Keywords
Homocysteine - Intracranial Aneurysm - Subarachnoid Hemorrhage - Stroke
Palavras-chave
Homocisteína - Aneurisma Intracraniano - Hemorragia Subaracnóidea - Acidente Vascular
Cerebral
INTRODUCTION
Homocysteine (homocysteine) is a non-essential amino acid involved in the methionine
metabolism pathways.[1] Homocysteine has been implicated in several vascular pathologic processes since
1969, with the observation of children with homocystinuria who had atherosclerosis,
myocardial infarction, stroke, and other atherothrombotic conditions at a very early
age.[2]
High levels of homocysteine are robustly associated with stroke.[3]
[4]
[5] Several meta-analyses have demonstrated an increased risk of ischemic stroke in
patients with hyperhomocysteinemia, which motivated the scientific community to undertake
randomized controlled trials of B12 and folate supplementation for stroke prevention.[5]
[6]
[7]
There is very little data regarding homocysteine's influence on the formation and
rupture of intracranial aneurysms, and the results are conflicting.[8]
[9]
[10]
[11] The objective of the present study is to compare serum levels of homocysteine between
patients with ruptured and unruptured IA. Furthermore, we sought to evaluate the possible
influences of this molecule on vasospasm and functional outcomes.
METHODS
This is a retrospective, case-control methodology. An exhaustive review of medical
records was conducted for all sequential patients who sought consultation at the cerebrovascular
neurosurgery clinic of Hospital das Clínicas da Faculdade de Medicina da Universidade
de Sao Paulo (HC-FMUSP). The research project received approval from the HC-FMUSP's
ethics review board, consistent with the principles articulated in the Helsinki Declaration,
and the necessity for informed consent was excused due to the retrospective nature
of the study (approval number 5.576.011, 2022).
Population
The study's inclusion criteria comprised patients diagnosed with intracranial aneurysms,
irrespective of rupture history, who also had homocysteine serum levels measured.
Exclusion criteria were set as patients below the age of 18, those with arteriovenous
malformations or concurrent cerebrovascular diseases excluding prior strokes, or individuals
in the acute post-stroke phase, defined as less than one-month post-discharge.
Demographic data were gathered, which incorporated variables such as age, gender,
pre-existing health conditions, current smoking habits, a history of ischemic stroke,
and statin usage. For patients with a history of ruptured aneurysms, inpatient records
were meticulously examined to detect any incidence of vasospasm, documented on angiotomography,
angiography, or transcranial Doppler. Neurological outcomes were subsequently evaluated
utilizing the modified Rankin Scale (mRS).[12]
Homocysteine levels were obtained in the outpatient environment, with a time frame
ranging from a minimum of one month to a maximum of six months post-discharge. For
patients with unruptured aneurysms, serum levels were obtained when first evaluated
in the outpatient clinic before any treatments were considered. Serum homocysteine
concentrations were considered as both a continuous and a dichotomous variable - hyperhomocysteinemia
was classified as a homocysteine level ≥ 15 µmol/L (mild), homocysteine level ≥ 30
µmol/L (moderate), or homocysteine level > 100 µmol/L (severe). Since folate and vitamin
B12 play important roles in the physiology of homocysteine, data on these vitamins
were collected, to correct for potential confounding.
Outcomes
The primary outcomes encompassed:
-
• disparities in serum homocysteine between patients with ruptured and unruptured
aneurysms;
-
• the impact of homocysteine concentrations on functional status;
-
• the effect of homocysteine levels on vasospasm occurrence.
Statistical analyses
Continuous data are depicted as either mean (±standard deviation) or median (interquartile
range), subject to the results of normality assessments (Shapiro-Wilk's test). T-tests
were employed for comparing normally distributed continuous variables, while non-parametric
variables were compared using the Mann-Whitney U tests. Categorical variables are
represented as frequencies (valid %) and underwent comparison using chi-squared tests.
To evaluate the effect of homocysteine on binary outcomes – aneurysm rupture, vasospasm,
and excellent functional outcome (defined as mRS ≤ 1), logistic regression analyses
were performed. Covariates were incorporated into multivariable analyses on the basis
of imbalances in baseline characteristics and the plausibility of their biological
impact. Interactions were tested and non-linear correlations were assessed in accordance
with diagnostic graphs for the models, where appropriate. All analyses were executed
using R (R Foundation for Statistical Computing, Vienna, Austria, 2021).
RESULTS
Population characteristics
The characteristics of the study population are summarized in [Table 1]. This study included a total of 348 participants, of which 114 (32.8%) had experienced
a previous aneurysm rupture and 234 (67.2%) had unruptured aneurysms. The average
age of patients with ruptured aneurysms was 53.9 (±13.0) years, while those with unruptured
aneurysms had a mean age of 58.6 (±12.0) years (P < 0.01). Females comprised 73.7%
and 78.2% of the ruptured and unruptured patient groups, respectively (P = 0.42).
