Vitamin D Deficiency and Hematological Parameters in People Living with HIV/AIDS

ansinet.com/pjn Science Alert scialert.net an open access publisher http://ansinet.com OPEN ACCESS Pakistan Journal of Nutrition ISSN 1680-5194 DOI: 10.3923/pjn.2020.217.224 Research Article Vitamin D Deficiency and Hematological Parameters in People Living with HIV/AIDS 1 Juli Yosa Mega, 1Elrica, 2Dina Keumala Sari and 3Dewi Indah Sari Siregar 1 Master Program in Tropical Medicine, Faculty of Medicine, Universitas Sumatera Utara, Medan 20155, North Sumatera, Indonesia Department of Nutrition, Faculty of Medicine, Universitas Sumatera Utara, Medan 20155, North Sumatera, Indonesia 3 Department of Clinical Pathology, Faculty of Medicine, Universitas Sumatera Utara, Medan 20155, North Sumatera, Indonesia 2 Abstract Background and Objective: Patients with Human Immunodeficiency Virus (HIV)/Acquired Immunodeficiency Syndrome (AIDS) infection experiences deficiency of vitamin D and abnormality of hematological parameters related to inflammatory and thrombotic activities. This study was designed to determine the relationship between changes in hematological parameters and vitamin D in HIV/AIDS infection. Materials and Methods: A cross-sectional study was conducted on 70 HIV/AIDS patients consuming Efavirenz (EFV)-based Antiretroviral Therapy (ART) for less than 6 months. Parameters including 25-hydroxy-vitamin D [25(OH)D] level and complete blood count. All parameters were measured in the Special Treatment Center (Pusat Pelayanan Khusus, Pusyansus) of the Voluntary Counseling and Testing (VCT) Clinic at Rumah Sakit Umum Pusat (RSUP) Haji Adam Malik, Medan, Indonesia. Results: There was a significant difference in terms of platelet count [mean±standard deviation: 329531.25 (79175.99) µLG1 vs 282710.53 (69895.25) µLG1, p = 0.011], Mean Platelet Volume/Platelet count (MPV/PLT) [Median (Interquartile Range): 2.86×10G5 (1.70×10G5-5.06×10G5) vs 3.27×10G5 (1.94×10G5-10.0×10G5), p = 0.022] and plateletcrit (PCT) [mean±standard deviation: 0.29 (0.08)% vs 0.25 (0.06)%, p = 0.018] in the group with 25(OH)D level of <21 ng mLG1 compared to the group with 25(OH)D level of $21 ng mLG1. There was a significant difference in terms of platelet count between the vitamin D sufficiency and insufficiency groups [mean±standard deviation: 300166.67 (71387.33) µLG1 vs 274653.85 (69095.84) µLG1, p = 0.036]. A significant difference was found in terms of platelet distribution width (PDW) between the vitamin D deficiency and insufficiency groups [mean±standard deviation: 9.02 (1.14)% vs 9.48 (1.03)%, p = 0.020]. Conclusion: Low level of vitamin D significantly correlated with platelet index in HIV/AIDS patients consuming EFV-based ART. Key words: 25-hydroxyvitamin D, complete blood count, deficiency, HIV/AIDS Received: January 12, 2020 Accepted: March 15, 2020 Published: April 15, 2020 Citation: Juli Yosa Mega, Elrica, Dina Keumala Sari and Dewi Indah Sari Siregar, 2020. Vitamin D deficiency and hematological parameters in people living with HIV/AIDS. Pak. J. Nutr., 19: 217-224. Corresponding Author: Dina Keumala Sari, Department of Nutrition, Faculty of Medicine, Universitas Sumatera Utara, Medan 20155, North Sumatera, Indonesia Tel: (+6261) 8212296/+6281397177693 Copyright: © 2020 Juli Yosa Mega et al. This is an open access article distributed under the terms of the creative commons attribution License, which permits unrestricted use, distribution and reproduction in any medium, provided the original author and source are credited. Competing Interest: The authors have declared that no competing interest exists. Data Availability: All relevant data are within the paper and its supporting information files. Pak. J. Nutr., 19 (5): 217-224, 