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Relation between plasma homocysteine, folate, vitamin b12 levels with blood pressure, residual renal function, anemia, dialysis duration and serum albumin in maintenance hemodialysis

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Objectives: To analyze relation between levels of plasma homocysteine (Hcy), folate, vitamin B12 with blood pressure (BP), residual renal function (RRF), anemia, dialysis duration and serum albumin in hemodialysis (HD) patients.

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RELATION BETWEEN PLASMA HOMOCYSTEINE, FOLATE,

VITAMIN B12 LEVELS WITH BLOOD PRESSURE, RESIDUAL

RENAL FUNCTION, ANEMIA, DIALYSIS DURATION

AND SERUM ALBUMIN

IN MAINTENANCE HEMODIALYSIS PATIENTS

Diem Thi Van*; Hoang Trung Vinh* Summary

Objectives: To analyze relation between levels of plasma homocysteine (Hcy), folate, vitamin B12 with blood pressure (BP), residual renal function (RRF), anemia, dialysis duration and serum albumin in hemodialysis (HD) patients Subjects and methods: 112 HD patients were clinically and subclinically examined including measurements of plasma Hcy, folate and vitamin B12 levels Assessments of Hcy, folate and vitamin B12 levels in HD patients were based on their levels in 56 healthy individuals Results: Patients with unaccepted BP control had higher level of plasma Hcy and rate of hyperhomocysteinemia but a lower rate of vitamin B12 deficiency than those with accepted BP control Patients with RRF had significantly lower rate of hyperhomocysteinemia and higher rate of vitamin B12 decrease than patients without RRF Levels of plasma Hcy, folate and vitamin B12 in HD patients were not significantly relative to anemia With dialysis duration, folate was negatively correlated (r = -0.47; p < 0.001) whereas vitamin B12 was positively correlated (r = 0.26; p < 0.01) Hcy was positively correlated with serum albumin (r = 0.27; p < 0.01) Conclusions: In HD patients, Hcy was significantly related to

BP control, RRF and serum albumin; folate was only related to dialysis duration; vitamin B12 had significant relation to BP control, RRF and dialysis duration

* Key words: Maintenance hemodialysis; Homocysteine; Folate; Vitamin B12; Relation

INTRODUCTION

Cardiovascular disease (CVD) is the

leading cause of death in HD patients,

accounting for 45 - 50% of causes of

death [5] Chronic kidney disease (CKD)

patients, who have the mortality rate due

to CVD were 16 times higher than that

of population [5] There are many risk

factors for CVD in HD patients Apart

from traditional risk factors known such

as hypertension, age, gender, smoking,

diabetes, dyslipidemia, physical inactivity, obesity HD patients also have renal disease-related risk factors such as chronic volume overload, hyperhomocysteinemia,

hypoalbuminemia, oxidative stress and chronic inflammation, among them, hyperhomocystenemia and anemia play

an important role in appearance and progress of CVD [5] Hcy is an independent risk factor for CVD Hcy is elevated in 85 - 100% of chronic kidney

* 103 Hospital

Corresponding author: Diem Thi Van (hoalantim1901@yahoo.com)

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disease (CKD) patients and negatively

correlated with glomerular filtration rate

(GFR) [8] In cases of advanced CKD and

hemodialysis, Hcy level is 1.5 - 2 fold higher

than that of normal people [9] Folate and

vitamin B12 are not only essential

cofactors in Hcy metabolism but also

important causes contributing to anemia

in hemodialysis patients [8] Therefore,

we conducted this study for purpose: To

analyze relation between levels of plasma

Hcy, folate, vitamin B12 with BP, residual

renal function, anemia, dialysis duration

and serum albumin in HD patients

SUBJECTS AND METHODS

1 Subjects

168 participants were enrolled in our

study and divided into 2 groups: the study

group consisted of 112 hemodialysis

patients at the Department of Nephrology

and Hemodialysis, 103 Hospital, from

June to October, 2016 The control group

consisted of 56 healthy individuals who

age, gender matched with those of the

study group

* Selected criteria for the study group:

