Little is known about the aetiology of renal cell carcinoma (RCC). Components of one-carbon (1C) metabolism, which are required for nucleotide synthesis and methylation reactions, may be related to risk of RCC but existing evidence is inconclusive. We conducted a systematic review and independent exposure-specific meta-analyses of dietary intake and circulating biomarkers of 1C metabolites and RCC risk.Medline and Embase databases were searched for observational studies investigating RCC or kidney cancer incidence or mortality in relation to components of 1C metabolism and 12 eligible articles were included in the meta-analyses. We used Bayesian meta-analyses to estimate summary relative risks (RRs) and 95% credible intervals (CrIs) comparing the highest versus lowest categories as well as the between-study heterogeneity.6, folate, vitamin B12, methionine, homocysteine, choline, or betaine) and RCC risk. However, vitamin B6 biomarker status did have a protective (RR = 0.62) but imprecise (95% CrI 0.39–1.14) effect estimate and folate intake had a notable association as well .We did not find convincing evidence of an association between any exposure  contains supplementary material, which is available to authorized users. Relativthionine . RCC is The aim of this review was to systematically examine the existing evidence on the association between components of 1C metabolism, both dietary intake and circulating biomarkers, and risk of RCC, and to present a quantitative summary of these relationships by conducting meta-analyses.2), vitamin B6, folate (vitamin B9), vitamin B12, methionine, homocysteine, choline, or betaine. The full search strategy for Medline is provided in Online Resource 1.Medline and Embase databases were searched on April 5th, 2019 for records of observational studies related to RCC or kidney cancer incidence or mortality and 1C metabolism. A medical school librarian was consulted for selection of search terms. The exposures were intake or circulating biomarkers of riboflavin (vitamin BResults from the database searches were combined and duplicates were removed. Two investigators (JLC and AKH) independently screened the titles and abstracts with a web-based screening software and removed ineligible papers . Papers Data collected for each paper included: author, publication year, study name and location, study design, number of cases, cohort size or number of controls, years of follow-up, sex distribution, age range, measured exposures of interest, outcome , covariates included in the most comprehensively adjusted model, effect estimate and 95% confidence interval (CI), and criteria for the Newcastle–Ottawa Scale (NOS) assessment. The summary measure of interest was the relative risk (RR); however, RCC is a sufficiently rare disease to also include odds ratios (ORs), hazard ratios (HRs), and standardised incidence ratios (SIRs) as reasonable approximations of the RR. If an article reported pooled results from more than one study, only the individual study results were used. All data were collected by one investigator (JLC) and verified by a second (AKH).After consideration of several quality assessment tools, the NOS scale was selected as the most appropriate for use with observational studies . In the 6, folate, vitamin B12, methionine, betaine, and choline) as well as homocysteine were analysed if data were available from at least two different studies (k ≥ 2). All effect estimates were log-transformed for analysis.Each exposure was modelled individually and dietary intake and biomarkers were assessed separately. All dietary components of 1C metabolism  is normal with a mean of 0 and standard deviation of 0.82 and the distribution for τ (the between-study standard deviation) is log-normal with a log mean of -3.27 and log standard deviation of 1.68. The distribution for µ is derived from an assumption that a relative risk of 5 is a generous upper limit for the expected estimated effect and a standard deviation of 0.82 allows for 95% of the distribution to be as or less extreme compared to this limit, assuming a normal distribution. We consider this a “weakly informative” prior distribution, in that it puts low probability on implausible parameter values, and substantial probability over the range of plausible parameter values. The distribution for τ is suggested by Turner et al. for use in a meta-analysis of a “major morbidity event” comparing non-pharmacological exposures [A Bayesian approach was used for the meta-analyses to estimate summary RRs and 95% credible intervals (CrIs) for the highest versus lowest category of the exposures. A CrI is analogous to a confidence interval in frequentist analysis, but it has the advantage of a more intuitive definition in that it represents the probability that the population parameter lies within the specified range conditional on the model and the data [his case . Since txposures . The priµ and a half-normal distribution for τ, which has been shown to be an appropriate distribution in meta-analyses with a small number of studies [µ and τ and it is intended to test the impact of extreme alteration of prior distributions. We avoided using a uniform prior because placing equal density across all real values would give too much weight to implausible values. Frequentist random-effects and fixed-effect models were run as well. The restricted maximum likelihood estimator was used in the frequentist random-effects models.Two other Bayesian models were run to assess sensitivity of the results to the prior distribution. The first uses the same distribution for  studies . The secThe primary outcome of interest was RCC. Because a majority of kidney cancers in adults are RCC, studies assessing risk of overall kidney cancer were included in a secondary analysis to increase the number of studies available.Multiple sensitivity analyses, specified a priori, were undertaken to assess the robustness of the results. Studies with a lower NOS score are assumed to be more susceptible to bias, so those with a NOS score less than seven were excluded for the first sensitivity analysis. For the next sensitivity analysis, all case–control studies were excluded for dietary intake exposures to avoid the risk of recall bias from retrospectively collected data. Finally, we conducted analyses on dietary intake exposures for food consumption only, excluding supplement use, because of the differences in bioavailability and data collection methods.τ2 and I2 based on the posterior distribution of τ. I2 is an intuitive measure of heterogeneity, indicating the percentage of total variation in the estimated associations due to between-study heterogeneity. We have also reported τ2, indicating the between-study variance because, unlike I2, it is not dependent on the number or size of included studies [Funnel plots were visually inspected to check the risk of publication bias and heterogeneity was assessed with  studies .Meta-analyses were done using R 3.6.0 , the bay6 intake, six included folate intake, three included vitamin B12 intake, two included methionine intake, two included choline intake, and two included betaine intake. For biomarkers, analyses from the same two studies, the European Prospective Investigation into Cancer and Nutrition (EPIC) and the Alpha-Tocopherol Beta-Carotene Cancer Prevention Study (ATBC), included riboflavin, folate, vitamin B12, and homocysteine, and these two studies plus the Melbourne Collaborative Cohort Study (MCCS) included vitamin B6. There was a mean of 2.8 studies per exposure. There were not enough studies examining mortality to be able to perform meta-analysis on this outcome.Figure The 13 papers were published between 1997 and 2018 and they included a total of 13 different study populations . The mean scores were 6.5 for case–control studies and 6.3 for cohort studies. The lowest scoring topics were ascertainment of exposure in case–control studies and ascertainment of exposure plus representativeness of the cohort for cohort studies.6, folate and choline, but estimates were accompanied by substantial uncertainty, particularly for riboflavin and choline. The direction of effect estimates varied between nutrients, with vitamin B12, methionine, and betaine having relative risks greater than 1. The pooled estimates for highest versus lowest category were as follows for intake exposure: RR = 0.89 (95% CrI 0.70–1.13) for riboflavin, RR = 0.86 (95% CrI 0.71–1.04) for vitamin B6, RR = 0.85 (95% CrI 0.71–1.01) for folate, RR = 1.14 (95% CrI 0.87–1.49) for vitamin B12, RR = 1.27 (95% CrI 0.89–1.82) for methionine, RR = 0.88 (95% CrI 0.62–1.26) for choline, and RR = 1.01 (95% CrI 0.69–1.49) for betaine.Results for the meta-analyses of intake are presented in Fig. 6, RR = 0.79 (95% CrI 0.54–1.15) for folate, RR = 0.73 (95% CrI 0.51–1.06) for vitamin B12, and RR = 0.88 (95% CrI 0.61–1.27) for homocysteine. All biomarkers showed estimates in the direction of a protective effect, however, the estimates were lacking precision. No papers examined biomarker status for choline or betaine and only one had results for methionine biomarker status, hence these three exposures could not be included in the meta-analyses.Figure τ indicated a lack of between-study heterogeneity for all exposures except vitamin B6 biomarker status . Estimates of between-study heterogeneity were less extreme for the main Bayesian model compared to the frequentist random-effects estimates which ranged from I2 = 0% for 9 exposure to I2 = 79.4% for vitamin B6 biomarker status . Vitamin BI2 was greater than 0%, namely folate intake and vitamin B6 biomarker status. The effect estimate for folate intake from the fixed-effect model compared to the random-effects model was identical with a slightly more precise CI  and the estimate for vitamin B6 biomarker status was slightly stronger  in the fixed-effect