2020 aggregative in nature, possess higher concentration of INTRODUCTION thromboxane A2 and express more glycoprotein (Gp) Ib and IIb/IIIa receptors16. PCT and PDW are platelet parameters that Decreased level of vitamin D in HIV infection may be due to a pre-existing deficiency of 25(OH)D which contributes to the incidence of HIV, or through chronic inflammation in HIV and induction of pro-inflammatory cytokine production may also play a role, leading to disruption in vitamin D metabolism. This decrease may also be caused by complications of infection, leading to less exposure to sunlight and inadequate nutritional intake1,2. Vitamin D deficiency may occur in both healthy individuals and individuals with illnesses. The incidence of vitamin D deficiency also increase in patients with tuberculosis (TB)3-5. Deficiency in healthy individuals is related to several risks, such as old age, living in areas with low ultraviolet radiation and a habit to avoid exposure to sunlight6. Medication may also play a role in decreasing vitamin D level. Medications such as EFV has been reported to contribute to decreased vitamin D level, due to it being a strong inducer of CYP2B6 and CYP3A enzymes, which were known to be associated with vitamin D metabolism. Induction of CYP3A causes catalysis of 4-hydroxylation 25(OH)D and contributes to vitamin D deficiency7. HIV/AIDS patients also experienced problems related to coagulation abnormalities. Age, CD4 concentration, viral load, opportunistic infections, medications and viral factors may play a role in the coagulation abnormalities experienced by HIV/AIDS patients. Complications due to these abnormalities may include cardiovascular diseases, stroke, thromboembolism and even HIV/AIDS mortality. Abnormalities in coagulation may be observed through changes in platelet counts, as well as changes in coagulation factors, both extrinsic and intrinsic8. Anti-thrombogenic, anti-inflammatory and anticoagulant activities may be affected by vitamin D level. Low-level vitamin D is often associated with inflammation, endothelial dysfunction, higher risk of cardiovascular diseases, higher risk of infection, HIV progressivity and mortality. Persistent inflammation is often associated with low-level vitamin D and an increase in MPV7,9-11. Several studies have reported a correlation between vitamin D level and platelet index 6,9,11-14. Platelets release inflammatory mediators, activate the complement factors and increase vascular permeability, contribute to blood coagulation, hemostasis and thrombosis, as well as atherosclerosis. Platelet consumption in chronic inflammation causes an increase in MPV, which is a marker of platelet activity, linking it to inflammatory and thrombotic processes. The higher the MPV, the more reactive and thrombogenic the hemostasis11,15. Compared to smaller platelets, larger platelets have more granules, are more provide thrombotic activities. PCT is a platelet concentration parameter and describe changes of platelet production and activity as well as thrombotic potentials6. In vitamin D deficiency, inflammatory response, pro-inflammatory cytokine (TNF-α and IL-6) levels, platelet reactivity and MPV are all increased13,16. Due to the fact and HIV infection is often accompanied by vitamin D deficiency and due to the fact that this deficiency is often correlated to abnormalities in hematological parameters, so, this study aimed to determine the relationship between changes in hematological parameters and vitamin D in HIV/AIDS infection. MATERIALS AND METHODS Study design: This is an analytical descriptive study using cross-sectional approach, conducted between August and October 2019 in the Special Treatment Center (Pusat Pelayanan Khusus, Pusyansus) of