Vitamin D levels do not predict the stage of hepatic fibrosis in patients with non-alcoholic fatty liver disease: A PRISMA compliant systematic review and meta-analysis of pooled data

Table 1 Search results of vitamin D and non-alcoholic fatty liver disease

Database/search	Search terms	Search results
EMBASE
1	("liver cirrhosis"/exp OR cirrhosis: ti, ab OR cirrhoses: ti, ab OR fibrosis: ti, ab OR fibroses: ti, ab)
2	("vitamin D"/exp OR "25 hydroxyvitamin d"/exp OR "vitamin d": ti, ab OR "ergocalciferols": ti, ab OR "ergocalciferol": ti, ab OR "25 hydroxy vitamin d": ti,ab OR "25 hydroxyvitamin d": ti, ab OR "25 hydroxy d": ti, ab OR "25(OH)D": ti, ab OR "25-hydroxyvitamin d 2": ti, ab)
3	("nonalcoholic fatty liver"/exp OR "Non-alcoholic Fatty Liver": ti, ab OR "nonalcoholic fatty liver": ti, ab OR "Non-alcoholic Fatty Livers": ti, ab OR "nonalcoholic fatty livers": ti, ab OR "NAFLD": ti, ab OR "NASH": ti, ab OR "nonalcoholic steatohepatitis": ti, ab OR "nonalcoholic steatohepatitides": ti,ab OR "fatty liver"/de OR "fatty liver": ti, ab OR "Steatohepatitis": ti, ab OR "Steatosis of Liver": ti, ab OR "Liver Steatosis": ti, ab OR "Liver Steatoses": ti, ab OR "hepatic steatosis": ti, ab OR "hepatosteatosis": ti, ab)
4	1 and 2 and 3	199
Web of science
1	("Non-alcoholic Fatty Liver" OR "nonalcoholic fatty liver" OR "Non-alcoholic Fatty Livers" OR "nonalcoholic fatty livers" OR "NAFLD" OR "NASH" OR "nonalcoholic steatohepatitis" OR "fatty liver" OR Steatohepatitis OR "Steatosis of Liver" OR "Liver Steatosis" OR "Liver Steatoses" OR "hepatic steatosis" OR "hepatosteatosis")
2	("liver cirrhosis" OR cirrhosis OR cirrhoses OR fibroses OR fibrosis)
3	("vitamin d" OR "ergocalciferols" OR "ergocalciferol" OR "25 hydroxy vitamin d" OR "25 hydroxyvitamin d" OR "25 hydroxy d" OR "25(OH)D" OR "25-hydroxyvitamin d 2")
4	1, 2 and 3	69
Cochrane
1	MeSH descriptor: [Non-alcoholic Fatty Liver Disease] explode all trees	181
2	MeSH descriptor: [Liver Cirrhosis] explode all trees	2462
3	MeSH descriptor: [Vitamin D] explode all trees	2907
4	"Non-alcoholic Fatty Liver" or "nonalcoholic fatty liver" or "Non-alcoholic Fatty Livers" or "nonalcoholic fatty livers" or "NAFLD" or "NASH" or "nonalcoholic steatohepatitis" or "fatty liver" or Steatohepatitis or "Steatosis of Liver" or "Liver Steatosis" or "Liver Steatoses" or "hepatic steatosis" or"hepatosteatosis": ti, ab, kw	1470
5	"liver cirrhosis" or cirrhosis or cirrhosis or fibrosis or fibroses: ti,ab,kw	13273
6	"vitamin d" or "ergocalciferols" or "ergocalciferol" or "25 hydroxy vitamin d" or "25 hydroxyvitamin d" or "25 hydroxy d" or "25(OH)D" or "25-hydroxyvitamin d 2": ti,ab,kw	6061
7	1 or 4	1470
8	2 or 5	13273
9	3 or 6	6722
10	7 and 8 and 9	13
PubMed
1	(("Non-alcoholic Fatty Liver Disease"[Mesh] OR "Non-alcoholic Fatty Liver"[tw] OR "nonalcoholic fatty liver"[tw] OR "Non-alcoholic Fatty Livers"[tw] OR "nonalcoholic fatty livers"[tw] OR "NAFLD"[tw] OR "NASH"[tw] OR "nonalcoholic steatohepatitis"[tw] AND "Fatty Liver"[Mesh: noexp] OR "fatty liver"[tw] OR Steatohepatitis[tw] OR "Steatosis of Liver"[tw] OR "Liver Steatosis"[tw] OR "Liver Steatoses"[tw] OR "hepatic steatosis"[tw] OR "hepatosteatosis"[tw])
2	("vitamin d"[mh] OR "vitamin d"[tw] OR "ergocalciferols"[tw] OR "ergocalciferol"[tw] OR "25 hydroxy vitamin d"[tw] OR "25 hydroxyvitamin d"[tw] OR "25 hydroxy d"[tw] OR "25(OH)D"[tw] OR "25-hydroxyvitamin d 2"[tw])
3	("liver cirrhosis"[mh] OR cirrhosis[tw] OR cirrhoses[tw] OR fibrosis[tw] OR fibroses[tw])
4	1 and 2 and 3	56
		Search results
	Total	337
	Duplicated	101
	Final total	236

