Role of Vitamin D in Systemic Lupus Erythematosus Disease Activity

Document Type : Original Article

Authors

1 Department of Clinical Pathology, Faculty of Medicine, Al-Azhar University

2 Al-Azhar University, Faculty of medicine, Clinical Pathology Department, Cairo, Egypt

3 Department of Rheumatology, Rehabilitation and Physical Medicine, Faculty of Medicine, Al-Azhar University

Abstract

Background:Systemic Lupus erythematous is systemic inflammatory disorder associated with immunological abnormalities that could affect many organs. Vitamin D has inhibitory properties on activity and multiplications of immune system cells associated with inflammation.
Objective:To evaluate vitamin D level in systemic lupus erythematosus cases, and its relation to activity of the disease.
Material and Methods:The study was done on 100 individuals in clinical pathology and rheumatology, rehabilitation and physical medicine departments at Al-Azhar University Hospitals. Subjects classified into 2 groups. Group I included 70 systemic lupus erythematosus patients either in active or inactive state. Group II comprised 30 matched healthy subjects as regard age and sex. All participants were subjected to complete rheumatological examination and recommended laboratory investigations.
Results:Vitamin D was significant low in systemic lupus erythematosus patients versus healthy control. Serum vitamin D was statistically lower in patients with active disease versus inactive disease. Significant correlation was found between vitamin D and activity index of the disease and laboratory markers of disease activity.
Conclusions:Vitamin D inadequacy is a finding among systemic lupus erythematosus patients in active disease state. Vitamin D level could be a biomarker of disease activity.

Keywords

Full Text

INTRODUCTION

Systemic lupus erythematosus is an autoimmune disease. Many environmental factors and many genes are involved in the pathogenesis of the disease. The disease is known to be associated with autoantibody production, dysfunction in of immune system and variable degree of inflammation that could affect more than one organ 1.

Environmental factors are assumed to share in clinical expression of the systemic lupus erythematosus diseases. Vitamin D level has been studied as one of the environmental factors. Vitamin D has an important role in various processes of the immune system, and its receptors  were found on immune cells; macrophage, dentritic cells and lymphocytes. Immune cells are able to synthesize the active vitamin D metabolite. Vitamin D has the capability to  modulate  innate and adaptive immune responses 2.

Vitamin D deficiency is prevalent in SLE patients due to photosensitivity,  renal impairment and the use of different drugs in treatment of the disease such as glucocorticoids   which alter the metabolism and functions of vitamin D. 3

 

 

PATIENTS AND METHODS

This work was done in Clinical Pathology and Rheumatology Departments at Al- AZHAR university hospitals (Al-Hussein and Bab El-Sharia).  The study was conducted on 100 individuals. Subjects enrolled in the study were categorized into two groups (Patient and Control groups).  

 Patient group (Group I): includes 70 SLE patients. They were 62 females and 8 males. Their ages were 29.3±5.4 years. Their disease duration was from 2 - 10 years.   Systemic lupus international collaborating clinics (SLICC) data were used to diagnose SLE disease 4. Assessment of SLE activity was determined using the modified SLE disease activity index (SLEDAI). 5

Exclusion criteria: Patients with other autoimmune diseases, chronic inflammatory diseases, systemic diseases (endocrine, cardiac, renal, hepatic, malignant) or under calcium or vitamin D therapy were excluded from the study.

The patient group was categorized retrospectively into: Active SLE patients, it comprised 44 SLE patients. They were 40 females and 4 males and inactive SLE patients ,it comprised 26 SLE patients. They were 22 females and 4 males.

Group II (Control group): 30   healthy  individuals( matching cases as regard age and sex) were enrolled in the study  as a control group.

Subjects involved in the study were informed orally about the procedures and  gave  written consent to share. The participants were subjected to thorough clinical examination and laboratory investigations.