the Voluntary Counseling and Testing (VCT) Clinic in Rumah Sakit Umum Pusat (RSUP) Haji Adam Malik Medan, Sumatera Utara, Indonesia. The study involved HIV/AIDS patients with the following inclusion criteria: aged $20 years and being treated with EFV-based ART for less than 6 months. Exclusion criteria include history of vitamin D supplement consumption, history of anticoagulant consumption, history of thromboembolism, history of chronic kidney disease, a history of cirrhosis, increased in liver function markers for more than 5 times normal values, history of blood transfusion and pregnancy. Patients fulfilling the inclusion criteria will be provided with a written explanation and subsequently asked for consent to participate as subjects in the study. This study has been approved by the Health and Medical Research Ethics Committee of the Faculty of Medicine of Universitas Sumatera Utara/ RSUP Haji Adam Malik Medan with an ethical clearance certificate. [Reference Number: 625/TGL/KEPK FK USU-RSUP HAM/2019]. Data collection and procedure: Data were collected using anthropometric measurements for weight, height, to obtain body mass index (BMI) and using a questionnaire. Data related to currently-consumed ART and its duration were obtained from the medical records. Measurements of serum vitamin D 25(OH)D level were conducted using chemiluminescent microparticle immunoassay (CMIA) ARCHITECT 25-OH Vitamin D. Levels of 25(OH)D were classified based on US 218 Pak. J. Nutr., 19 (5): 217-224, 2020 We found a statistically significant difference in terms of platelet count, MPV/PLT and PCT (p = 0.011; p = 0.022 and p = 0.018, respectively). Platelet and PCT were higher in the group with 25(OH)D level of <21 ng mLG1 while MPV/PLT was higher in the group with 25(OH)D level of $21 ng mLG1. There was no statistically significant difference in terms of hemoglobin concentration, hematocrit, WBC count, platelet distribution width (PDW), neutrophil to lymphocyte ratio (NLR) and platelet to lymphocyte ratio (PLR) between the two study groups (Table 2). We found a statistically significant difference in terms of age (p = 0.034), platelet count (p=0.024), MPV/PLT (p = 0.027) and PDW (p = 0.034) when comparing between vitamin D deficiency, insufficiency and sufficiency (Table 3). There was no statistically significant difference between vitamin D status and hemoglobin concentration, Total Lymphocyte Count (TLC), platelet count, MPV and PCT (p>0.05); however, we found a statistically significant difference between vitamin D status and PDW (p = 0.024) (Table 4). A total of subjects experienced anemia, one in the vitamin D deficiency group and 1 in the vitamin D sufficiency group. Twelve subjects had low TLC across the three vitamin D status groups. Higher platelet count was observed in 2 subjects belonging to the vitamin D deficiency group. High MPV was also observed in 13 subjects belonging to the vitamin D deficiency and insufficiency groups. Low PDW was observed in 54 subjects across the three vitamin D status groups. Low PCT was observed in 1 subject from the vitamin D insufficiency group. We also found no relationship between BMI, age, hemoglobin concentration, hematocrit, Neutrophil to Lymphocyte Ratio (NLR) and Platelet to Lymphocyte Ratio (PLR) with platelet count. We found a positive correlation between BMI and MPV and negative correlation between NLR-PLR and MPV (p<0.05). We found negative correlation between age and PCT and positive correlation between WBC count as well as PLR and PCT (p<0.05). BMI, hemoglobin concentration and hematocrit were positively correlated with PDW (p<0.05) (Table 5). Across the study groups, we found statistically significant difference in terms of vitamin D level with duration