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Isolation, Characterization, and Purification of Macrophages from Tissues Affected by Obesity-related Inflammation
Published on: April 3, 2017
Difference in gene expression of macrophage between normal spleen and portal hypertensive spleen identified by cDNA
1Postdoctoral Station, Department of Hepato-Biliary Surgery, Zhongshan Hospital, Xiamen University, Xiamen 361004, Fujian Province, China. yanyisheng_ricky@tom.com
Aim:
To identify the difference in gene expression of microphage (Mphi) between normal spleen and portal hypertensive spleen using cDNA microarrays and find new gene functions associated with hypersplenism in portal hypertension.
Methods:
The Biostar-H140s chip containing 14112 spots of cDNAs were used to investigate the difference of the expression. The total RNA extracted from macrophages isolated from both normal spleen and portal hypertensive spleen was reversely transcribed to cDNA with the incorporation of fluorescent (cy3 and cy5) labeled dCTP to prepare the hybridization probes. After hybridization, the gene chip was scanned for the fluorescent intensity. The differentially expressed genes were screened. That was repeated three times, and only the genes which had differential expression in all three chips were considered to be associated with hypersplenism in portal hypertension.
Results:
Eight hundred and ninety-six, 1330 and 898 genes were identified to be differentially expressed in three chips, respectively. One hundred and twenty-one genes (0.86%) were identified to be differentially expressed in all three chips, including 21 up-regulated genes and 73 down-regulated genes. The differentially expressed genes were related to ionic channel and transport protein, cyclin, cytoskeleton, cell receptor, cell signal conduct, metabolism, immune, and so on. These genes might be related to the hypersplenism in portal hypertension.
Conclusion:
The investigations based on cDNA microarray can screen differentially expressed genes of macrophages between normal spleen and portal hypertensive spleen, thus may provide a new idea in studying the pathogenesis of hypersplenism in portal hypertension.
Insights
This study used cDNA microarrays to find gene expression differences in macrophages from normal and portal hypertensive spleens. It identified 121 differentially expressed genes potentially linked to hypersplenism in portal hypertension.
Area of Science:
- Genomics
- Molecular Biology
- Immunology
Background:
- Portal hypertension is a condition characterized by increased pressure in the portal venous system.
- Hypersplenism, or an overactive spleen, is a common complication of portal hypertension.
- The specific molecular mechanisms underlying hypersplenism in portal hypertension remain incompletely understood.
Purpose of the Study:
- To identify differences in gene expression profiles of macrophages between normal and portal hypertensive spleens.
- To discover novel genes and pathways associated with the development of hypersplenism in portal hypertension.
- To utilize cDNA microarray technology for high-throughput gene expression analysis.
Main Methods:
- Macrophages were isolated from both normal and portal hypertensive spleens.
- Total RNA was extracted and converted to fluorescently labeled cDNA probes.
- Differential gene expression was analyzed using the Biostar-H140s cDNA microarray chip (14112 spots).
- Genes showing consistent differential expression across three independent experiments were identified.
Main Results:
- A total of 121 genes (0.86%) exhibited differential expression in macrophages from portal hypertensive spleens compared to normal spleens.
- Among these, 21 genes were up-regulated and 73 were down-regulated.
- The identified genes are involved in various functions, including ionic channel transport, cell cycle regulation (cyclin), cytoskeleton organization, cell signaling, and immune responses.
Conclusions:
- cDNA microarray analysis is effective in identifying differentially expressed genes in macrophages from spleens affected by portal hypertension.
- The identified genes provide potential molecular targets and insights into the pathogenesis of hypersplenism.
- This research offers a novel approach to understanding the complex mechanisms of hypersplenism in portal hypertension.