Transcriptomic characteristics of lipopolysaccharide-induced cardiac microvascular endothelial cells

doi:10.3877/cma.j.issn.2095-1221.2020.06.002

Abstract

Abstract:

Objective

To explore the role of lipopolysaccharide (LPS) in regulating the transcriptome of rat cardiac microvascular endothelial cells (rCMECs) .

Methods

Control group (rCMECs under normal culture condition) , LPS group (rCMECs treated with 100 ng/mL LPS for 6 h) , each group was subjected to 3 repeatly transcriptome sequencing. After identified differentially expressed genes, quantitative real-time PCR (q-PCR) was used to verify mRNA expression for some of differentially expressed genes. GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment were carried out in up-regulated and down-regulated genes respectively and co-expression network analysis was conducted fordifferentially expressed genes. Statistical analysis was performed using the unpaired t-test.

Results

265 genes were up-regulated and 118 down-regulated after LPS treatment. The 10 most significant up-regulated genes were Mt2a, Cyp7b1, Sod2, Icam1, Ccl2, AC128848.1, Mt1, Cebpd, Serpinb2 and Tnfrsf11b. while the 10 most significant down-regulated genes were Cavin2, Ankrd1, Edn1, Prss35, Lmod1, Dhrs3, Ttc22, Sema6a, Map2k3 and Sema7a. The results of q-PCR showed that the expressions of Mt2a, Sod2, Ccl2, Cxcl1, Icam1 and Vcaml indeed up-regulated (P < 0.01) , and the expressions of Cavin2、Ankrd1、Edn1 and Prss35 were down regulated (P < 0.05) . The enrichment of GO and KEGG showed that up-regulated genes and inflammatory immune cellswere related tochemotaxis and adhesion of endothelial lated to calcium ion signal and G proteinsicells. The down-regulated were related to calcium signaling and G protein-related pathways as well as increased endothelial permeability. In addition, the co-expression network analysis of differentially expressed genes showed that Sod2 was the key gene, which suggested that it may be related to the changes of rCMECs induced by LPS.

Conclusion

Lipopolysaccharide regulates the large numbers of genes of inflammatory immune cells entering myocardial tissue in rCMECs.

Key words: Lipopolysaccharide, Rat, Cardiac microvascular endothelial cells, Endothelial function, Transcriptome

Yingxing Yue, Zhouxin Yang, Yan Lu, Jing Yan. Transcriptomic characteristics of lipopolysaccharide-induced cardiac microvascular endothelial cells[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2020, 10(06): 328-335.

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Figures/Tables 8

References 16

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基因	序列（5'→3'）	预期扩增产物长度（bp）
Mt2a	上游AAATGCACCTCCTGCAAGAAA	200
	下游AAGTGTGGAGAACCGGTCA
Sod2	上游GTAGAGCCTTTGCCTGTCTTAT	214
	下游CTTTGTCCATCTTGAGCACTTTC
Ccl2	上游ATCTCTCTTCCTCCACCACTAT	281
	下游AATGTACTTCTGGACCCATTCC
Cxcl1	上游ACTGTGCTAGAAGGTGTTGAG	245
	下游GACGAGAAGGAGCATTGGTTA
Icam1	上游CGGTAGACACAAGCAAGAGAAG	469
	下游GCTAAAGGTGAGCGTCCATATT
Vcaml	上游GACAGGAGACATGGTGCTAAA	226
	下游CATCCCAATGGCGGGTATTA
Gapdh	上游GGTCGGTGTGAACGGATTT	219
	下游TGGAAGATGGTGATGGGTTTC
Cavin2	上游TGCCGATCTGACCATTGAAG	103
	下游CTCCTTGGAGTTGAGCATGTAG
Ankrd1	上游GCGGACCTCAATGTCAAGAA	121
	下游CTCCTTGAGGCTGTCGAATATC
Edn1	上游TAGGAGAAGAAGGCAGGTGTA	229
	下游ACAGCAAGAAGAGGCAAGAG
Prss35	上游TAGGAGAAGAAGGCAGGTGTA	132
	下游GACAGCTTCACCGCTGTATTA

基因	序列（5'→3'）	预期扩增产物长度（bp）
Mt2a	上游AAATGCACCTCCTGCAAGAAA	200
	下游AAGTGTGGAGAACCGGTCA
Sod2	上游GTAGAGCCTTTGCCTGTCTTAT	214
	下游CTTTGTCCATCTTGAGCACTTTC
Ccl2	上游ATCTCTCTTCCTCCACCACTAT	281
	下游AATGTACTTCTGGACCCATTCC
Cxcl1	上游ACTGTGCTAGAAGGTGTTGAG	245
	下游GACGAGAAGGAGCATTGGTTA
Icam1	上游CGGTAGACACAAGCAAGAGAAG	469
	下游GCTAAAGGTGAGCGTCCATATT
Vcaml	上游GACAGGAGACATGGTGCTAAA	226
	下游CATCCCAATGGCGGGTATTA
Gapdh	上游GGTCGGTGTGAACGGATTT	219
	下游TGGAAGATGGTGATGGGTTTC
Cavin2	上游TGCCGATCTGACCATTGAAG	103
	下游CTCCTTGGAGTTGAGCATGTAG
Ankrd1	上游GCGGACCTCAATGTCAAGAA	121
	下游CTCCTTGAGGCTGTCGAATATC
Edn1	上游TAGGAGAAGAAGGCAGGTGTA	229
	下游ACAGCAAGAAGAGGCAAGAG
Prss35	上游TAGGAGAAGAAGGCAGGTGTA	132
	下游GACAGCTTCACCGCTGTATTA

分组	实验次数	Mt2a	Sod2	Ccl2	Cxcl1	Icam1
对照组	3	0.084473±0.004400	0.007357±0.000623	0.000378±0.000039	0.001666±0.000144	0.019167±0.002541
LPS组	3	1.445459±0.372817	0.076643±0.003449	0.084866±0.004514	0.239796±0.027300	0.075535±0.006341
t值		6.322	34.230	32.410	15.110	14.290
P值		0.003	< 0.001	< 0.001	0.001	0.001
分组	实验次数	Vcaml	Cavin2	Ankrd1	Edn1	Prss35
对照组	3	0.000021±8.48873E-06	0.041950±0.018844	0.115589±0.014128	0.026018±0.007298	0.001844±0.000018
LPS组	3	0.002501±0.000030	0.009144±0.000279	0.057983±0.003549	0.005615±0.001817	0.000632±0.000297
t值		137.200	3.015	7.290	5.003	5.472
P值		< 0.001	0.039	0.005	0.015	0.012

分组	实验次数	Mt2a	Sod2	Ccl2	Cxcl1	Icam1
对照组	3	0.084473±0.004400	0.007357±0.000623	0.000378±0.000039	0.001666±0.000144	0.019167±0.002541
LPS组	3	1.445459±0.372817	0.076643±0.003449	0.084866±0.004514	0.239796±0.027300	0.075535±0.006341
t值		6.322	34.230	32.410	15.110	14.290
P值		0.003	< 0.001	< 0.001	0.001	0.001
分组	实验次数	Vcaml	Cavin2	Ankrd1	Edn1	Prss35
对照组	3	0.000021±8.48873E-06	0.041950±0.018844	0.115589±0.014128	0.026018±0.007298	0.001844±0.000018
LPS组	3	0.002501±0.000030	0.009144±0.000279	0.057983±0.003549	0.005615±0.001817	0.000632±0.000297
t值		137.200	3.015	7.290	5.003	5.472
P值		< 0.001	0.039	0.005	0.015	0.012

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