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Adverse myocardial effects of B-vitamin therapy in subjects with chronic kidney disease and hyperhomocysteinaemia
Nutrition, Metabolism & Cardiovascular Diseases (2013) 23, 836e842
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/nmcd
Adverse myocardial effects of B-vitamin therapy in subjects with chronic kidney disease and hyperhomocysteinaemia Z. Rafeq a, J.D. Roh c,d, P. Guarino f, J. Kaufman a,e, J. Joseph a,b,d,e,* a
Veteran Affairs Boston Healthcare System, Department of Medicine, Boston, MA, USA Brigham and Women’s Hospital, Department of Medicine, Boston, MA, USA c Beth Israel Deaconess Medical Center, Department of Medicine, Boston, MA, USA d Harvard Medical School, Boston, MA, USA e Boston University School of Medicine, Boston, MA, USA f Cooperative Studies Program Coordinating Center, VA Medical Center, West Haven, CT, USA b
Received 19 March 2012; received in revised form 18 June 2012; accepted 3 July 2012 Available online 16 August 2012
KEYWORDS Homocysteine; Heart failure; Diastolic dysfunction; Chronic kidney disease
Abstract Background & aims: Hyperhomocysteinaemia (HHCY), a common finding in patients with chronic kidney disease (CKD), has been shown to contribute to adverse cardiac remodelling and failure. We hypothesised that in human subjects with CKD, HHCY would be associated with myocardial dysfunction, and that homocysteine (HCY)-lowering therapy would improve myocardial remodelling and heart-failure (HF) outcomes. Methods and results: Post hoc analysis of the Homocysteinemia in Kidney and End Stage Renal Disease (HOST) trial (n Z 2056) was performed to determine if HCY-lowering therapy with high dose B vitamins affects HF outcomes in patients with CKD. In addition, effects on myocardial remodelling were assessed in a subgroup of 220 trial subjects who had transthoracic echocardiograms done before study randomisation and during the course of the study as part of their routine clinical care. HF outcomes were not significantly affected by treatment compared to the placebo. HCY levels were inversely correlated with diastolic function (R Z 0.21; p Z 0.038). Vitamin therapy resulted in a significant increase in left atrial size (þ0.15 0.8 cm vs. 0.13 0.07 cm; p Z 0.0095). No other echocardiographic parameters were significantly associated with baseline HCY levels or changes with vitamin therapy. Conclusion: HHCY is associated with diastolic dysfunction in patients with CKD. However, B-vitamin therapy did not improve HF outcomes despite lowering of plasma HCY levels, and
* Corresponding author. Cardiology Section (111), VA Boston Healthcare System, 1400 VFW Parkway, West Roxbury, MA 02132, USA. Tel.: þ1 857 203 6841; fax: þ1 857 203 5550. E-mail address: [email protected] (J. Joseph). 0939-4753/$ - see front matter Published by Elsevier B.V. http://dx.doi.org/10.1016/j.numecd.2012.07.002
Adverse myocardial effects of B-vitamin therapy
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was associated with an increase in left atrial size, which is a surrogate for worsening left ventricular diastolic dysfunction. These findings suggest that high-dose B vitamin therapy may be harmful in patients with CKD. Published by Elsevier B.V.
In 1969, Kilmer McCully proposed the theory that a perturbation of the methionineehomocysteine cycle, manifested biochemically as Hyperhomocysteinaemia (HHCY), increased the risk for cardiovascular disease [1]. Numerous observational studies have demonstrated positive correlations between elevated plasma homocysteine (HCY) levels and adverse cardiovascular outcomes, including myocardial infarction (MI) and stroke [2,3]. However, no large-scale prospective trial has convincingly shown that effectively lowering plasma levels of HCY decreases rates of MI, stroke or mortality [4e7]. Interestingly, recent findings from preclinical animal studies have suggested that diet-induced HHCY may directly affect the myocardium, independent of its effects on the vascular system. Work done in our laboratory and those of others has demonstrated that HHCY increases myocardial collagen content, adverse myocardial remodelling, and diastolic dysfunction in rat models [8e10]. While these findings have not been directly confirmed in humans, small observational studies suggest that a similar phenomenon may exist. HHCY correlates with increases in left ventricular (LV) mass and wall thickness [11,12], and is associated with increased risk and severity of clinical heart failure (HF) [13,14]. Whether circulating HCY serves as a biomarker for worsening HF or is a causal factor in its pathophysiology, however, remains unclear. Patients with chronic kidney disease (CKD) represent a unique cohort for studying the potential relationship between HHCY and HF given the increased prevalence of both of these disease states in this patient population [15e17]. Elevation in HCY levels in patients with CKD has been attributed to reduced renal clearance of circulating HCY and impaired HCY metabolism [18,19]. Likewise, similar mechanisms of deteriorating renal function in HF have been proposed to be the link between HF and HHCY [20,21]. To further investigate the relationship between HHCY and HF in human subjects, we performed post hoc analysis of the Homocysteinemia in Kidney and End Stage Renal Disease (HOST) trial, a study which evaluated cardiovascular outcomes of HCY-lowering vitamin therapy in patients with concomitant CKD and HHCY [7]. We specifically examined the relations of echocardiographic evidence of myocardial remodelling and clinical HF outcomes (which had not been evaluated in the HOST trial) with baseline HCY levels and vitamin therapy.
Methods Study population Details of the HOST study have been previously published [7]. In brief, the study was a prospective, randomized, double-blind study designed to determine if HCY-lowering therapy, that is, a combination of high doses of folic acid,
vitamin B12 and vitamin B6, could reduce all-cause mortality and cardiovascular outcomes in patients with CKD. Participants (n Z 2056) were recruited from 36 US Department of Veterans Affairs medical centres from the years 2001 to 2006, and followed for a median of 3.2 years on either placebo or vitamin treatment. Inclusion criteria included age >21 years, advanced CKD (eGFR 30 ml min1) (n Z 1305) or end-stage renal disease (ESRD; n Z 751), and high plasma HCY levels (15 mM). HF outcomes were not evaluated in this study, but were available for review through the electronic Computerized Patient Record System (CPRS) of the Veterans Affairs Healthcare System. All procedures in this study were approved by the HOST Executive Committee and the Institutional Review Board of Veterans Affairs Boston Healthcare System.
