Archives of Academic Emergency Medicine. 2019; 7 (1): e1 OR I G I N A L RE S E A RC H The Correlation between Serum Level of Vitamin D and Outcome of Sepsis Patients; a Cross-Sectional Study Majid Shojaei1, Anita Sabzeghabaei2, Helia Valaei Barhagh1∗, Soheil Soltani1 1. Emergency Department, Imam Hossein Hospital, Shahid Beheshti University of Medical Sciences, Tehran, Iran. 2. Emergency Department, Shohadaye Tajrish Hospital, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Received: September 2018; Accepted: September 2018; Published online: 10 January 2019 Abstract: Introduction: The effect of vitamin D deficiency in manifestation of sepsis and its role as an important mediator in the immune system has received attention. The present study was done with the aim of evaluating the corre- lation between serum levels of vitamin D and outcome of sepsis patients. Methods: The present cross-sectional study was performed on patients over 18 years of age suspected to sepsis presenting to an emergency depart- ment during 1 year using non-probability convenience sampling. For all eligible patients, blood sample was drawn for measurement of serum level of vitamin D3 and the correlation of this vitamin with outcomes such as mortality, renal failure, liver failure and etc. was assessed. Results: 168 patients with the mean age of 70.8 ± 13.3 (43.0 – 93.0) years were studied (56.0% male). Mean serum level of vitamin D3 in the studied patients was 19.03 ± 13.08 (4.0 – 85.0) ng/ml. By considering 20 – 50 ng/ml as the normal range of vitamin D, 61.6% of the patients had vitamin D deficiency. Only age (r=-0.261, p=0.037) and mortality (r=-0.426, p=0.025) showed a significant correlation with mean vitamin D. Sepsis patients with older age and those who died had a lower level of vitamin D. Area under the ROC curve of serum vitamin D level regarding 1-month mortality of the sepsis patients was 0.701 (95%CI: 0.439 – 0.964). Conclusion: Based on the results of the present study, the prevalence of vitamin D deficiency in sepsis patients presenting to the ED was estimated as 61.6%. A significant and indirect correlation was found between the serum level of vitamin D3 and mortality as well as older age. It seems that consump- tion of vitamin D supplements might be helpful in decreasing the prevalence of infection, sepsis, and mortality caused by it, especially in older age. Keywords: Systemic inflammatory response syndrome; sepsis; patient outcome assessment; 24, 25-dihydroxyvitamin d3; mortality Cite this article as: Shojaei M, Sabzeghabaei A, Valaei Barhagh H, Soltani S. The Correlation between Serum Level of Vitamin D and Outcome of Sepsis Patients; a Cross-Sectional Study. Arch Acad Emerg Med. 2019; 7(1): e1. 1. Introduction Sepsis is a clinical syndrome caused by immune response to various infections. Despite the existence of proper antimi- crobial and palliative care, due to the high prevalence of mul- tiple organ failure, being affected with sepsis is associated with a high rate of mortality. More than 31 million sepsis cases, 19 million severe sepsis cases, and 5.3 million deaths a year is a rough estimation of the burden of this disease in high-income countries (1-3). Recent studies have shown the correlation between vitamin D deficiency and severe infec- tions and sepsis. In addition, it has been confirmed that vita- ∗Corresponding Author: Helia Valaei Barhagh; Emergency Department, Imam Hossein Hospital, Madani Avenue, Imam Hossein Square, Tehran, Iran. Tel: 00989111325336 E-mail: valaeebarhagh@gmail.com min D is an important mediator in the immune system and its inhibitory role in sepsis pathogenesis has been evaluated (4-6). Vitamin D can regulate acquired and innate immune responses (7). This vitamin prevents overexpression of in- flammatory cytokines and is an important mediator in aggre- gation of leukocytes, formation of local inflammation, and anti-bacterial responses in innate immunity. Vitamin D defi- ciency has been linked to an increase in the risk of initiation and development of viral and bacterial infections (8-10). Low serum level of vitamin D on admission of patients to the in- tensive care unit has correlated with an increase in mortality risk and blood infections (11). Additionally, a higher risk of developing sepsis has been shown in patients who have had vitamin D deficiency before hospital admission (7, 10). How- ever, studies in this regard are still ongoing to reach a final de- cision and the correlation between vitamin