model.Frequentist fixed-effect model results differed from the random-effects models only for exposure where the estimate of There were no notable differences when analyses were run with the other two Bayesian models (Online Resources 6–9). The point estimates were substantively and qualitatively unchanged for all exposures and the 95% CrIs were slightly wider for the first alternate Bayesian model and much wider for the second alternate Bayesian model.12 biomarker concentration (Online Resource 10). There was not a considerable change for riboflavin intake  or folate intake  compared to the RCC-only results. The pooled estimate for vitamin B12 biomarker status changed most substantially, from RR = 0.73 (95% CrI 0.51–1.06) to RR = 1.07 (95% CrI 0.55–2.24), and there was considerable heterogeneity between primary studies (I2 = 75% for the posterior median of τ).We also conducted an analysis with any kidney cancer as the outcome rather than RCC. Two publications reporting kidney cancers replaced two older publications from the same studies reporting RCC only (Nicodemus 2004 replaced Prineas 1997 and Tavani 2012 replaced Bosetti 2006). Two additional kidney cancer publications were added, so a total of ten studies were available for analyses. Meta-analyses were possible for only three exposures: riboflavin intake, folate intake, and vitamin B6 biomarker status, folate biomarker status, vitamin B12 biomarker status, and homocysteine biomarker status.When including only studies with a NOS score of seven or greater, only folate intake met the criteria of having a different set of two or more studies available compared to those included in the original analysis (Online Resource 11). The pooled estimate was marginally weaker than that from the main analysis . Exposures already having only studies with a NOS score of at least seven in the original analyses were riboflavin biomarker status, vitamin B6 intake, the two most protective estimates were from case–control studies in the original model, so the pooled estimate was weaker when these were dropped . Similarly, three of the four most extreme estimates for folate intake were removed and the resulting pooled estimate was weaker . One case–control study was removed for vitamin B12 intake, leaving only the two estimates from the Cho 2013 paper. Our pooled result  was similar to that presented by Cho et al.  [12 intake was stronger than from our main analysis.Four studies examining dietary intake were included after removing case–control studies (Online Resource 12). For vitamin B6 , and folate  all had a slightly weaker pooled estimate compared to the original models (Online Resource 13). The estimate for vitamin B12 intake from food was slightly stronger than that from the main analysis, but still accompanied by substantial uncertainty .For estimates of food intake only, excluding supplements, riboflavin , vitamin B6, and folate), which all had pooled estimates suggestive of a protective association. This advocates for a role by nucleotide synthesis in a mechanistic explanation of association because of its direct link with the folate cycle.The purpose of this systematic review and meta-analysis was to assess the existing evidence for an association between 1C metabolites and RCC risk. None of the individual intake or biomarker exposures analysed had a pooled estimate with enough precision to indicate a clear association with RCC risk. This lack of precision is largely attributable to the low number of studies and in some cases to between-study heterogeneity. There was a range of two to six studies per exposure, and additional exposures of interest were left out because they had fewer than two studies available. Areas warranting further investigation were identified, including the three B vitamins involved in the folate cycle  and each of these had a stronger pooled RR for biomarker status over intake. There are multiple factors that affect the link from intake to circulating concentration of a nutrient including biosynthesis by the microbiome, variable rates of absorption and de novo synthesis regulated by feedback mechanisms, uneven distribution between tissues and in circulation, and measurement error, particularly in the measurement of dietary intake. Vitamin B12 was the only nutrient with a qualitative difference between intake (RR greater than 1) and biomarker status (RR less than 1). Unlike the RCC-specific estimate, the biomarker estimate including any kidney cancer (RR = 1.07) was in the same direction as the intake estimate; however, there is considerable between-study heterogeneity for this estimate largely driven by the Arendt 2013 study. This study differed from the others in that the median follow-up time after blood draw was only 3.5 years, so their results may be more strongly affected by reverse causation.There were four metabolites which were included in both the intake and biomarker analyses  only included data from the NHS and HPFS studies within the Cho 2013 paper, which also presented pooled results from the two studies. The estimates found from our model were very similar to those presented by Cho et al. .12 biomarker status [6, and vitamin B12 biomarker statuses are inversely associated with RCC risk. Whilst we did not complete a dose–response analysis because of differences in intake adjustment methods and biomarker measurement methods between studies, our results are broadly consistent with those previously published.Our analysis included three more RCC-specific studies than the most recently published meta-analysis, and also assessed additional key 1C metabolites choline and betaine. Overall, our results are similar to those from the previous meta-analysis, which found no clear associations with RCC for any of the 1C metabolism intake or biomarker exposures examined, though it did provide weak evidence for a protective effect of vitamin Br status . The dos6 and increased risk for higher circulating homocysteine [12 concentration and the evidence suggests a positive association with circulating folate as well [1C metabolism may play a parallel role in kidney cancer and liver cancer, because these are the only two organs where betaine can be used rather than only folate as the methyl donor . Researccysteine . This mecysteine . In cont as well . Folate  as well . Our res6 intake, but a specific RR was not given so these exposures could not be included in meta-analyses. We did not attempt to contact authors to obtain data not presented in the papers included in this review.A major risk of bias in meta-analyses comes from selective reporting of results within studies. Some included papers stated that all measured associations were selected a priori, but in other papers it is not clear if there may have been bias in the selection of reported results. The Nicodemus 2004 paper stated that no association was found for folic acid supplementation or vitamin BOne limitation of this meta-analysis is the inconsistency of covariates included in the primary analyses. Some models were only adjusted for basic demographics and did not account for key established risk factors for RCC including sex and body mass index. Some primary analyses in our meta-analysis did not adjust for concurrent nutrient intakes, while those that did are likely affected by residual confounding from unidentified nutritional components.Population diversity is lacking in the studies included in this meta-analysis, with a strong overrepresentation of participants of European descent, largely from affluent countries. Bock et al. did compare European-American and African-American participants and found similar associations for the two groups . The AunA strength of this meta-analysis was the use of a Bayesian model complemented by several additional models to assess the impact of implied assumptions and our choice of priors. This allowed for a more robust estimation of the between-study heterogeneity compared to using frequentist methods as well as the propagation of uncertainty through to the effect estimates. This is especially important when the number of studies is small. Further, we conducted relevant sensitivity analyses, chosen a priori, to check the robustness of our results against common sources of bias.Because tumours as well as healthy cells rely on 1C metabolism, the related risk factors for RCC prognosis may differ from those for RCC incidence. Unfortunately, despite their inclusion in the literature search, prognostic outcomes could not be included in the meta-analysis due to a lack of existing publications. This largely unexplored area will become increasingly important following predictions of a rising number of RCC cases .In summary, the results of this systematic review and meta-analysis do not provide overwhelming evidence for the role of any single component of 1C metabolism in RCC risk, but the findings are based on sparse data. This is consistent with an overall lack of consensus on the role of 1C metabolism in multiple types of cancer. The suggestive evidence of inverse associations for both intake and circulating concentrations of several 1C metabolism components warrant further investigation.Supplementary file1 (PDF 843 kb)Below is the link to the electronic supplementary material.