Table 2 Inclusion and exclusion criteria of studies on vitamin D in non-alcoholic fatty liver disease

Inclusion criteria

Patients ≥ 18 yr

Studies that evaluated vitamin D in NAFLD

Studies that evaluated the liver fibrosis stage, only based on liver biopsy

Studies that reported serum or plasma 25(OH)D levels

Exclusion criteria

Age < 18 yr

Liver diseases other than NAFLD

Studies that used non-invasive methods to evaluate liver fibrosis

Studies with inadequate data

25-OH(D): 25-hydroxyvitamin D; NAFLD: Nonalcoholic fatty liver disease.

Table 3 Characteristics of patients’ studies for vitamin D status in non-alcoholic fatty liver disease

Citation	Patel et al[32]	Luger et al[30]	Barchetta et al[28]	Anty et al[27]	Dasarthy et al[24]	Bril et al[29]	Nelson et al[31]	Targher et al[26]
Year	2016	2016	2012	2016	2014	2015	2016	2007
Country	United States	Austria	Italy	France	United States	United States	United States	Italy
Subjects (M, F)	293 (195, 98)	50 (10, 40)	45 (22, 23)	398 (64, 334)	187 (51, 136)	239 (204, 35)	190 (89, 101)	120 (80, 40)
Population	Suspected NAFLD undergoing liver biopsy	Gastric bypass patients	Suspected NAFLD	Morbidly obese referred for bariatric surgery	Biopsy proven NAFLD, normal controls	Overweight patients	Biopsy proven NAFLD	Biopsy proven NAFLD
Mean BMI	36.1 ± 7.8	43.8 ± 4.3	30.5 ± 5.5	42.8 ± 5.0	35.7 ± 7.0	34.6 ± 0.4	35.6 ± 10.8	26.3 ± 2.0
Subjects	NAFLD	All	NASH	All	NAFLD	NASH	NAFLD	NAFLD
Mean ± SD ALT IU/L	66.5 ± 51.2	36.4 ± 20.8	87.5 ± 46.6	35.2 ± 24.5	45.9 ± 30.0	64.0 ± 4.0	77.0 ± 48.2	105 ± 42.0
Subjects	NAFLD	All	NASH	Morbidly Obese	NAFLD	NASH	NAFLD	NAFLD
Study design	Cross sectional	Cross sectional	Cross sectional	Cross sectional	Cross sectional	Cross sectional	Cross sectional	Cross sectional
Vitamin D analysis	CLIA	Not described	CLIA	CLIA	CLIA	CLIA	GC-MS	CLIA
Mean/SD 25(OH)D (ng/mL), (n) subjects	27.6 ± 11.8	15.6 ± 5.6	22.0 ± 12.4	19.2 ± 9.0	21.2 ± 10.4	21.8 ± 1.0	20.9 ± 4.0	20.4 ± 8.8
Mean/SD 25(OH)D (ng/mL) Non-NAFLD Controls	27.9 ± 12.8	NA	52.9 ± 11.02	21.5 ± 10/2	35.7 ± 6.0	24.5 ± 2.1	NA	30.0 ± 6.0
P value; NAFLD vs controls	0.878	NA	Not significant	0.13	< 0.01	0.18	NA	< 0.001

n: Number of subjects; 25-OH(D): 25-hydroxylvitamin D; SD: Standard deviation; NAFLD: Nonalcoholic fatty liver disease; NASH: Nonalcoholic steatohepatitis; BMI: Body mass index; CLIA: Chemiluminescence; GC-MS: Gas chromatography mass spectroscopy; M: Male; F: Female.

