRI) of the brain or cerebral computer tomography (CT), cerebrospinal fluid (CSF) analysis, Mini-Mental State Evaluation (MMSE), and electroencephalography (EEG). Clinicians had access to detailed clinical files, including the medical histories by proxy and referral letters from the general practitioners and routine blood analysis (including differential blood cell count, levels of C-reactive protein, glucose, lipids, liver enzymes, and thyroid hormones).

20 MCI patients (MMSE: 25-26; 61–92 years, mean age 79 years; 15 females, 5 males), 9 FTD patients (55–83 years; mean age 77 years; 3 females, 6 males), 26 VaD patients (55–96 years; mean age 78 years; 14 females, 12 males), and 42 AD patients (65–95 years; mean age 79 years; 27 females, 15 males). Demographic data of the study cohort, including body mass index (BMI) and blood values of hemoglobin, ferritin, and creatinine were given in Table 1A. These patients were further subdivided according to the results obtained in MMSE into mild AD (MMSE: 24-20; 15 patients; 65–95 years; mean age 78 years; 11 females, 4 males), moderate AD (MMSE: 10–19; 17 patients; 67–90 years; mean age 80 years; 10 females, 7 males), and severe AD (MMSE: 0–9; 10 patients; 70–92 years; mean age 79 years; 6 females, 4 males). CSF characteristics including phospho-tau (ptau), whole tau (htau), Amyloid-β1-40, Amyloid-β1-42, Amyloid-β ratio, and quotient of serum/CSF albumin (Q Albumin) of the patients participating in the study are provided in Table 1B.

Table 1A

Demographic data of study cohort. The number, mean age, sex, hemoglobin, ferritin, and creatinine level in serum and body mass index (BMI) of the study cohort, neuropsychiatric healthy elderly volunteers (controls), AD patients, further subdivided according to the severity of the disorder, VaD, FTD, and LBD patients are presented in Table 1A. Overview of markers obtained by analyzing CSF, including htau, ptau, Amyloid-β 1–40, Amyloid-β 1–42, Amyloid-β ratio, and Q Albumin in the patients suffering from AD, VaD, FTD, and LBD and AD stages as well as p-values were shown in Table 1B. Kruskal-Wallis and Fisher's exact test were used to calculated differences between control subjects and patients

Characteristics	reference values		controls	VaD	FTD	LBD	AD	p	MCI	AD			p
	male	female					total			mild	moderate	severe
total (n)			54	26	9	9	42		20	15	17	10
age (years; mean)			72.00	78.19	71.56	76.5	78.83	0.0002	78.75	77.8	79.59	79.1	0.0009
sex (female/male)			41 / 13	14 / 12	3 / 6	1 / 5	27 / 15	0.0082	15 / 5	11 / 4	10 / 7	6 / 4	0.7197
hemoglobin (mmol/L)	8.7–10.9	7.5–9.6	8.308	7.988	8.238	8.583	8.244	0.66	7.942	8.193	8.200	8.411	0.5938
ferritin (μg/L)	4–665	13–651	231.6	277.6	331.3	266.7	267.1	0.2088	205.3	403.8	224.9	102.0	0.2155
creatinine (μmol/L)	53.1–106.2	44.3–88.2	75.76	89.64	75.00	74.67	87.66	0.3884	81.47	92.13	91.47	73.00	0.2714
BMI			26.16	28.12	25.11	24.4	25.57	0.5091	23.29	23.01	24.15	20.28	0.3778

Table 1B

Table1B

CSF biomarker	VaD	FTD	LBD	AD	p	MCI	AD			p
				total			mild	moderate	severe
ptau (mean)	39.71	60.80	62.50	95.64	0.0001	43.15	96.64	90.77	103.8	0.0001
htau (mean)	361.5	362.2	395.3	642.9	0.0002	271.2	560.6	619.4	872.2	< 0.0001
Amyloid-β1–40 (mean)	7503	8741	8101	10917	0.1178	8068	10358	11192	11625	0.2658
Amyloid-β1–42 (mean)	623.5	723.5	660.5	630.8	0.4417	799.8	617.2	720.3	468.5	0.0653
Amyloid-β ratio (mean)	0.914	1.000	0.900	0.609	0.0025	1.123	0.657	0.631	0.450	0.0032
Q Albumin (mean)	10.48	9.940	13.23	9.782	0.2351	8.700	9.671	9.333	10.77	0.5402

54 elderly control persons without dementia (43–89 years; mean age 72 years; 41 females, 13 males) participated in the study and donated blood. Some of these persons were disease control individuals, they suffered from other psychiatric disorders such as depressive disorder, somatic symptom disorder or personality disorder, that did not impair their cognitive abilities.

