Vitamin D and Its Association with Tumor Necrosis Factor-alpha

Level in Active Tuberculosis Patients in North Sumatera, Indonesia

Dina Keumala Sari

, Ridha Dharmajaya

, Mutiara Indah Sari

and Dewi Masyithah

Nutrition Department, Faculty of Medicine, Universitas Sumatera Utara, Medan, Indonesia

Neurosurgery Department, Faculty of Medicine, Universitas Sumatera Utara, Medan, Indonesia

Biochemistry Department, Faculty of Medicine, Universitas Sumatera Utara, Medan, Indonesia

Parasitology Department, Faculty of Medicine, Universitas Sumatera Utara, Medan, Indonesia

Keywords: Vitamin D, Tumor, Tuberculosis

Abstract: Background: Mycobacterium tuberculosis is one of bacteria that caused tuberculosis disease. This disease

related to macro and micronutrient deficiency, also to inflammation process. One of deficiency micronutrient

is vitamin D deficiency, host cell response in including deficiency vitamin D, this would affect tumor necrosis

factor (TNF)-alpha level. Objective: We sought to determine whether there is an association between vitamin

D status and TNF-alpha status in active tuberculosis patients. Design: We conducted a cross-sectional study

of 25 man and women aged 18-60 years with active tuberculosis in North Sumatera, Indonesia. Parameters

were 25(OH)D, TNF-alpha serum level, biomolecular parameters such as albumin and hs-CRP also assessed.

The association was analysis using chi-square or fischer test. Results: the mean of study subjects age were

37.2±14.9 years old, there were 81.2% subjects categorized into vitamin D deficiency-insufficiency and

18.8% categorized into vitamin D sufficiency. All of the subjects had high TNF alpha level, There is an

association between vitamin D and TNF alpha serum level. Conclusions: based on this result, there was an

association between vitamin D and TNF-alpha serum level inactive tuberculosis.

1 INTRODUCTION

Tuberculosis had become a global health problem in

developed countries, including Indonesia, which have

economic and social problem. Previous research re-

ported higher macro and micronutrient deficiency, in-

cluding vitamin D deficiency in normal and active tu-

berculosis patient who lived in abundant sunlight ex-

posure (Sari et al., 2017a, 2017b). Vitamin D defi-

ciency related to higher inflammatory effect, host cell

respons with tuberculosis infecton showed higher

proinflammatory agent including TNF alpha, inter-

leukin (IL)-1, IL-2, IL-8, interferon (IFN) gamma,

and granulocyte-macrophage colony stimulating fac-

tor. The association between vitamin D deficiency

and TNF alpha in tuberculosis remain poorly under-

stood. Previous research focused on TNF alpha, be-

cause of higher inflammatory effect of this cytokine.

One of the theories is vitamin D would inhibit the

growth of M. tuberculosis in the macrophages

(Crowle et al., 1987; Rook et al., 1986). Vitamin D

also promote anti-M. tuberculosis responses through

many mechanisms in the macrophage. The mediators

that involved in the macrophage were nitrit oxide

(NO) (Rockett et al., 1998), NADPH oxidase (Sly et

al., 2003; Sly et al., 2001), cathelicidin (P. T. Liu et

al., 2006; Philip T et al., 2007; Yuk et al., 2009), and

autophagy (Yuk et al., 2009) mechanisms. The vita-

min D action that would inhibit progression of tuber-

culosis is impaired M. tuberculosis activity in the

macrophage and inhibit proinflammatory signalling,

including TNF alpha signalling.

The aims of this study were to determine whether

there is an association between vitamin D status and

TNF-alpha status in active tuberculosis patients in

Medan, North Sumatera, Indonesia.

2 METHOD

We conducted a cross-sectional study of 25 man and

women aged 18-60 years in North Sumatera, Indone-

sia April to July 2018, during the dry season (dry sea-

son in Indonesia is between April and October, higher

sunlight exposure, lower rainy season). The location

of recruitment was in Sumatera Island (Medan, North

856

Sari, D., Dharmajaya, R., Sari, M. and Masyithah, D.

Vitamin D and Its Association with Tumor Necrosis Factor-alpha Level in Active Tuberculosis Patients in Nor th Sumatera, Indonesia.

