Low vitamin D levels and the long‐term functional outcome of stroke up to 5 years

Abstract Background and purpose Previous studies have established that vitamin D was associated with stroke. The purpose of this study was to investigate the relationship between vitamin D and 5‐year outcome of patients with stroke including acute ischemic stroke (AIS) and intracranial hemorrhage (ICH) stroke. Methods Serum 25‐hydroxyvitamin D levels were prospectively analyzed in patients admitted to the First Affiliated Hospital of Wenzhou Medical University from 2013 to 2015. Modified Rankin scale (mRS) was used to evaluate their 5‐year functional outcome, and univariate and multivariate logistic regressions were applied to evaluate the effects of vitamin D on stroke outcome. Results In total, 668 patients diagnosed with stroke were recruited, and 420 completed the 5‐year follow‐up. Ninety‐five patients experienced poor outcome in the 5 years since stroke onset. Vitamin D levels in patients with poor outcome showed significant differences compared to good outcome patients (p < .001). In multivariable logistic regression analysis, after adjusting the potential confounders, the 5‐year functional outcome was significantly associated with vitamin D levels. Stroke patients with vitamin D levels less than 38.4 nmol/L had a higher risk for poor outcome compared with those with vitamin D level over 71.4 nmol/L at 5‐year (odds ratio [OR] = 3.66, 95% confidence interval [CI] = 1.42–9.45, p = .007), which was consistent with AIS patients (OR = 6.36, 95% CI = 1.89–21.44, p = .003). Conclusion Vitamin D level less than 38.4 nmol/L at admission is a potential risk biomarker for poor functional outcome at 5‐year prognosis in AIS patients, which might provide new ideas for the prognostic assessment of stroke.


INTRODUCTION
Stroke is generally considered as the second leading cause of death and a first leading cause of adult disability worldwide. Around 2.5 million people are suffering from stroke each year in China, laying heavy burden on the families, communities, and healthcare systems (Holick et al., 2011;Wang et al., 2014). Survivors often need long-term rehabilitation, but most stroke survivors still lead to poor outcomes. Therefore, early identification of risk factors for poor outcome and improving the functional prognosis are essential.
The recent research on stroke outcomes suggests that several factors related to the outcome of stroke had been identified, including age, sex, stroke severity, etiology, and cardiovascular risk factors (Robinson et al., 2019;Siegler et al., 2013;Torres-Aguila et al., 2019;Werner et al., 2005). Other studies also suggested different stroke outcomes varied depending on the age stratification (Buell & Dawson-Hughes, 2008;Razmara et al., 2017;Ueno et al., 2018). However, there is limited research on the relationship between vitamin D and long-term functional outcomes of stroke.
Cholecalciferol (Vitamin D3) is the main form of vitamin D in nature, which is formed by the animals and human skin as a reaction to sunlight and could be obtained through diet (Krishna, 2019). Given that the cutaneous production of vitamin D decreases with age and the older are less exposed to sun, vitamin D deficiency becomes more common in stroke patients (Krishna, 2019;MacLaughlin & Holick, 1985). Previous studies have shown that lower 25-hydroxyvitamin D (25(OH)D) levels were associated with poorer functional outcomes and all-cause mortality in stroke patients (Chen et al., 2018;Daubail et al., 2013;Park et al., 2015;Tu et al., 2014;Wei & Kuang, 2018). Recent evidence has also suggested the negative results of low 25(OH)D levels on the survival of patients with different diseases, including myocardial infarction (Correia et al., 2013), heart failure (Liu et al., 2011), and chronic kidney disease (Pilz, Iodice, et al., 2011). Most studies on vitamin D and its outcomes have only focused on the short-term prognosis after stroke.
Large-scale studies are in high demand concerning the functional prognostic value of stroke patients with serum 25(OH)D after 5 years.
To date, the relationship between vitamin D and functional outcome 5 years after stroke has not been illustrated, and it still remains unclear whether vitamin D has a strong effect on long-term prognosis. As a result, this study was designed to investigate the association between vitamin D stratification and outcome among acute ischemic stroke (AIS) and intracranial hemorrhage (ICH) stroke after 5 years.  (Sun et al., 2006). According to the 17-item Hamilton Depression Scale (HAMD) to assess depression status after admission, all subjects were classified as having either post-stroke depression (PSD) (HAMD score ≥ 7) or non-PSD (PSD) (HAMD score < 7) (Zhang et al., 2017).

Serum 25(OH)D
The serum concentration of 25(OH)D was measured using a compet-

Outcome assessment
The primary targeting outcome was assessed by modified Rankin scale (mRS) on follow-up. The previous study has defined poor outcome as an mRS>2 (Pilz, Iodice, et al., 2011). Therefore, we defined the mRS as a dichotomous variable, an mRS score in the 0-2 range, which indicates no symptoms to a minimal disability, reflects good outcome, whereas the 3-6 range reflects moderate to severe disability or death, reflecting a poor outcome.

Statistical analysis
The relation of patient's baseline information and risk factors were statistically analyzed.

Baseline characteristics
After application of the eligibility criteria, 668 of 839 screened patients with stroke met the inclusion criteria and were enrolled in the study.

