Association of whole blood viscosity with non-alcoholic fatty liver disease

Abstract

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is an independent risk factor for increased cardiovascular disease. Altered hemorheological parameters have also been shown to play a crucial role in atherogenesis. Moreover, increased viscosity is observed in insulin resistance, metabolic syndrome, hypertension, diabetes, ischemic heart disease, and stroke. A recent study confirmed that whole blood viscosity (WBV) is a predictor of cardiovascular events.

AIMS: This study aimed to investigate the association of WBV with NAFLD.

METHODS: In this cross-sectional study, we investigated the relationship between WBV and NAFLD in 1329 subjects (962 men and 367 women) in a general health examination.

RESULTS: WBV at low shear stress was elevated in patients with NAFLD. In addition, the prevalence of metabolic syndrome and NAFLD increased as WBV quartiles increased both in men and in women. Multiple regression analysis further identified WBV as an independent and significant determinant for NAFLD.

CONCLUSIONS: The findings showed that WBV at low shear stress is increased in NAFLD. Moreover, WBV at low shear stress is independently associated with NAFLD even after adjusting other cardiovascular risk factors.

Keywords

Whole blood viscosity non-alcoholic fatty liver disease metabolic syndrome

1 Introduction

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver condition in the western countries, affecting 20–40% of the general population [12]. NAFLD includes liver diseases varying from steatosis and steatohepatitis to cirrhosis [19]. Epidemiological studies have reported that NAFLD is associated with obesity, dyslipidemia, hypertension, hypercholesterolaemia, metabolic syndrome and type 2 diabetes [11]. Clinical and experimental studies suggested that insulin resistance plays a key role in the pathology of NAFLD [8]. Recent studies showed that NAFLD is an independent risk factor for increased cardiovascular disease [3]. Evidence also indicated a graded association between NAFLD severity and increased vascular risk [2]. However, there are no suitable noninvasive biomarkers of NAFLD severity to allow better risk stratification based on cardiovascular outcomes.

Altered hemorheological parameters have also been shown to play a crucial role in atherogenesis. Many cardiovascular risk factors, including aging, obesity, carotid intima-media thickness, are associated with changes in hemorheological parameters [1 , 18]. Moreover, increased viscosity is observed in insulin resistance, metabolic syndrome, hypertension, diabetes, ischemic heart disease, and stroke [16]. In addition, a study confirmed that whole blood viscosity (WBV) is a predictor of cardiovascular events in apparently healthy individuals [10].

However, there is scarcity of data that explore the relationship of blood viscosity with NAFLD. The purpose of the study was to investigate whether rheological parameters levels are associated with NAFLD.

2 Methods

2.1 Participants

The study included 1329 subjects (962 men and 367 women) who received general health examination in our hospital from January 2010 to December 2010. There were 353 subjects with NAFLD (mean age 48.7 ± 5.0 years) and 976 controls (mean age 46.9 ± 5.7 years). We obtained written informed consent from all subjects. The study protocol was approved by the Ethics Committee of the Second Hospital of Harbin Medical University, China.

2.2 Clinical examination

Clinical data, including medical history, smoking status and medication use were recorded for each participant. All the subjects underwent physical examination, which included anthropometric and blood pressure measurements, and an ultrasound scan of the liver. Blood pressure was determined using a mercury-gravity sphygmomanometer in a sitting position after a 15-min rest. Systolic and diastolic blood pressures were measured twice on the same day and mean values were used in the analysis.

2.3 Biochemical analyses

Fasting venous blood samples were drawn for the analysis. The values included total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL), low-density lipoprotein cholesterol (LDL), aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyl transpeptidase (GGT), and fasting plasma glucose (FPG). All assays were performed at the Laboratory of Analytical Biochemistry at the Second Hospital of Harbin Medical University, Harbin, using a biochemical analyzer (Modular Analytics, Roche, Mannheim, Germany) using standard methods.

2.4 Hemorheological parameters

Whole blood viscosity was measured at shear rates between 3 s^–1 and 200 s^–1 corrected hematocrit of 45% at 37°C using a viscometer (Succeeder SA-9000, Beijing, China). Plasma viscosity was determined by Harkness method and hematocrit was evaluated by microcentrifugation. Plasma fibrinogen concentrations were assayed on the Beckman Coulter ACL-TOP analyzer (Instrumentation Laboratory, Lexingtion, MA, USA). All measurements were performed within 2 h of sampling.