Data collection Data on baseline history of HF and HF-related hospitalisations during the study period were collected for all subjects included in the original study. Discharge summaries were reviewed and classified as evidence of HF at baseline or during follow-up if they included any of the following ICD9 codes: any 428.X, 276.6, 518.4, and 514. In addition, all clinical variables, that is, mortality, used in the original HOST study, were also used for analysis. For the echocardiographic subgroup analysis, we identified 220 subjects (out of a total of 2056 subjects in the HOST study) who, as a component of their routine clinical care, had an echocardiogram within 6 months before randomisation, and 6 months or longer after randomisation to placebo or vitamin treatment. The following echocardiographic parameters were collected and analysed as continuous variables: LV ejection fraction (EF); LV wall thicknesseposterior wall (PW) and interventricular septum (IVS); LV end-diastolic dimension (LVEDD); LV end-systolic dimension (LVESD); and left atrial (LA) size. Diastolic function was evaluated as a categorical variable (normal or abnormal) since all medical centres involved did not report individual components of diastolic function assessment.
Statistical analysis Relationship of plasma HCY level and vitamin treatment to HF outcomes We first analysed the data to determine if there was a correlation between baseline HF prevalence and baseline plasma HCY levels by using analysis of covariance (ANCOVA). Subjects were stratified into advanced chronic kidney disease (ACKD) or ESRD based on the dialysis status at HOST study enrollment. For all study subjects, we used Cox proportional hazards regression survival analyses to compare vitamin and placebo groups for differences in the primary composite endpoint of HF-related hospitalisation
838 or mortality. A secondary endpoint of HF-related hospitalisation, alone, was also evaluated. Analysis was adjusted for the following confounding variables: age, gender, race, strata (ACKD vs. ESRD), diabetes mellitus, haemoglobin levels, hypertension, history of MI, baseline medication use (angiotensin-converting enzyme inhibitors, angiotensin receptor blockers, beta-blockers and aldosterone inhibitors), and baseline HF. An interaction term of treatment with baseline HF status was included in the model to test if the whole cohort could be analysed together or whether the effect of treatment needed to be examined separately in those with and without a history of HF. Relation of plasma HCY level and vitamin treatment to cardiac structure and function (echocardiographic substudy) Baseline clinical and detailed echocardiographic data during the study period were available for a total of 220 patients. We compared the baseline clinical and treatment characteristics for the subgroup population (n Z 220) versus those not included (n Z 1836). Analysis was then conducted to determine if baseline echocardiographic parameters correlated with plasma HCY levels for the whole subgroup. Univariate linear regression was performed on each echocardiographic parameter to examine its correlation to the baseline plasma HCY level. For diastolic dysfunction, logistic regression was used. Subsequently, we determined if a change in each echocardiographic parameter between values obtained before randomisation and during the study were different between the placebo and vitamin groups (separate tests for each variable). Parameters evaluated were changes in LV wall thickness, LVEF, LVEDD, LVESD, diastolic function, and LA size. Since the date of echocardiogram was available, analysis was adjusted for differences in the time from randomisation when a repeat echocardiogram was performed. ANCOVA was then performed with baseline echocardiogram and days from the date of randomisation until the date of repeat echocardiogram included as covariates, i.e., Change in echo Z baseline echo þ days þ treatment group þ error.
Z. Rafeq et al. Table 1 Relation of baseline plasma HCY levels and vitamin treatment to clinical HF outcomes. Outcome
Baseline HCY
Vitamin treatment
Hazard ratio (95% C.I.)
Hazard ratio (95% C.I.)
Entire group HF admissions þ mortality HF admissions
1.00 (0.99e1.01); p Z 0.49 0.98 (0.96e1.00); p Z 0.09 Advanced chronic kidney disease HF admissions þ 1.01 (1.00e1.02); mortality p Z 0.16 HF admissions 0.98 (0.95e1.01); p Z 0.26 End stage renal disease HF admissions þ 1.00 (0.99e1.01); mortality p Z 0.93 HF admissions 0.98 (0.96e1.01); p Z 0.21
1.11 (0.95e1.29); p Z 0.18 0.90 (0.64e1.26); p Z 0.54 1.07 (0.91e1.26); p Z 0.40 0.75 (0.50e1.13); p Z 0.16 1.08 (0.88e1.32); p Z 0.53 0.98 (0.67e1.43); p Z 0.93
Hazard ratios with 95% confidence intervals generated from Cox proportional hazards analysis to compare vitamin and placebo groups for differences in the primary composite endpoint of HFrelated hospitalization or mortality or the secondary endpoint of HF-related hospitalizations. HCY, homocysteine; HF, heart failure.
Baseline history of HF was significantly more prevalent in the subgroup, along with higher prevalence of beta-blocker therapy, ESRD and diabetes. However, within the subgroup, there were no significant differences between subjects who had been randomised to the vitamin group (45%) and those who had been randomised to the placebo group (55%). As shown in Table 3, no significant differences in baseline age, body mass index, serum albumin, haemoglobin, diagnosis of ACKD vs. ESRD, HF, hypertension, or use of angiotensin converting enzyme inhibitors or beta-blockers were detected between the two groups.
Results Relation of plasma HCY and vitamin treatment to HF outcomes There was no significant association of baseline plasma HCY levels with baseline history of HF (HR 0.995, C.I. 0.983e1.007). As shown in Table 1, treatment with high-dose vitamin therapy did not affect HF-related hospitalisations or mortality in ACKD or ESRD groups during the course of the study. The total number of events in the study population was 986 HF þ mortality events and 207 HF events.
Characteristics of the echocardiographic subgroup analysis Table 2 demonstrates the significant differences in baseline characteristics between the echocardiographic subgroup and the subjects of the HOST trial who were excluded from this substudy based on the lack of echocardiograms.
Table 2 Baseline demographic data of echocardiographic subgroup vs. original HOST study. Variable
HOST subjects included in subgroup (n Z 220)
HOST subjects excluded from subgroup (n Z 1836)
p-Value
Age (year) ESRD Serum albumin (g/dl) Baseline HF Diabetes Beta-blocker use
64.2 11.6 115 (52%) 3.9 0.5
66.0 11.8 636 (35%) 4.0 0.5
0.034 <0.0001 0.004
73 (33%) 136 (62%) 155 (71%)
413 (23%) 993 (54%) 1194 (57%)
0.0005 0.033 0.0001
Data shown as mean SD or total number subjects with percentage of subgroup indicated in parentheses. ESRD, end stage renal disease; HF, heart failure.