Blood was drawn after about 12 h fasting at 7 a.m. and immediately transported to the institute for clinical chemistry and pathobiochemistry of our university hospital, where all analysis were performed. The levels of vitamin B1 was measured in whole blood, pyridoxal-5’-phosphate (PLP) as indicator for the status of vitamin B6 in plasma, total homocysteine, folate, and total B12 were measured in stored serum and plasma samples using commercial electrochemiluminescence immunoassays (ECLIA) from Roche Diagnostics that were run on a COBAS 8000 e 602 analyzer (Roche Diagnostics, Switzerland).

Patients with a history of immune disease, immunomodulating treatment, cancer, chronic terminal disease, substance abuse, and severe trauma or clinical/paraclinical findings indicating these disorders were excluded.

Statistical analysis

Data were first analyzed using the Shapiro-Wilk normality test (the obtained results were shown in Supplementary Figures 1 and 2). To determine differences between the indicated groups, Kruskal-Wallis test, followed by post-hoc tests (Bonferroni or Dunn's multiple comparison test), and Spearman correlation were performed. Analysis of covariance was performed using aligned rank transformation of data (ART; http://www.r-project.org, nonparametric). Significance was defined as ^*p < 0.05, ^**p < 0.005, or ^***p < 0.001.

RESULTS

Deficiency of vitamin B1, B6, B12, and folate in patients with different forms of dementia

Reference values for vitamin B1 is 28–85 mg/dL, for vitamin B6 > 4.5 mg/dL, for vitamin B12 197–771 ng/L, and for folate 3.89–26.8 ng/mL.

Compared to non-demented elderly persons, the levels of vitamin B1 (median 46 mg/dL; p = 0.0009), vitamin B6 (median 6.4 mg/dL; p = 0.0496), vitamin B12 (median 248 ng/L; p = 0.0006), and folate (median 6.55 ng/mL; p = 0.0281) were decreased in AD patients.

Patients suffering from VaD showed a 40% reduction in vitamin B6 level (median 4.5 mg/dL; p =0.0053) compared to the control group (Supplementary Figure 3); all other vitamins were unaltered. Compared to the control persons, vitamin B12 (median 325 mg/dL; p = 0.0207) was reduced in LBD patients. In FTD patients, no alterations in blood vitamin levels were detected (Table 2A, B). In order to determine whether the patients’ sex influences the vitamin level, we investigated the level of vitamins in females and males separately. However, no differences were found between any analyzed vitamin and sex (Table 3A). Moreover, we analyzed whether the sex of the non-demented controls influenced the vitamin level. We found no undersupply of vitamin B1, but more than 20% of controls had less vitamin B6 in serum than the normal range (Table 3B), mostly attributed to women (> 90%). Vitamin B12 and folate were decreased in 5.7% of control individuals and also these vitamins were associated with female sex.

Table 2A

Influence of age and BMI on the level of homocysteine and vitamin level in serum in dementia. Given are the median values of homocysteine, vitamin B1, B6, B12, and folate in serum of controls and patients with AD, VaD, FTD, and LBD (A) and the results of the Kruskal-Wallis test and analysis of covariance using aligned rank transformation of data (ART; nonparametric) without and with age included as single covariate to determine diagnosis-related differences of vitamins; given are the p-values. In (B) are given the p-value results of the Dunn’s multiple comparisons test as post-hoc test following the Kruskal-Wallis test between controls and patients with AD, VaD, FTD, and LBD. No significant differences were obtained between AD, VaD, FTD, and LBD patients, therefore these p-values were not shown

groups (median values)					Kruskal-Wallis Test	ART analysis
variables	controls	AD	VaD	FTD	LBD	p	p	p (covariate age)
vitamin B1	54.50	46.00	47.00	47.00	53.00	0.077	0.825	< 0.001
vitamin B6	7.500	6.400	4.500	5.900	4.250	0.073	0.641	< 0.001
folate	8.700	6.550	7.900	6.700	5.950	0.021	0.029	0.379
vitamin B12	415.0	248.0	325.0	262.5	223.5	< 0.001	0.003	0.082
homocysteine	16.45	23.00	21.20	15.35	23.60	0.224	0.392	0.595