2.5 Ultrasonography

An experienced ultrasonographist performed the abdominal ultrasonography using a sonography machine (Voluson E8, GE, America) with a 3.5-MHz probe. Hepatic steatosis was diagnosed by characteristic echo patterns, such as diffuse hyperechogenicity of the liver relative to the kidneys, ultrasound beam attenuation, and poor visualization of intrahepatic structures (11).

2.6 Diagnostic criteria

Diagnosis of type 2 diabetes (DM) was based on American Diabetes Association criteria such as fasting plasma glucose ≥7.0 mmol/L, current treatment with a hypoglycemic agent, or casual glucose ≥11.1 mmol/L. For the controls or the patients with impaired fasting glucose, DM was diagnosed if a 2-hr post-glucose level after a 75-g oral glucose tolerance test ≥11.1 mmol/L. Hypertension was diagnosed if systolic blood pressure ≥140 mmHg and diastolic pressure ≥90 mmHg, or as antihypertensive treatment. Two readings were taken, with a 5-minute interval between measurements. The mean of the two readings was recorded. Metabolic syndrome was defined by the presence of 3 or more of the following risk factors: (1) obesity with BMI ≥25.0 kg/m²; (2) high triglycerides ≥1.7 mmol/L; (3) low HDL-cholesterol <1.04 mmol/L for men and <1.30 mmol/L for women; (4) elevated systolic blood pressure ≥130 mmHg or elevated diastolic blood pressure ≥85 mmHg; and (5) high fasting plasma glucose ≥6.1 mmol/L. Subjects who reported taking anti-hypertensive or anti-diabetic medications were considered to have elevated blood pressure or high fasting plasma glucose.

2.7 Exclusion criteria

The exclusion criteria included: 1) an alcohol intake of more than 20 g/day; 2) viral, autoimmune, or hereditary liver disease; 3) a prior history of taking medication that could cause steatosis; 4) a prior history of any kind of liver disease; 5) tumor, infection, hematological disorders, and rheumatoid arthritis; 6) medical treatment with lipid-lowering agents, hormone replacement, and anticoagulant.

2.8 Statistical analysis

The data were expressed as means ± SD or median (IQR) or percentage. The Chi-square statistical test was used for all categorical variables, while the student’s t-test or Mann-Whitney U test was used for continuous variables in two independent samples. One-way ANOVA or Kruskal-Wallis H test was performed for continuous variables according to WBV quartiles. The odds ratios (ORs) and 95% confidence intervals (95% CIs) for NAFLD were calculated after adjusting for confounding variables across WBV quartiles using multivariate logistic regression analysis. All above tests were considered significant at P < 0.05 (two tailed). Statistical analyses were performed using the SPSS software package version 17.0 (SPSS Inc., Chicago, IL, USA).

3 Results

Clinical and laboratory data of participants with and without NAFLD were shown in Table 1. The group with NAFLD had higher age, BMI, SBP, DBP, FPG, TG, LDL, AST, ALT, GGT, WBV, fibrinogen and lower HDL levels compared to the group without NAFLD. Male, smoker, the proportion of patients with hypertension, diabetes and metabolic syndrome were more prevalent in NAFLD group. TC, hemoglobin, hematocrit, and PV in the two groups had no difference.

The clinical and laboratory characteristics of subjects were shown in Tables 2 and 3 according to WBV 3 s^–1 quartiles. In men, mean age, BMI, SBP, DBP, ALT, GGT, FPG, TC, TG, LDL, WBV 200 s^–1, fibrinogen, hematocrit, PV, and the proportion of smokers, hypertension and diabetes increased gradually as WBV increased. There was no significant difference in AST and hemoglobin. In women, mean age, BMI, SBP, DBP, FPG, TG, LDL, WBV 200 s^–1, fibrinogen, hematocrit, and the proportion of smoker and hypertension increased gradually as WBV increased. TC, AST, ALT, GGT, hemoglobin, PV, and the proportion of diabetes were not significantly different among the subjects with all four WBV quartiles in women. The levels of HDL decreased as WBV quartiles increased both in men and in women.