 The following laboratory investigations were done for all participants: Complete blood picture,  erythrocyte sedimentation rate , C-reactive protein  on automated spectrophotometry Au 480 system, USA, Complement C3 and C4 level , Anti-nuclear antibody (ANA) by immunofluorescence  (IIF) technique,  and by enzyme immunosorbent  assay (ELISA) ,Anti- DNA  by IIF technique on Crithedia Lucilia substrate, DiaSorin, Italy and by ELISA technique using Kit Invoa Diagnostics USA. Serum calcium (Ca), creatinine and urinary  proteins of 24 hours by colorimetric assay using kits from Spinreact, Spain. Serum 25-hydroxychloecalciferol vitamin D 25(OH)D by ELISA technique Abcam, USA

Statistical Analysis:

Recorded data were analyzed using the statistical package of social science (SPSS) version 24 . Quantitative data expressed as Median (IQR) as the data was abnormally distributed. Qualitative data expressed as frequency and percentage. To compare between two means (for abnormal distributed data) Mann- Whitney test was used.  To compare between non-parametric data Chi-square test was used. Kruskal Willis test (KW): when comparing more than two means (for abnormal distributed data).  For correlating data Pearson's correlation coefficient test was used. Probability (P-value):  P-value > 0.05 was considered insignificant, P- value < 0.05 was considered significant and P-value < 0.001 was considered highly significant.

RESULTS

 

Groups

Parameters 

Cases

Control

MW

P-value

Hemoglobin (g/dl)

Median

10

11.9

0.0

< 0.001

IQR

9 - 10

11.8 - 12.7

PLTs (x10³/ul)

Median

180

205

602.5

0.001

IQR

160 - 203

179.8 – 230

24 h urine protein (mg/24h)

Median

68

35.5

315

< 0.001

IQR

50 - 835

21.5 - 70.3

Creatinine (mg %)

Median

0.80

0.6

571.5

< 0.001

IQR

0.60 - 0.90

0.5 - 0.7

Ca (mg/dl)

Median

8.45

8.7

651

0.003

IQR

8 - 8.9

8.5 - 9.02

ESR (mm/h)

Median

39

7

0.0

< 0.001

IQR

28 - 47

6 - 9

ANA titer (U/ml)

Median

80

9.5

10.5

< 0.001

IQR

68 - 100

7 - 15.3

Anti-DNA titer (U/ml)

Median

340.5

59.5

0.0

< 0.001

IQR

232.8 - 415

35.3 - 70.3

C3 (mg/dl)

Median

110

158

32

< 0.001

IQR

100 - 131.3

149 - 168

C4 (mg/dl)

Median

16

38.5

0.0

< 0.001

IQR

10 - 20

36 - 41.3

 

MW: Mann Whitney U test.                                                                                   

Table 1: Comparison between patient and control group as regard studied laboratory data.

 

There was statistical significant decrease in platelet count and serum calcium (ca) in cases group compared to control group. There were highly statistical decrease in hemoglobin level, complement C3 and complement C4 in cases group versus control group. There was highly statistical increase in 24 hours total urinary proteins, serum creatinine, erythrocyte sedimentation rate ,  ANA and anti- DNA  in cases group versus control group.

 

Groups

Vitamin D 

Cases

(N = 70)

Control

(N = 30)

Stat. test

P-value

Vitamin D (ng/ml)

Median

9

36

MW = 80.5

< 0.001 HS

IQR

7 - 16

25.8 - 46.5

Vitamin D Status

Sufficient

0

0%

17

56.7%

X2 = 53.9

< 0.001 HS

Insufficient

14

20%

8

26.7%

Deficient

56

80%

5

16.7%


Table 2: Comparison between patients and control group as regard vitamin D.


There was highly statistical significant decreased vitamin D in cases group versus control cases.

 

 

 


Groups

Parameters 

Inactive

(N = 26)

Active

(N = 44)

test

P-value

Hemoglobin (gm % )

Median

10.10

9.75

MW =406

0.043 S

IQR

9.30 - 10.8

9 - 10.3

24 h urine protein (mg/24 h)

Median

52.5

760

MW =214

< 0.001 HS

IQR

39 - 67

60.5 - 995

Creatinine (mg %)

Median

0.7

0.90

MW =317.5

0.002 S

IQR

0.5 - 0.8

0.70 – 1.0

ESR (mm/h)

Median

27

45

MW =3.5

< 0.001 HS

IQR

23 - 30

40 - 52

ANA titer (U/ml)