The etiologies of chronic neurological diseases, which heavily contribute to global disease burden, remain far from elucidated. Despite available umbrella reviews on single contributing factors or diseases, no study has systematically captured non-purely genetic risk and/or protective factors for chronic neurological diseases.meta-umbrella) published until September 20th, 2018, using broad search terms in MEDLINE, SCOPUS, Web of Science, Cochrane Database of Systematic Reviews, Cumulative Index to Nursing and Allied Health Literature, ProQuest Dissertations & Theses, JBI Database of Systematic Reviews and Implementation Reports, DARE, and PROSPERO. The PRISMA guidelines were followed for this study. Reference lists of the identified umbrella reviews were also screened, and the methodological details were assessed using the AMSTAR tool. For each non-purely genetic factor association, random effects summary effect size, 95% confidence and prediction intervals, and significance and heterogeneity levels facilitated the assessment of the credibility of the epidemiological evidence identified.We performed a systematic analysis of umbrella reviews  were eligible. The median number of primary studies per meta-analysis was 7 (interquartile range (IQR) 7) and that of participants was 8873 . The search yielded 115 distinctly named non-genetic risk and protective factors with a significant association, with various strengths of evidence. Mediterranean diet was associated with lower risk of dementia, Alzheimer disease (AD), cognitive impairment, stroke, and neurodegenerative diseases in general. In Parkinson disease (PD) and AD/dementia, coffee consumption, and physical activity were protective factors. Low serum uric acid levels were associated with increased risk of PD. Smoking was associated with elevated risk of multiple sclerosis and dementia but lower risk of PD, while hypertension was associated with lower risk of PD but higher risk of dementia. Chronic occupational exposure to lead was associated with higher risk of amyotrophic lateral sclerosis. Late-life depression was associated with higher risk of AD and any form of dementia.meta-research).We identified several non-genetic risk and protective factors for various neurological diseases relevant to preventive clinical neurology, health policy, and lifestyle counseling. Our findings could offer new perspectives in secondary research ( Nowases, see ). This ises, see –7 ], whileses, see –11.phenome/ exposome/ environment-wide association studies), relevant statistical tools , and epidemiological approaches  and genome (GWAS) and transcriptome-wide association studies have revealed numerous possibly related SNPs and mechanistic clues –14. Fielproaches ), this iproaches –32. For proaches .The contribution of environmental factors to neurological disorders is in many cases modifiable (further discussed in ). These uniform approach for all factors, considering the expected variability in their quality, focus-of-interest, and degree of evidence reliability , alongside their corresponding confidence interval (CI), and the number of participants in each study. We, also, reviewed whether the included studies performed any quality control over the reviews and meta-analyses used.For data extracted from studies where one non-purely genetic risk or protective factor or biomarker was reviewed en face of multiple health outcomes, we extracted only data that were relevant to neurological diseases.A Measurement Tool to Assess Systematic Reviews (AMSTAR) method to assess the quality of reviews .The minimum and maximum number of studies included in the systematic reviews and meta-analyses of any of the risk or protective factors or biomarkers was 1 and 67, respectively. In Additional file In Additional file 12 were associated with a reduced risk of developing MS and PD. Vitamin E and non-aspirin nonsteroidal anti-inflammatory drugs (NSAIDs) were protective factors for PD and AD. Serum vitamin D levels were associated with a lower risk of MS, AD, PD, cognition, and stroke. Physical activity and alcohol intake were associated with a reduced risk of developing AD, PD, vascular dementia, and all types of dementia  effect on neurological disorders Table . On the In Additional file Being overweight in late-life  was significantly associated with a lower risk of developing dementia. Contrarily, a higher frequency of social contacts, loneliness, social participation, tooth loss, rheumatoid arthritis, benzodiazepine use, and atrial fibrillation were significantly associated with a higher risk of dementia. Diphtheria and tetanus vaccination, as well as a higher anti-Epstein-Barr virus (anti-EBV) IgG seronegativity, were significantly associated with a lower risk of developing MS. Conversely, anti-Epstein Barr nuclear antigen (anti-EBNA) IgG seropositivity, infectious mononucleosis, appendectomy at an age ≤ 20 years, EBV DNA in serum and mononuclear cells, tonsillectomy at an age ≤ 20 years, traumatic injury, anti-viral capsid antigen IgG seropositivity, chronic cerebrospinal venous insufficiency, serum homocysteine, and Chlamydia pneumoniae  were significantly associated with a higher risk of developing MS.Herpesviridae infection, aluminum exposure, stroke, and neuroticism were significantly associated with a higher risk of developing AD  perspective on risk and protective factors and biomarkers, albeit not with quantitative approaches. In contrast, field-wide meta-analyses using quantitative approaches have assessed the entire field of putative risk and protective factors but for a specific disease, not an entire spectrum of diseases or body organ system  [Increasing accretion of data has led to recent calls for comprehensive, field-wise analyses of risk and protective factors for many human disorders . Our stu system) .next-generation systematic reviews [third-generation systematic review. It is an approach that aims to offer a new perspective of secondary research (meta-research), a field hallmarked by the need to provide the most integrated evidence possible, and in which several novel study designs have appeared during the last years, e.g., series of systematic reviews and meta-analyses in a single publication, where the analytical unit is the umbrella review study design [field-wide meta-analyses, in which a meta-analysis of observational studies is conducted on the sum of risk factors under consideration [systematic reviews of systematic reviews [overviews of systematic reviews [meta-reviews [systematic meta-reviews, comprehensive reviews [research-on-research [meta-meta-analyses [meta-umbrella review could represent another study design added to the armamentarium of meta-research [Notably, following previous characterizations of umbrella reviews as  reviews , our appy design . Likewisderation , or to tderation ), system reviews , overvie reviews , meta-re-reviews , systema reviews , researcresearch , and metanalyses , 96; theresearch .meta-research for global health neurology, namely neurological diseases of resource-poor countries , as well as for rare/orphan neurological disorders.Although our primary aim was to study the largest possible number of neurological conditions , we discovered that the umbrella reviews had studied only 12 neurological conditions. For instance, we could not find umbrella reviews on risk or protective factors for some common or major chronic neurological disorders such as migraine, headache, brain cancer, or epilepsy. Therefore, future umbrella reviews should be considered regarding the non-purely genetic risk factors of these conditions. Interestingly, almost all studies had focused on neurological disorders with high prevalence and in resource-rich countries, which could be indicative of the disproportionally lower number of publications regarding We studied 115 distinctly named risk/protective factors with a marked association with chronic non-communicable neurological disorders, including biomarkers, habits, dietary factors, medical history or/and comorbid diseases, drugs, and exposure to toxic environmental agents. Fourteen factors exhibited a decreased risk for an extensive number of non-communicable neurological disorders, with these factors ranging in strength from class I to class IV. Below, we provide our insight into some of these findings.Notably, the following associations appeared with class I evidence: (a) in neurodegenerative diseases, dementia, and AD, Mediterranean diet was a protective factor; (b) in MS, smoking, anti-EBNA IgG seropositivity, as well as infectious mononucleosis were risk factors; (c) in ALS, lead was a risk factor; (d) in PD, physical activity was protective, while constipation was a risk factor ; (e) inI2 metric, heterogeneity was present in published reports, and few studies were consistent with non-heterogeneous evidence when data had a prediction interval excluding the null.While several risk and protective factors had class III and IV evidence of being significant in the occurrence of these neurological conditions, three of them, namely tobacco smoking, hypertension, and serum uric acid, exerted a mixed risk and beneficial effect. Based on the With regard to dietary factors, we found substantial evidence highlighting the potential role of Mediterranean diet in lowering the risk of dementia, AD, cognitive impairment, neurodegenerative diseases, and stroke. Until now, several meta-analyses have reported quite solid evidence of the beneficial effect of Mediterranean diet in AD and other dementias, i.e., major categories of neurodegenerative disorders . HowIn parallel, negative associations between coffee consumption and PD and AD  have beeThis systematic review of umbrella reviews revealed counterintuitively a significant association of low serum uric acid levels with a decreased risk of several neurological diseases , while diabetic peripheral neuropathy and stroke mortality were associated with an increased risk. Our credibility assessment revealed that, with the exception of PD (with class II evidence) and stroke mortality (with class I evidence), these significant associations were within class IV evidence . Hence, According to class I–IV evidence, physical activity was found to exert a beneficial effect against PD, AD, and all types/vascular dementia. Physical exercise can increase serum uric acid levels, which has been associated with a lower risk of developing PD and dementia , 104. Ho12, C, and D levels were associated