Table 2B

Table2B

KW	controls versus
variables	p	AD	VaD	FTD	LBD
vitamin B1	0.077	0.0009	0.1845	0.4744	0.8968
vitamin B6	0.073	0.0496	0.0053	0.6421	0.0587
folate	0.021	0.0281	0.4896	0.2796	0.0773
vitamin B12	< 0.001	0.0006	0.0535	0.0549	0.0207
homocysteine	0.224	0.0399	0.2613	0.7033	0.1390

Table 3A

Sex-associated differences in vitamin levels. Given are the mean serum level of vitamin B1, B6, folate, and B12, separated by sex and the p-value comparing male and female study participants, analyzed by Mann-Whitney test

	female	male
	mean	mean	p
B1	51.50	54.77	0.0947
B6	8.778	9.077	0.6637
folate	8.134	7.369	0.7977
B12	338.6	325.1	0.7106
homocysteine	22.59	22.19	0.4398

Table 3B

Table3B

	decreased level
vitamin	total	female	male
B1	0	0	0
B6	22%	92%	8%
B12	5.7%	100%	0
folate	5.7%	66.6%	33.3%

Vitamin levels in MCI and AD stages

MCI is associated with an increased risk to develop AD. According to severity, AD is subdivided into mild, moderate, and severe AD. In MCI patients, decreased vitamin B6 (median 4.2 mg/dL; p = 0.0342) and folate (median 4.5 ng/mL; p = 0.0225) were detected.

Vitamin B1 was reduced in mild AD (median 45.00 mg/dL; p = 0.0217). The vitamin B12 level was decreased in mild (median 272.0 mg/dL; p = 0.0442), moderate (median 223.0 mg/dL; p = 0.0079), and severe AD (median 243.0 mg/dL; p = 0.0325). The level of vitamin B6 and folate was not dependent on AD stage (Table 4A, B).

Table 4A

Influence of age and BMI on the level of homocysteine and vitamin level in serum in MCI and AD stages. Given are the median values of homocysteine, vitamin B1, B6, B12, and folate in serum of controls and patients with MCI, mild, moderate, and severe AD (B) and the results of the Kruskal-Wallis test and analysis of covariance using aligned rank transformation of data (ART; nonparametric) without and with age included as single covariate to determine diagnosis-related differences of vitamins; given are the p-values. In (B) are given the p-value results of the Dunn’s multiple comparisons test as post-hoc test following the Kruskal-Wallis test between controls and patients with MCI, mild, moderate, and severe AD. No significant differences were obtained between MCI, mild, moderate, and severe AD patients, therefore these p-values were not shown

	groups (median values)					Kruskal-Wallis Test	ART analysis
variables	controls	MCI	mild AD	moderate AD	severe AD	p	p	p (covariate age)
vitamin B1	54.50	53.00	45.00	52.00	43.00	0.109	0.661	0.007
vitamin B6	7.500	4.200	6.600	6.400	5.500	0.295	0.748	0.197
folate	8.700	6.300	6.300	7.000	6.700	0.020	0.037	0.774
vitamin B12	415.0	255.0	272.0	223.0	243.0	0.004	0.014	0.179
homocysteine	16.45	25.20	24.10	20.80	28.50	0.035	0.055	0.170

Table 4B

Table4B

KW	controls versus
variables	p	MCI	mild AD	moderate AD	severe AD
vitamin B1	0.109	0.0877	0.0217	0.3441	0.0682
vitamin B6	0.295	0.0342	0.1452	0.5229	0.0556
folate	0.020	0.0225	0.1499	0.3441	0.0682
vitamin B12	0.004	0.0900	0.0442	0.0079	0.0325
homocysteine	0.035	0.0066	0.0322	0.1035	0.0013

Homocysteine level in demented patients

The total homocysteine level was, compared to that present in the control individuals (median 16.45μmol/L), increased in all forms of dementia, but only significant in AD (median 23μmol/L; p = 0.0399; Table 2A, B), in particular in patients suffering from MCI (median 25.2μmol/L; p = 0.0066), mild (median 24.10μmol/L; p = 0.0322), and severe AD (median 28.5μmol/L; p = 0.0013, Table 4A, B).