The prevalence of metabolic syndrome and NAFLD were calculated by the quartiles of WBV levels (Figs. 1–4). For men, the WBV quartiles were quartile 1 (Q1) (≤7.26 mPa.s), quartile 2 (Q2) (7.27–7.84 mPa.s), quartile 3 (Q3) (7.85–9.53 mPa.s), quartile 4 (Q4) (≥9.54 mPa.s). For women, the WBV quartiles were quartile 1 (Q1) (≤6.78 mPa.s), quartile 2 (Q2) (6.79–7.82 mPa.s), quartile 3 (Q3) (7.83–8.61 mPa.s), quartile 4 (Q4) (≥8.62 mPa.s). For men, the PR% of metabolic syndrome in Q1, Q2, Q3, and Q4 was 7.79% (19/244), 9.66% (23/238), 13.22% (32/242) and 43.28% (103/238), respectively. For women, the PR% of metabolic syndrome in Q1, Q2, Q3, and Q4 was 10.53% (10/95), 13.33% (12/90), 18.56% (18/97) and 25.88% (22/85), respectively. For men, the PR% of NAFLD in Q1, Q2, Q3, and Q4 was 10.66% (26/244), 11.34% (27/238), 41.32% (100/242) and 49.58% (118/238), respectively. For women, the PR% of NAFLD in Q1, Q2, Q3, and Q4 was 7.37% (7/95), 11.11% (10/90), 31.96% (31/97) and 40.00% (34/85), respectively. The results indicated that the prevalence of metabolic syndrome and NAFLD increased as WBV quartiles increased.

The risks of NAFLD according to WBV quartiles are shown in Table 4. After adjusting for age, BMI, smoking status, the prevalence risk of NAFLD for the highest quartile of WBV in men and women were 6.314 (3.834–10.396) and 4.989 (1.934–12.872), respectively (P < 0.001). These associations were similar after additional adjustment for SBP, DBP, FPG, TC, TG, HDL, LDL, hypertension, diabetes, AST, ALT, GGT, WBV 200 s^–1, PV, hematocrit, and fibrinogen.

4 Discussion

In this study, we found that WBV at low shear stress is increased in NAFLD. Moreover, multiple regression analysis further identified WBV as an independent and significant determinant for NAFLD.

Our study indicated that WBV at low shear stress is associated with NAFLD. Possible mechanisms linking WBV with NAFLD may be insulin resistance, subclinical inflammation, and dyslipidaemia, which are implicated in the pathophysiology of NAFLD. Firstly, the development and progression of insulin resistance play a primary role in the initiation and propagation of NAFLD. Furthermore, insulin resistance and metabolic syndrome are associated with hyperviscosity syndrome [4]. Our results reported that WBV in low shear stress is increased in NAFLD. In low-shear stress, flow alterations cause reduced shear stress in focal areas, leading to endothelial remodeling and altered physiological responses [6]. Elevated WBV has shown to lead to vascular remodeling, altered lipid metabolism, and endothelial inflammation [13, 17]. WBV-induced endothelial changes could further increase atherothrombotic risk. Secondly, The liver plays a central role in mediating systemic inflammation and atherothrombosis. C-reactive protein (CRP), a common inflammatory marker, is elevated in NAFLD, with levels increasing as the severity of NAFLD progresses [15]. Recent studies reported that CRP and leucocyte count are positively correlated with blood viscosity. Additionally, increased blood viscosity is associated with metabolic syndrome, hypertension, diabetes, ischemic heart disease, and stroke, which all are associated with chronic inflammation. Finally, dyslipidaemia is a common feature of NAFLD, including high TG levels, low HDL cholesterol, and high LDL levels. A large body of evidence had documented that WBV is positively correlated with LDL cholesterol and TG concentrations, and negatively correlated with HDL cholesterol [7, 14]. Dyslipidaemia and high viscosity accelerate the development of atherosclerosis in a synergistic fashion.

Recently, higher blood viscosity and hematocrit were observed in patients with metabolic syndrome [5]. Consistent with the results, our study showed that WBV level is associated with metabolic syndrome and NAFLD.

Some limitations of the present study are noteworthy. Firstly, due to the cross-sectional nature of the study, no causal relationships can be established. Prospective studies are required to draw firm conclusions. Secondly, the diagnosis of NAFLD was based on ultrasound and was not confirmed with liver biopsy.

In conclusion, WBV at low shear stress is increased in NAFLD. Moreover, WBV at low shear stress is independently associated with NAFLD even after adjusting other cardiovascular risk factors.

Conflicts of interest

None.

Footnotes

Acknowledgments

We wish to thank the staff of International Physical Examination and Healthy Center for their help with data collection.

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