Non invasive blood flow measurement in cerebellum detects minimal hepatic encephalopathy earlier than psychometric tests

Table 1 Composition of the groups and clinical data (mean ± SD)

	Range	Control	Patients without MHE	Patients with MHE
Total individuals		14	24	16
Age		55 ± 9	58 ± 6	65 ± 9
Alcohol			24	16
Ascitis			2	5
Child Pugh A/B/C			21/3/0	11/3/2
MELD			8.9 ± 3.5	9.5 ± 3.4
AST (mU/mL)	1-37	20 ± 4.0	73 ± 56b	82.5 ± 58b
ALT (mU/mL)	1-41	18 ± 6.0	77 ± 24b	90.1 ± 24b
GGT (mU/mL)	10-49	26.7 ± 5.0	86.4 ± 60b	106 ± 64b
Uric acid (mg/dL)	2.5-7	4.0 ± 1.0	6.2 ± 2.0	5.73 ± 2.3
Creatinine (mg/dL)	0.5-1.3	0.92 ± 0.1	1.1 ± 0.2	1.2 ± 0.2
Cholesterol (mg/dL)	140-200	172 ± 22	175 ± 44	167 ± 55
Triglycerides (mg/dL)	40-160	95 ± 32	111 ± 64	119 ± 64
Bilirubin (mg/dL)	0.1-1	0.6 ± 0.2	1.7 ± 0.7b	2.3 ± 0.6b
Albumin (g/dL)	3.5-5	4.4 ± 0.2	3.7 ± 0.6b	2.9 ± 0.6bc
Prothrombin time (s)		13 ± 1.3	24 ± 4b	30 ± 4b
Fibrinogen (g/L)	2-4	3.1 ± 1.0	3.3 ± 1.3	3.6 ± 1.2
Alkaline phosphatase (mU/mL)	50-250	147 ± 53	216 ± 77b	314 ± 96bc
Erythrocytes	4.2-6.1	4.6 ± 0.4	4.3 ± 0.7	3.4 ± 0.6
Leucocytes	4.8-10.8	6.5 ± 1.3	6 ± 2.6	5.5 ± 2.0
Neutrophils (%)	55-75	55 ± 7.4	54 ± 6.2	59 ± 9.3
Lymphocytes (%)	17-45	35 ± 6.0	29 ± 10	27 ± 9.4
Monocytes (%)	2-8	6.0 ± 1.3	8.4 ± 3.0b	10 ± 2.6b
Eosinophils (%)	1-4	3.3 ± 2.0	2.4 ± 1.2	1.7 ± 1.0
Basophils (%)	0.05-0.5	0.5 ± 0.2	0.6 ± 0.3	0.6 ± 0.1

^bP < 0.01 vs control,

^cP < 0.05 vs without minimal hepatic encephalopathy (MHE). MELD: Model for end stage liver disease. Values that are significantly different from controls are indicated by superscripts.

Table 2 Biochemical parameters in blood (mean ± SE)

Parameter	Controls(n = 14)	Patients without MHE(n = 24) P vs control	Patients with mHE(n = 16) P vs control	Patients with MHEPvs without MHE	P values
cGMP in plasma (pmol/mL)	0.64 ± 0.1	6.1 ± 0.6	8.7 ± 1.1
		P < 0.0001	P < 0.0001	< 0.05	< 0.0001
Nitrates + Nitrites (μmol/L)	18 ± 1	38 ± 4	44 ± 9
		P = 0.004	P = 0.004	NS	0.001
3-Nitro-tyrosine (nmol/L)	3.5 ± 1.2	10 ± 1	43 ± 6	< 0.0001	< 0.0001
			P < 0.0001
IL-6 (pg/mL)	0.7 ± 0.2	2.3 ± 0.3	4.4 ± 0.5	0.001	< 0.0001
		P < 0.05	P < 0.0001
IL-18 (pg/mL)	103 ± 13			< 0.05	< 0.0001
		268 ± 39	404 ± 44
		P = 0.02	P < 0.0001
Ammonia (μmol/L)	39 ± 4	84 ± 14	103 ± 33	NS	0.005
		P = 0.04	P = 0.006

One-way analysis of variance (ANOVA) followed by post-hoc Bonferroni’s multiple comparisons test were performed. Adjusted P values are shown. NS: Not significant.