Influence of age and BMI on vitamin levels

Since age varied between the non-demented control individuals and patients with AD and VaD (Table 1), the correlation between age and vitamin level was calculated. In AD patients, and in particular patients with mild AD, increasing age was correlated with diminished vitamin B1 (AD total: r = –0.380; p = 0.019; mild AD: r = –0.680; p = 0.011) and B6 level (AD: r = –0.329; p = 0.044; mild AD: r = –0.608; p = 0.027; Table 5A). However, we determined also an age-dependent influence of the B1 level in the control group. Age did not influence the level of folate and vitamin B12 in any group. Moreover, we performed an analysis of covariance using aligned rank transformation of data (ART; nonparametric) including age as single covariate to determine diagnosis-related differences of vitamins (Table 2) [33, 34]. We determined that vitamins B1 and B6 strongly dependent on the age of the study participants. Vitamin B12 and folate were significantly different using Krukal-Wallis and ART analysis between patients suffering from AD, VaD, FTD, and LBD and non-demented persons, but were not influenced by age (Table 2A).

Table 5A

Associations between age and BMI with vitamin level. The correlation between age (A) and BMI (B) with vitamin B1, B6, folate, and B12 in non-demented volunteers and patients with MCI, AD (total, mild, moderate, severe), VaD, FTD, and LBD were determined using are the Spearman correlation, r and the p-value (bold = significant value) are given

age	controls	MCI	AD	AD			VaD	FTD	LBD
vitamin			total	mild	moderate	severe
B1	r	–0.296	–0.198	–0.380	–0.680	–0.202	0.458	–0.140	–0.072	0.203
	p	0.030	0.478	0.019	0.011	0.454	0.219	0.556	0.906	0.714
B6	r	–0.136	0.346	–0.329	–0.608	–0.305	–0.322	–0.390	–0.679	0.600
	p	0.328	0.173	0.044	0.027	0.251	0.385	0.076	0.110	0.242
folate	r	–0.011	0.284	–0.187	–0.046	–0.479	0.433	0.360	–0.086	–0.054
	p	0.939	0.268	0.255	0.876	0.061	0.250	0.444	0.919	0.807
B12	r	–0.194	–0.114	–0.165	–0.224	–0.001	–0.033	–0.200	–0.543	–0.053
	p	0.164	0.663	0.314	0.441	0.996	0.948	0.714	0.297	0.809

In addition, among patients with MCI or different stages of AD, the level of vitamin B1 was strongly age dependent. Levels of homocysteine, vitamin B12, and folate were different between patients and controls and not age-dependent influenced (Table 4A).

A marker for the nutritional status of a person is the body mass index (BMI). No differences in BMI were detected among patients and the control group and also no correlation was determined between the level of vitamins and BMI (Table 5B).

Table 5B

Table5B

BMI	controls	MCI	AD	AD			VaD	FTD	LBD
vitamin			total	mild	moderate	severe
B1	r	0.106	0.230	0.272	0.542	0.275	–0.234	0.194	–0.086	–0.564
	p	0.665	0.914	0.120	0.056	0.362	0.595	0.506	0.919	0.506
B6	r	0.151	0.233	0.098	–0.129	0.470	0.270	–0.714	–0.500	–0.437
	p	0.538	0.368	0.582	0.674	0.105	0.556	0.136	0.450	0.118
folate	r	0.083	–0.293	0.175	–0.187	0.491	0.464	0.075	0.464	0
	p	0.737	0.254	0.315	0.523	0.088	0.302	0.799	0.356	1.000
B12	r	0.266	–0.206	–0.084	–0.205	0.080	0.036	–0.084	0	–0.600
	p	0.271	0.428	0.632	0.483	0.795	0.963	0.776	1.000	0.350

Correlation between vitamins and Q Albumin and homocysteine levels

In order to find out whether the vitamins influence homocysteine levels, we correlated the serum levels of vitamin B1, B6, B12, and folate with the corresponding homocysteine level and detected correlation between vitamin B6 (r = –0.3664; p = 0.0089), vitamin B12 (r = –0.4007; p = 0.0039), and folate (r = –0.5653; p < 0.0001) with homocysteine level (Table 6A).