of sunlight exposure (p = 0.007) with mean 25(OH)D level of 19.97 ng ng mLG1 in the group with <30 min sunlight exposure/day and 25.39 ng mLG1 in the group with $30 min sunlight exposure/day. In relation to frequency of sunlight exposure, we found no statistically significant difference in terms of vitamin D level across the three sunlight exposure frequency groups (p = 0.280). Mean 25(OH)D levels were 21.45, 21.15, 21.15 and 24.63 ng mLG1 in the groups with rare Endocrine Society Classification, as vitamin D deficiency (#20 ng mLG1), vitamin D insufficiency (21-29 ng mLG1) and vitamin D sufficiency ($30 ng mLG1)17. Complete blood count was measured with Hematology Analyzer using flow cytometer principles. Results were considered normal if: hemoglobin concentration is $10 g dLG1, White Blood Cells(WBC) 4.000-11.000 µLG1, platelet count between 150,000-450,000 µLG1, MPV 6.5-9.5 fL, PCT 0.10-0.50% and PDW between 10.0-18.0%. Statistical analyses: Statistical analyses were performed using IBM SPSS Statistics version 25.0. Variables were described using proportion for categorical variables and using mean and standard deviation as well as median and IQR for numerical variables with normal and non-normal distribution, respectively. Statistical significance between study groups was determined using non-paired t-test and Mann-Whitney test for numerical variables with normal and non-normal distributions, respectively; as well as χ2 and Fisher exact tests for categorical variables with normal and non-normal distribution, respectively. Comparative statistics analyses were conducted on more than 2 unpaired groups, with One-way ANOVA test used on data with normal distribution and Kruskal-Wallis test used on data with non-normal distribution followed by post-hoc analysis. An unpaired analysis was performed on categorical variables using crosstabs and MannWhitney test. Correlation tests using Pearson and Spearman correlations were conducted on numerical variables with normal and non-normal distributions, respectively. p-value of less than 0.05 was considered significant. RESULTS Table 1 presents a comparison between 32 subjects with 25(OH)D level of <21 ng mLG1 and 38 subjects with 25(OH)D level of $21 ng mLG1. Sociodemographic and clinical characteristics were also presented in Table 1. There was no statistically significant difference in terms of gender (p = 0.785) and age (p = 0.300) between the two study groups. We found no significant difference in terms of WHO HIV stage between the group with 25(OH)D level of <21 ng mLG1 [stage I/II (12.5%) and stage III/IV (87.5%)] and the group with 25(OH)D level of $21 ng mLG1 [stage I/II ( 23.7%)] and III/IV [(76.3%); p = 0.373]. There was no significant difference in terms of BMI between the two groups (p = 0.262). No significant difference was observed in terms of sunlight exposure duration between the two groups (p = 0.144). The frequency of sunlight exposure was similar between the two groups (p = 0.364). 219 Pak. J. Nutr., 19 (5): 217-224, 2020 Table 1: Sociodemographic and clinical characteristics of HIV/AIDS subjects with 25(OH)D levels of <21 and $21 ng mLG1 Parameters 25(OH)D <21 ng mLG1 (n = 32) 25(OH)D $21 ng mLG1 (n = 38) --------------------------------------- --------------------------------------- No. Percentage No. Percentage p-value 27 84.4 5 15.6 >40 years old 5 #40 years old 27 Higher Lower OR 30 78.9 0.785 1.440 8 21.1 15.6 11 28.9 0.300 0.455 84.4 27 71.1 29 90.6 30 78.9 0.314 2.578 3 9.4 8 21.1 9 28.1 10 26.3 1.000 1.096 23 71.9 28 73.7 - - 0.005 4.686 0.373 0.460 0.262 2.160 1.000 0.881 0.144 2.292 0.364 1.761 Sex Male Female Age Education Occupation None Working Mode of transmission Homosexual 5 15.6 3 7.9 Heterosexual 17 53.2 21 60.6 IDU