- Patients with 3 hemodialysis sessions

per week

- Using the same dialyzers with

low-flux membranes

- Using hypertensive drugs, erythropoietin,

tardyferol B9 in treatment

- ESRD patients with different age and

gender and dialysis duration > 1 month

* Excluded criteria for the study group:

- Patients were treated by HDF online method

- Patients with currently serious complications or malabsorption syndrome

2 Methods

* Study design: a cross-sectional

description, in comparison with control group

* Study contents:

- For the study group: We collected

history of disease, performed clinical examinations and laboratory tests Patients were asked to provisionally stop using folate and vitamin B12-containing drugs at least one day before taking plasma homocysteine, folate and vitamin B12 tests Blood sample was drawn under fasting condition and before the first dialysis session of the week Plasma Hcy, folate and vitamin B12 levels were measured by microparticle chemiluminescent immunoassay in the Department of

Biochemistry, 103 Military Hospital

- For the control group: We also asked for history of health and performed clinical examinations of organs After assessing heatlthy individuals, they also measured plasma Hcy, folate and vitamin B12 levels

by the same method as in the study group

* Statistical analysis: Stata 12.0 were

used to analyse data Mean Hcy, folate and vitamin B12 levels of the study group were considered as increases when > X +

SD and decreases when < X - SD of the control group

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RESULTS

Table 1: General characteristics of the study group (n = 112)

Mean age (year) 49.93 ± 14.74 Mean dialysis duration (months) 48.62 ± 47.26

Uncontrolled BP

Anemia

Urine conservation

Serum albumin < 35 g/L

Table 2: Features of Hcy, folate, vitamin B12 levels in HD patients

Table 3: Relation between Hcy, folate, vitamin B12, and blood pressure control (n = 112)

Variable Unaccepted control

(n = 71)

Accepted controlled

Hcy

(µmol/L)

Folate

Vit B12

+ Mean Hcy level and rate of hyperhomocysteinemia of group with unaccepted BP control were significantly higher than those of the other group

+ Unaccepted BP control group had a lower folate level and a higher rate of folate deficiency than the other group, but these differences had no satistical significance + There were no significant differences in mean vitamin B12 levels between two groups (p > 0.05)

+ Rate of vitamin B12 deficiency in unaccepted BP control group was lower than that in the other group (p < 0.01)

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Table 4: Relation between Hcy, folate, vitamin B12 and residual kidney function (n = 112)

Hcy (µmol/L)

Folate

Vit B12

+ Mean Hcy level of patients with RRF was lower than that of patients without RRF but had no statistical significance (p > 0.05)

+ Rate of hyperhomocysteinemia of RRF group was significantly higher than that of the other group (p < 0.05)

+ Mean folate level and rate of folate deficiency were not different between the two groups

+ RRF group had a lower mean vitamin B12 level and a higher rate of vitamin B12 deficiency than the other group

Table 5: Relation between Hcy, folate, vitamin B12 and anemia (n = 112)

Variable Anemia (n = 106) Non-anemia (n = 6) p

Hcy

Folate

Vit B12

Mean Hcy, folate, vitamin B12 levels, rates of high Hcy, low folate and low vitamin B12 were not different between two groups of anemia (p > 0.05)

Table 6: Correlation between Hcy, folate, vitamin B12 and dialysis duration, serum albumin

Hcy (µmol/L) Folate (ng/mL) Vitamin B12 (pg/mL) Variable

+ With dialysis duration: folate had moderately negative correlation (r = -0.47, p < 0.001), vitamin B12 had slightly positive correlation (r = 0.26, p < 0.01), Hcy had no correlation