D and decrease in This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem M. Shojaei et al. 2 mortality has not been confirmed in all studies (12-14). Iran is among the countries that have a considerable prevalence of vitamin D deficiency in all age groups. Iran’s ministry of health has strongly advised using vitamin D supplements for all age groups (15, 16). On the other hand, we are faced with a day by day increase in the number of sepsis patients in emer- gency departments (ED) and intensive care units. Therefore, the present study was designed and performed with the aim of evaluating the correlation between serum levels of vitamin D and outcome of sepsis patients presenting to ED. 2. Methods 2.1. Study design and setting The present cross-sectional study was performed on patients suspected to sepsis presenting to the ED of Imam Hossein Hospital, Tehran, Iran, from 2015 to 2016. For all the eligi- ble patients, blood sample was drawn for measuring serum level of vitamin D and finally the correlation between the level of this vitamin and the studied outcomes was evalu- ated. The study was performed by adhering to the principles of Helsinki Declaration and approval of the ethics committee of Shahid Beheshti University of Medical Sciences. Inclusion of patients was done after obtaining informed consent from the patient or their relative. Keeping patient data confiden- tial and publishing findings anonymously were among other measures for ethical conduct in this study. 2.2. Participants The studied sample included patients over 18 years of age suspected with sepsis presenting to the emergency depart- ment in various working shifts, who were selected via non- probability convenience sampling. Sepsis diagnosis was con- firmed after clinical examinations, meeting the required cri- teria and performance of laboratory tests. Patients with a history of neuroendocrine diseases, history of mental disor- ders, shock due to reasons other than sepsis, consumption of corticosteroid drugs, recent and constant use of vitamin D or other supplements including it, pregnant women, those who did not give their consent for participation in the study, and those whose sepsis diagnosis was not confirmed in the end based on the mentioned criteria were eliminated from the study. Sepsis was defined as the presence of at least 2 of the symptoms of systemic inflammatory response syndrome (SIRS) with the origin of a suspected or confirmed infectious source (17). 2.3. Sampling Venous blood was drawn from the median cubital vein of the patients for measuring the serum level of vitamin D. Blood samples were kept in room temperature for 15-30 minutes and centrifuged (1500 rpm) for 10-15 minutes. Consequently, serums were separated and kept in -20◦C until the time of evaluation. After completion of samples, the concentration of serum 25-hydroxy vitamin D3 was measured. Measure- ment of serum concentration of 25-hydroxy vitamin D3 was done with radioimmunoassay method using DIASource kit made in Belgium and based on the guidelines of the kit. 2.4. Data gathering Data gathering was done by a trained senior emergency medicine resident using a pre-designed checklist including demographic data (age, sex), laboratory findings, vital signs, level of consciousness, origin of the infection, sepsis severity based on Mortality in Emergency Department Sepsis (MEDS) score, and finally, the outcome of patients (death, renal fail- ure, liver failure, need for mechanical ventilation and etc.). 30-day outcome of the patients (mortality) was gathered by follow-up via phone calls. Based on MEDS score, the patients were divided into 5 groups based on the probability of death including: 1- very low probability, 2- low probability, 3- mod- erate probability, 4- high probability, and 5- very high proba- bility. 2.5. Statistical Analysis Sample size required for the present study was determined to be 195 cases, based on the study by Rech et al. (6), and considering 54% prevalence of vitamin D deficiency in sep- sis patients, 95% confidence interval (CI) and maximum ac- ceptable error in estimation of prevalence as 7%. To describe data, frequency and percentage or mean ± standard devia- tion were reported. To assess the correlation between various factors and vitamin D deficiency, chi-square, Fisher’s exact and ANOVA tests were applied. All analyses were performed using SPSS 21.0 statistical software. Calculation of area under the receiver operating characteristic (ROC) curve was applied for determining the cut-off point of serum level of vitamin D in correlation with mortality. In this study, p<0.05 was con- sidered as the level of significance. 