Adverse myocardial effects of B-vitamin therapy Table 3 Baseline demographic data of placebo and vitamin therapy groups within the echocardiographic subgroup. Variable
Placebo
Vitamin treatment
p-Value (t-test)
Treatment Age (years) BMI (kg/m2) Hemoglobin (g/dl) Serum albumin (g/dl)
121 (55%) 64.5 9.9 27.2 4.9 11.4 1.6
99 (45%) 63.8 10.5 27.3 4.1 11.8 1.4
0.62 0.85 0.12
3.8 0.5
3.8 0.5
0.78
Variable
Placebo
Vitamin treatment
p-Value (Chi square test)
ACKD ESRD HF Hypertension Beta-blockers ACE inhibitor
60 (49.6%) 61 (50.4%) 42 (34.7%) 115 (95%) 85 (70.3%) 60 (49.6%)
45 54 31 93 70 39
0.54
(45.5%) (54.5%) (31.3%) (93%) (70.7%) (39.4%)
0.59 0.72 0.94 0.13
Data shown as mean SD or total number of subjects with percentage of subgroup indicated parentheses. BMI, body mass index; ACKD, advanced chronic kidney disease; ESRD, end stage renal disease; HF, heart failure; ACE, angiotensin converting enzyme.
Lowering of plasma homocysteine level in the echocardiographic subgroup All subjects in the echocardiogram subgroup had a plasma HCY level >15 mM, which was an inclusion criterion for the HOST trial. The mean plasma HCY levels in subjects with
Table 4
ACKD (n Z 105) was 23.9 mM (range 16e45), while that in the ESRD patients (n Z 115) was slightly higher (25.1 mM; range 15e67). Table 4 demonstrates the plasma HCY and folate levels in placebo and vitamin groups at 3 months and 1 year after randomisation. Since measurements were done only in a selected subset of patients after the 3-month assessments, only a small number of the substudy subjects had measurements at 1 year. Compared to baseline values, plasma HCY levels were reduced significantly in the vitamin group at 3 months and at 1 year after initiation of treatment. Plasma folate values were also increased significantly in the vitamin group at 3 months and 1 year. The placebo group did not demonstrate a significant change in plasma HCY or folate levels at either time point.
Effects of vitamin therapy on echocardiographic parameters Univariate regression analysis of the correlations of baseline echocardiographic measurements and pre-treatment HCY levels demonstrated a significant inverse relationship between plasma HCY levels and diastolic function (R Z 0.21, p Z 0.038). In the echocardiographic subgroup, 86% of the subjects had abnormal diastolic function at baseline. An inverse correlation of borderline significance was seen with LA size (R Z 0.14, p Z 0.08). There were no significant correlations between plasma HCY level and the other echocardiographic parameters measured. We examined whether there were any significant changes in any echocardiographic parameter during the study period compared to baseline values, in the placebo and vitamin groups. Table 5 shows the results of multivariate regression analyses that included determinants of changes in echocardiographic parameters during the study period, including baseline value, days to follow-up
Plasma total homocysteine and folate at baseline and follow-up in substudy population.
Total homocysteine e mmol/L Placebo group: N Median (Q1, Q3) Vitamin group: N Median (Q1, Q3) p-Valuea Folate e ng/mLb Placebo group: N Median (Q1, Q3) Vitamin group: N Median (Q1, Q3) p-Valuea
839
Baseline
3 Months e baseline
1 Year e baseline
121 22.0 (19.0, 26.0)
116 0.6 (3.7, 2.3)
16 0.3 (2.7, 3.9)
99 22.0 (18.3, 26.7)
94 5.4 (9.2, 2.3)
12 4.3 (7.3, 0.7)
<0.0001
0.04
116 18.4 (11.7, 25.0)
111 1.5 (5.3, 14.5)
16 4.2 (11.0, 19.1)
97 17.4 (9.6, 25.0)
92 1616 (88, 3736)
12 1954 (2, 5688)
<0.0001
0.01
Q1, Q3 denotes values of the first and third quartile. To convert values for homocysteine to mg/liter divide by 7.397. To convert values for folate to nanomoles per liter multiply by 2.266. a Wilcoxon rank sum test p-value. b Tests results reported at baseline as >25 were analyzed as 25. This was because baseline values were obtained from different laboratories, several of which had cutoffs of 25. Subsequent analyses were done in a single laboratory with no cutoff.
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Table 5 Associations of B-vitamin therapy with changes in echocardiographic parameters during the study period. Echocardiographic parameter
Treatment effect
DLVEF DLVIVS DLVPW DLVEDD DLVESD DLA size DDiastolic function
0.28 0.08 0.08 0.09 0.15 0.27 0.09
(2.0) (0.04) (0.04) (0.13) (0.18) (0.1)*** (0.08)
Multivariate regression analysis including baseline value, days to follow-up echocardiogram and treatment group as variables. Data shown as parameter estimate (SE). ***p Z 0.0095. LV, left ventricle; EF, ejection fraction; IVS, intraventricular septal wall thickness; PW, posterior wall thickness; EDD, end-diastolic dimension; ESD, end-systolic dimension; LA, left atrium.
echocardiogram and treatment groups as covariates. The only change during the study period that was independently associated with HCY-lowering vitamin therapy was an increase in LA size. Further multivariate regression analysis adjusted for clinical variables and variables that affect LA size, such as baseline LV and LA size, days to follow up echocardiogram, baseline mitral regurgitation (MR) and change in degree of MR during follow-up, demonstrated a significant increase in LA size during the study period in the vitamin group (þ0.15 0.08 cm) compared to the placebo group (0.13 0.07 cm; pZ0.0095).