Table 4 Relationship of vitamin D to liver fibrosis in non-alcoholic fatty liver disease

Author/year/Ref	(n), 25-OH(D) Mean ± SD F0	(n), 25-OH(D) Mean ± SD F1	(n), 25-OH(D) Mean ± SD F2	(n), 25-OH(D) Mean ± SD F3	(n), 25-OH(D) Mean ± SD F4	P value
Patel et al[32], 2016	(39) 24.4 ± 10.4	(78) 26.5 ± 8.9	(55) 29.1 ± 12.5	(63) 30.7 ± 14.1	(9) 20.2 ± 20.2	0.028
Targher et al[26], 2007	(16) 20.8 ± 8.4	(10) 14.4 ± 9.2	(7) 10.0 ± 10.0	(6) 6.0 ± 10.8	0	0.01
Anty et al[27], 2016	(50) 20.04 ± 7.81	(233) 19.91 ± 9.12	(98) 18.28 ± 9.58	(15) 16.71 ± 9.86	(2) 25 ± 10.18	0.01
Luger et al[30], 2016	(2) 15.6 ± 5.2	(30) 15.2 ± 6.0	(8) 15.6 ± 4.4	(4) 17.6 ± 7.6	(2) 20.4 ± 4.4	0.792
Bril et al[29], 2015	(61) 20.5 ± 10.4	(75) 24.2 ± 15.1	(22) 20.8 ± 12.1	(22) 25.5 ± 12.2	(5) 21.1 ± 6.9	0.27
Barchetta et al[28], 2012	(1) 20.5	(10) 23.5 ± 14.4	(7) 16.25 ± 6.1	(6) 28.8 ± 14.9	(1) 17.3	0.56

n: Number of subjects; 25-OH(D): 25-hydroxylvitamin D; SD: Standard deviation; F0-F4: Severity score of hepatic fibrosis.

Table 5 Relationship of vitamin D to liver fibrosis in non-alcoholic fatty liver disease by high vs low fibrosis score

Author	(n), 25-OH(D) Mean ± SD F0-F2	(n), 25-OH(D) Mean ± SD F3-F4
Patel et al[32], 2016	(172) 26.9 ± 10.7	(72) 29.4 ± 15.4
Targher et al[26], 2007	(33) 16.6 ± 10.0	(6) 6.0 ± 10.8
Anty et al[27], 2016	(381) 19.5 ± 9.1	(17) 17.7 ± 10.0
Luger et al[30], 2016	(40) 15.2 ± 5.6	(6) 18.6 ± 6.4
Bril et al[29], 2015	(158) 22.3 ± 13.2	(27) 24.7 ± 11.5
Barchetta et al[28], 2012	(7) 20.2 ± 11.07	(18) 26.7 ± 14.2

n: Number of subjects; 25-OH(D): 25-hydroxylvitamin D; SD: Standard deviation; F0-F4: Severity score of hepatic fibrosis.

Table 6 Levels of sIL-2R, ALT, and HBV DNA in the sera of patients with chronic HBV infection (mean ± SD)

Study	Fibrosis stage used
NASH Clinical Research Network Scoring System Definition	Seven stages:
Kleiner et al[53], 2005	F0: No fibrosis
	F1a: Mild zone 3 sinusoidal fibrosis
	F1b: Moderated zone 3 sinusoidal fibrosis
	F1c: Peri-portal sinusoidal fibrosis
	F2: Zone 3 sinusoidal fibrosis and peri-portal sinusoidal fibrosis
	F3: Bridging fibrosis
	F4: Cirrhosis
Brunt et al[54], 1999	Stage 1: Zone 3 perisinusoidal/pericellular fibrosis; focally or extensively present
	Stage 2: Zone 3 perisinusoidal/pericellular fibrosis with focal or extensive periportal fibrosis
	Stage 3: Zone 3 perisinusoidal/pericellular fibrosis and portal fibrosis with focal or extensive bridging fibrosis
	Stage 4: Cirrhosis

NASH: Nonalcoholic steatohepatitis.

Table 7 Methodology for grading of hepatic fibrosis utilized by the authors of the six included studies

Study	Fibrosis stage used
Anty et al[27], 2016	Kleiner et al[53], 2005
Barchetta et al[28], 2012	Brunt et al[54], 1999
Bril et al[29], 2015	Kleiner et al[53], 2005
Luger et al[30], 2016	Kleiner et al[53], 2005
Patel et al[32], 2016	Kleiner et al[53], 2005
Targher et al[26], 2007	Brunt et al[54], 1999