with a lower risk of different neurological conditions, such as MS , which cautioned against false interpretations. The observed heterogeneity most likely arose from different comparison groups in prospective, retrospective, and case-control studies, causing some of the meta-analyses to be derived from studies with diverse, contrasted categories of serum vitamin B12, C, and D levels [Serum vitamins Brecently ), AD, deD levels –51. FurtD levels ).C. pneumonia infection with the occurrence of several neurological conditions . However, most of these associations were based on class III and IV evidence, which could have resulted from the substantial heterogeneity among the primary studies. Hence, these associations warrant cautious interpretation. We also suggest that the findings on chronic cerebrospinal venous insufficiency should be interpreted with caution, considering both the wide range of the corresponding confidence intervals and previous reports in the field [Our meta-umbrella review provides some evidence for a positive association of exposure to farming, pesticides, low-frequency electromagnetic fields, organic solvents, and he field , 111.meta-umbrella review into the broader context of studies reviewing risk and protective factors for neurological disorders, we noticed that, in another comprehensive review of systematic reviews, for example, exposure to pesticides was identified as the commonest risk factor for AD, ALS, and PD, whereas smoking was associated with AD and MS [Framing our D and MS .the poster child of causal relations [snuff) usage, which was considered to be associated with a lower risk of MS, potentially through nicotine-mediated effects on subunits of immune cells expressing acetylcholine receptor [Below, we discuss the findings on the effects of tobacco smoking in a separate section. This choice was made because we consider that, with all the body of evidence surrounding this field, smoking should represent an exemplar for studying risk and protective factors for neurological disorders, or, as other authors have previously claimed, represents elations . We founelations , even thelations , 115). Melations . Of notemeta-umbrella approach. Perhaps, in this specific field, it would have been probably wiser to select only the meta-analyses of prospective studies among the umbrella reviews. Similarly, another possibility could be to consider only umbrella reviews that have examined credibility ceilings [credibility ceilings, which take into consideration limitations regarding the methodology of the studies) [meta-umbrella approach. Besides, it is commonly known that extensively performing sub-analyses in many subgroups could be linked to artificially increasing events of statistical significance. In every case, the teaching example of cross-sectional studies on lung cancer and smoking  for causing inverse causation should always be kept in mind [Another possibility could be that people suffering from a certain neurological disorder, such as MS, prefer to smoke, whereas those unaffected choose to stop smoking more easily, as previously observed in patients with schizophrenia . Therefoceilings , in ordestudies) , 69, 119 in mind .suggesting that longer duration of smoking is needed for a risk reduction, as cited in the above study) from core public health messages.With regard to PD, the potential underlying genetic and non-genetic roots (or/and bias) of the association between tobacco smoking and PD are reviewed elsewhere . HoweverIn every case, we feel that the core message of promoting tobacco smoking cessation as an effective public health intervention should remain undiluted because of its several well-established positive health effects , 122, irin vacuum but rather in association with other heavy metals and welding, even though the latter two are classified in lower levels of evidence . Moreover, when lead levels are taken into account, the association of another heavy metal, i.e., copper, with ALS risk becomes attenuated, suggesting a chief role of lead [Below, we wish to highlight some additional features with class I evidence. Chronic occupational exposure to lead presented a higher risk for ALS. Arguably, lead toxicity represents a major underlying mechanism in ALS-related pathogenicity , 127. In of lead , even th of lead . Interes of lead ; thus, iConstipation was positively associated with PD (class 1 evidence). A prospective cohort study reported a significant association with a similar effect size in meta-analyses (reviewed in ). AnotheOur meta-umbrella review assessed specific risk factors related to dementia and AD. While only late-life depression and type 2 diabetes mellitus were positively associated with AD, depression at any stage in life was linked to all types of dementia. In fact, late-life depression was markedly associated with both dementia  and AD . It is sbrain reserve, which describes an individual’s ability to not develop the disease phenotype despite brain pathological changes that are either age- or disease-specific [Low levels of social interaction markedly affected the occurrence of dementia. Thus, social networking, along with educational and leisure activities, are modifiable protective factors, which might aid in the maintenance of cognitive function with increasing age . Tspecific , 141.Lastly, while serum 25-hydroxy-vitamin D has been investigated in umbrella reviews of neurological disorders, the same does not hold true for 1,25-hydroxy-vitamin D, as the latter has only been assessed in cancer .health impact pyramid, in which the main social and economic determinants of health, such as education, race, housing, and income, are included [Overall, despite this extensive body of evidence, we wish to emphasize that the majority of epidemiologically identified risk and protective factors do not lie at the bottom of the included  (for an included ). Intereincluded . Thus, cincluded –146.Family-Wide Association Studies [epidemiological and environmental neuroscience [In addition, there are potentially less appreciated or less easily quantifiable risk and protective factors, such as (a) the family environment (now-studied through  Studies ); (b) th Studies ; (c) liv Studies –151; and Studies . These foscience ).meta-umbrella study provides the opportunity to (a) stimulate more comprehensive, patient-centered approaches, allowing truly informed decisions during genetic counseling or/and coaching for lifestyle changes [window of opportunity still exists [meta-umbrella review approaches as the highest level of evidence in the meta-research field; (h) serve as teaching material for courses on preventive neurology offered by the relevant medical education institutions; (i) contribute to helping physicians understand the contribution of environmental elements as risk factors for neurological disorders, to assist environmental health specialists in the appreciation of the ties between the nervous system and environmental health; and (k) to address these factors  by taking action starting from early and middle adulthood, thus ultimately reducing to some extent the incidence and, hence, the prevalence of neurological disorders.Major implications for several target groups, namely patients and their caregivers, healthy subjects, clinicians, researchers, environmental health specialists, policy makers, and educational institutions, could be anticipated from this meta-umbrella review. In a way similar to umbrella reviews in other fields , 154, th changes ; (b) enh changes , and, inl exists ; in thatholistic policy making , and it could also serve as an impetus in developing transdiagnostic approaches in neurology, similarly to psychiatry [transdiagnostic theory, see [Although others have argued that deciphering how the mechanistic effects of certain risk and protective factors are different between distinct neurological disorders  , we feelychiatry  .publication of protocol for umbrella reviews, in alignment with previous calls [In addition, our approach would advocate the development of appropriate statistical tools to account for the fractions of affected neurological populations vs. risk and protective factors, in alignment with previous approaches . In theus calls . Furtherus calls ).scientists should stop advocating the need for yet another clinical trial on the cognitive benefits of healthy lifestyles and lobby decision-makers to implement societal polices to actively promote propitious lifestyles. This approach will substantially produce benefit not only for the brain but for the society at large. Complicated situations in which a factor has both a beneficial and a risk effect  provide an opportunity to highlight the broader potential discrepancies between public health and precision medicine [implementation science research to guide health policy and to be a major component of the so-called population health science [Many of the class I evidence results in our study re-affirm previous opinions on implementation science . As premedicine . Interes science .meta-research-oriented; thus, it represents a call-for-action, similar to similar calls in other diseases [overarching and up-to-date knowledge on a wide array of contributing epidemiological factors. In this context, our approach attempts to address the challenge of evaluating evidence provided by a number of high-quality meta-analyses and, in turn, umbrella reviews [This study has several strengths, as it is public health policy-, clinical science-, and diseases . The firdiseases , and who reviews .overarching approach allows studying the top evidence. In general, umbrella designs are of special value when applied to provide an overall picture that can inform guidelines. For instance, rather than examining one risk factor or disease, a meta-umbrella review can consider multiple risk factors for multiple disease