Discussion Our post hoc analysis of the HOST trial used echocardiographic data and clinical HF outcomes to investigate the relationship between HHCY and HF in human subjects with CKD. The major findings of this study are: (1) HHCY is correlated with myocardial diastolic dysfunction, (2) highdose B-vitamin therapy does not improve HF outcomes despite lowering of plasma HCY levels and (3) HCY-lowering B-vitamin therapy is associated with cardiac structural changes (i.e., left atrial enlargement), suggestive of worsening diastolic dysfunction, in patients with CKD. The correlation detected between HHCY and baseline diastolic dysfunction corroborates findings of previous studies, which have proposed that an alteration of the methionineehomocysteine cycle, manifested as HHCY, may be an independent risk factor for the development of both systolic and diastolic forms of HF [13,21,22]. The mechanism by which this occurs, however, is not well elucidated. Preclinical animal studies have suggested that an alteration in the methionineehomocysteine cycle has direct effects on the myocardium, leading to increased fibrosis and collagen deposition, and eventual myocardial stiffening and pump failure [8e10]. To ascertain if HCY directly contributes to HF pathophysiology in humans, one would have to demonstrate improvements in either myocardial remodelling or clinical HF outcomes with HCY-lowering therapy in patients with known HHCY. We focussed on both these parameters in our post hoc
analysis of the HOST trial. Similar to the HOPE-2 trial [6], we did not detect a significant improvement in HF-related hospitalisations or mortality with vitamin therapy in the HOST study. Even though veterans generally receive all of their care in the Veterans Affairs Health System, there is a small possibility that some events may not have been captured in this analysis. However, our echocardiographic subgroup analysis did reveal an unexpected finding contrary to our original hypothesis. Rather than improving myocardial remodelling, aggressive HCY-lowering vitamin therapy in patients with CKD was associated with a significant increase in LA size, a surrogate marker of worsening diastolic dysfunction. While the available echocardiographic data of the HOST trial subjects were not adequate to analyse quantitative changes in diastolic function, LA dimensions have been reported to be an accurate determinant of progression of diastolic HF [23,24]. Hence, it is quite likely that this was a reliable indicator that adverse cardiac remodelling was occurring in association with vitamin therapy. To our knowledge, this is the first study to investigate the effects of HCY-lowering treatment on myocardial remodelling in HF. Thus, we feel this finding raises an important question of whether unrestricted vitamin supplementation is safe in patients with CKD. The unforeseen harmful effects of vitamin therapy on cardiac remodelling detected in our study underscores the fact that an elevated plasma HCY level likely represents more than just a single defect in a complex process of sulphur aminoacid metabolism and affects redox balance and methylation reactions. HCY metabolism occurs via two interrelated processes of remethylation, which requires folic acid and vitamin B12 as cofactors, and transulphuration, which requires vitamin B6 as a cofactor. While it is difficult to determine from our study why HCY-lowering vitamin therapy resulted in adverse cardiac remodelling in our study population, we propose that patients with advanced CKD may be more susceptible to acute changes in HCY levels given intrinsic deficiencies in their ability to metabolise HCY. Patients with CKD have underlying defects in both DNA and protein methylation, and have specifically been shown to demonstrate impairments in HCY remethylation [25,26]. Thus, acutely increasing HCY remethylation with folic acid and vitamin B12 supplementation could affect any one of multiple intracellular methylation processes which are dependent on the methionineehomocysteine cycle. In addition, given their underlying impairments in methylation, patients with CKD are likely more dependent upon the alternative pathway of HCY transulphuration, which notably plays a key role in the generation of the anti-oxidant glutathione (via cysteine). Patients with CKD and HF have increased basal levels of chronic inflammation and oxidant stress [27,28]. Thus, it is possible that by redirecting HCY metabolism towards remethylation via folic acid and vitamin B12 supplementation, glutathione production would be particularly suppressed in patients with CKD, which could result in increased oxidant stress in the myocardium. Lastly, it is possible that vitamin therapy may directly affect the myocardium in CKD patients, independent of its effects on HCY-lowering. Folic acid therapy in patients with ESRD has been shown to affect DNA methylation and gene expression without altering plasma HCY levels [29].
Adverse myocardial effects of B-vitamin therapy While further work is necessary to elucidate why vitamin therapy potentially promotes adverse cardiac remodelling in patients with advanced CKD, it is also important to recognise that certain patient populations likely respond to seemingly benign vitamin therapies differently, and that these variable responses can be harmful. A large, metaanalysis investigating the effects of HCY-lowering folate therapy found potential harm in terms of cardiovascular disease in patients with elevated HCY levels, but not in those with normal-to-minimally elevated HCY levels [30]. Whether these patients with higher HCY levels represented a similar demographic to subjects studied in the HOST trial is unclear. However, if so, this would support the notion that folic acid therapy in patients with advanced CKD has potentially harmful effects on cardiac function. While our post hoc analysis of the HOST study does raise interesting concerns about unrestricted vitamin use, there are multiple limitations to our study which must be noted. First, the HOST trial was limited to a select cohort of patients with advanced or end stage kidney disease, which does not allow us to apply our findings to a more general population. Furthermore, in contrast to prior studies, we did not detect a significant correlation between baseline HCY levels and HF. This was likely influenced by the fact that all subjects in the HOST trial had significantly elevated plasma HCY levels, an inclusion criterion of the trial, which did not allow us to compare HF-related outcomes in patients with normal or low plasma levels of HCY. Second, echocardiograms were done as part of routine clinical care and, thus, were not standardised for this study. Thus, multiple additional parameters could not be evaluated. Specifically, we could not quantify changes in diastolic dysfunction since various methods to evaluate diastolic functions were used by different institutions, and LA size, as opposed to a more specific LA volume, had to be used to assess for changes in diastolic dysfunction. In addition, since the echocardiograms were done for clinical indications, there may be some selection bias, with those with more severe clinical manifestations more likely to undergo echocardiography. Another limitation of our study is that we do not know how long the subjects have been exposed to high levels of HCY before randomisation to vitamin therapy or placebo, which could have an influence on the effects of therapy. In summary, our post hoc analysis of the HOST trial showed that HHCY correlates with diastolic dysfunction. However, in patients with advanced CKD, aggressive HCYlowering vitamin therapy does not improve clinical HF outcomes, and may actually be detrimental in terms of worsening adverse cardiac remodelling and diastolic dysfunction. Given the complexity of HCY metabolism in kidney disease, further mechanistic and prospective studies need to be conducted before recommendations regarding HCY-lowering vitamin therapy can be established in patients with CKD.