+ With serum albumin: plasma Hcy level had slightly positive correlation (r = 0.27;

p < 0.01), plasma folate and vitamin B12 levels had no correlation

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DISCUSSION

1 Relation with blood pressure

control

* Hcy: Lim and Cassano (2002) explored

the relationship between Hcy and blood

pressure by analyzing data in the NHANES

III study from 1988 to 1994 The results

indicated that the average diastolic and

systolic blood pressure measurements

increased by 3.7 and 9.3 mmHg,

respectively, from the lowest to the

highest quintile of Hcy, unadjusted for

age An increase of 5 µmol/L in Hcy was

associated with increases in diastolic and

systolic blood pressure of 0.5 and 0.7

mmHg, respectively, in men and of 0.7

and 1.2 mmHg in women [6] Mechanisms

that could explain the relationship between

homocysteine and blood pressure are

homocysteine-induced arteriolarconstriction,

renal dysfunction and increased sodium

reabsorption, and increased arterial stiffness

[10] Because of relation between Hcy and

blood pressure, blood pressure control is

affected by plasma Hcy level Our study

demonstrated that patients with unaccepted

BP control had higher level of plasma Hcy

and rate of hyperhomocysteinemia than

patients with accepted BP control (table 3)

In hemodialysis patients, Huynh Van

Nhuan (2009) also showed that plasma

Hcy level in patients without hypertension

was 18.04 ± 8.47 µmol/L, significantly lower

than that of patients with hypertension

(27.63 ± 12.70 µmol/L, p < 0.05) [2]

Ha Van Hung (2016) indicated that

non-hypertensive patients had mean plasma

Hcy level of 23.76 ± 8.10 µmol/L and rate

of elevated homocystein of 18.1%, which

were significantly lower than those in hypertensive patients (30.76 ± 7.36 µmol/L and 81.9%, respectively) [1]

* Folate and vitamin B12: folate and

vitamin B12 are the main nutrition factors affecting Hcy levels and have a protective role against cardiovascular disease Protective effect of these B vitamins are not only due

to their ability of lowering Hcy level but also due to the ability to act as an antioxidant, to increase the concentration of

NO - a vasodilator of endothelial cells

Many studies have shown that treatment with B vitamins may reduce BP and cardiovascular events A study by Scazzone (2014) in 116 hypertensive patients and

81 healthy individuals reported that average level of folate in the hypertensive group (6.7 ± 5.0 ng/mL) was significantly lower than that of the group control (9.0 ± 4.4 ng/mL), while vitamin B12 level was

not different between the two groups (440

± 223 pg/mL in the hypertensive group versus 491 ± 185 pg/mL in the control group) [9] Our study indicated that no differences in folate and vitamin B12 levels were observed between the group with accepted BP control and the group

with unaccepted BP control (table 3) We

believe that our results could not find out the relation between folate, vitamin B12 and blood pressure because in HD patients, blood pressure was influenced

by many factors, in which folate and vitamin B12 play only a small role

2 Relation with residual renal function (or urine conservation)

Residual renal function is assessed by volume of 24-hour urine while patients still

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remain dialysis If 24-hour urine output

< 500 mL is considered to have no residual

renal function or no urine conservation In

contrast, 24-hour urine output ≥ 500 mL is

considered as having residual renal

function or urine conservation RRF plays

an important role for patients on dialysis,

especially peritoneal dialysis patients

RRF contributes significantly to the overall

health and well-being of dialysis patients It

does not only provide small solute

clearance but also plays an important role

in maintaining fluid balance, phosphorus

control, and removal of middle molecular

uremic toxins such as beta 2 microglobulin

Decline of RRF also contributes significantly

to anemia, inflammation, and malnutrition

in patients on dialysis [12] Plasma Hcy,

folate and vitamin B12 levels are all

influenced by residual renal function

because all of them are filtered through

the glomeruli Therefore, if patients still

has residual renal function, levels of these

substances will be lower than those of

patients without RRF Our results showed

that patients with RRF had lower rate of

hyperhomocysteinemia than patients

without RRF We also found that patients

with RRF had lower vitamin B12 levels

and higher rate of vitamin B12 deficiency

than patients without RRF But regarding

folate, our study revealed that no differences

in folate level and rate of folate deficiency

were observed between the two groups

(table 4) In our study, folate was not

related to RRF, which can be attributable

to folate level in hemodialysis patients

influenced by many factors such as diet,

using folate-containing drugs in treatment

before

3 Relation to anemia

* Hcy: Anemia is a common manifestation

in hemodialysis patients and has multifactorial etiology, in which lack of materials for hematopoiesis such as folic acid and vitamin B12 also significantly contributed