Table 3 Psychometric tests and attention and coordination tasks in the different groups of cirrhotic patients and in controls (mean ± SD)

Test	Sub-test	Controls(n = 14)	Patients without MHE(n = 24)P vs Control	Patients with MHE(n = 16)P vs control	Patients with MHEP vs without MHE	P values
CFF		43.2 ± 0.7	40.5 ± 0.7	36 ± 1	P = 0.001	< 0.001
				P < 0.001
PHES Global score		-0.13 ± 0.25	-1.23 ± 0.30	-6.31 ± 0.72
				P < 0.001	P < 0.001	< 0.001
	Symbol digit test	-0.07 ± 0.12	-0.11 ± 0.08	-0.87 ± 0.12
				P < 0.001	P < 0.001	< 0.001
	NCT-A	-0.07 ± 0.15	-0.04 ± 0.07	-1.19 ± 0.28
				P < 0.001	P < 0.001	< 0.001
	NCT-B	-0.07 ± 0.18	-0.27 ± 0.14	-2.07 ± 0.30
				P < 0.001	P < 0.001	< 0.001
	Serial dotting test	0.00 ± 0.00	-0.57 ± 0.17	-1.19 ± 0.29	NS
				P = 0.001		0.002
	Line tracing test	-0.07 ± 0.07	-0.15 ± 0.07	-1.25 ± 0.33	P < 0.001
				P < 0.001		< 0.001
Stroop	Congruent task	102 ± 5	90 ± 2	76 ± 4
			P = 0.03	P < 0.0001	P = 0.009	< 0.0001
	Neutral Task	75 ± 3	67 ± 2	54 ± 3
				P < 0.0001	P = 0.003	< 0.0001
	Incongruent Task	42 ± 2	36 ± 2	28 ± 2	P = 0.030	0.001
				P = 0.001
Bimanual coordination		1.73 ± 0.05	2.16 ± 0.05	2.83 ± 0.15
			P = 0.05	P < 0.0001	P < 0.0001	< 0.0001
Visuo-motor coordination		2.15 ± 0.08	2.61 ± 0.08	3.89 ± 0.28		< 0.0001
				P < 0.0001	P < 0.0001

Statistical analysis: One-way analysis of variance (ANOVA) followed by post-hoc Bonferroni’s multiple comparisons test , except for Bimanual and Visuo-motor coordination tests, in which univariate analysis of covariance (ANCOVA) was performed, with age included as covariate, followed by post-hoc Bonferroni. Adjusted P values are shown. Stroop test: Congruent task: number of words read in 45 s; Neutral task: Number of colours read in 45 s; Incongruent task: Number of items completed in 45 s. Bimanual and Visuo-motor coordination tests: Score in minutes. CFF: Critical Flicker Frequency; PHES: Psychometric Hepatic Encephalopathy Score; NCT-A, NCT-B: Number Connection Test A and B: respectively; NS: Not significant.

Table 4 Correlations between tests and biochemical parameters and blood flow in the brain areas

Studied parameter/test	Sub-test	Frontal lobe	Parietal lobe	Occipital	Vermis	Cerebellar hemisphere	Thalamus
CFF		NS	NS	r = 0.353	r = -0.326	NS	NS
				P = 0.017	P = 0.025
PHES Global score		r = -0.36	NS	r = 0.309	r = -0.504	r = -0.348	NS
		P = 0.007		P = 0.031	P < 0.001	P = 0.01
	Symbol digit test	NS	NS	NS	NS	NS	NS
	NCT-A	r = -0.469	NS	NS	r = -0.530	r = -0.320	NS
		P < 0.001			P < 0.001	P = 0.018
	NCT-B	NS	NS	NS	r = -0.531	r = -0.307	NS
					P < 0.001	P = 0.027
	Serial dotting test	NS	NS	NS	NS	NS	NS
	Line tracing test	NS	NS	NS	r = -0.330	r = -0.364	NS
					P = 0.018	P = 0.007
STROOP	Congruent task	NS	NS	NS	r = -0.320	r = -0.354	NS
					P = 0.027	P = 0.012
	Neutral task	NS	NS	NS	NS	NS	NS
	Incongruent task	NS	NS	NS	NS	NS
							NS
Bimanual coordination		NS	NS	NS	NS	r = 0.362	NS
						P = 0.011
Visuo-motor coordination		NS	NS	NS	NS	r = 0.335	NS
						P = 0.017
cGMP in plasma (pmoles/mL)		NS	NS	NS	NS	r = 0.301	NS
						P = 0.034
Nitrates + Nitrites (mmol/L)		NS	NS	NS	r = 0.339	r = 0.362	NS
					P = 0.043	P = 0.023
3-Nitro-tyrosine (nmol/L)		NS	NS	NS	r = 0.358	NS	NS
					P = 0.011
IL-6 (pg/mL)		NS	NS	NS	NS	NS	NS
IL-18 (pg/mL)		NS	NS	r = -0.353	NS	NS	r = -0.405
				P = 0.041			P = 0.032
Ammonia (mmol/L)		NS	NS	NS	NS	NS	NS

PHES: Psychometric Hepatic Encephalopathy Score; NCT-A, NCT-B: Number Connection Test A and B, respectively; CFF: Critical flicker frequency; NS: No significant correlation.