or tattoo 1 3.1 1 2.6 Blood transfusion 1 3.1 1 2.6 Unknown 8 25.0 10 26.3 Marital status Married 21 65.6 11 28.9 Unmarried 11 34.4 27 71.1 WHO stage I/II 4 12.5 9 23.7 28 87.5 29 76.3 BMI $25 kg mG2 5 15.6 3 7.9 BMI <25 kg mG2 27 84.4 35 92.1 III/IV Body mass index (BMI) Duration of ART >90 days 21 65.6 26 68.4 #90 days 11 34.4 12 31.6 Duration of exposure to sunlight <30 minutes dayG1 20 62.5 16 42.1 $30 minutes dayG1 12 37.5 22 57.9 Frequency of exposure to sunlight 1-3 times weekG1 and 4-5 times weekG1 22 68.8 21 55.3 6-7 times weekG1 10 31.3 17 44.7 25(OH)D: 25-hydroxycholecalciferol/Calcidiol, IDU: Injection drug users, WHO: World health organization, BMI: Body mass index, ART: Antiretroviral therapy, EFV: Efavirenz, RIF: Rifampicin, p#0.05 is considered statistically significant Table 2: Laboratory examination characteristics of HIV/AIDS subjects with 25(OH)D levels of <21 and $21 ng mLG1 Parameters 25(OH)D <21 ng mLG1 (n = 33) 25(OH)D $21 ng mLG1 (n = 37) Hemoglobin (g dLG1), mean (SD) 13.12 (1.99) 13.58 (1.86) p-value 0.320 Hematocrit (%), mean (SD) 38.97 (4.99) 39.61 (4.82) 0.590 WBC (µLG1), median (IQR) 6750 (3050-11240) 6620 (3230-16250) 0.883 Neutrophil (%), mean (SD) 51.65 (10.46) 56.29 (12.27) 0.097 PLT (µLG1), mean (SD) 329531.25 (79175.99) 282710.53 (69895.25) 0.011 Lymphocytes (%), mean (SD) 32.71 (9.80) 28.07 (9.92) 0.054 TLC, mean (SD) 2206.28 (922.75) 1861.17 (755.05) 0.090 MPV (fL), mean (SD) 8.96 (0.72) 9.04 (0.66) 0.642 MPV/PLT, median (IQR) 2.86×10G5 (1.70×10G5-5.06×10G5) 3.27×10G5 (1.94×10G5 -10.0×10G5) 0.022 PCT (%), mean (SD) 0.29 (0.08) 0.25 (0.06) 0.018 0.534 PDW (%), mean (SD) 9.02 (1.14) 9.18 (1.05) NLR, median (IQR) 1.66 (0.52-4.46) 1.92 (0.84-89.44) 0.191 PLR, median (IQR) 8970.16 (5405.41-21497.01) 8844.92 (4594.38-439325.84) 0.962 IQR: Interquartile Range, SD: Standard deviation, WBC: White blood cell, TLC: Total lymphocyte count, MPV: Mean platelet volume, MPV/PLT: Mean platelet volume/platelet count, PCT: Plateletcrit, PDW: Platelet distribution width, 25(OH)D: 25-hydroxycholecalciferol/Calcidiol, NLR: Neutrophil to lymphocyte ratio, PLR Platelet to lymphocyte ratio, p#0.05 is considered statistically significant 220 Pak. J. Nutr., 19 (5): 217-224, 2020 Table 3: Demographic, clinical and laboratory examination characteristics in vitamin D deficiency, insufficiency and sufficiency groups Age (years), median (IQR) BMI (kg mG2), mean (SD) Duration of ART treatment (days), mean (SD) Hemoglobin (g dLG1), mean (SD) Hematocrit (%) WBC (µLG1), median (IQR) Neutrophil (%), mean (SD) PLT (µLG1), mean (SD) Lymphocytes (%), mean (SD) TLC, mean (SD) MPV (fL), mean (SD) MPV/PLT, median (IQR) PCT (%), mean (SD) PDW (%), mean (SD) 25(OH)D (ng mLG1), mean (SD) NLR, median (IQR) PLR, median (IQR) 25(OH)D (ng mLG1) -------------------------------------------------------------------------------------------------------------------------------------Deficiency (#20) (n = 32) Insufficiency (21-29) (n = 26) Sufficiency ($30) (n = 12) p-value 28.00 (22.00-52.00) 21.65 (3.25) 34.00 (24.00-68.00) 22.09 (2.87) 36.50 (22.00-66.00) 21.73 (1.95) 0.034a 0.848 111.06 (57.05) 13.12 (1.99) 38.97 (4.99) 6750 (3050-11240) 51.65 (10.46) 329531.25 (79175.99) 32.71 (9.80) 2206.28 (922.75) 8.96 (0.72) 2.86×10G5 (1.70×10G5-5.06×10×5) 0.29 (0.08) 9.02 (1.14) 15.53 (4.09) 1.66 (0.52-4.46) 8970.16 (5405.41-21497.01) 108.23 (55.79) 13.86 (1.78) 40.38 (4.29) 6800 (4180-16250) 56.47 (12.26) 274653.85 (69095.84) 28.67 (10.19) 1993.99 (803.06) 9.19 (0.68) 3.50×10G5 (1.94×10G5-10.0×10G5) 0.25 (0.06) 9.48 (1.03) 25.10 (2.30) 1.71 (0.84-89.44) 8576.82 (4595.38-439325.84) 126.08 (56.00) 0.651 12.97 (1.95) 0.253 37.92 (5.65) 0.305 6035 (3230-9580) 0.312 55.89 (12.83) 0.252 300166.67 (71387.33) 0.024b 26.78 (9.60) 0.137 1573.39 (565.57) 0.085 8.72 (0.53) 0.134 2.80×10G5 (2.15×10G5-4.78×10G5) 0.027c 0.26 (0.06) 0.058 8.53 (0.80) 0.034d 36.05 (5.04) <0.001 2.16 (0.84-8.44) 0.390 11888.20 (4826.18-18541.67) 0.326 Kruskal-Wallis Test. Mann-Whitney Analysis: Deficiency vs. Insufficiency p = 0.073, Deficiency vs. Sufficiency p = 0.020, Insufficiency vs. Sufficiency p = 0.256. bOne-way Anova Test. Bonferroni post-hoc analysis: Deficiency vs. Insufficiency p = 0.299, Deficiency vs. Sufficiency p = 0.534, Insufficiency vs. Sufficiency p = 0.036. cKruskal-Wallis Test. Mann-Whitney Analysis: Deficiency vs. Insufficiency p = 0.009, Deficiency vs. Sufficiency p = 0.474, Insufficiency vs. Sufficiency p = 0.137. dOne-way anova test. Bonferroni post-hoc analysis: Deficiency vs. Insufficiency p = 0,020, Deficiency vs. Sufficiency p = 0.741, Insufficiency vs. Sufficiency p = 0.986. IQR: Interquartile Range, SD: Standard deviation, BMI: Body mass index, WBC: White blood cell, TLC: Total lymphocyte count, MPV: Mean platelet volume, MPV/PLT: Mean platelet volume/platelet count, PCT: Plateletcrit, PDW: Platelet distribution width, 25(OH)D: 25-hydroxycholecalciferol/ Calcidiol, NLR: Neutrophil to lymphocyte ratio, PLR Platelet to lymphocyte ratio, p#0.05 is considered statistically significant a Table 4: Hematological parameter categories based on vitamin D status across study groups 25(OH)D (ng mLG1) ---------------------------------------------------------------------------------------------------------------#20 21-29 $30 -------------------------------------------------------------------------------------No. Percentage No. Percentage No. Percentage Hemoglobin (g dLG1) TLC Platelet (µLG1) MPV (fL) PDW (%) PCT (%) Anemia (<10) No anemia ($10) Low (<1200) Normal ($1200) Low (<150,000) Normal (150,000-450,000) High (>450,000) Low (<6.5) Normal (6.5-9.5) High (>9.5) Low (<10) Normal (10-18) High (>18) Low (<0,1) Normal (0,1-0,5) High (>0,5) 1 31 4 28 0 30 2 0 26 6 26 6 0 0 32 0 3.1 96.9 12.5 87.5 0.0 93.8 6.3 0.0 81.3 18.8 81.3 18.8 0.0 0.0 100.0 0.0 0 26 4 22 1 25 0 0 19 7 16 10 0 1 25 0 0.0 100.0 15.4 84.6 3.8 96.2 0.0 0.0 73.1 26.9 61.5 38.5 0.0 3.8 96.2 0.0 1 11 3 9 0 12 0 0 12 0 12 0 0 0 12 0 8.3 91.7 25.0 75.0 0.0 100.0 0.0 0.0 100.0 0.0 100.0 0.0 0.0 0.0 100.0 0.0 p-value 0.702 0.372 0.381 0.111 0.024 1.000 25(OH)D: 25-hydroxycholecalciferol/Calcidiol, TLC: Total lymphocyte count, MPV: Mean platelet volume, PCT: Plateletcrit, PDW: Platelet distribution width, p#0,05 is considered statistically significant with BMI of <25 kg mG2. No statistically significant difference was found in terms of vitamin D levels with duration of ART treatment (p = 0.342) with mean 25(OH)D level of 23.27 ng mLG1 in the group with duration of ART treatment >90 days and 21.23 ng mLG1 in the group with duration of ART treatment #90 days. exposure (1-3 times weekG1), moderate exposure (4-5 times weekG1) and frequent exposure (6-7 times weekG1), respectively. We also found no statistically significant difference in terms of vitamin D levels with BMI of $25 kg mG2 (p = 0.190) with mean 25(OH)D level of 18.93 ng mLG1 in the group with BMI of $25 kg mG2 and 23.08 ng mLG1 in the group 221 Pak. J. Nutr., 19 (5): 217-224, 2020 Table 5: Correlation between platelet index and independent variables PLT MPV PCT PDW ------------------------------ ----------------------------- ----------------------------- ----------------------------- p-value r p-value p-value r p-value BMI -0.155 r 0.201 0.337 0.004 -0.053 0.662 0.445 <0.001 Age -0.210 0.081 0.010 0.936 -0.268 0.025 0.039 0.750 Duration of ART treatment (days) -0.058 0.631 0.143 0.239 -0.020 0.869 0.198 0.100 Hemoglobin -0.210 0.081 0.035 0.772 -0.205 0.088 0.344 0.004 Hematocrit -0.145 0.231 0.031 