Acknowledgements The HOST study was supported by the Cooperative Studies Program, Department of Veterans Affairs Office of Research and Development; PamLab; and Abbott Laboratories.
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Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/nmcd
Adverse myocardial effects of B-vitamin therapy in subjects with chronic kidney disease and hyperhomocysteinaemia Z. Rafeq a, J.D. Roh c,d, P. Guarino f, J. Kaufman a,e, J. Joseph a,b,d,e,* a
Veteran Affairs Boston Healthcare System, Department of Medicine, Boston, MA, USA Brigham and Women’s Hospital, Department of Medicine, Boston, MA, USA c Beth Israel Deaconess Medical Center, Department of Medicine, Boston, MA, USA d Harvard Medical School, Boston, MA, USA e Boston University School of Medicine, Boston, MA, USA f Cooperative Studies Program Coordinating Center, VA Medical Center, West Haven, CT, USA b
Received 19 March 2012; received in revised form 18 June 2012; accepted 3 July 2012 Available online 16 August 2012
KEYWORDS Homocysteine; Heart failure; Diastolic dysfunction; Chronic kidney disease
Abstract Background & aims: Hyperhomocysteinaemia (HHCY), a common finding in patients with chronic kidney disease (CKD), has been shown to contribute to adverse cardiac remodelling and failure. We hypothesised that in human subjects with CKD, HHCY would be associated with myocardial dysfunction, and that homocysteine (HCY)-lowering therapy would improve myocardial remodelling and heart-failure (HF) outcomes. Methods and results: Post hoc analysis of the Homocysteinemia in Kidney and End Stage Renal Disease (HOST) trial (n Z 2056) was performed to determine if HCY-lowering therapy with high dose B vitamins affects HF outcomes in patients with CKD. In addition, effects on myocardial remodelling were assessed in a subgroup of 220 trial subjects who had transthoracic echocardiograms done before study randomisation and during the course of the study as part of their routine clinical care. HF outcomes were not significantly affected by treatment compared to the placebo. HCY levels were inversely correlated with diastolic function (R Z 0.21; p Z 0.038). Vitamin therapy resulted in a significant increase in left atrial size (þ0.15 0.8 cm vs. 0.13 0.07 cm; p Z 0.0095). No other echocardiographic parameters were significantly associated with baseline HCY levels or changes with vitamin therapy. Conclusion: HHCY is associated with diastolic dysfunction in patients with CKD. However, B-vitamin therapy did not improve HF outcomes despite lowering of plasma HCY levels, and
* Corresponding author. Cardiology Section (111), VA Boston Healthcare System, 1400 VFW Parkway, West Roxbury, MA 02132, USA. Tel.: þ1 857 203 6841; fax: þ1 857 203 5550. E-mail address: [email protected] (J. Joseph). 0939-4753/$ - see front matter Published by Elsevier B.V. http://dx.doi.org/10.1016/j.numecd.2012.07.002
Adverse myocardial effects of B-vitamin therapy
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was associated with an increase in left atrial size, which is a surrogate for worsening left ventricular diastolic dysfunction. These findings suggest that high-dose B vitamin therapy may be harmful in patients with CKD. Published by Elsevier B.V.
In 1969, Kilmer McCully proposed the theory that a perturbation of the methionineehomocysteine cycle, manifested biochemically as Hyperhomocysteinaemia (HHCY), increased the risk for cardiovascular disease [1]. Numerous observational studies have demonstrated positive correlations between elevated plasma homocysteine (HCY) levels and adverse cardiovascular outcomes, including myocardial infarction (MI) and stroke [2,3]. However, no large-scale prospective trial has convincingly shown that effectively lowering plasma levels of HCY decreases rates of MI, stroke or mortality [4e7]. Interestingly, recent findings from preclinical animal studies have suggested that diet-induced HHCY may directly affect the myocardium, independent of its effects on the vascular system. Work done in our laboratory and those of others has demonstrated that HHCY increases myocardial collagen content, adverse myocardial remodelling, and diastolic dysfunction in rat models [8e10]. While these findings have not been directly confirmed in humans, small observational studies suggest that a similar phenomenon may exist. HHCY correlates with increases in left ventricular (LV) mass and wall thickness [11,12], and is associated with increased risk and severity of clinical heart failure (HF) [13,14]. Whether circulating HCY serves as a biomarker for worsening HF or is a causal factor in its pathophysiology, however, remains unclear. Patients with chronic kidney disease (CKD) represent a unique cohort for studying the potential relationship between HHCY and HF given the increased prevalence of both of these disease states in this patient population [15e17]. Elevation in HCY levels in patients with CKD has been attributed to reduced renal clearance of circulating HCY and impaired HCY metabolism [18,19]. Likewise, similar mechanisms of deteriorating renal function in HF have been proposed to be the link between HF and HHCY [20,21]. To further investigate the relationship between HHCY and HF in human subjects, we performed post hoc analysis of the Homocysteinemia in Kidney and End Stage Renal Disease (HOST) trial, a study which evaluated cardiovascular outcomes of HCY-lowering vitamin therapy in patients with concomitant CKD and HHCY [7]. We specifically examined the relations of echocardiographic evidence of myocardial remodelling and clinical HF outcomes (which had not been evaluated in the HOST trial) with baseline HCY levels and vitamin therapy.
Methods Study population Details of the HOST study have been previously published [7]. In brief, the study was a prospective, randomized, double-blind study designed to determine if HCY-lowering therapy, that is, a combination of high doses of folic acid,
vitamin B12 and vitamin B6, could reduce all-cause mortality and cardiovascular outcomes in patients with CKD. Participants (n Z 2056) were recruited from 36 US Department of Veterans Affairs medical centres from the years 2001 to 2006, and followed for a median of 3.2 years on either placebo or vitamin treatment. Inclusion criteria included age >21 years, advanced CKD (eGFR 30 ml min1) (n Z 1305) or end-stage renal disease (ESRD; n Z 751), and high plasma HCY levels (15 mM). HF outcomes were not evaluated in this study, but were available for review through the electronic Computerized Patient Record System (CPRS) of the Veterans Affairs Healthcare System. All procedures in this study were approved by the HOST Executive Committee and the Institutional Review Board of Veterans Affairs Boston Healthcare System.