to appearance and progression of anemia

Hcy level is affected by folate and vitamin B12 because they act as cofactors in the process of Hcy re-methylation into methionine Thus between Hcy and anemia may have a relation with each other because they are all affected by the status of folate, vitamin B12 in the body

In our study, no significant differences were observed in Hcy level and the rate of hyperhomocysteinemia between anemia

patients and non-anemia patients (table 5)

Similar to our results, Ha Van Hung (2016) reported that Hcy levels of patients with anemia was 28.64 ± 7.65 µmol/L, which was not different from that of patients without anemia (34.02 ± 8.07 µmol/l) (p > 0.05) There were no differences in

rate of elevated Hcy between group with anemia and group without anaemia [1]

Tayebi (2016) showed that no significant correlation was found between Hcy level and hemoglobin [11]

* Folate and vitamin B12: Folate and

vitamin B12 are essential for the process

of hematopoiesis so that the lack of them will cause macrocytic anemia However,

in this study we did not observe any significant relations between folate,

vitamin B12 levels and anemia (table 5)

This could be explained by anemia in HD patients attributable to many causes such

as shortened red blood cell survival, blood loss, inflammation and the deficiency of

EPO and vitamin

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4 Correlation with dialysis duration

* Hcy: Hcy has a molecular weight of

135 dalton within range of glomerular

filtration Absorption of Hcy in renal tubular

was clearly seen through kinetic studies in

rat renal cortex Kidneys have an important

role in the metabolism of homocysteine

Total homocysteine concentration increases

as impaired renal function and achieves

the highest level in end-stage renal disease

Many studies have demonstrated that after

each hemodialysis session, Hcy levels

may decrease partly but then rise again

until the next session Hence the longer

dialysis duration is, the more level of Hcy

accumulates De Vecchi (2000) indicated

that plasma Hcy level was positively

correlated with dialysis duration (r = 0.32;

p < 0.0001) [4] Moustapha A et al (1999)

reported that there was a positive correlation

between Hcy level and dialysis duration

(r = 0.15; p < 0.05) [7] However, our study

demonstrated that plasma Hcy level was

not correlated with dialysis duration (table

6) We could not find out the correlation

between Hcy level and dialysis duration,

which could be explained by plasma Hcy

level influenced by many factors

* Folate: Folate is a water-soluble vitamin,

small molecular weight of 441 dalton It

exists in plasma mainly in the free form or

loosely bound with non-specific proteins,

so that folate may be lost through the

membrane Hence, the longer dialysis

duration is, the higher the risk of folate

deficiency is Our study showed that

plasma folate level was inversely correlated

with dialysis duration (table 6)

* Vitamin B12: Vitamin B12 has a greater

molecular weight than folate (1,355 dalton)

Moreover, in blood, vitamin B12 exists in

the form bound to the carrier proteins (transcobalamin and haptocorrin), so vitamin B12 is hardly lost through low-flux dialysis membrane Our research showed that a positive correlation was observed between vitamin B12 level and dialysis duration

(table 6), which may be firstly explained

by vitamin B12 level not affected by hemodialysis; Moreover, patients with long dialysis duration often have more nervous complications than patients with short dialysis duration, so they often use group

B vitamins including vitamin B12 more

5 Correlation with level of serum albumin

Serum albumin is an important marker

of Hcy level because in plasma most Hcy binds with protein (mainly albumin) Meanwhile, only free form of Hcy is filtered through the glomeruli Thus, when the level of serum albumin increases, the rate of albumin-bound Hcy will increase, leading to decreased elimination of Hcy

by kidneys, which results in elevated level

of plasma Hcy Our study indicated that there was a positive correlation between

Hcy level and serum albumin (table 6)