TA B L E 3
Multivariate adjusted odds ratios for the association between serum levels of 25 (OH) D and poor outcome in all stroke patients

DISCUSSION
To date, it has been recognized that there were few large-scale research to examine vitamin D levels and functional outcome of stroke patients with 5-year follow-up. In the present prospective observational study, we found 22.7% of acute stroke patients had poor outcome, and decreased 25(OH)D levels at baseline were independently associated with increased risk of poor outcome at 5 years after stroke in AIS, but not the ICH patients. This association was weakened after adjusting for HAMD score at admission, thus the effect of vitamin D levels has on functional outcome may be partially mediated by altering depression symptoms.
There were several population-based reports on the relationship between vitamin D deficiency and cardiovascular disease, obesity,  (Vila et al., 2003). Vitamin D produces a series of reactions referring to the anti-inflammatory reaction by Treg and Th2, cytokine production, and pro-inflammatory cytokine suppression, for instance, interleukin-1, interleukin-6, tumor necrosis factor-alpha, and Toll-like receptors.
On the other hand, Vitamin D has been shown to reduce ischemiainduced brain damage and suppress dysregulation of the inflammatory response after ischemia (Di Rosa et al., 2012;Wang et al., 2000).
Consistently, in vivo studies also revealed that vitamin D deficiency exacerbates the proinflammatory state and suppressed neuroprotectants such as insulin-like growth factor-I (IGF-I) (Balden et al., 2012).
Second, the main function of vitamin D is to regulate bone metabolism and maintain calcium-phosphate homeostasis. Deficiency of vitamin D contributes to altered bone mineralization and low bone mass and accelerates bone resorption and reduction of bone density in patients with stroke (Lips & van Schoor, 2011;Pilz, Tomaschitz, et al., 2011;Yoshida & Stern, 2012).
Third, in vivo studies revealed that vitamin D3 supplementation decreased the volume of cerebral infarct, suggesting that vitamin D could exert neuroprotective effects (Milionis et al., 2014;Wang et al., 2000). Meanwhile, Vitamin D also upregulates the neurotrophic factors, such as nerve growth factor, neurotrophin-3, neurotrophin-4, and IGF-I (Bogazzi et al., 2011). IGF-I plays a significant role in post-stroke recovery through regulating the repair of the central nervous system by promoting the germination and regeneration of axons and dendrites after stroke, and supporting the growth and survival of nerve progenitors, astrocytes, and microglial cells . It is also reported that IGF-I may modulate the immune environment of the ischemic brain (Balden et al., 2012).
Additionally, Vitamin D deficiency increases the risk of cardiovascular disease by activating the renin-angiotensin system, leading to hypertension (Ginde et al., 2009;Lee et al., 2008). Vitamin D is also linked to diabetes, insulin resistance, and high BMI, which are risk factors for cardiovascular disease and all-cause mortality. On the other hand, Vitamin D deficiency could give rise to poorer cognitive function and dementia, which affect the executive function and daily life of patients (Chen et al., 2018;Goodwill & Szoeke, 2017;Slinin et al., 2010).
Previous research have shown that lower vitamin D levels are associated with depression and AIS (Berridge, 2017;Hoogendijk et al., 2009;Larsson et al., 2018). Our team's previous study also have reported that vitamin D is an important biomarker for the occurrence of PSD (Han et al., 2015). In this study, after adjusting for HAMD score at admission, the effect of vitamin D levels on functional outcome was weakened. Therefore, the risk of poor outcome may be influenced by the interaction between vitamin D levels and depressive state.
However, in a latest double-blind placebo-controlled trial, vitamin D3 supplementation did not improve the recovery after acute stroke (Momosaki et al., 2019). There are no robust clinical trials regarding vitamin D supplementation to alter stroke outcome. Therefore, the underlying mechanism of the effect imposed by 25(OH) D on poor outcome may involve several convoluted pathological pathways, which remain to be elucidated.
There are still limitations in this study. The data were collected from a single study center and the sample size is still limited; a larger sample size would provide better sensitivity for the detection between the vitamin D levels and outcome. In addition, patients enrolled in the study were mild in most (median of NIHSS score was 3); the present findings could not be extrapolated to all AIS patients, and future research should add sample size to simultaneously assess the impact of vitamin D on different severity of stroke. Third, because the data were from past 5 years and most of the patients were mild, the number of patients involved in rehabilitation was small. Therefore, we could not evaluate the impact of rehabilitation in the observed associations. We did not have data on living habits such as exercise and diet, which may confound our conclusions.

CONCLUSION
In conclusion, this study has demonstrated that lower vitamin D levels are of long-term prognostic significances after AIS within a 5- year follow-up. Vitamin D lower than 38.4 nmol/L is an independent prognostic marker for poor outcome after adjusting for possible confounding factors. Further analyses are anticipated to investigate the pathophysiological mechanisms and the role of vitamin D supplementation in the long-term prognosis of stroke patients.

ACKNOWLEDGMENTS
We thank all collaborators for data collection. We also thank all patients who participated.

CONFLICT OF INTEREST
The authors declare no conflict of interest.

AUTHOR CONTRIBUTIONS
Ya-Ying Zeng and Jin-Cai He designed the study. Ya-Ying Zeng and Cheng-Xiang Yuan wrote the manuscript. Ya-Ying Zeng did the statistical analyses. Meng-Xuan Wu, Lin Cheng, Sheng-Nan Zhou, Kai-Li Fan, and Ping-lang Hu screened and extracted data. Jin-Cai He and Wen-Jie Tang supervised study. All authors have made an intellectual contribution to the manuscript and approved the submission.

DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions.

PEER REVIEW
The peer review history for this article is available at https://publons. com/publon/10.1002/brb3.2244