Data collection Data on baseline history of HF and HF-related hospitalisations during the study period were collected for all subjects included in the original study. Discharge summaries were reviewed and classified as evidence of HF at baseline or during follow-up if they included any of the following ICD9 codes: any 428.X, 276.6, 518.4, and 514. In addition, all clinical variables, that is, mortality, used in the original HOST study, were also used for analysis. For the echocardiographic subgroup analysis, we identified 220 subjects (out of a total of 2056 subjects in the HOST study) who, as a component of their routine clinical care, had an echocardiogram within 6 months before randomisation, and 6 months or longer after randomisation to placebo or vitamin treatment. The following echocardiographic parameters were collected and analysed as continuous variables: LV ejection fraction (EF); LV wall thicknesseposterior wall (PW) and interventricular septum (IVS); LV end-diastolic dimension (LVEDD); LV end-systolic dimension (LVESD); and left atrial (LA) size. Diastolic function was evaluated as a categorical variable (normal or abnormal) since all medical centres involved did not report individual components of diastolic function assessment.
Statistical analysis Relationship of plasma HCY level and vitamin treatment to HF outcomes We first analysed the data to determine if there was a correlation between baseline HF prevalence and baseline plasma HCY levels by using analysis of covariance (ANCOVA). Subjects were stratified into advanced chronic kidney disease (ACKD) or ESRD based on the dialysis status at HOST study enrollment. For all study subjects, we used Cox proportional hazards regression survival analyses to compare vitamin and placebo groups for differences in the primary composite endpoint of HF-related hospitalisation
838 or mortality. A secondary endpoint of HF-related hospitalisation, alone, was also evaluated. Analysis was adjusted for the following confounding variables: age, gender, race, strata (ACKD vs. ESRD), diabetes mellitus, haemoglobin levels, hypertension, history of MI, baseline medication use (angiotensin-converting enzyme inhibitors, angiotensin receptor blockers, beta-blockers and aldosterone inhibitors), and baseline HF. An interaction term of treatment with baseline HF status was included in the model to test if the whole cohort could be analysed together or whether the effect of treatment needed to be examined separately in those with and without a history of HF. Relation of plasma HCY level and vitamin treatment to cardiac structure and function (echocardiographic substudy) Baseline clinical and detailed echocardiographic data during the study period were available for a total of 220 patients. We compared the baseline clinical and treatment characteristics for the subgroup population (n Z 220) versus those not included (n Z 1836). Analysis was then conducted to determine if baseline echocardiographic parameters correlated with plasma HCY levels for the whole subgroup. Univariate linear regression was performed on each echocardiographic parameter to examine its correlation to the baseline plasma HCY level. For diastolic dysfunction, logistic regression was used. Subsequently, we determined if a change in each echocardiographic parameter between values obtained before randomisation and during the study were different between the placebo and vitamin groups (separate tests for each variable). Parameters evaluated were changes in LV wall thickness, LVEF, LVEDD, LVESD, diastolic function, and LA size. Since the date of echocardiogram was available, analysis was adjusted for differences in the time from randomisation when a repeat echocardiogram was performed. ANCOVA was then performed with baseline echocardiogram and days from the date of randomisation until the date of repeat echocardiogram included as covariates, i.e., Change in echo Z baseline echo þ days þ treatment group þ error.
Z. Rafeq et al. Table 1 Relation of baseline plasma HCY levels and vitamin treatment to clinical HF outcomes. Outcome
Baseline HCY
Vitamin treatment
Hazard ratio (95% C.I.)
Hazard ratio (95% C.I.)
Entire group HF admissions þ mortality HF admissions
1.00 (0.99e1.01); p Z 0.49 0.98 (0.96e1.00); p Z 0.09 Advanced chronic kidney disease HF admissions þ 1.01 (1.00e1.02); mortality p Z 0.16 HF admissions 0.98 (0.95e1.01); p Z 0.26 End stage renal disease HF admissions þ 1.00 (0.99e1.01); mortality p Z 0.93 HF admissions 0.98 (0.96e1.01); p Z 0.21
1.11 (0.95e1.29); p Z 0.18 0.90 (0.64e1.26); p Z 0.54 1.07 (0.91e1.26); p Z 0.40 0.75 (0.50e1.13); p Z 0.16 1.08 (0.88e1.32); p Z 0.53 0.98 (0.67e1.43); p Z 0.93
Hazard ratios with 95% confidence intervals generated from Cox proportional hazards analysis to compare vitamin and placebo groups for differences in the primary composite endpoint of HFrelated hospitalization or mortality or the secondary endpoint of HF-related hospitalizations. HCY, homocysteine; HF, heart failure.
Baseline history of HF was significantly more prevalent in the subgroup, along with higher prevalence of beta-blocker therapy, ESRD and diabetes. However, within the subgroup, there were no significant differences between subjects who had been randomised to the vitamin group (45%) and those who had been randomised to the placebo group (55%). As shown in Table 3, no significant differences in baseline age, body mass index, serum albumin, haemoglobin, diagnosis of ACKD vs. ESRD, HF, hypertension, or use of angiotensin converting enzyme inhibitors or beta-blockers were detected between the two groups.
Results Relation of plasma HCY and vitamin treatment to HF outcomes There was no significant association of baseline plasma HCY levels with baseline history of HF (HR 0.995, C.I. 0.983e1.007). As shown in Table 1, treatment with high-dose vitamin therapy did not affect HF-related hospitalisations or mortality in ACKD or ESRD groups during the course of the study. The total number of events in the study population was 986 HF þ mortality events and 207 HF events.
Characteristics of the echocardiographic subgroup analysis Table 2 demonstrates the significant differences in baseline characteristics between the echocardiographic subgroup and the subjects of the HOST trial who were excluded from this substudy based on the lack of echocardiograms.
Table 2 Baseline demographic data of echocardiographic subgroup vs. original HOST study. Variable
HOST subjects included in subgroup (n Z 220)
HOST subjects excluded from subgroup (n Z 1836)
p-Value
Age (year) ESRD Serum albumin (g/dl) Baseline HF Diabetes Beta-blocker use
64.2 11.6 115 (52%) 3.9 0.5
66.0 11.8 636 (35%) 4.0 0.5
0.034 <0.0001 0.004
73 (33%) 136 (62%) 155 (71%)
413 (23%) 993 (54%) 1194 (57%)
0.0005 0.033 0.0001
Data shown as mean SD or total number subjects with percentage of subgroup indicated in parentheses. ESRD, end stage renal disease; HF, heart failure.
