Intrahepatic expression of genes related to metabotropic receptors in chronic hepatitis.
OBJECTIVE
To screen for genes related to metabotropic receptors that might be involved in the development of chronic hepatitis.


METHODS
Assessment of 20 genes associated with metabotropic receptors was performed in liver specimens obtained by punch biopsy from 12 patients with autoimmune and chronic hepatitis type B and C. For this purpose, a microarray with low integrity grade and with oligonucleotide DNA probes complementary to target transcripts was used. Evaluation of gene expression was performed in relation to transcript level, correlation between samples and grouping of clinical parameters used in chronic hepatitis assessment. Clinical markers of chronic hepatitis included alanine and aspartate aminotransferase, γ-glutamyltranspeptidase, alkaline phosphatase and cholinesterase activity, levels of iron ions, total cholesterol, triglycerides, albumin, glucose, hemoglobin, platelets, histological analysis of inflammatory and necrotic status, fibrosis according to METAVIR score, steatosis, as well as anthropometric body mass index, waist/hip index, percentage of adipose tissue and liver size in ultrasound examination. Gender, age, concomitant diseases and drugs were also taken into account. Validation of oligonucleotide microarray gene expression results was done with the use of quantitative real-time polymerase chain reaction (qRT-PCR).


RESULTS
The highest (0.002 < P < 0.046) expression among genes encoding main components of metabotropic receptor pathways, such as the α subunit of G-coupled protein, phosphoinositol-dependent protein kinase or arrestin was comparable to that of angiotensinogen synthesized in the liver. Carcinogenesis suppressor genes, such as chemokine ligand 4, transcription factor early growth response protein 1 and lysophosphatidic acid receptor, were characterized by the lowest expression (0.002 < P < 0.046), while the factor potentially triggering hepatic cancer, transcription factor JUN-B, had a 20-fold higher expression. The correlation between expression of genes of protein kinases PDPK1, phosphoinositide 3-kinase and protein kinase A (Spearman's coefficient range: 0.762-0.769) confirmed a functional link between these enzymes. Gender (P = 0.0046) and inflammation severity, measured by alanine aminotransferase activity (P = 0.035), were characterized by diverse metabotropic receptor gene expression patterns. The Pearson's coefficient ranging from -0.35 to 0.99 from the results of qRT-PCR and microarray indicated that qRT-PCR had certain limitations as a validation tool for oligonucleotide microarray studies.


CONCLUSIONS
A microarray-based analysis of hepatocyte metabotropic G-protein-related gene expression can reveal the molecular basis of chronic hepatitis.
Department of Gastroenterology, Hepatology and Infectious Diseases, Jagiellonian University Medical College, 31-521 Kraków, Poland. aciesla@su.krakow.pl
Andrzej
A
Cieśla
Maciej
M
Kuśmider
Agata
A
Faron-Górecka
Marta
M
Dziedzicka-Wasylewska
Monika
M
Bociąga-Jasik
Danuta
D
Owczarek
Irena
I
Ciećko-Michalska
Dorota
D
Cibor
Tomasz
T
Mach
D064413:3-Phosphoinositide-Dependent Protein Kinases; D000328:Adult; D019390:Arrestins / Q000235:genetics / Q000378:metabolism; D001706:Biopsy; D002864:Chromogranins; D005260:Female; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000235:genetics / Q000378:metabolism; D019694:Hepatitis B, Chronic / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D019698:Hepatitis C, Chronic / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D022781:Hepatocytes / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008099:Liver / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000235:genetics* / Q000378:metabolism*; D012189:Retrospective Studies; D059467:Transcriptome; D000071557:beta-Arrestins
D019390:Arrestins; D002864:Chromogranins; D043562:Receptors, G-Protein-Coupled; D000071557:beta-Arrestins; D064413:3-Phosphoinositide-Dependent Protein Kinases; C578562:PDPK1 protein, human; D017346:Protein Serine-Threonine Kinases; C406732:GNAS protein, human; D019205:GTP-Binding Protein alpha Subunits, Gs