processes. For this reason, we ensured that four researchers participated in the search and quality assessment of published studies, thus, enhancing the validity of our analyses. Notably, whereas earlier non-umbrella reviews examined 14 neurological disorders (and the associated risk and protective factors) as part of National Health Guidelines [Our systematic, idelines , our stumeta-umbrella approach, can detect irregular patterns in the associations observed . In thicides in ). To enhthird-generation systematic review study design, allowing, among others, to discuss research gaps and to determine the potential mechanistic underpinnings of such associations. In other words, because of the advent in the wealth of umbrella reviews conducted, this meta-umbrella review should be seen as a logical next step, where available umbrella reviews will serve as the analytical unit of the review, in which the meta-umbrella approach will allow selecting and including the highest level of evidence, notably other umbrella reviews .p values and CIs. Indeed, when a p value is within the non-significant range, this means that either the groups have no difference or, alternatively, that the participants’ number is too low to show if such a difference exists. Similarly, when a confidence interval lies in both sides of the line of no effect, this means that either the groups have no difference, or, alternatively, that the number of participants (sample size) is too small to show if such a difference exists. Conversely, a confidence interval not crossing the line of no effect can provide reassurance on the strength or weakness of the evidence, and importantly, whether the study’s results are definitive, with no further need to perform additional studies on the particular issue. Therefore, these notions should be considered while interpreting the results of this meta-umbrella study.This study obviates the need to perform further research on many different factors demonstrating no significant association, as shown in previous studies . In thatoverarching goal. Also, the wide presentation of risk and protective factors could be perceived as attenuated if not followed by investigations on their biological plausibility, thus calling for studies to bridge the gap between epidemiology and pathobiology of neurological diseases.Some limitations should be acknowledged when considering this meta-umbrella study. First, the population and interventions discussed could be perceived as too broad. Our study, however, was in alignment with its transferred to the meta-analyses, umbrella reviews, and herein, our meta-umbrella review . For exrther in . This is reviews . Likewis reviews , 54. Col choices .meta-umbrella study. For instance, the role of physical and emotional stress is not discussed in (or even captured) umbrella reviews, while the same holds true for pregnancy-related maternal health, epilepsy, myasthenia, Tourette syndrome, and Huntington disease, to name a few examples . Likewi factors . Similarfined in ), such afined in , could hfined in –173, or fined in ) play crfined in , 57, 175That said, with regard to the majority of risk and protective factors, which cannot be examined using standard epidemiological approaches or are not frequently assessed, the level of evidence pertaining to these factors will be most likely low given the scarcity of the existing relevant data (as further discussed in ). Similameta-umbrella approach, are inherently biased in favor of commonly assessed factors, towards more common neurological disorders (perhaps with the exception of neuromyelitis optica), or towards those of adult but not pediatric populations, because, so far, all umbrella reviews have been conducted on these disorders. This is because of the lack of umbrella reviews examining factors that are less common in rare or adult neurological disorders, besides the conditions included. Assessing the input of genetic and environmental factors in rare diseases by comparing evidence from common diseases could be hampered by the heterogeneity of studies . Perhap infancy . Likewis infancy –181.Despite our efforts to be as thorough as possible in employing a sensitive literature search strategy, no umbrella reviews except in the English language were found in our systematic literature search. However, we cannot exclude the likelihood that some relevant papers in other languages might have been overlooked, even though Moher et al. have shown that omitting studies in non-English languages may not introduce considerable bias . Similarcorrelates nowadays requires sophisticated methods , the description and usage of which exceed the aims of the present study .vibration of effects in observational studies, which is linked to how the selection of adjusting variables leads to results’ variability, should be considered while assessing our results [Another source of bias could refer to the heterogeneity of the data in combining risk and protective factors for multiple different disease processes. Multiple risk factors, which may be ill-defined or based on studies that are observational in design, may be associated with very distinct disease processes, with differing pathophysiologies. For example, PD is particularly prone to recall bias as it tends to have a longer prodromal phase with symptoms that might not be diagnosed as PD until manifestation of the later characteristic motor symptoms. Likewise, the comparator groups for different neurological disorders included in this meta-umbrella review are expected to be different. Importantly, the so-called  results .overarching character of presenting more-than-one protective and risk factors for the sum of neurological conditions should be regarded through the public health and clinical counseling lenses rather than from a mere epidemiological perspective. One of our chief goals was to reduce reporting bias—this is, indeed, one of the main reasons for which systematic reviews (in our case a systematic review of umbrella reviews) are performed. In doing so, we wish to highlight (a) the challenges preventive medicine specialists can face  during counseling services, especially with regard to how well they can communicate their message, e.g., when the same factor is protective for one neurological disease and risk factor for the other, and (b) the potential inherent difficulties of forming public health guidelines.The primordial factors , therefore representing risk markers, or that there is an apparent heterogeneity in the definition of these factors [pragmatic approach by showing confidence to the choice of the primary systematic reviewers—and, this should be recognized [In the same context, other limitations could include (a) the potentially different, or, in some cases, potentially inaccurate definitions regarding healthy control groups among meta-analyses or/and individual studies leading to inaccuracies, thus introducing another source of bias . Ideally, extracting detailed information from the primary meta-analyses  would be needed, as it is difficult to assess these particularities at the level of the meta-umbrella review; however, such a task could not be easily performed in our study because (a) our initially designed pragmatic approach followed the definition of the umbrella reviews, (b) there were no informaticians at our library facilities to assist in this task, (c) umbrella reviews, in most cases, do not include the inclusion criteria of the original studies, and (d) if we had decided to set out for the above task, then we would have had to list the inclusion criteria of all the primary studies, which were presumably different.Corrected Covered Area, to assess overlapping .risk factor with predictor and explanatory factor for risk stratification and causal studies, respectively [risk markers to reflect that many risk factors, such as immigration and ethnicity, may stem from the interaction of other risk factors [meta-umbrella studies should agree on the effect size to be used for comparisons and on the preferred terminology regarding risk and protective factors , let alone when these effect sizes stemmed from different study designs  or when distinct effect size metrics are not converted to a single metric of reference. Thus, our study’s results should be interpreted more through a qualitative rather than a quantitative lens . Additioectively . Similar factors . Collectogy, see ). In themeta-umbrella review was not registered for its goal or protocol at a database like PROSPERO, although (a) our search criteria were predefined and typical of systematic reviews, (b) our exclusion criteria were not intending to alter the number or nature of umbrella reviews identified and (c) no re-synthesis or quantitative synthesis with regard to outcomes of interest or of any other data in any sort of meta-analysis (as discussed in [Another limitation is that our ussed in ) took plmerging of distinct study design types, which holds an undeniable bias in principle, has not impacted our results, because of (a) our pragmatic approach, (b) the non-quantitative approach applied herein, and (c) the lack of evident internal discrepancy among the findings reviewed. Besides, both study types offer real-word results . Nevertheless, selecting for retrospective studies to include can affect, in principle, the meta-analyses  and not a meta-umbrella review. In this sense, performing a sensitivity analysis may not be feasible for a meta-umbrella review. In addition, the fact that umbrella reviews only use statistical testing to show the existence of bias and cannot provide evidence of their nature and extent should be taken into account, alongside with the fact that umbrella reviews cannot supply any comparative ranking, as is done in a network meta-analysis.Other limitations could be that (a) we merged the systematic reviews referring to observational