Median

66.5

97

MW = 61

< 0.001 HS

IQR

37.5 - 69.3

80.3 - 107.3

ANA IIF

Negative

7

26.9%

0

0%

X2 = 23.8

< 0.001 HS

1/40

19

73.1%

25

56.8%

1/80

0

0%

19

43.2%

Anti-DNA titer (U/ml)

Median

205

410

MW = 0.0

< 0.001 HS

IQR

150 - 239.3

350 - 555.5

Anti-DNA IIF

1/40

26

100%

8

18.2%

X2 = 43.8

< 0.001 HS

1/80

0

0%

19

43.2%

1/160

0

0%

17

38.6%

C3 (mg/dl)

Median

135.5

100

MW = 0.0

< 0.001 HS

IQR

130 - 140

88.5 - 109.8

C4 (mg/dl)

Median

20.5

12

MW = 32.5

< 0.001 HS

IQR

20 - 26

9 - 15


Table 3: Comparison between active versus inactive SLE patients as regard studied laboratory data.


There was statistical significant decrease in hemoglobin level and statistical significant increase in serum creatinine level in active SLE patients versus inactive SLE patients. There was highly statistical significant increase in 24 hours total urinary proteins, ESR, ANA done by ELISA and done by IIF, anti-DNA done by ELISA and done by IIF technique in active SLE cases versus inactive SLE cases. There were highly statistical significant decrease in serum level of complement C3 (C3) and complement C4 (C4) in active SLE versus inactive SLE group .

 

Groups

Vitamin D

Activity

Stat. test

P-value

Inactive

(N = 26)

Active

(N = 44)

Vitamin D (ng/ml)

Median

17

7

MW = 0.0

< 0.001 HS

IQR

14.8 - 23

7 - 9

Vitamin D Status

Insufficient

14

53.8%

0

0%

X2 = 29.6

< 0.001 HS

Deficient

12

46.2%

44

100%


Table 4: Relation between vitamin D and activity in cases group.


There was highly statistical significant decreased vitamin D level in active SLE patients versus inactive SLE patients.

 

Variables

Vitamin D

r

p-value

Duration of disease

- 0.37

0.001

DAI

- 0.915

< 0.001

PLT

0.319

0.007

24 hour urine proteins

- 0.573

< 0.001

Creatinine

- 0.415

< 0.001

Erythrocyte sedimentation rate

- 0.724

< 0.001

ANA

- 0.734

< 0.001

Anti ds-DNA Titer

- 0.778

< 0.001

Complement C3

0.752

< 0.001

Complement C4

0.752

< 0.001


Table 5:  Vitamin D correlations with DAI and other data in cases group.

 

There was highly statistical negative correlation between vitamin D and index of disease activity (DAI), creatinine, ESR, ANA titer, anti-DNA titer and 24 hour urine protein. There was significant correlation between vitamin D level and disease duration. There was statistically positive correlation between vitamin D and PLTs. There were statistical correlation between vitamin D and C3 & C4.

 

DISCUSSION

Measurement of Systemic lupus Erythematosus disease activity is central to evaluate outcomes, differentiate between SLE patients, response to therapy and to improve morbidity and mortality rate of SLE disease.  Fluctuation levels of disease activity are present among systemic lupus erythematosus patients. As there is no standard laboratory marker that properly reflect disease activity many composite clinical indices have been developed for the evaluation of disease activity 6,7.

25 hydroxycholecalciferol (25 (OH) vitamin D is the main circulating form of vitamin D.  25 (OH) vitamin D reflects vitamin D supply to the body from skin synthesis and nutritional intake. It has been found that many immune cells express both vitamin D receptor and 1α, hydroxylase needed for synthesis of the active form of vitamin D  with potential of autocrine or paracrine effect in additional to endocrine effect. Vitamin D has inhibitory properties on cell proliferation, anti-inflammatory and immunomodulation. Vitamin D inadequacy could direct the immune system to a loss of tolerance. Supplementation of vitamin D could be of great value in patients with lupus 3,8.

The target of this study was to evaluate vitamin D level in Egyptian patients with SLE disease and to  explore if it could be used as a biomarker reflecting the degree of disease activity.