Table 6A

Associations with homocysteine. Spearman correlation analysis was performed to identify associations between serum homocysteine and the levels of the serum levels of the vitamins B1, B6, folate, and B12 or Q Albumin (A). Spearman analysis were also used to determine correlations between hemoglobin, ferritin, and creatinine and the level of vitamin B1, B6, folate, and B12 (B). Given are the Spearman r and the p-value

Parameter	vitamin B1	vitamin B6	vitamin B12	folate	Q Albumin
Spearman r	–0.2790	–0.3664	–0.4007	–0.5653	0.4940
p (two-tailed)	0.0522	0.0089	0.0039	<0.0001	0.0228
p summary	ns	^**	^**	^***	^*

Q Albumin (quotient of albumin in the cerebrospinal fluid and serum) is a marker for the integrity of the blood-cerebrospinal fluid-barrier (B-CSF-B). We detected a positive correlation between Q Albumin and the homocysteine level (r = 0.4940; p = 0.0228; Table 6A).

Correlation between vitamins and homocysteine with hemoglobin, ferritin and creatinine and multivariate analysis

Decreased level of vitamins, in particular B12 and folate, could also be associated with anemia or renal impairment. Therefore, we correlated the vitamin level with hemoglobin and ferritin as markers for anemia and creatinine as marker for renal function. The hemoglobin level was positively correlated with the vitamins B1 (p = 0.0121) and B6 (p = 0.0318), but not with B12 or folate, ferritin was not associated with any B vitamin (Table 6B). Creatinine was correlated with vitamin B6 (p = 0.0031), B12 (p = 0.0249), and homocysteine (p = 0.0011; Table 6B). Furthermore, there was a correlation between homocysteine and Q Albumin, with creatinine as confounding factor (r = 0.370; p = 0.036). Finally, we conducted a multivariate analysis to determine independent predictors of homocysteine and included the vitamins B1, B6, B12, and folate, creatinine, the diagnosis “dementia”, age and sex. We found that only folate was significant and thereby an independent predictor of homocysteine (Table 6C).

Table 6B

Table6B

hemoglobin	B1	B6	folate	B12	homocysteine
Spearman r	0.2353	0.2012	0.1567	–0.0088	–0.0936
p (two-tailed)	0.0121	0.0318	0.0901	0.9253	0.4378
p summary	^*	^*	ns	ns	ns
ferritin	B1	B6	folate	B12	homocysteine
Spearman r	0.0915	0.0641	–0.0442	–0.0253	–0.0545
p (two-tailed)	0.3486	0.5116	0.6468	0.7932	0.6794
p summary	ns	ns	ns	ns	ns
creatinine	B1	B6	folate	B12	homocysteine
Spearman r	–0.1217	–0.2745	–0.0974	–0.2074	0.3806
p (two-tailed)	0.1990	0.0031	0.2941	0.0249	0.0011
p summary	ns	^**	ns	^*	^**

Table 6C

Table6C

	F value	Pr(< F)
vitamin B1	0.592	0.464
vitamin B6	1.525	0.252
folate	6.186	0.038
vitamin B12	1.281	0.290
creatinine	0.187	0.677
diagnosis	3.585	0.077
age	3.862	0.085
sex	3.418	0.102

DISCUSSION

In our study, we found that recently diagnosed AD patients had decreased levels of the vitamins B1, B6, B12, and folate and increased homocysteine in serum. LBD patients had diminished vitamin B12 level and VaD was the only form of dementia with lowered vitamin B6 in serum. FTD patients had no alterations in vitamin and homocysteine serum levels. Correlation analyses in AD patients showed associations between vitamin deficiencies and Q Albumin and homocysteine.

Homocysteine acts as an agonist at the glutamate binding site of the N-methyl-D-aspartate receptor (NMDA-R) which is relevant for the memory function of patients with AD, but also as a partial antagonist of the glycine co-agonist site [35]. Memantine, a drug used to treat AD patients, binds at the NMDA-R as a non-competitive and non-selective antagonist [36]. Thereby, it prevents the binding of glutamate, reduces overactivation of NMDA-R and improves cognitive functions. We have recently shown that NMDA-R autoantibodies influence the course of several forms of dementia and are associated with a poor prognosis [37 –39]. Chung et al. had shown that also increased homocysteine levels can exacerbate AD pathology [40].