Table 1
Baseline characteristics
|
Characteristics
|
Ruptured (n =114)
|
Unruptured (n = 234)
|
P-value
|
|
Age
|
53.9 (±13.0)
|
58.6 (±12.0)
|
< 0.01
|
|
Female
|
84 (73.7%)
|
183 (78.2%)
|
0.42
|
|
Smoking
|
35 (30.7%)
|
71 (30.3%)
|
1.00
|
|
Hypertension
|
31 (27.2%)
|
81 (34.6%)
|
0.20
|
|
Previous ischemic stroke
|
0 (0%)
|
12 (5.1%)
|
0.03
|
|
Dyslipidemia
|
26 (22.8%)
|
63 (26.9%)
|
0.49
|
|
Statin use
|
4 (3.5%)
|
22 (9.4%)
|
0.08
|
Notes: Data presented as mean (±standard deviation) or count (valid percentage). P-values
refer to Welch t-test or Chi-squared tests.
Current smoking was reported by 30.7% and 30.3% (P = 1.0), a diagnosis of hypertension
was established in 27.2% and 34.6% (P = 0.2), dyslipidemia was identified in 22.8%
and 26.9% (P = 0.49), and statin usage was noted in 3.5% and 9.4% (P = 0.08) of the
patients with ruptured and unruptured aneurysms, correspondingly. A history of prior
ischemic stroke was confirmed in 12 (5.1%) patients with unruptured aneurysms, with
no recorded instances within the ruptured aneurysm group (P = 0.03). With the exception
of age and the incidence of prior ischemic stroke, no significant discrepancies were
observed between the groups in relation to the baseline characteristics.
Ruptured versus unruptured aneurysms
The median homocysteine level was 10.75 µmol/L (IQR = 4.59) in the ruptured aneurysm
group, and 11.5 µmol/L (IQR = 5.84) in the unruptured group (OR 0.98, 95% CI 0.94-1.02).
[Table 2] further delineates median levels of B12 and folate, both of which were scrutinized
to account for potential confounding effects on homocysteine levels. Neither were
found to be associated with aneurysm rupture: OR 1.0 (95% CI 0.99-1.01) for B12 and
OR 0.99 (0.94-1.03) for folate.
Table 2
Homocysteine, B12 and folate comparison between ruptured and unruptured aneurysms:
univariate
|
Ruptured
|
Unruptured
|
OR
|
95% CI
|
|
Homocysteine (µmol/L)
|
10.75 (4.59)
|
11.5 (5.84)
|
0.98
|
0.94-1.02
|
|
B12 (pmol/L)
|
442.5 (198)
|
393.5 (211.5)
|
1.00
|
0.99-1.01
|
|
Folate (ng/mL)
|
11.5 (5.65)
|
11.5 (7.5)
|
0.99
|
0.94-1.03
|
Notes: Median (interquartile range) of homocysteine, B12 and folate serum levels.
Odds ratios (OR) refer to univariate logistic regression models comparing serum levels
of these molecules (continuous) between patients with ruptured and unruptured aneurysms.
Multivariable analyses, compensating for disparities between groups, were conducted
([Table 3]). No significant association was detected between homocysteine levels and rupture
status (OR 0.99, 95% CI 0.96-1.04), after adjustments for age, prior ischemic stroke,
and statin use. Tested interaction terms between homocysteine and the covariates revealed
no statistical significance.
Table 3
Influence of homocysteine continuous and dichotomized on rupture of intracranial aneurysms:
multivariable analyses
|
OR
|
95% CI
|
|
Continuous
|
Homocysteine*
|
0.99
|
0.96–1.04
|
|
Dichotomized
|
Hcy > 30 µmol/L**
|
1.25
|
0.32-4.12
|
|
Hcy > 15 µmol/L**
|
1.00
|
0.54-1.81
|
|
Hcy, B12, and Folate on the same model
|
Homocysteine
|
0.98
|
0.93-1.02
|
|
B12
|
1.00
|
0.99-1.01
|
|
Folate
|
0.98
|
0.93-1.02
|
Abbreviations: CI, confidence interval; Hcy, homocysteine; OR, odds ratio.
Notes: *Adjusted for age, previous ischemic stroke, and statin use. Interaction terms
were also non-significant; ** Age-adjusted.
When homocysteine was evaluated as a dichotomous variable and adjusted for age, neither
mild (> 15 µmol/L; OR 1.25, 0.32-4.12) nor moderate (> 30 µmol/L; OR 1.0, 0.54-1.81)
hyperhomocysteinemia demonstrated significant correlations with ruptured aneurysms.
Lastly, a multivariable model incorporating homocysteine, B12, and folate was executed
([Table 3]). None of these biochemical entities revealed any significant associations with
ruptured aneurysms. Interactions terms were tested, yielding non-significant results
as well.