DOI: 10.5220/0010095408560860

In Proceedings of the International Conference of Science, Technology, Engineering, Environmental and Ramiﬁcation Researches (ICOSTEERR 2018) - Research in Industry 4.0, pages

856-860

ISBN: 978-989-758-449-7

Sumatera, Indonesia) with latitude: 3.67 N and longi-

tude 97.55 E, average temperature: ±32

C (90

F).

This study was carried out after ethical approval was

obtained from the Health Research Ethics Committee

of Sumatera Utara University Medical School (No.

96/TGL/KEPK FK USU-RSUP HAM/2018) and all

participants were given written informed consent to

the study procedures. and all participants were given

written informed consent to the study procedures.

2.1 Study Participants

The subjects of this study consisted tuberculosis

patients in two community health centers with the

higher tuberculosis prevalence in Medan, North

Sumatera, Indonesia. Two community health centers

were in Puskesmas Teladan and Puskesmas Amplas.

There were man and women with various occupa-

tions, and taken purposively, there were 25 subjects

that include in this study. The inclusion criteria were

active tuberculosis patients within the range of 18-60

years old. Exclusion criteria were subjects with his-

tory of diabetes mellitus, myocardial infarction, renal

or liver dysfunction. In addition to those exclusion

criterias, subjects who were HIV, pregnant and lactat-

ing were also excluded.

2.2 Laboratory Analysis

We measured 25(OH)D serum concentration by

chemiluminescent immunoassay (CLIA) technology

(Diasorin, Stillwater, MN), measures were between

4.0 and 150 ng/mL. The lowest value was 4.0 ng/mL

which is based on an inter-assay precision 3.90% CV.

Reference range were <20 ng/mL categorized defi-

ciency, 20-30ng/mL (insuficiency), 30-100 ng/mL

(suficiency) (Sharma & Meena, 2017). To convert

ng/mL to nmol/L is multiply with 2.496. We meas-

ured albumin serum using architect C800 with color-

imetry method and to measured hs-CRP, we also use

architect C800 with different method which are turbi-

dimetry/immunoturbidimetry. Tumor necrosis factor

alpha analysis using ELISA (Enzyme-Linked Im-

munosorbent Assay) using Quantikine ELISA Hu-

man TNF alpha, R&D, USA.

2.3 Statistical Analysis

Continuous variables were expressed as continuous

variables as means±SDs. Categorical variables were

expressed as percentage proportions, using chi-square

to expressed significancy difference between two

groups, and Fischer test if the data did not met the

criterias. The p values <0.05 were considered statisti-

cally significant. We used SPSS program (version

11.5; SPSS Inc, Chicago, IL) to perform the analysis.

3 RESULTS AND DISCUSSIONS

The results will be discussed in 4 subsections, they

are clinical characteristic of study participants, vita-

min D and biomolecular parameters, and tumor ne-

crosis factor alpha serum level and its association.

3.1 Clinical Characteristic of Study

Participants

The aim of this study was todetermine whether

there is association between 25(OH)D with TNF al-

pha serum in active tuberculosis patients in tropical

country. In this study showed that the higher preva-

lence of age classification was in 31-40 years (64%,

mean±SD: 35.3±5.9 years), most of the subjects were

male (80%), and based on BCG scar, the subjects

were had clear BCG scar (Table 1). Based on this re-

sult, BCG scar seemed had no effect on tuberculosis

incidence, probably because of long state of malnutri-

tion or unsuccessful immunization. Protein and fat

had influence in nutrition status in tuberculosis pa-

tient. Low protein intake would lower immunity cell

activity, this would affect Mycobacterium tuberculo-

sis infection in malnutrition patients.

Table 1. Clinical characteristics

Variables

n(%)

Distribution by severity class/TB score

Class I

Class II

Class III

25 (100)

BCG scar

Clear

No scar

Dubious

17 (68)

3 (12)

5 (20)

Abbreviations:

SD: Standart Deviation

BCG: Bacille Calmette-Guérin

Vitamin D and Its Association with Tumor Necrosis Factor-alpha Level in Active Tuberculosis Patients in North Sumatera, Indonesia

857

3.2 Vitamin D and Biomolecular

Parameters

Based on this study, most of the active tuberculosis

patients had deficiency-insufficiency vitamin D

(90.2%), the effects of vitamin D to tuberculosis pro-

gression through vitamin D receptor (VDR). This

VDR activity, which is the signaling would induces a

series of antimicrobial responses such as induction of

autophagy, phagolysosomal fusion, release, and acti-

vation of the antimicrobial peptide cathelicidin, and

killing of intracellular Mycobacterium tuberculosis

(Gibney et al., 2008; Yoon et al., 2013). There also

many research about VDR gene, especially polymor-

phism of VDR gene which were TaqI and FokI. The

patient with polymorphism would resulting lower vit-

amin D serum level and higher infection of Mycobac-

terium tuberculosis. Previous studies also report

about polymorphism of TNF alpha polymorphism,

that would resulting ineffective of TNF alpha activity.