Hyperhomocysteinemia (HHcy) can lead to neurotoxicity since it leads to the production of the neurotoxic products homocysteic acid and cysteine sulfinic acid which act as NMDA-R agonist and exhibit neurotoxic effects on dopaminergic neurons [41]. Furthermore, HHcy could result in damage on vascular cells [42] and increase the B-CSF-B [43]. Taken Q Albumin as a marker for the integrity for the B-CSF-B, our data support this finding: increased Q Albumin were associated with increased homocysteine levels.

HHcy can be caused by genetic mutations in metabolizing enzymes, but also by smoking, aging, renal failure, or deficiency in B vitamins. In the metabolism of homocysteine, B vitamins are necessary for the interaction and conversion of all mediate products. B vitamins have the potential to reduce homocysteine [42] which is known to interfere with the cerebral blood flow and therefore the cognitive function [44]. In our study, we found increased levels of homocysteine in all forms of dementia but significant only in AD. Patients with VaD had no significant elevated levels of homocysteine compared with the control group. This result is contrary to another study, showing higher homocysteine levels in VaD patients than in AD patients [45]. A possible explanation could be that the patients with VaD included in our study suffered from cerebral microangiopathy and not from an infarction. Increased homocysteine levels were also described in other studies investigating patients with dementia [46, 47]. Increased homocysteine levels and decreased vitamins B12 and folate were described to be correlated with cognitive decline in the elderly population, shown in 607 and 100 patients [48, 49]. We detected decreased levels of the vitamins B1, B6, B12, and folate in AD patients. In patients with MCI, known to have an increased risk to develop AD, have decreased vitamin B6 and folate, and an increased homocysteine level. As also shown by others, the most common causes of homocysteine elevation are deficiencies in vitamins B6, folate, or B12 [50, 51].

It might be that an increased homocysteine level is counter-regulated by an increased requirement of vitamins due to inflammation (for vitamin B6) or malabsorption (for vitamin B12) [52, 53] leading to a deficiency of vitamins and in turn enhanced homocysteine levels. Therefore, despite the same supply of vitamins, a deficiency might develop. Nevertheless, it was shown that supplementation of B vitamins could decrease the homocysteine level and brain atrophy in 271 older people with MCI [54]. Moreover, preclinical studies have shown that administration of folate can inhibit amyloid toxicity [55, 56]. This is of particular interest since in our multivariate analysis including the vitamins B1, B6, B12, and folate, the form of dementia, creatinine, age, and sex, folate was the only independent predictor of homocysteine.

In VaD, the homocysteine level was increased and the vitamin B6 was reduced. Others had shown that vitamin B6 was unaltered in VaD and AD [57]. In a study investigating patients with AD without vascular disease, decreased level of vitamin B1, B6, and B12 were found compared to controls sharing the same age, BMI (“normal weight”) and MMSE [58], indicating that these nutrients might contribute to the disorder. We detected in mild AD an association between increased age and decreased vitamin B1 and B6. Vitamin B12 deficiency is often not diagnosed, but this vitamin might be important for the development of dementia as well [59].

Patients with LBD showed similarly to AD decreased folate and vitamin B12 concentrations, FTD showed no alterations in vitamin and homocysteine levels. However, other studies have shown that in FTD, like in AD, folate was reduced, while it was unchanged in LBD [60]. In addition, vitamin B12 was found to be reduced in FTD [61 –63]. Since this is in contrast to our data, we have to conclude that at least in our study cohort, neither vitamins nor homocysteine seemed to play a major role in FTD.

It is well established that sex influences the susceptibility toward the different forms of dementia. Therefore, it might be reasonable to ask whether there were differences in the vitamin level between male and female study participants. However, no differences in the vitamin levels were obtained between females and males.

Low levels of vitamins in blood might be due to physiological conditions. It was shown that the level of vitamin B6 is inversely correlated with the CRP level as marker for inflammation [52]. Since inflammation enhances the requirement of this vitamin and chronic inflammation was determined in several disorders in advanced age such as dementia, it is likely that inflammation contributes the low level of vitamin B6.

Another important factor that might contribute to the low level of vitamin B12 is malabsorption. Underlying reasons might include gastric or intestinal disorders or drug-induced malabsorption due to drug-nutrient interactions [53].

Pernicious anemia is the classical form of vitamin B12 deficiency, food-bound vitamin B12 malabsorption is a consequence of atrophic gastritis and common in older people. This chronic inflammation of the stomach leads to an atrophy of the mucosa, thereby reducing the gastric acid secretion (hypochlorhydria). Consequently, the absorption of vitamin B12 decreases since hydrochloric acid and pepsin are essential for the release of this vitamin from food proteins [64]. Moreover, hypochlorhydria might induce an overgrowth of bacteria in the stomach and small intestine which then utilize vitamin B12. As another consequence, the amount of the vitamin B12 which is available for absorption is further reduced [65].