Adverse myocardial effects of B-vitamin therapy Table 3 Baseline demographic data of placebo and vitamin therapy groups within the echocardiographic subgroup. Variable
Placebo
Vitamin treatment
p-Value (t-test)
Treatment Age (years) BMI (kg/m2) Hemoglobin (g/dl) Serum albumin (g/dl)
121 (55%) 64.5 9.9 27.2 4.9 11.4 1.6
99 (45%) 63.8 10.5 27.3 4.1 11.8 1.4
0.62 0.85 0.12
3.8 0.5
3.8 0.5
0.78
Variable
Placebo
Vitamin treatment
p-Value (Chi square test)
ACKD ESRD HF Hypertension Beta-blockers ACE inhibitor
60 (49.6%) 61 (50.4%) 42 (34.7%) 115 (95%) 85 (70.3%) 60 (49.6%)
45 54 31 93 70 39
0.54
(45.5%) (54.5%) (31.3%) (93%) (70.7%) (39.4%)
0.59 0.72 0.94 0.13
Data shown as mean SD or total number of subjects with percentage of subgroup indicated parentheses. BMI, body mass index; ACKD, advanced chronic kidney disease; ESRD, end stage renal disease; HF, heart failure; ACE, angiotensin converting enzyme.
Lowering of plasma homocysteine level in the echocardiographic subgroup All subjects in the echocardiogram subgroup had a plasma HCY level >15 mM, which was an inclusion criterion for the HOST trial. The mean plasma HCY levels in subjects with
Table 4
ACKD (n Z 105) was 23.9 mM (range 16e45), while that in the ESRD patients (n Z 115) was slightly higher (25.1 mM; range 15e67). Table 4 demonstrates the plasma HCY and folate levels in placebo and vitamin groups at 3 months and 1 year after randomisation. Since measurements were done only in a selected subset of patients after the 3-month assessments, only a small number of the substudy subjects had measurements at 1 year. Compared to baseline values, plasma HCY levels were reduced significantly in the vitamin group at 3 months and at 1 year after initiation of treatment. Plasma folate values were also increased significantly in the vitamin group at 3 months and 1 year. The placebo group did not demonstrate a significant change in plasma HCY or folate levels at either time point.
Effects of vitamin therapy on echocardiographic parameters Univariate regression analysis of the correlations of baseline echocardiographic measurements and pre-treatment HCY levels demonstrated a significant inverse relationship between plasma HCY levels and diastolic function (R Z 0.21, p Z 0.038). In the echocardiographic subgroup, 86% of the subjects had abnormal diastolic function at baseline. An inverse correlation of borderline significance was seen with LA size (R Z 0.14, p Z 0.08). There were no significant correlations between plasma HCY level and the other echocardiographic parameters measured. We examined whether there were any significant changes in any echocardiographic parameter during the study period compared to baseline values, in the placebo and vitamin groups. Table 5 shows the results of multivariate regression analyses that included determinants of changes in echocardiographic parameters during the study period, including baseline value, days to follow-up
Plasma total homocysteine and folate at baseline and follow-up in substudy population.
Total homocysteine e mmol/L Placebo group: N Median (Q1, Q3) Vitamin group: N Median (Q1, Q3) p-Valuea Folate e ng/mLb Placebo group: N Median (Q1, Q3) Vitamin group: N Median (Q1, Q3) p-Valuea
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Baseline
3 Months e baseline
1 Year e baseline
121 22.0 (19.0, 26.0)
116 0.6 (3.7, 2.3)
16 0.3 (2.7, 3.9)
99 22.0 (18.3, 26.7)
94 5.4 (9.2, 2.3)
12 4.3 (7.3, 0.7)
<0.0001
0.04
116 18.4 (11.7, 25.0)
111 1.5 (5.3, 14.5)
16 4.2 (11.0, 19.1)
97 17.4 (9.6, 25.0)
92 1616 (88, 3736)
12 1954 (2, 5688)
<0.0001
0.01
Q1, Q3 denotes values of the first and third quartile. To convert values for homocysteine to mg/liter divide by 7.397. To convert values for folate to nanomoles per liter multiply by 2.266. a Wilcoxon rank sum test p-value. b Tests results reported at baseline as >25 were analyzed as 25. This was because baseline values were obtained from different laboratories, several of which had cutoffs of 25. Subsequent analyses were done in a single laboratory with no cutoff.
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Table 5 Associations of B-vitamin therapy with changes in echocardiographic parameters during the study period. Echocardiographic parameter
Treatment effect
DLVEF DLVIVS DLVPW DLVEDD DLVESD DLA size DDiastolic function
0.28 0.08 0.08 0.09 0.15 0.27 0.09
(2.0) (0.04) (0.04) (0.13) (0.18) (0.1)*** (0.08)
Multivariate regression analysis including baseline value, days to follow-up echocardiogram and treatment group as variables. Data shown as parameter estimate (SE). ***p Z 0.0095. LV, left ventricle; EF, ejection fraction; IVS, intraventricular septal wall thickness; PW, posterior wall thickness; EDD, end-diastolic dimension; ESD, end-systolic dimension; LA, left atrium.
echocardiogram and treatment groups as covariates. The only change during the study period that was independently associated with HCY-lowering vitamin therapy was an increase in LA size. Further multivariate regression analysis adjusted for clinical variables and variables that affect LA size, such as baseline LV and LA size, days to follow up echocardiogram, baseline mitral regurgitation (MR) and change in degree of MR during follow-up, demonstrated a significant increase in LA size during the study period in the vitamin group (þ0.15 0.08 cm) compared to the placebo group (0.13 0.07 cm; pZ0.0095).
