Nonalcoholic fatty liver disease (NAFLD) is a condition that
causes fatty hepatic parenchymal cell degeneration without a history of
alcohol consumption. The pathogenic variations of NAFLD include
simple steatosis, nonalcoholic steatohepatitis (NASH), and NASH-related
cirrhosis, which may progress to hepatocellular carcinoma (HCC).
The introduction of reliable non or minimal invasive accurate
biomarkers is urgently needed to avoid the hazardous complications of
invasive liver biopsy technique.
Adiponectin has been shown to reduce hepatic and systemic insulin
resistance, as well as liver inflammation and fibrosis. Adiponectin is a
hormone that predicts the degree of steatosis and NAFLD. Despite the
lack of a proven pharmacotherapy for NAFLD, current therapeutic efforts
have focused on the indirect upregulation of adiponectin via the
administration of various therapeutic agents and/or lifestyle changes.
The aim of this study was to investigate adiponectin promotor
methylation status in patients with NAFLD and to evaluate the correlation
between the adiponectin promotor methylation status and the
clinicopathological characteristics of NAFLD patients
The study included 49 subjects of both sex selected from
Department of Endemic Medicine, Faculty of Medicine, Cairo University
Hospital.
The subjects were categorized into 3 groups:
simple steatosis group: included 5 patients, diagnosed by clinical,
radiological and histopathological examinations (NAS score < 4)
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NASH group: included 29 patients, diagnosed as NAFLD patients
by clinical, radiological and histopathological examinations(NAS
score ≥ 4)
Control group: included 15 apparently healthy subjects, age and
sex matched, with apparently normal liver.
All individuals were subjected to:
Full history taking.
General and local abdominal examinations.
Investigations include:
- Routine Laboratory investigations
- Radiological investigations
Liver biopsy for histopathology (from patients’ group).
Molecular biology investigations: SYBR Green methylation
specific polymerase chain reaction (qMSP) for detection of
adiponectin promotor methylation status.
Blood samples were collected into EDTA vacutainers from all
individuals and stored at -80°C. Adiponectin promotor qMSP assay was
performed as follows:
Extraction of genomic DNA from peripheral blood samples.
Bisulfite treatment of genomic DNA.
SYBR Green Methylation specific PCR (qMSP) using specific
primer sets for either methylated or non-methylated products of
adiponectin gene promoter. The PCR product was separated by
gel electrophoresis, stained with ethidium bromide and visualized
by UV irradiation, for detection of specific bands. Adiponectin
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methylation percentage was calculated according to the following
equation:
While ΔCt = adiponectin Ct – GAPDH Ct
The current study showed that patients with NASH had significantly
higher results when compared to simple steatosis regarding, pain as
clinical symptom and body mass index.
Importantly, there was a significant increase in adiponectin
promotor methylation in NASH patients with higher fibrosis grades as
compared to those with lower liver fibrosis grades
There was significant positive correlation between adiponectin
promotor methylation and height, body mass index, serum level of AST
and prothrombin concentration in NASH patients.
ROC curve analysis showed that AUC of adiponectin promotor
methylation in detection of NAFLD was 0.767with Sensitivity 55.9%,
Specificity was 100%, PPV was 100%, NPV was 69.4%, and Accuracy
was 78.0%, with significant prediction (P |
