Identification of disease-specific genes in chronic pancreatitis using DNA array technology

Abstract

Objective: To use DNA arrays to analyze the differential gene expression patterns in the normal pancreas and in pancreatic diseases.

Summary background data: Genome-wide gene expression analysis will provide new insights into gene function and cause of disease.

Methods: RNA was extracted from eight normal pancreatic specimens, eight specimens with chronic pancreatitis (CP), and eight pancreatic cancer (PCa) tissues. Poly A(+) RNA was purified, reverse-transcribed, and converted into cRNA using biotinylated nucleotides. The HuGeneFL DNA array containing 5,600 full-length human genes was used for analysis.

Results: First, normal pancreatic tissues were analyzed in comparison with a panel of other normal tissues (colon, liver, prostate, lung, lymph node). This analysis revealed 11 signature genes that were selectively expressed in the pancreas (e.g., pancreatic elastase-IIA). Comparison of the expression of 5,600 genes between the normal pancreas, CP, and PCa specimens showed that the expression of 34 genes was decreased in CP tissues compared with normal pancreatic tissues, and that the expression of all of these genes was simultaneously decreased in PCa. In addition, the expression of 157 genes was increased in CP tissues compared with the normal pancreas. Of those, 152 genes were simultaneously increased in PCa. Thus, only 5 of 5,600 genes were significantly overexpressed in CP compared with both normal pancreas and PCa.

Conclusions: The majority of alterations observed in CP are present in PCa, and the number of genes whose expression is selectively deregulated in CP is surprisingly small. These results may provide new insight into the pathobiology of CP and help identify certain molecular alterations that might serve as targets for new diagnostic tools and disease-specific therapy.

Figure 1. Example of a DNA array probed with RNA isolated from a normal pancreatic specimen (left panel) and a chronic pancreatitis specimen (right panel). Magnifications of corresponding areas are shown on the top. The depicted gene is SPARC/osteonectin, which exhibited an average 51-fold increase in chronic pancreatitis compared with normal tissues when all eight samples were compared.

Figure 2. Expression analysis of chronic pancreatitis and normal pancreatic tissue samples. Relative expression levels of the 152 genes with increased (A) and the 34 genes with decreased (B) expression levels in chronic pancreatitis are shown. Arbitrary expression units: green, less than 100; brown, 100 to 500; dark red, 500 to 2,500; bright red, more than 2500. The GenBank accession numbers are depicted on the right.

Figure 3. Northern blot analysis of germline oligomeric matrix protein (COMP) (A) and cysteine-rich secretory protein-3 (CRISP-3) (B) mRNA in pancreatic tissue samples obtained from normal controls (normal) and chronic pancreatitis and pancreatic cancer tissues. Total RNA (20 μg) was subjected to Northern blot analysis and probed with the ³²P-labeled COMP and CRISP-3 cDNA, respectively. The blot was subsequently rehybridized with a 7S cDNA probe to verify equivalent RNA loading. The approximate sizes of the COMP and CRISP-3 mRNA transcripts are indicated on the right.