Discussion Our post hoc analysis of the HOST trial used echocardiographic data and clinical HF outcomes to investigate the relationship between HHCY and HF in human subjects with CKD. The major findings of this study are: (1) HHCY is correlated with myocardial diastolic dysfunction, (2) highdose B-vitamin therapy does not improve HF outcomes despite lowering of plasma HCY levels and (3) HCY-lowering B-vitamin therapy is associated with cardiac structural changes (i.e., left atrial enlargement), suggestive of worsening diastolic dysfunction, in patients with CKD. The correlation detected between HHCY and baseline diastolic dysfunction corroborates findings of previous studies, which have proposed that an alteration of the methionineehomocysteine cycle, manifested as HHCY, may be an independent risk factor for the development of both systolic and diastolic forms of HF [13,21,22]. The mechanism by which this occurs, however, is not well elucidated. Preclinical animal studies have suggested that an alteration in the methionineehomocysteine cycle has direct effects on the myocardium, leading to increased fibrosis and collagen deposition, and eventual myocardial stiffening and pump failure [8e10]. To ascertain if HCY directly contributes to HF pathophysiology in humans, one would have to demonstrate improvements in either myocardial remodelling or clinical HF outcomes with HCY-lowering therapy in patients with known HHCY. We focussed on both these parameters in our post hoc
analysis of the HOST trial. Similar to the HOPE-2 trial [6], we did not detect a significant improvement in HF-related hospitalisations or mortality with vitamin therapy in the HOST study. Even though veterans generally receive all of their care in the Veterans Affairs Health System, there is a small possibility that some events may not have been captured in this analysis. However, our echocardiographic subgroup analysis did reveal an unexpected finding contrary to our original hypothesis. Rather than improving myocardial remodelling, aggressive HCY-lowering vitamin therapy in patients with CKD was associated with a significant increase in LA size, a surrogate marker of worsening diastolic dysfunction. While the available echocardiographic data of the HOST trial subjects were not adequate to analyse quantitative changes in diastolic function, LA dimensions have been reported to be an accurate determinant of progression of diastolic HF [23,24]. Hence, it is quite likely that this was a reliable indicator that adverse cardiac remodelling was occurring in association with vitamin therapy. To our knowledge, this is the first study to investigate the effects of HCY-lowering treatment on myocardial remodelling in HF. Thus, we feel this finding raises an important question of whether unrestricted vitamin supplementation is safe in patients with CKD. The unforeseen harmful effects of vitamin therapy on cardiac remodelling detected in our study underscores the fact that an elevated plasma HCY level likely represents more than just a single defect in a complex process of sulphur aminoacid metabolism and affects redox balance and methylation reactions. HCY metabolism occurs via two interrelated processes of remethylation, which requires folic acid and vitamin B12 as cofactors, and transulphuration, which requires vitamin B6 as a cofactor. While it is difficult to determine from our study why HCY-lowering vitamin therapy resulted in adverse cardiac remodelling in our study population, we propose that patients with advanced CKD may be more susceptible to acute changes in HCY levels given intrinsic deficiencies in their ability to metabolise HCY. Patients with CKD have underlying defects in both DNA and protein methylation, and have specifically been shown to demonstrate impairments in HCY remethylation [25,26]. Thus, acutely increasing HCY remethylation with folic acid and vitamin B12 supplementation could affect any one of multiple intracellular methylation processes which are dependent on the methionineehomocysteine cycle. In addition, given their underlying impairments in methylation, patients with CKD are likely more dependent upon the alternative pathway of HCY transulphuration, which notably plays a key role in the generation of the anti-oxidant glutathione (via cysteine). Patients with CKD and HF have increased basal levels of chronic inflammation and oxidant stress [27,28]. Thus, it is possible that by redirecting HCY metabolism towards remethylation via folic acid and vitamin B12 supplementation, glutathione production would be particularly suppressed in patients with CKD, which could result in increased oxidant stress in the myocardium. Lastly, it is possible that vitamin therapy may directly affect the myocardium in CKD patients, independent of its effects on HCY-lowering. Folic acid therapy in patients with ESRD has been shown to affect DNA methylation and gene expression without altering plasma HCY levels [29].
Adverse myocardial effects of B-vitamin therapy While further work is necessary to elucidate why vitamin therapy potentially promotes adverse cardiac remodelling in patients with advanced CKD, it is also important to recognise that certain patient populations likely respond to seemingly benign vitamin therapies differently, and that these variable responses can be harmful. A large, metaanalysis investigating the effects of HCY-lowering folate therapy found potential harm in terms of cardiovascular disease in patients with elevated HCY levels, but not in those with normal-to-minimally elevated HCY levels [30]. Whether these patients with higher HCY levels represented a similar demographic to subjects studied in the HOST trial is unclear. However, if so, this would support the notion that folic acid therapy in patients with advanced CKD has potentially harmful effects on cardiac function. While our post hoc analysis of the HOST study does raise interesting concerns about unrestricted vitamin use, there are multiple limitations to our study which must be noted. First, the HOST trial was limited to a select cohort of patients with advanced or end stage kidney disease, which does not allow us to apply our findings to a more general population. Furthermore, in contrast to prior studies, we did not detect a significant correlation between baseline HCY levels and HF. This was likely influenced by the fact that all subjects in the HOST trial had significantly elevated plasma HCY levels, an inclusion criterion of the trial, which did not allow us to compare HF-related outcomes in patients with normal or low plasma levels of HCY. Second, echocardiograms were done as part of routine clinical care and, thus, were not standardised for this study. Thus, multiple additional parameters could not be evaluated. Specifically, we could not quantify changes in diastolic dysfunction since various methods to evaluate diastolic functions were used by different institutions, and LA size, as opposed to a more specific LA volume, had to be used to assess for changes in diastolic dysfunction. In addition, since the echocardiograms were done for clinical indications, there may be some selection bias, with those with more severe clinical manifestations more likely to undergo echocardiography. Another limitation of our study is that we do not know how long the subjects have been exposed to high levels of HCY before randomisation to vitamin therapy or placebo, which could have an influence on the effects of therapy. In summary, our post hoc analysis of the HOST trial showed that HHCY correlates with diastolic dysfunction. However, in patients with advanced CKD, aggressive HCYlowering vitamin therapy does not improve clinical HF outcomes, and may actually be detrimental in terms of worsening adverse cardiac remodelling and diastolic dysfunction. Given the complexity of HCY metabolism in kidney disease, further mechanistic and prospective studies need to be conducted before recommendations regarding HCY-lowering vitamin therapy can be established in patients with CKD.
Acknowledgements The HOST study was supported by the Cooperative Studies Program, Department of Veterans Affairs Office of Research and Development; PamLab; and Abbott Laboratories.
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