Malnutrition might be another important factor that could result in a decreased uptake of vitamins by patients suffering from dementia. We used the BMI as a marker for the nutritional state. Despite the trend that patients with severe AD (BMI 20.28) have a reduced uptake of a sufficient amount of nutrients compared to controls (BMI 26.16) or milder AD stages, this did not reach significance. However, a BMI from 25 is a cutoff point between normal and overweight [66] which means that most of the participants in our study are overweight. An increased body weight at midlife was associated with and increased risk for the development of late-life dementia [67]. Moreover, overweight has several negative metabolic consequences (e.g., type II diabetes mellitus) that are likely to contribute to cognitive decline and dementia [9]. Insulin resistance seemed to account for a significant proportion of the relationship between cognitive performance and obesity [68]. In regard to this, the increased BMI in the study cohort showed a general trend for an overweight population and underlines the importance of the daily quantity and quality of consumed food, containing the recommended quantity of B vitamins. It was shown that AD and VaD patients had significantly higher energy intake than their energy demands, characterized by an excess uptake of polyunsaturated fatty acids, combined with relative deficiencies in B vitamins [69]. This might trigger oxidative stress, chronic inflammation and result in an exacerbation of dementia. In this context it is important that a correlation between the intake of the vitamins B1 and B12 and blood level of the total antioxidant capacity were described in VaD, but not in AD or LBD [70]. For FTD it was shown that the patients have altered eating habits or increased appetite [71], and also LBD patients suffer from several eating problems [72]. However, the patients included in our study had no changes in BMI that goes in line with no significant alterations in the vitamin level.

We also found an association between hemoglobin and the level of vitamin B1 and B6. Older people with low hemoglobin level were found to have an increased risk for the development of dementia [73, 74]. In particular folate and vitamin B12 associated with low hemoglobin were found to be involved in dementia [75 –77]. Some reports also showed that vitamin B1 and vitamin B6 might be involved in low hemoglobin level [78 –81]. However, the meaning of the correlation between both vitamins and hemoglobin remains further investigation.

Besides, we determined a correlation between creatinine and vitamin B6, B12, and homocysteine. This observation was in line with other studies showing a relation between reduced kidney function, homocysteine, and vitamin deficiency [82 –84]. Therefore, also a decline of renal function should be taken into account when determine the association between B vitamins and homocysteine.

Our study has some limitations that have to be considered: 1) We investigated only the time point when the patients were admitted to our clinic. It might be very interesting to follow the patients with the progression of the disorder to analyze alterations in body weight, vitamin levels, and homocysteine level and to compare them with MMSE and Q Albumin values. 2) We have no reliable data upon the nutrition behavior of the patients within the last weeks or months. It might be helpful to know whether this was changed due to the disorder. 3) This is a retrospective study. We have no data concerning the genetic background of the patients (e.g., ApoE, APP, Presenilins) that might influence the observations. 4) Our control group was significantly younger. Whether age is an important factor, these volunteers should be re-examined at the age comparable to the patients’ groups. Moreover, these persons did not suffer from dementia or MCI, but some of them suffered from depressive disorder, somatic symptom disorder, or personality disorder. Whether any of these illnesses contribute to the nutritional state and therefore the vitamin level remains to be investigated. 5) We measured total vitamin B12, but not active B12 (holotranscobalamin II). 6) The cut-off values for vitamin B12 published by different authors from different countries vary, they range between 100 pmol/L to 350 pmol/L [85]. It was shown that low-normal levels may already conceal a subclinical deficiency.

Taken together, our study showed that the levels of several B vitamins are decreased in dementia, but homocysteine is increased already patients with mild cognitive decline. Therefore, our data underline the importance to check the level of B vitamins in elderly people since they might help to slow down or even prevent cognitive decline. In following research studies, it might be interesting to combine a therapy of AD with memantine and the supplementation of B vitamins.

Footnotes

ACKNOWLEDGMENTS

We thank Henrik Dobrowolny for performing statistical analysis.

Authors’ disclosures available online ().

The supplementary material is available in the electronic version of this article: .

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