3. Results 168 patients with the mean age of 70.8 ± 13.3 (43.0 – 93.0) years were studied (56.0% male). Tables 1 and 2 depict the baseline characteristics and laboratory findings of the stud- ied patients. Mean serum level of vitamin D3 in the studied patients was 19.03 ± 13.08 (4.0 – 85.0) ng/ml. By consider- ing 20 – 50 ng/ml as the normal range of vitamin D, 61.6% of the patients had vitamin D deficiency. CRP (p=0.176) and platelet (p=0.951) levels, as 2 important inflammatory in- dices, were not significantly different between the patients with vitamin D deficiency and others. Table 3 shows the cor- relation between mean serum level of vitamin D and differ- ent variables. Only age (r=-0.261, p=0.037) and mortality (r=- This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem 3 Archives of Academic Emergency Medicine. 2019; 7 (1): e1 Table 1: Baseline characteristics of studied patients Variable Value Sex Male 94.0 (56.0) Female 74.0 (44.0) Age (year) 40 – 60 39 (23.0) ≥ 60 129 (77.0) Vital signs Systolic blood pressure (mmHg) 89.41 ± 18.63 Pulse rate (/minute) 103.6 ± 9.5 Temperature (C) 38.4 ± 0.7 Saturation O2 (%) 85.1 ± 9.4 GCS 3 - 7 16 (9.6) 8 - 14 87 (50.7) 15 67 (39.7) Source of infection Pneumonia 46 (31.1) Urinary tract infection (UI) 44 (29.7) Pneumonia + UI 32 (21.6) Bed sore 14 (9.5) Multi source 10 (6.9) Gastrointestinal 2 (1.2) Mechanical ventilation Yes 56 (43.1) No 74 (56.9) Septic shock Yes 28 (26.9) No 76 (73.1) 30-day mortality Yes 106 (63.1) No 62 (36.9) MEDS score 1 31 (18.5) 2 25 (14.9) 3 62 (36.9) 4 33 (19.6) 5 17 (10.1) Data are presented as mean ± standard deviation or number (%). There is missing data in some variables. GCS: Glasgow coma scale. 0.426, p=0.025) showed a significant correlation with mean serum vitamin D level. Sepsis patients with older age and those who died had a lower level of vitamin D. Area under the ROC curve of vitamin D regarding 1-month mortality of the sepsis patients was 0.701 (95%CI: 0.439 – 0.964) (figure 1). Patients whose vitamin D level was less than 25 ng/ml were significantly more at risk of death (p=0.016). Relative risk of mortality in patients with a vitamin D level less than 25 ng/ml was estimated to be 1.42 (95%CI: 1.027- 1.970). 4. Discussion Based on the results of the present study, the prevalence of vitamin D deficiency among sepsis patients presenting to the Figure 1: Area under the curve of serum level of vitamin D and 1- month mortality of the sepsis patients presenting to emergency de- partment. ED was estimated as 61.6%. A significant indirect correlation was observed between the serum level of vitamin D and mor- tality as well as older age. In line with the present study, previ- ous studies have suggested that vitamin D concentration cor- relates with the rate of mortality in patients (4-6). In the study by Parekh et al. there was a significant correlation between mean serum level of vitamin D in patients with sepsis who died and those who survived (18). The results of a study by Rech et al. showed that vitamin D deficiency in patients with severe sepsis or septic shock was significantly associated with increase in 30-day mortality (6). In the study by Nguyen et al. also low serum level of vitamin D was considered related to increase in 23-day mortality among 91 patients with sepsis (5). In Arnson et al. study the correlation between vitamin D deficiency and mortality rate was confirmed in critically ill patients (11). In 2399 patients, vitamin D deficiency from 365 days before hospitalization was reported as a reliable predic- tor for both short term and long term mortality and positive blood culture for microorganism growth (19). The correla- tion between sufficient vitamin D and reduced mortality has not been reported in all studies (13, 14). In one study, the rate of mortality in sepsis patients correlated with vitamin D level upon admission, but this correlation was not significant (20). Based on the National Health and Nutrition Examina- tion Survey, level of 25-hydroxy vitamin D had an inverse cor- relation with occurrence of upper respiratory tract infection (21). The evidence is also indicative of the effect of vitamin D level on influenza and invasive pneumococcal disease (22). Two meta-analyses performed on observational studies have confirmed that there is a significant correlation