Pooled genetic analysis in ultrasound measured non-alcoholic fatty liver disease in Indian subjects: A pilot study

Table 1 List of single nucleotide polymorphisms included in the study

SNP No	rsID	Risk allele	Associated gene	Associated with	Ref.
1	rs738409	G	PNPLA3	Hepatic steatosis	[6]
2	rs4240624	A	PPP1R3B	Hepatic steatosis	[6]
3	rs2228603	T	NCAN	Hepatic steatosis	[6]
4	rs780094	A	GCKR	Hepatic steatosis	[6]
5	rs12137855	C	LYPLAL1	Hepatic steatosis	[6]
6	rs2645424	C	FDFT1	NAFLD activity score	[8]
7	rs343062	T	-	Degree of fibrosis	[8]
8	rs1227756	G	COL13A1	Lobular inflammation	[8]
9	rs6591182	G	-	Lobular inflammation	[8]
10	rs887304	A	EFCAB4B	Lobular inflammation	[8]
11	rs2499604	A	Intronic ZP4-TRNAP23P	Serum levels of alanine aminotransferase	[8]
12	rs6487679	C	PZP	Serum levels of alanine aminotransferase	[8]
13	rs1421201	C	-	Serum levels of alanine aminotransferase	[8]
14	rs2710833	T	-	Serum levels of alanine aminotransferase	[8]
15	rs2854116	A	APOC3	Hypertriglyceridemia in Asians	[9]
16	rs2854117	G	APOC3	Hypertriglyceridemia in Asians	[9]
17	rs2143571	A	SAMM50	NAFLD	[7]
18	rs2073080	T	PARVB	NAFLD	[7]
19	rs1390096	A	HS3ST1-HSP90AB2P	NAFLD	[7]
20	rs11206226	A	YIPF1	NAFLD	[7]

PNPLA3: Patatin-like phospholipase domain containing 3; NCAN: Neurocan; GCKR: Glucokinase regulatory protein; LYPLAL1: Lysophospholipase-like protein 1; PPP1R3B: Protein phosphatase 1, regulatory subunit 3B; PZP: Pregnancy zone protein.

Table 2 Demographic and clinical characteristics of the study group

Parameter	Controls	Patients	P value
	(n = 150)	(n = 156)
Mean ± SD	39.78 ± 10.83	36.63 ± 8.20	0.004
Age, yr	18-63	19-62
Range
Males	110 (73.33%)	138 (88.46%)	-
Females	40 (26.66%)	18 (11.53%)	-
Waist circumference
Males	80.13 ± 10.35	94.09 ± 10.53	0.0001
Females	81.77 ± 13.65	92.53 ± 8.27	0.01
BMI (kg/m2)	24.04 ± 7.77	27 ± 5.86	0.001
Triglycerides (mg/dL)	135.18 ± 7.77	184.40 ± 84.31	0.0001
HDL (mg/dL)	41.86 ± 9.70	39.56 ± 13.02	0.2923
ALT (IU/L)	25.39 ± 14.73	110.20 ± 67.05	0.0001
AST (IU/L)	25.99 ± 8.46	69.14 ± 37.77	0.0001
Hypertensives	4 (2.6%)	18 (11.53%)	-
Diabetics	19 (12.66%)	27 (17.3%)	-

BMI: Body mass index; HDL: High density lipoprotein; ALT: Alanine transaminase; AST: Aspartate amino transferase.