studies with those of clinical trials, given that observational studies should be treated with high cautiousness when referring to risk and protective factors (commented in ); (b) weented in ). Anothesystematic reviews of systematic reviews, meta-reviews, systematic meta-reviews , symples in , 213–216s manner .meta-research could be made, in alignment with previously expressed opinions [Moreover, an a posteriori search strategy using similar search strings revealed only one study with potential applicability . Under opinions , in partopinions .overarching outline of different aspects of the assumed risk and protective factors in the field. Such comprehensive approaches provide a way to systematically analyze the qualitative and quantitative characteristics of the existing literature, the validity of evidence, and how these studies could provide readers with information on knowledge gaps. In doing so, this meta-umbrella review allows to identify the sum, so far, of preventable or/and modifiable risk factors, which may be relevant—either as common to most or specific to certain—neurological disorders. To all these goals, standardizing how a study is designed and how exposure to certain factors is defined would be essential [Notwithstanding potential considerations that the topic under study may appear broad, this exposure-wide assessment of risk and protective factors of chronic neurological disorders, conducted using a systematic review of umbrella reviews , offers an ssential . Likewismeta-umbrella approach provides a wealth of discussion on potential biases that could occur in reporting umbrella reviews, and it provides a rationale to develop reproducible robust methodologies while performing umbrella studies. To enhance extensive analyses of risk/protective factors, we recommend developing standardized and reproducible methods to identify these factors—performing a field-based systematic review of umbrella reviews might simplify the process. Equally importantly, as almost all umbrella review studies have so far focused on neurological disorders of resource-rich countries, a call for intensifying meta-research on neurological disorders affecting resource-poor countries could be addressed.In addition, our meta-umbrella review offers new perspectives in the meta-research field. The fact that not all risk factors or/and diseases were uniformly mentioned could per se serve as a call-for-action to develop guidelines and a uniform framework for Systematic reviews of Umbrella reviews. In the same context, our meta-umbrella review could serve as a call-for-action to conduct umbrella reviews using more harmonized criteria of evidence and strength of evidence; this could be performed by grouping studies by the criteria used to assess evidence, and then reaching conclusions in this regard.Lastly, this disease network neighborhoods) [Being distant from any overly expectations on reaching final conclusions on causal vs. casual associations, this study attempted to highlight the commonality of certain risk and protective factors, and the relevant level of evidence surrounding these factors . Emphasiorhoods) , 220; hometa-umbrella review) should be jointly assessed with studies coupling mechanistic and biological insights with statistical methods on causal inference [mediator variable between an independent and dependent variable) and multivariable Mendelian randomization studies, which can render some risk factors as causal determinants [living systematic review [Under all circumstances, we feel that to be able to reap further epidemiological rewards, this sort of study , which,c review ). In doic review , should Neurological Diseases Atlas, in which every risk and protective factor will be both qualitatively and quantitively linked to all associated diseases . In doing so, this Atlas—if appropriately assessing mean percentage population attributable fractions of each risk or protective factor for the sum of diseases . This As  risk and protective factors for chronic neurological diseases. Table S2-Methodological quality of included umbrella reviews based on the AMSTAR criteria and score.Additional file 3. Updated search of umbrella reviews.


VDR gene variants ApaI (rs7975232), BsmI (rs1544410), FokI (rs2228570), and TaqI (rs731236) and urolithiasis susceptibility, we performed the present study through meta-analysis. The PubMed, Cochrane Library, China National Knowledge Infrastructure, EMBASE, Web of Science, and Wanfang databases were searched to retrieve qualified case-control studies. Finally, 31 reports were selected for the present meta-analysis. The results demonstrated that the VDR gene TaqI TT genotype was related to decreased risk of urolithiasis in the overall population . In ethnicity subgroup analysis, we found that the TaqI variant was obviously correlated to urolithiasis risk among Asians and Caucasians (P < 0.05). Additionally, significant urolithiasis risk was identified in adults. However, the FokI, BsmI, and ApaI variants did not have an increased risk of developing urolithiasis. Trial sequential analysis results were on a sufficiently large number of participants and did not require more research to confirm associations. Our research suggested that the VDR gene variant TaqI was correlated with urolithiasis susceptibility and that the t-allele might be the risk gene and T-allele the protective gene in VDR TaqI variant.The relationship between urolithiasis and vitamin D receptor (VDR) gene variants is still under debate according to the available published literature. To assess correlations between  Urolithiasis is a disease with a prevalence rate of 1 to 5% around the world, and relapse is common after treatment. The 10 year recurrence rate of urolithiasis is up to 50%  belongs to the superfamily of transcription factor nuclear receptors and is a soluble protein that exists in many nuclei and cell membranes  and many tumor cells , TaqI (rs731236), BsmI (rs1544410), and ApaI (rs7975232) are most intensively studied. VDR gene is located in 12q13.11 on the chromosome. Among the four VDR variants , three of them occur in the intron sections: the TaqI, ApaI, and BsmI variants, while only the FokI variant changes the codon  AND  AND (variant OR SNP OR genotype OR polymorphism). We searched published studies up to Jan 30, 2020. In addition, we also traced back the relevant references of relevant studies and manually searched relevant articles as well as degree papers.A comprehensive search of the PubMed, Cochrane Library, China National Knowledge Infrastructure, EMBASE, Web of Science, and Wanfang databases and manual search were carried out to find relevant studies. Search strategies were:  research included VDR gene BsmI, ApaI, TaqI and FokI variants; (3) frequencies of alleles or genotypes in control groups and case groups could be exacted from the articles; (4) the distribution of genotypes of controls were in accord with Hardy-Weinberg equilibrium (HWE).(1) Case-control studies that focused on the relationship of VDR FokI, BsmI, ApaI, and TaqI variants; (4) works with incomplete data or where data were not available.(1) Reviews, meta-analysis, letters, case reports; (2) studies published repeatedly; (3) results not based on VDR gene variants, and genotyping methods. The genetic nomenclatures used in this meta-analysis were according to the recommendations of the Human Genome Variation Society (HGVS) and the American College of Medical Genetics and Genomics (ACMGG)  was applied to estimate the quality of relevant articles. If the score was more than or equal to 5 points, the quality of the article was considered to be high. Two people independently extracted data from included documents depending on the formal criteria and then carefully checked the results against each other. If disagreement arose, then the document would be sent to another person for evaluation and review. The data extraction included the first author's name, population ethnicity and area of origin, publication date, sample size of total cases and controls, hypercalciuria in the urolithiasis group, age group, genotype distribution of four MGG) see .I2 were applied to measure the heterogeneity. If I2 < 50% and P > 0.1, it was considered that the studies had no heterogeneity overall, and a fixed-effect model was used; otherwise, a random-effect model was applied. The contrast models were the allele model, dominant model, co-dominant model, homozygote model, and recessive model. Associations of VDR variants and urolithiasis under different genetic models were, respectively, analyzed by STATA 12.0 software.The odds ratio (OR) and 95% confidence interval (95% CI) were used for the analyses. The Cochran's Q-Test and heterogeneity coefficient P-value of Egger's test is more than 0.05 and the funnel plot is symmetrical, it can be considered that there is no significant evidence of publication bias.We used Egger's test to evaluate the publication bias. If the We performed a sensitivity analysis to assess whether individual studies affected the overall results. In the end, it was found that after removing each one of the studies in turn, the combined odds ratio (OR) of the remaining studies was within the 95% CI in the meta-analysis. The above results showed that the combined OR of this meta-analysis had good stability.For subgroup analysis, we divided all subject data into different subgroups so that comparisons could be made between subgroups. Subgroup analysis can be performed on different types of subjects . Ethnicity was categorized as Caucasian, Asian, or African. In the present study, we performed subgroup analysis by different ethnicity, age, and calciuria level group.The TSA was performed using TSA v0.9.5.10 Beta software developed by the Copenhagen Clinical Trial Center in Denmark. This study sets the OR reduction to 20%, the first type of