Vasospasm and functional outcomes
In total, fifteen patients were diagnosed with vasospasm, comprising 13.2% of the
ruptured aneurysm group. Neither univariate (OR 0.86; 95% CI 0.71-1.0) nor multivariable
age-adjusted (OR 0.91; 95% CI 0.75-1.05) models evidenced a significant association
when assessing correlations between homocysteine levels and vasospasm.
Excellent functional outcomes at 6-month follow-up (mRS ≤ 1) were achieved by 91 patients
(79.8%) within the ruptured aneurysm cohort. The logistic regression analysis did
not reveal a significant association between homocysteine levels and excellent functional
outcomes (OR 1.04; 95% CI 0.94-1.16).
DISCUSSION
There was no significant discrepancy in homocysteine levels between patients with
ruptured and those with unruptured intracranial aneurysms. Moreover, homocysteine
did not demonstrate a correlation with either vasospasm or functional outcomes post-aneurysm
rupture.
Cardiovascular risk
A potent association exists between homocysteine and vascular dysfunction, and, consequently,
cardiovascular risk. A panoply of molecular mechanisms have been implicated in homocysteine-mediated
vascular damage, including oxidative stress, production of reactive oxygen species,
thrombolysis impairment, mitochondrial modifications, and endothelial dysfunction.[1]
[13]
[14]
[15]
[16] Clinically, a 25% reduction in serum homocysteine levels has been linked with an
11% and 19% decrease in the risk of ischemic heart disease and stroke, respectively.[4] Furthermore, a study by Fan et al. (2017) even uncovered a significant dose-response
surge in all-cause mortality tied to this molecule.[17]
Stroke is also significantly associated with elevated homocysteine levels.[3]
[18]
[19] As such, clinical trials have been instigated to assess the benefits of supplementing
folic acid, B6, and B12 in order to decrease homocysteine levels and potentially reduce
stroke risk. However, the outcomes of these trials are far from consensus.[5] While some studies and meta-analyses, including the Heart Outcomes Prevention Evaluation
(HOPE) 2, SU.FOL.OM3, and the China Stroke Prevention Trial (CSPPT), indicate a reduction
in stroke risk,[5]
[20]
[21]
[22] others, such as the Vitamin Intervention for Stroke Prevention (VISP)[23] and the Norwegian Vitamin Trial (NORVIT),[24] show no such benefits or even suggest potential harm. Consequently, current guidelines
do not routinely recommend vitamin supplementation.
Intracranial aneurysm
Investigations exploring the relationship between homocysteine and intracranial aneurysms
(IA) are scarce. Preclinical evidence suggests a methionine-rich diet may correlate
with an elevated risk and expedited formation of IA,[25] and specific genetic variants of the homocysteine cycle have been linked with an
increased IA risk.[26]
[27]
Retrospective studies offer contradictory results.[28]
[29] Ren et al. (2017)[29] identified an association between homocysteine and IA patients [OR 2.2 (95%CI 1.19-4.06)],
but both patients and controls exhibited exceptionally high homocysteine levels. As
for subarachnoid hemorrhage (SAH), associations have been made with delayed cerebral
ischemia,[10] disability,[30] and SAH itself.[31] Yet, some research contradicts these findings,[9]
[32] even demonstrating an inverse correlation between homocysteine levels and mortality
after SAH.[8] Caution is urged when interpreting retrospective association studies due to potential
publication bias and an inability to establish causality.[33] This study further fortifies the notion that homocysteine may not be associated
with aneurysmal subarachnoid hemorrhage.
Strengths and limitations
This study has limitations. Its retrospective nature, although apt for exploratory
issues, inherently limits its capacity to provide robust causality information. The
decision to analyze homocysteine in an ambulatory setting post-discharge was taken
to control potential confounders associated with the acute inflammatory state, but
this also introduces bias by excluding patients who did not survive their hospital
stay.
Nonetheless, to the best of our knowledge, this is the inaugural study comparing patients
with SAH to controls with unruptured aneurysms concerning these biomolecules. The
relatively large sample size and adjusted analyses, which account for potential confounders,
constitute key strengths of this study.
In conclusion, there were no differences regarding homocysteine serum levels between
patients with ruptured and unruptured intracranial aneurysms. In patients with ruptured
aneurysms homocysteine levels did not correlate to vasospasm or functional outcomes
after discharge.
Bibliographical Record
João Paulo Mota Telles, Jefferson Rosi Junior, Vitor Nagai Yamaki, Nicollas Nunes
Rabelo, Manoel Jacobsen Teixeira, Eberval Gadelha Figueiredo. Homocysteine serum levels
in patients with ruptured and unruptured intracranial aneurysms: a case-control study.
Arq Neuropsiquiatr 2024; 82: s00441779270.
DOI: 10.1055/s-0044-1779270
