A study of the Role of 25 Hydroxy-Cholecalciferol Level on Non Alcoholic Fatty Liver Disease ( NAFLD ) in a Cohort of Egyptian Patients Elsayed

Patients and Methods: We studied 78 consecutive patients with biopsy-proven NAFLD. Patients were divided into 2 groups according to serum level of 25 (OH) D; group I; have deficient 25 (OH) vitamin D (<50nmo/L) and group II; have sufficient 25 (OH) vitamin D (50-70 nmol/L). Liver injury profile (ALT, AST), lipid profile (LDL, HDL and triglycerides), inflammatory marker (CRP) as well as histopathological assessment according to NAS scoring were evaluated at baseline. Vitamin D supplementation for 24 weeks was given for both populations with follow up evaluation of laboratory parameters at the end of the study.


INTRODUCTION
Non-alcoholic fatty liver disease (NAFLD) has become the most common chronic liver disease worldwide [1], with an estimated prevalence of 20-30%in general population and 75% in patients with type 2 DM, obesity and metabolic syndrome [2,3].NAFLD is a pathological condition characterized by aberrant triglycerides accumulating in the hepatocytes, in some cases accompanied by necro-inflammatory activity and fibrosis (steatohepatitis) and potentially evolving into liver cirrhosis [4].However, beside lifestyle intervention, no established therapy of NAFLD has been identified yet [5].Non-alcoholic fatty liver disease is the hepatic component of metabolic syndrome where the insulin resistance increases non esterified fatty acids release from adipose tissue and favor their deposition into hepatocytes [6].
Low vitamin D levels have been associated with the histological severity of NAFLD/NASH [7].Overall So, the aim of the present study was to evaluate the effect of vitamin D supplementation on specific sub-populations of patients with fatty liver not studied in the previous trials, namely NAFLD patients with moderate to severe liver disease.

PATIENTS AND METHODS
Study population was recruited among patients referred to hepatology clinic of Zagazig university hospitals, Egypt.Between December 2016 and June 2017, 78 patients proved to have NAFLD were divided according to 25 (OH) cholecalciferol serum level into 2 groups: Group I: patients have deficient 25 (OH) cholecalciferol level (<50 nmol/L).Group II: patients have sufficient but sub-optimal 25 (OH) cholecalciferol level (50-75 nmol/L).
To be eligible for the study, patients had to satisfy the following criteria: fatty liver detected by abdominal ultrasonography (US) and confirmed by histopathology, ALT is more than AST, negative tests for HBsAg and HCV Abs.

Exclusion criteria include :
History of alcohol abuse (as defined by an average daily consumption of alcohol >30 g/day in men and >20 g/day in women), cirrhosis, autoimmune hepatitis and other causes of liver disease (hemochromatosis, Wilson's disease), hyper/ hypoparathyroidism, known hypersensitivity to cholecalciferol, hypercalcemia, hypercalciuria, nephrolithiasis, nephrocalcinosis.
All patients were subjected to: 1-Thorough history taking with full clinical examination and all medications were carefully recorded at baseline visits and drug alterations regarding anti-diabetic agents, anti-hypertensive treatments.Statins were not allowed throughout the study.

2-
The body mass index (BMI) was calculated as weight in kilograms divided by the square of the height in meters.

3-Serum 25 (OH)-cholecalciferol was measured as an indicator of vitamin D status [16].
4-Liver ultrasound scanning was performed to assess the presence of hepatic steatosis.
6-Liver biopsy for all patients at base line, to confirm the diagnosis and for histological grading of NAFLD, using Kleiner NAFLD activity score (NAS).This score is the sum of three histological components; steatosis (0-3), lobular inflammation (0-3) and ballooning degeneration (0-2

Ethical Considerations
Since the patients have elevated liver enzymes with no obvious cause, some patients had family history of liver diseases and liver malignancy and imaging evident of focal fat and focal sparing, liver biopsies were done to detect the underlying cause and written consent was obtained from all patients before the study.
The study protocol was reviewed and approved by the Institutional Review Board of the faculty of Medicine, Zagazig University, Egypt.After 24 weeks of vitamin D supplementation, there was statistically significant improvement in LDL, HDL, triglycerides, ALT and AST in group I (Table 2), while there was non-significant change in group II (Table 2).

Out
Finally, it was noted that at baseline there was non-significant difference between both groups regarding the anti-inflammatory marker (CRP); 3.4±1.3vs. 3.4±1.3,p value 0.213.After vitamin D supplementation there was statistically significant improvement of CRP in both populations.
When comparing both groups after vitamin D supplementation, there were non-significant differences regarding hepatic indicators, lipid profile and HbA1c%, but there were significant differences in CRP (Table 3).
Moreover, we performed a correlation analysis that showed a significant association between NAFLD scores, serum 25(OH)-cholecalciferol levels, lipid profile and histological grading (Table 4).

Figure 1 :
Figure 1: vitamin D levels among both populations Furthermore, data from meta-analyses point towards the existence of an association between low circulating vitamin D levels and NAFLD [8].However, Sharifi et al. [14] and Barchetta et al. [15] reported no beneficial effect of vitamin D supplementation in NAFLD patients.
No major adverse events occurred during the study, highlighting safety of the dosing and no change in the ongoing therapies throughout the study.Regarding the cause of NAFLD, there was no statistically significant difference between both populations (data were not shown).

Table ( 1
); Baseline clinical and biochemical characteristics of both groups:

Table 2 :
Comparison of characteristics before and after study treatment in both groups Figure 2: Distribution of NASH according to activity grade.

Table 3 :
Biochemical characteristics after vitamin D supplementation in both groups

Table 4 :
[14]elation between vitamin D and different parameters before vitamin D supplementation In this study, the intervention was limited to 4 months and the dose of cholecalciferol was 50.000IU/2weeks.Moreover the disease activity was mild.These differences may explain the discrepancy with the recent study.On the other hand, although there was no statistically significant difference at baseline regarding CRP, there was significant improvement in CRP among all patients, high lightening the anti-inflammatory effect of vitamin D[21].In agreement with this result, significant improvement of the inflammatory marker (hs.CRP) has been reported after vitamin D supplementation in NAFLD patients[14].