0.799 -0.107 0.380 0.343 0.004 WBC r 0.201 0.096 -0.098 0.418 0.247 0.040 0.104 0.393 -0.117 0.334 0.011 0.925 -0.108 0.372 -0.101 0.405 Lymphocytes 0.016 0.897 0.057 0.637 0.029 0.813 0.221 0.066 TLC 0.147 0.225 0.016 0.895 0.151 0.212 0.282 0.018 NLR 0.113 0.350 -0.244 0.042 0.030 0.803 -0.196 0.104 PLR 0.185 0.125 -0.252 0.036 0.584 <0.001 -0.202 0.093 Neutrophil r: Clinical significance, PLT: Platelet, MPV: Mean platelet volume, PCT: Plateletcrit, PDW: Platelet distribution width, WBC: White blood cell, TLC: Total lymphocyte count, ART: Antiretroviral therapy, NLR: Neutrophil to lymphocyte ratio, PLR: Platelet to lymphocyte ratio, p#0.05 is considered statistically significant DISCUSSION no statistically significant difference was found in terms of platelet count between the group with vitamin D deficiency and vitamin D insufficiency. Continuous inflammation in vitamin D deficiency triggers the release of pro-inflammatory cytokines (TNF-α and IL-6), leading to oxidative stress and megakaryopoiesis, which results in the production of larger sized platelets9,11. The current study found no statistically significant difference in terms of MPV between the groups with 25(OH)D levels of <21 and $21 ng mLG1 and between the groups with vitamin D deficiency, sufficiency and insufficiency. A contrasting result was reported in a study by Erkus et al.9 where a statistically significant difference was found in terms of MPV between the group with vitamin D deficiency and the group with normal vitamin D level (p<0.001). The study stated that MPV value (>6.22 fL) may be used to predict vitamin D deficiency (AUC 0.77). A study by Korzonek-Szlacheta et al.13 on patients with stable coronary artery disease found that MPV value was highest in the group with 25(OH)D <10 ng mLG1 and lowest in the group with 25(OH)D 20-30 ng mLG1. It was stated that MPV (>10.5 fL) had moderate ability (AUC 0.70) to predict vitamin D deficiency and the level of 25(OH)D (#15.5 ng mLG1) had low ability to (AUC 0.65) predict an increase in MPV. A study by Park et al.11 on general population reported that MPV was significantly higher in the group with vitamin D deficiency compared to the groups with vitamin D sufficiency and insufficiency. Another study by Cure et al.16 found that MPV was significantly lower in the groups with vitamin D deficiency and insufficiency and MPV was negatively correlated with vitamin D level. There was a significant difference in terms of PCT between the groups with 25(OH)D level of <21 ng mLG1 and 25(OH)D $21 ng mLG1 in the current study. Esen et al.6 conducted a regression analysis and reported that PCT was Vitamin D has anti-inflammatory properties due to a direct connection between vitamin D and antimicrobial peptide hepcidin which regulates systemic iron concentration. In addition, pro-inflammatory cytokines may disrupt erythropoiesis through inhibition of erythropoietin production as well as through the disruption of erythroid progenitor cell differentiation and proliferation18. The current study did not find significant difference in terms of hemoglobin concentration across the study groups. This is similar to a study by Ernst et al.19 which reported no statistically significant difference in terms of hemoglobin concentration before and after supplementation of 2800 IU cholecalciferol for 8 weeks in patients with hypertension and vitamin D deficiency. However, a study by Kim et al.20 on patients with End-Stage Renal Disease reported a significant positive correlation ($ = 0.292; p = <0.001) between hemoglobin concentration and serum level of 25(OH)D. They also found that the risk of anemia was higher in patients with 25(OH)D level of <10 ng dLG1 compared to patients with 25(OH)D level of $10 ng