Low albumin serum showed lower immunity cell ac-

tivity, interestingly in this study showed most of the

subject had normal albumin serum level, but yet still

had active inflammation. This would need another ex-

planation to this phenomenon.

Table 2. Serum levels of 25-hydroxyvitamin D and biomol-

eculer parameters

Parameters Mean±SD;

n(%)

25-hydroxyvitamin D serum levels

(ng/mL)#

21.8±7.6

Vitamin D status, n (%)

Deficiency-insufficiency

Sufficiency

22 (88)

3 (22)

Albumin (g/dL)

Low

Normal

High

3.8±0.8

6(24)

19(76)

hs-CRP (mg/dL)

Low inflammation

Active inflammation

49.1±15.7

25(100)

Continues variable: mean ± SD; categorical variable: n (%);

SD=standard deviation

3.3 Tumor Necrosis Factor-alpha

Serum Level

Tumor Necrosis Factor-alpha (TNF-alpha) is a

17.5kDa, 157 amino acid protien that is a potent lym-

phoid factor, which produced cytotoxic effects on a

wide range of tumor cells and other target cells. In the

cell, TNF alpha is a pro-inflammatory and the pri-

mary mediator of immune regulation. The biosynthe-

sis of TNF alpha is a major secreted factor in activated

infected cells besides other proinflammatory cyto-

kines (P. D. Davies, Brown, & Woodhead, 1985; P.

D. O. Davies, 1985; Wilkinson et al., 2000). In tuber-

culosis infection, human macrophages exposed to M.

tuberculosis demonstrated a robust release of TNF-al-

pha degradation, including other inflammation pa-

rameter such as NF-kappa-B nuclear translocation

and other interleukin (P. D. Davies et al., 1985; P. D.

O. Davies, 1985).

In this study showed as presented in Table 3,

higher TNF alpha serum level, this probably because

of status of the study subjects. All of the study subject

were new cases and active tuberculosis. They also had

lower nutrition status, macro and micronutrient defi-

ciency including lower protein status and vitamin D

deficiency.

Table 3. TNF alpha serum levels

Parameters Mean±SD; n

(%)

Tumor necrosis factor alpha serum

levels (pg/mL)#

167.15±45.5

TNF alpha categorized, n (%)

Low

Normal

High

25 (100)

Continues variable: mean ± SD; categorical variable: n

(%);

SD=standard deviation

3.4 Association between Vitamin D and

TNF-alpha

There was an association between vitamin D status

and TNF alpha status in active tuberculosis patient,

this study reported that when there was a vitamin D

deficiency and insufficiency, it was an association

with higher TNF alpha serum level (Table 4). Other

previous results showed there was an association also

between vitamin D level and TNF alpha serum level

(Anandaiah et al., 2013).

ICOSTEERR 2018 - International Conference of Science, Technology, Engineering, Environmental and Ramiﬁcation Researches

858

Table 4. The analysis of vitamin D and TNF alpha status

TNF alpha

status, n(%)

Nor-

mal

High

Vitamin D

status,

n(%)

Deficiency-

insuficiency

(88)

(100)

0.02

Sufficiency 3 (22) 0

Total 5

(100)

Using Fischer test, * significant

Limitation of this study was we have not yet get

the information about genetic polymorphism of vita-

min D receptor gene and TNF alpha gene. We also

had a limitation in the lower sample size.

4 CONCLUSIONS

Based on this result, there was an association between

vitamin D and TNF-alpha serum level in active tuber-

culosis.

ACKNOWLEDGEMENTS

The authors gratefully acknowledge that the present

research is supported by Ministry of Research and

Technology and Higher Education Republic of Indo-

nesia, Research and Community Service, Universitas

Sumatera Utara. The support is under the research

grant DRPM of Year 2018 Contract Number

263/UN5.2.3.1/PPM/KP-DRPM/2018.

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