between vita- min D status and risk of being affected with sepsis, infection severity and 30-day mortality (23, 24). An important limita- This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem M. Shojaei et al. 4 Table 2: Laboratory findings of studied patients Variable Mean ± SD pH 7.38 ± 0.08 HCO3 (mEq/L) 23.81 ± 6.58 PaCO2 (mmHg) 38.52 ± 11.51 Hemoglobin (g/dl) 11.02 ± 2.21 Hematocrit (%) 34.21 ± 7.90 White blood cell (109 /L) 12256.46 ± 8597.33 Platelet count (109 /L) 234675.67 ± 136736.51 Polymorphonuclear (%) 80.62 ± 8.33 Blood urea nitrogen (mg/dL) 81.59 ± 47.09 Creatinine (mg/dL) 1.49 ± 1.26 Aspartate aminotransferase (U/L) 44.61 ± 89.99 Alanine aminotransferase (U/L) 53.77 ± 109.80 Alkaline phosphatase (U/L) 266.41 ± 134.47 Bilirubin total (mg/dL) 0.77 ± 0.40 Bilirubin direct (mg/dL) 0.34 ± 0.28 Vitamin D3 (ng/ml) 19.03 ± 13.08 C-reactive protein (mg/dL) 66.69 ± 28.57 SD: standard deviation. tion present in all observational studies is also present in our study and it is not known if vitamin D deficiency is a marker for sepsis or if it causes sepsis, in other words if vitamin D deficiency is the cause for sepsis or its effect. In an experi- mental study on mice the causality of vitamin D as the reason for mortality in sepsis was evaluated. The findings of this ex- perimental study showed that imposing vitamin D deficiency on the mice through their diet led to sepsis and increase in inflammation and cellular disorders and bacterial growth as well as disturbing the balance of apoptotic neutrophil aggre- gation. These experimental studies support the hypothesis that vitamin D deficiency might be a major cause in mani- festation of sepsis (18, 25). It seems that with all the present evidence, consumption of vitamin D supplement might be able to reduce infection, sepsis and mortality caused by it, especially in older ages. 5. Limitation Controlling all the probable confounding factors in observa- tional studies such as the present one is not possible and this can affect the findings of the study. In addition, this study was a single centered one with a relatively small sample size, which makes data generalization hard. Another limitation of the study was that some data affecting the level of vitamin D such as cytokine level could not be recorded. 6. Conclusion Based on the results of the present study, the prevalence of vitamin D deficiency in sepsis patients presenting to the ED was estimated as 61.6%. A significant and indirect correlation was found between the serum level of vitamin D and mortal- Table 3: Relationship between mean serum vitamin D3 level and different factors Variable Vitamin D3 P Sex Male 18.67 ± 13.62 0.726 Female 19.43 ± 12.54 Age (year) 40 – 60 14.44 ± 7.08 0.037 ≥ 60 20.18 ± 14.40 30-day mortality Yes 17.4 ± 9.9 0.025 No 21.4 ± 11.5 Source of infection Pneumonia 20.3 ± 13.0 Urinary tract infection (UI) 18.1 ± 17.0 Pneumonia + UI 18.1 ± 9.4 0.825 Bed sore 18.1 ± 10.1 others 18.3 ± 3.4 MEDS score 1 18.9 ± 7.7 2 21.6 ± 11.2 3 18.4 ± 11.7 0.642 4 18.8 ± 11.5 5 16.7 ± 8.8 Septic shock Yes 21.53± 4.70 0.375 No 18.83 ± 12.87 Mechanical ventilation Yes 20.09 ± 15.73 0.428 No 18.01 ± 12.71 Acute renal failure (Cr>1.6 mg/dl) Yes 21.92 ± 18.20 0.109 No 17.95 ± 10.66 Liver failure (AST>45 U/L) Yes 20.07 ± 15.87 0.613 No 18.66 ± 12.86 MEDS: Mortality in Emergency Department Sepsis. ity as well as older age. It seems that consumption of vitamin D supplements might be helpful in decreasing the prevalence of infection, sepsis, and mortality caused by it, especially in older age. 7. Appendix 7.1. Acknowledgements The staff of emergency department and laboratory of Imam Hossein Hospital are thanked for their cooperation in carry- ing out this project. This article is derived from the thesis of Dr. Helia Valaei to achieve her specialist degree in emergency medicine. 7.2. Author contribution All the authors met the criteria of authorship based on the recommendations of the international committee of medical journal editors. This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem 5 Archives of Academic Emergency Medicine. 2019; 7 (1): e1 Authors ORCIDs Majid Shojaei: 0000-0001-6819-9237 7.3. 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Inter- This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem M. Shojaei et al. 6 national journal of medical sciences. 2007;4(4):190. This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem Introduction Methods Results Discussion Limitation Conclusion Appendix References