Similar to our result, many other researchers also reported a positive correlation between Hcy and serum albumin such as Huynh Van Nhuan (2009) (r = 0.353; p < 0.01) [2]; Arnadottir (1999) (r = 0.28; p < 0.05) [3] We did not find any correlation between folate, vitamin B12 and serum albumin

(table 6), which may be also suitable

because folate in plasma exists mainly in the free form and vitamin B12 is bound with two transport proteins (not albumin) (transcobalamin and haptocorrin), so that the level of folate and vitamin B12 are not affected by serum albumin

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CONCLUSIONS

- Patients with unaccepted BP control

had higher level of plasma Hcy and rate

of hyperhomocysteinemia whereas a lower

rate of vitamin B12 decrease than those

with accepted BP control

- Patients with RRF had significantly

lower rate of hyperhomocysteinemia and

higher rate of vitamin B12 decrease than

patients without RRF

- Levels of plasma Hcy, folate and

vitamin B12 in HD patients were not

significantly relate to anemia

- Folate was negatively correlated with

dialysis duration (r = -0.47; p < 0.001)

whereas vitamin B12 was positively

correlated with dialysis duration (r = 0.26;

p < 0.01)

- Hcy was positively correlated with

serum albumin (r = 0.27; p < 0.01)

REFERENCES

1 Ha Van Hung Researching on change

in serum hs-CRP and homocysteine levels

before and after HDF online in chronic renal

failure patients on hemodialysis Specialist

level 2 thesis VietNam Military Medical

University Ha Noi 2016

2 Huynh Van Nhuan Research on change

of blood homocysteine levels and effect of

treatment in patients with chronic renal failure

on hemodialysis Medical doctoral thesis, Hue

University of Medicine and Pharmacy 2009

3 Arnadottir M, Berg A.L, Hegbrant J et al

Influence of haemodialysis on plasma total

homocysteine concentration, Nephrol Dial

Transplant 1999, 14 (1), pp.142-146

4 De Vecchi A.F, Bamonti-Catena F,

Finazzi S et al Homocysteine, vitamin B12,

and serum and erythrocyte folate in peritoneal

dialysis and hemodialysis patients Perit Dial Int 2000, 20 (2), pp.169-173

5 Herzog C.A, Asinger RW, Berger A.K et

al Cardiovascular disease in chronic kidney

disease A clinical update from kidney disease: Improving Global Outcomes (KDIGO) Kidney Int 2011, 80 (6), pp.572-586

6 Lim U, Cassano P.A Homocysteine and

blood pressure in the Third National Health and Nutrition Examination Survey, 1988 -

1994 American Journal of Epidemiology

2002, 156, pp.1105-1113

7 Moustapha A, Gupta A, Robinson K et

hyperhomocysteinemia in hemodialysis and peritoneal dialysis Kidney Int 1999, 55 (4), pp.1470-1475

8 Nand N, Sharma M, Mittal N

Prevalence of hyperhomocysteinaemia in chronic kidney disease and effect of supplementation of folic acid and vitamin B12

on cardiovascular mortality JIACM 2013, 14 (1), pp.33-36

9 Scazzone C, Bono A, Tornese F et al

Correlation between low folate levels and hyperhomocysteinemia, but not with vitamin B12 in hypertensive patients Annals of Clinical & Laboratory Science 2014, 44 (3), pp.286-290

10 Stehouwer C.D, Van G.C Does

homocysteine cause hypertension? Clin Chem Lab Med 2003, 41 (11), pp.1408-1411

11 Tayebi A, Biniaz V, Savari S et al

Effect of vitamin B12 supplementation on serum homocysteine in patients undergoing hemodialysis: A Randomized Controlled Trial Saudi Journal of Kidney Diseases and Transplantation 2016, 27 (2), pp.256-262

12 Wang A.Y, Lai K.N The importance of

residual renal function in dialysis patients Kidney Int 2016, 69 (10), pp.1726-1732

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