error α = 0.05, and the second type of error β = 0.2 to calculate the required information size (RIS). When the size of the population is accumulating but is less than the expected amount, a trial sequential monitoring boundary (TSMB) is set based on the RIS. We performed this analysis according to the RIS and TSMB. When the cumulative Z-value crosses the TSMB, the results are considered statistically significant. At the same time, it can be considered that the sample size is sufficient.VDR ApaI, BsmI, FokI, or TaqI gene variants and urolithiasis susceptibility were recruited through the search strategy  considering the relationship between susceptibility to urolithiasis and VDR BsmI was again not correlated with urolithiasis susceptibility  < OR <1.37 (Upper Limit)], which suggested that our results were reliable .P = 0.661 for ApaI; P = 0.836 for BsmI; P = 0.953 for FokI; P = 0.464 for TaqI). The final results indicated no publication bias for results relating urolithiasis risk to VDR gene variants in the included studies.Egger's test and Begg's funnel plots were applied to assess publication bias . Therefore, it can be asserted that TaqI variant TT carriers had a lower risk of urolithiasis than tt carriers and that the evidence was reliable. However, for the ApaI, BsmI, and FokI variants, the sample size did not reach the required information size . Hence, more case-control studies are still needed to confirm the relationship between the BsmI, ApaI, and FokI variants and urolithiasis susceptibility.We carried out TSA to reduce the risk of type I error and to assess the RIS. The final results based on the Urolithiasis is a common urological disease. It is affected by a variety of genetic and external factors, including natural factors, social environment, dietary habits, local diseases of the urinary system, and systemic metabolic disorders. Many of these are difficult to control, such as climate, latitude and sunlight, water quality, sex, etc.  is a fat-soluble substance that is produced due to exposure of the skin to ultraviolet rays from sunlight and is therefore called a “sunshine vitamin” (Philippa et al., VD is also a steroid hormone in the human body. Normal people can produce VD from sunlight exposure when they exercise outdoors. However, some groups of people, such as the white-collar workers, especially those living in the Northern hemisphere, have limited exposure to the sun, and their main source of VD is food (Nelson et al., Vitamin D (VDR is a pronuclear protein and belongs to a member of a superfamily that is composed of steroid hormone/thyroid hormone receptors (Ergon et al., VDR (Ergon et al., VDR is likely to be associated with susceptibility to urolithiasis.VDR variants. For the TaqI variant, the TT genotype was significantly associated with decreased urolithiasis risk, while the Tt and tt genotypes elevated urolithiasis susceptibility. Therefore, the t-allele might be the risk gene, and the T-allele might be the protective gene. For the ApaI, BsmI, and FokI variants, we did not find any associations with urolithiasis risk.Our meta-analysis systematically investigated correlations between urolithiasis susceptibility and TaqI variant. The results suggested that TT genotype carriers were at a lower risk compared to tt genotype and Tt genotype carriers in adults. Thus, the TT genotype might be the proactive factor for urolithiasis susceptibility, while the Tt and tt genotype might be risk factors in urolithiasis.In the age subgroup analysis, we found a decreased risk of susceptibility to urolithiasis associated with the VDR TaqI variant does not modify the VDR protein structure but can influence the translation efficiency and/or stability of the RNA, which might affect the development of urolithiasis (Jurutka et al., VDR mRNA levels of allele t were significantly higher than those of allele T. Furthermore, Carling et al. (VDR mRNA levels than those with the TT genotype (Carling et al., In terms of the mechanism, the g et al.  found thApaI and TaqI variants. Nevertheless, we did not find any significant association between urolithiasis susceptibility and the ApaI variant. Even in the racial subgroup analysis, there was no significant correlation among the Asian and Caucasian populations. Our meta-analysis included more studies than Zhang's study. Furthermore, in TSA, the cumulative Z-curve crossed the boundary of the TSA, which showed that our meta-analysis had a sufficient number of cases and controls. This may cause the difference between our results and those of Zhang.This meta-analysis included 31 case-control studies with 4,144 controls and 3,782 patients, while a previous study by Zhang et al.  only hadVDR variants.Moreover, our meta-analysis also had some advantages. Firstly, our research carried out TSA to analyze the results of the study and the adequacy of the evidence. The TSA results showed that the results of the effect tests on the whole population, Asians, and Caucasians under different genetic models were based on sufficient evidence. Secondly, we carried out subgroup analysis on the basis of a range of related factors. Thirdly, we included more than 30 articles in our meta-analysis. Compared with previous studies, this meta-analysis contained a larger sample size, containing 7,926 subjects, with 3,782 urolithiasis patients and 4,144 non-urolithiasis participants, which was sufficient to draw a reliable conclusion. Fourthly, the subgroup analysis was sufficient. In addition, after Begg's test and sensitivity analysis, the pooled results and conclusions are proved to be credible to evaluate the relationship between urolithiasis risk and ApaI and FokI variants had significant heterogeneity. Thus, we carried out the subgroup analysis of other related factors in order to decrease the sources of heterogeneity. The heterogeneity decreased in subgroup analysis by ethnicity and age.Heterogeneity is very important and may affect meta results and result in error. In the present meta-analysis, the contrast model of VDR variants.There were still some limitations to this study that cannot be avoided. First, some unpublished studies with negative results might not be included. Second, this study did not reveal gene–environment and gene–gene interactions due to a lack of enough original information from included studies. Therefore, more multicenter investigations with a large sample are still needed in future to study the association between urolithiasis risk and TaqI variant. The Tt and tt genotypes could elevate the incidence of urolithiasis, while the TT genotype decreased urolithiasis risk. Therefore, the t-allele might be the risk gene and T-allele the protective gene in the VDR gene variant TaqI. However, no associations were observed in the ApaI, BsmI, and FokI variants. Our results also remind us of the necessity of early screening for urolithiasis in TaqI variant t-allele carriers.In conclusion, the results from this systematic review and meta-analysis indicated that urolithiasis susceptibility was associated with the The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation, to any qualified researcher.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Fetuin-A has been associated with the progression of metabolic syndrome, but previous studies found inconsistent results on the relationship between metabolic syndrome and the concentration of fetuin-A. The aim of this study was to perform a meta-analysis to summarize previous findings on this relationship.This study was registered with the International Prospective Register of Systematic Reviews PROSPERO (CRD42019129566). Studies examining the relationship between metabolic syndrome and the concentration of circulating fetuin-A were identified using a systematic search in the electronic databases of Embase, PubMed, Web of Science, and Cochrane Library before March 2019. A random effects model was used to summarize the effect size of the association in terms of the standardized mean difference (SMD).p<0.001). Besides, circulating fetuin-A was a risk factor for metabolic syndrome .Fourteen eligible studies compared fetuin-A concentrations between 4,551 metabolic syndrome patients and 8,805 controls. The circulating fetuin-A concentration was significantly higher in the metabolic syndrome patients than in the controls : 0.48 to 0.83, Z = 7.18, These findings suggest that fetuin-A may be an important indicator for metabolic syndrome, in which case this may lead to new perspectives in early diagnosis, identification of novel biomarkers, and providing novel targets for pharmacological interventions. Metabolic syndrome (MetS) is a new epidemic worldwide and it is associated with an increased risk for developing other chronic disease such as diabetes mellitus . The carSeveral hypothetical mechanisms that play major roles in patients with MetS are being revealed. For example, it has been proposed that circulating fetuin-A modulates the insulin sensitivity and insulin resistance in organs such as adipose, liver, and other tissues . AccordiTaken together, the preceding hypotheses suggest that circulating fetuin-A is a complex metabolic phase reactant with a debated role in MetS. However, studies investigating the association between circulating fetuin-A and MetS found inconsistent results , 9. TherThis meta-analysis was conducted in accordance with the rules and regulations for meta-analysis in the Cochrane Handbook version 5.1.0. In addition, it was registered with the International Prospective Register of Systematic Reviews PROSPERO (CRD42019129566). The results were reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) checklist . With thStudies meeting the following criteria were included in this meta-analysis: (1) case-control studies; (2) studies which reported diagnostic criteria for MetS; (3) studies that provided mean and standard deviation (SD) or odds ratio (OR) of circulating fetuin-A, or these could be obtained from authors as required; (4) studies were peer-reviewed before publication; and (5) studies were published in English. On the other hand, studies meeting the following criteria were excluded: (1) reviews or case reports; (2) MetS was studied in combination with other diseases; (3) the circulating fetuin-A was pharmacologically challenged before circulating fetuin-A measurement; (4) studies of animals; and (5) grey literature (unpublished literature). After independently screening articles, two reviewers [KA and AK] selected eligible studies and submitted them to the third reviewer [AL] who made the final decision.Using a standardized data extraction form, methodological and outcome variables of interest were collected from each study as follows: (1) the first author's last name and year of publication; (2) characteristics of subjects such as Body Mass Index (BMI), mean age , and gender; (3) region of study; (4) laboratory measures such as fasting plasma glucose (FPG), diastolic and systolic blood pressure, HDL, and triglycerides; (5) biological sample characteristics such as material of sample, and mean concentration of circulating fetuin-A  or OR associated with the concentration of circulating fetuin-A; and (6) assay methods and storage temperatures. Two reviewers [BP and AK] used EpiData 3.0 and Excel 2007 to organize and save the extracted data. Any discrepancies were resolved by consensus.To assess the quality of the eligible studies for this meta-analysis, the Newcastle-Ottawa Scale (NOS) was used . This isI2 statistic. The heterogeneity was considered high when the I2 was greater than 75%, moderate when the I2 was 25%~75% and low when the I2 was less than 25% [The ‘meta’ and ‘metafor’ packages in R software (version R 3.4.3) were used to perform meta-analysis. Given the variation in the populations and criteria used to define outcomes, a random effects model was used to pool the estimates from the eligible studies . Using tthan 25% .p values were calculated as two-sided and considered significant if less than 0.05.When heterogeneity was high, meta-regression analysis was conducted to explore the source of heterogeneity . The folMetS was defined when 3 or more of the following were satisfied: hypertriglyceridemia (≥150 mg/dL) or taking lipid-lowering drugs; elevated BP  or taking antihypertensive drugs; low high-density lipoprotein cholesterol ; hyperglycemia (fasting glucose ≥100 mg/dL) or taking hypoglycemic agents drugs or insulin; and increased waist circumference . p<0.001). However, heterogeneity was considerable (I2= 92.3%).I2 = 71.7%) in the analysis of non-linear dose–response relationship between circulating fetuin-A concentration and the risk of MetS , diastolic blood pressure (DBP), and triglycerides reporting were not significantly different. However, after introducing HDL into the meta-regression analysis model, results showed that sources of heterogeneity could be explained by HDL as the difference was significant. .In the non-linear dose–response relationship, there was a slight trend towards an increment in the risk of MetS with an increment in the circulating fetuin-A concentration, although the increment was not statistically significant (p = 0.06). There was high heterogeneity . Moreover, subgroup analyses were conducted to explore the impact of HDL, which showed that circulating fetuin-A concentrations were significantly higher in the MetS patients than in the controls for the two subgroups, high HDL (≥40mg/dL) concentrations subgroup  and low HDL (<40mg/dL) concentrations subgroup , but with high heterogeneity .The results of the analysis of dose-response relationship between HDL and SMD are shown in 2 = 41.5%). Besides, the concentration of circulating fetuin-A was significantly higher in the MetS patients with DBP ≥ 85mmHg than in the control group, and most of the heterogeneity was explained (I2 = 7.7%).t = 1.21, p = 0.245).Sensitivity analysis showed that the SMD and corresponding 95% CI changed little after each individual study was excluded sequentially, indicating that the overall results were relatively stable. The Egger funnel plot for circulating fetuin-A concentrations was symmetrical, and the Egger’s test did not reject the hypothesis that there was no publication bias  was associated with significantly higher heterogeneity. Thus, it may be suggested that the activity of circulating fetuin-A, or the molecular mechanisms of circulating fetuin-A bioconversion were impaired in the adult MetS patients. Further investigations are necessary to understand the molecular mechanisms that account for the changes of circulating fetuin-A concentrations for different age groups among the MetS patients.Furthermore, subgroup analysis showed that, for MetS patients with DBP≥85mmHg, the concentration of circulating fetuin-A was significantly higher than that of the control group, and this subgroup explained most of the heterogeneity. Considering that there were only three studies reporting DBP≥85mmHg, which represents limited available data thus far as regards MetS patients with DBP≥85mmHg, future research should expand the sample size to verify whether DBP plays a regulatory role in the MetS patients and circulating fetuin-A.Many published reviews have suggested that circulating fetuin-A may play a role in the risk of type 2 diabetes mellitus, cardiovascular disease, and nonalcoholic fatty liver disease , 19. ForFor instance, free fatty acids and circulating fetuin-A play important roles in the interaction and metabolic processes between the liver and adipose tissues. According to some experiments, free fatty acids can increase the activity of nuclear factor κB (NF-κB) by restraining the AMP protein-kinase (AMPK), which in turn is a consequence of ERK 1/2 activation , 21. LikEvidence from animal experiments has shown that elevated circulating fetuin-A concentrations could lead to impaired glucose control due to migration and activation of macrophage, impaired signaling of insulin receptors, triglyceride accumulation in hepatocytes, and dysfunction of adipocytes . When thIt has been proven from many clinical studies that circulating fetuin-A concentration in individuals with centripetal obesity is usually elevated. Given that centripetal obesity is closely related to insulin resistance and dyslipidemia, it is considered as a major sign of MetS . The expThe hypothesis of an association between MetS and circulating fetuin-A was supported not only by clinical and experimental studies, but also genetic epidemiological studies. For instance, it is widely accepted that the AHSG gene is located on chromosome 3q27, a region that has been identified as a MetS susceptibility locus. Thus, AHSG gene variant associated with lower circulating fetuin-A was more common in normal-weight men than in their overweight counterparts . TherefoIn this study, it was also shown that HDL played an important role in the relationship between MetS and circulating fetuin-A, which may be due to the fact that HDL is a possible component of dyslipidemia in MetS. However, it was further shown, through subgroup analysis, that there was significant heterogeneity among groups with different HDL concentrations and this was contrary to the results of the meta-regression analysis of HDL. The preceding contradiction may be caused by treating HDL differently in the subgroup and meta-regression analyses. For example, HDL was considered as a categorical variable in the subgroup analysis, whereas it was considered as a continuous variable in the meta-regression analysis. That is, taking HDL as a continuous variable might have increased the statistical efficiency of meta-regression analysis than considering it as a categorical variable in the subgroup analysis.Thus, the results of subgroup analysis due to HDL should be seen as exploratory and interpreted cautiously. Nevertheless, recent studies also revealed that there was a correlation between HDL and circulating fetuin-A . MoreoveThis meta-analysis and meta-regression are subject to several limitations. First, although there was a dose-response relationship between circulating fetuin-A and the risk of MetS, all the eligible studies analyzed were case-control studies, which could not make causality inference. In future, prospective cohort studies should be considered to explore the link between circulating fetuin-A and MetS. Second, majority of the eligible studies did not adjust for the possible confounding variables. Third, heterogeneity was high when synthesizing evidence on the relationship between the concentration of circulating fetuin-A and MetS. Therefore, future studies should consider investigating the association between the concentration of circulating fetuin-A and MetS in homogeneous populations using same methods. Last, only articles published in English were included. Therefore, this meta-analysis may be more prone to bias since some negative results are generally reported in regional journals in native languages.Circulating fetuin-A concentration in the MetS patients was significantly higher than that in the control groups, and there was a slight trend towards an increment in the risk of MetS with an increment in the circulating fetuin-A concentration. Also, HDL may play a possible regulating role between circulating fetuin-A and MetS. Therefore, these results provide a rationale for evaluating circulating fetuin-A to see if it could affect the pathophysiological process of the risk of MetS. Specifically, this evaluation might lead to new perspectives in the early diagnosis, identification of novel biomarkers, and discovery of novel targets for pharmacological interventions.S1 FileDetails of search strategy.(DOC)Click here for additional data file.S1 Table(DOC)Click here for additional data file.