Vitamin D inadequacy could be present in healthy populations. In our study 43.4% of control group showed inadequate vitamin D levels [16.7% deficient and 26.7% insufficient]. This finding is going with Squance and coworkers. 9 and Khairallah and coworkers. 10.

In our work, in SLE group, 80% showed deficient level, 20% showed insufficient level and 0% showed normal vitamin D level. There was statistical significant decrease in vitamin D in SLE group versus control group. This finding despite sunny days are present most of the year in Egypt. This is going with Korah and coworkers 11, Elsaid and coworkers 12 and Khairallah and coworkers. 10. Other authors reported 25(OH) vitamin D inadequacy in up to 90% of SLE cases in Saudi Arabia; 82% Norway 71% Poland. However much lower rates have been reported in other studies 18% Canada, 27%. Hong Kong 20% United Status and 15% Spain 13. They explained lower levels and high prevalence of vitamin D inadequacy as most patients are females with low outdoor activities or due to clothing, lower body surface area and hormonal effects. Other contributing factors could be lupus nephritis due to decrease 1 α hydroxylase activity, usage of drugs as glucocorticoids, hydroxychloroquire and anticonvulsants that enhance vitamin D catabolism and the presence of vitamin D antibodies that probably enhance vitamin D clearance.

In our study, vitamin D in active SLE group was statistically low versus inactive SLE group. There were statistical correlation between vitamin D and activity index of the disease . These findings going with Yap and coworkers. 14, Zheng and coworkers. 15 and Khairallah and coworkers 10. So vitamin D inadequacy could predict SLE disease activity. This finding is expected due to the immunosuppressive and the anti-inflammatory properties of vitamin D. The inflammatory process in SLE enhance vitamin D catabolism [Bidirectional relationship]. This finding contradictory to other studies 16.The conflicting results could be due to diverse study populations, the retrospective nature of the study, methodological variations, statistical power differences between studies and heterogeneity of treatment.

Anti-ds-DNA antibody is specific for SLE and its level fluctuates with disease activity. In this study, DNA antibodies were statistically high in active SLE group versus inactive SLE group. There were highly statistical correlation between vitamin D levels and anti-ds-DNA levels. This finding going with Mok and coworkers. 17, Attar and Siddiqui 13 and Nerviani et al.18, but was in disagreement with Khairallah and coworkers. 10.

In our study,  both complement C3 and complement C4 were statistically low in active SLE group versus inactive SLE group. There were highly statistical   correlation between vitamin D levels and C3 and C4 levels. These findings are in agreement with study done by Giles and Boackle 19, but contradictory to studies done by Attar and Siddiqui13 who found correlation between vitamin D and C4 only. They explained this finding by the fact that in SLE the main triggering of complement activity is mediated mainly by the classical pathway. In addition our findings were contradictory to study done by Khairallah et al.10. These conflicts between studies could be due to variations in the compensatory effect of the liver between populations, variation in DAI, heterogeneity of drugs used for treatment of SLE and difference in method of assay.

In our study, ANA test was positive in 73.1% in inactive SLE group but was 100% positive in active SLE group. The level of ANA was statistically high in active SLE group versus inactive SLE group. There were statistical correlation between vitamin D level and ANA levels. On contrary Squance et al. 9 did not find any correlation between vitamin D and ANA levels. They stated that 2 steps are needed to develop SLE overt disease. The first step is the conversion from tolerance to benign [serological] autoimmunity. Vitamin D levels have impact upon this step. The second step is the conversion from serological SLE to overt SLE disease. Other factors [environmental or hormone] could trigger this step 2. So vitamin D deficiency may predispose to disease expression without influencing its severity or behavior once established.

In our study ESR levels were statistically high in SLE group versus control group. ESR was found  significantly high in active SLE group versus inactive SLE group. There were significant correlation between vitamin D levels and ESR levels. These findings are going with studies done by Miskovic and coworkers. 20 who explained this fact due to the effect of cytokines in the induction of acute phase reactant.

CONCLUSION

Vitamin D is statistically decreased in SLE patients versus healthy individuals and showed more significant decrease in active SLE versus inactive SLE patients. So vitamin D level could predict the activity state of the disease in patients with systemic lupus erythematosus.

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