Table 3 Comparison of variant carriers between patients and controls

SNP-	Allele frequency controls (n = 150)		Allele frequency patients (n = 156)		χ2	P value	Corrected P value1	OR	95%CI
gene name	Major	Minor	Major	Minor					lower-upper
rs1227756	0.49	0.51	0.44	0.56	0.02	0.88	0.93	1.04	0.54-2.03
COL13A1
rs12137855	0.77	0.23	0.73	0.27	0.41	0.51	0.62	1.20	0.68-2.12
LYPLAL1
rs1390096	0.68	0.32	0.66	0.34	0.007	0.93	0.93	0.97	0.55-1.70
HS3ST1-HSP
rs1421201	0.88	0.12	0.85	0.15	1.60	0.20	0.36	1.55	0.78-3.10
intronic
rs2073080	0.81	0.19	0.69	0.31	8.42	0.003	0.02	2.36	1.31-4.22
PARVB
rs2143571	0.80	0.20	0.69	0.31	6.25	0.01	0.05	2.07	1.16-3.69
SAMM50
rs2228603	0.97	0.03	0.92	0.08	4.09	0.04	0.12	3.29	1.10-9.84
NCAN
rs2499604	0.53	0.47	0.60	0.40	3.76	0.05	0.13	1.92	0.98-3.75
intronic
rs2645424	0.5	0.50	0.56	0.44	1.21	0.27	0.40	0.69	0.36-1.32
FDFT1
rs2710833	0.60	0.40	0.66	0.34	3.09	0.07	0.17	0.56	0.30-1.07
intronic
rs2854117	0.58	0.42	0.55	0.45	4.24	0.03	0.12	1.83	1.02-3.28
APOC3
rs343062	0.55	0.45	0.52	0.48	1.96	0.16	0.32	1.53	0.84-2.80
intronic
rs4240624	0.94	0.06	0.91	0.09	1.21	0.27	0.40	1.56	0.69-3.52
PPP1R3B
rs6487679	0.89	0.11	0.75	0.25	9.65	0.001	0.01	2.81	1.44-5.48
PZP
rs6591182	0.63	0.37	0.59	0.41	0.99	0.31	0.44	1.39	0.72-2.67
intronic
rs738409	0.92	0.08	0.60	0.40	46.37	0.0001	0.001	12.66	5.45-29.38
PNPLA3
rs780094	0.76	0.24	0.74	0.26	0.48	0.48	0.62	1.22	0.69-2.15
GCKR
rs887304	0.82	0.18	0.83	0.17	0.19	0.65	0.73	0.87	0.47-1.59
EFCAB4B
rs11206226	1.00	0.00	1.00	0.00	-	-		-	-
YIPF1

¹Benjamini and Hochberg false discovery rate correction was applied to the “P value”. PNPLA3: Patatin-like phospholipase domain containing 3; NCAN: Neurocan; GCKR: Glucokinase regulatory protein; LYPLAL1: Lysophospholipase-like protein 1; PPP1R3B: Protein phosphatase 1, regulatory subunit 3B; PZP: Pregnancy zone protein.

Table 4 Significant single nucleotide polymorphisms associated with higher odds of body mass index, triglycerides and alanine transaminase based on clinical characteristics

Variable	SNP-gene	OR	P value	95%CI
				lower-upper
BMI (obese and non-obese)	rs2073080-PARVB	1.81	0.0470	1.00-3.25
BMI (obese and non-obese)	rs343062-intronic	2.25	0.0100	1.21-4.21
BMI (obese and non-obese)	rs6591182-intronic	2.05	0.0340	1.05-4.00
TG (abnormal and normal)	rs2854117-APOC3	2.31	0.0040	1.29-4.13
TG (abnormal and normal)	rs738409-PNPLA3	1.94	0.0170	1.12-3.37
ALT (abnormal and normal)	rs2073080-PARVB	1.92	0.0200	1.10-3.36
ALT (abnormal and normal)	rs2143571-SAMM50	1.77	0.0200	1.10-3.36
ALT(abnormal and normal)	rs2228603-NCAN	3.23	0.0180	1.22-1.37
ALT (abnormal and normal)	rs6487679-PZP	1.92	0.0300	0.05-3.51
ALT (abnormal and normal)	rs738409-PNPLA3	5.07	0.0001	0.03-10.92

BMI: Body mass index; TG: Triglycerides; ALT: Alanine transaminase; SNP: Single nucleotide polymorphisms.

Table 5 PARVB-SAMM50-PNPLA3 haplotype data

Total counts	Haplotype	Number in controls	Number in patients
138	000	90	48
31	001	6	25
10	010	10	0
4	011	2	2
4	100	2	2
48	110	32	16
71	111	8	63

“Total Counts” column consists of the number of haplotypes generated in both controls and patient group combined. In the haplotype column, 0 stands for wild type allele and 1 for either heterozygous or homozygous variant carrier. The sequence of the genes is PARVB, SAMM50 and PNPLA3. The individuals count for the haplotypes for controls and patient group is given in the Number in controls and Number in patients column.

Table 6 Results of multiple logistic regression analysis

Variable	Regression	Standard	P value	OR	95%CI
	coefficient	error			Lower	Upper
PZP	0.880	0.415	0.034	2.41	1.05	5.44
PNPLA3	2.289	0.480	< 0.0001	9.86	3.85	25.29
Triglyceride levels	1.502	0.391	0.000	4.48	2.08	9.67
Constant	-0.619

PNPLA3: Patatin-like phospholipase domain containing 3; PZP: Pregnancy zone protein.