dLG1. A decrease in platelet count in HIV infection may be due to autoimmune destruction as a result of direct infection to megakaryocytes, as well as coagulopathy consumption in AIDS8. Vitamin D deficiency may also trigger platelet reactivity13,16. A study by Park et al.11 on general population found that platelet counts were higher in the group with vitamin D deficiency compared to groups with vitamin D sufficiency and insufficiency. A similar result was found in the current study, where platelet count was significantly higher in the group with 25(OH)D level of <21 ng mLG1 compared to the group with 25(OH)D level of $21 ng mLG1. In post-hoc analysis, 222 Pak. J. Nutr., 19 (5): 217-224, 2020 most significantly correlated with vitamin D level in patients with End-Stage Renal Disease (ESRD) receiving Renal Replacement Therapy (RRT). Al-Nimer and Salih12 reported that vitamin D3 supplementation in women with iron deficiency anemia may significantly reduce platelet count, plateletcrit and MPV by 7.4, 18.1 and 11.5%, respectively. The current study found a significant difference in terms of PDW between the groups with vitamin D insufficiency and sufficiency, after post-hoc analysis. MPV/PLT was significantly different between the study groups with 25(OH)D level of <21 and $21 ng mLG1. A similar result was observed when comparing the groups with vitamin D insufficiency and deficiency during post-hoc analysis. In a study by Korzonek-Szlacheta et al.13 on patients with stable coronary artery disease, PDW was found to be highest in the groups with vitamin D deficiency and moderate vitamin D deficiency. MPV/PLT ratio was highest in the groups with vitamin D deficiency and moderate vitamin D deficiency. In contrast, CoÕkun and Ôahin14. in their study on healthy children aged 0-18 years reported that there was no correlation between 25(OH)D level and platelet index (platelet, PCT, PDW and MPV). The current study found no significant difference in terms of NLR and PLR between the groups with 25(OH)D level of <21 ng mLG1 and 25(OH)D $21 ng mLG1. However, Akbas et al.21 in their study reported a significant difference in terms of PLR and NLR between the group with vitamin D level of <20 and $20 ng mLG1. According to a study by Erkus et al.9 NLR (>1.69) was found to be able to predict vitamin D deficiency (AUC 0.72). In addition, vitamin D3 supplementation may significantly lower PLR in the group of women with iron deficiency anemia receiving ferrous sulfate and vitamin D3 supplementation compared to the group receiving ferrous sulfate only12. The current study has several limitations. First, this is a cross-sectional study and therefore unable to determine the cause of lack of vitamin D. Second, the study disregards coinfections in the study groups. Third, the sample size was not adequate to illustrate correlation. Fourth, most of the subjects were below 40 years of age. patients consuming EFV-based ART. Further studies with a larger sample size are needed to generate a more precise conclusion. ACKNOWLEDGMENT We would like to thank Tambar Kembaren, Franciscus Ginting, Restuti Hidayani Saragih and Lenni Evalina Sihotang from Pusyansus VCT Clinic RSUP Haji Adam Malik Medan for their contribution in the initial survey, data collection and feedback provided throughout the study. We would also like to thank Dinas Kesehatan Kota Medan for their help in the initial survey. This study was funded by research grants from Direktorat Riset dan Pengabdian Masyarakat (DRPM) Kementerian Riset, Teknologi, dan Pendidikan Tinggi (KEMENRISTEKDIKTI) [Reference No: 10/UN5.2.3.1/PPM/KP-DRPM/2019]. REFERENCES 1. Musarurwa, C., L.S. Zijenah, D.Z. Mhandire, T. Bandason and K. 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