Investigation of differentially expressed genes in liver tissues of patients with alveolar echinococcosis

doi:10.12140/j.issn.1000-7423.2021.01.007

Abstract

Abstract:

Objective To investigate the expression profile of differentially expressed genes in liver lesions of alveolar echinococcosis patients, based on the findings that these genes were indentified in mice infected with alveolar metacestode. Methods The gene chip dataset (No. GSE24376) in the high-throughput gene expression database was selected from the gene expression omnibus (GEO), to compare the genes with differential expression between infected and uninfected mice. The differentially expressed genes underwent gene ontology (GO) analysis. Lesion tissues and adjacent tissues 2 cm from the lesion were collected from 6 patients with E. multilocularis cyst infection. Fluorescence quantitative PCR was performed to detect the relative mRNA levels of two highly expressed differential genes. Forty-five paraffin-embedded liver tissue samples from patients with multilocular hydatid infection (29 samples with severe inflammatory response and liver fibrosis, and 16 with relatively normal liver tissue with less inflammatory cell infiltration and liver fibrosis) underwent immunohistochemistry to analyze the expression of the two highly expressed differential genes. Statistical analysis was performed on the data with SPSS 21.0 statistical software. Results As compared to the normal mice, 21 differentially expressed genes were found, including 9 overexpressed genes and 12 downregulated genes in liver tissues of infected mice. The results of GO analysis showed that the differentially expressed genes were expressed in vesicle lumen, cytoplasmic vesicle lumen and extracellular matrix, participated in biological processes of neutrophil degranulation and activation, and possessed CC chemokine receptor binding activity and chemokine activity. The fluorescence quantitative PCR results showed that the mRNA expression levels of CHI3L3 (CHI3L1 in humans) and CCL8, which had higher differential expression, were higher in the liver lesion tissues than in the surrounding tissues of the 6 patients with multilocular hydatid infection (P < 0.01), reaching an utmost 7 and 9 folds, respectively. The results of immunohistochemistry showed that among the 29 lesion samples with severe inflammatory reaction and liver fibrosis, 20 samples (69%) showed high expression of CHI3L1 protein, with staining mainly located in the cytoplasm, and 22 samples showed high levels of CCL8 protein (76%). Of the 16 relatively normal liver tissue samples, only 2 (13%) were positive for CHI3L1 protein, and only 4 were positive for CCL8 protein (33%). In the lesions with severe inflammatory response and liver fibrosis, the positive expression rate of CHI3L1 and CCL8 proteins reached 96% and 97%, respectively, which were higher than 3% and 8% in normal liver tissues (P < 0.01). Conclusion The expression of CHI3L1 and CCL8 was upregulated in the liver tissues of patients with E. multilocularis, suggesting that they play a role in the immune response to parasites.

Key words: Echinococcus multilocularis, Infection, Differentially expressed genes, CHI3L3, CCL8

CLC Number:

R383.33

ZHU Ji-hai, CAO De-ping, ZHAO Jun, LIU Jun, SHI Hu-xiang, LIU Yan. Investigation of differentially expressed genes in liver tissues of patients with alveolar echinococcosis[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2021, 39(1): 48-54.

Figures/Tables 5

Table 1

Table 2

Differentially expressed genes in GSE24376 gene chip between Echinococcus multilocularis-infected and healthy mice

差异表达基因 Differentially expressed genes	倍数变化 Fold change	P值 P value	注释 Annotation
CHI3L3	6.53	5.57E-07	几丁质酶3-3 Chitinase 3-like 3
CCL8	4.84	4.42E-06	C-C基序趋化配体8 C-C motif chemokine ligand 8
RETNLA	4.82	2.85E-05	抵抗素-α Resistin like alpha
SLPI	3.31	2.58E-04	分泌白细胞肽酶抑制剂 Secretory leukocyte peptidase inhibitor
CCL12	2.77	3.80E-04	趋化因子(C-C基序)配体12 Chemokine (C-C motif) ligand 12
CLEC4D	2.04	6.35E-04	c型凝集素域家族4成员D C-type lectin domain family 4 member D
EAR11	2.02	3.45E-04	嗜酸粒细胞相关,核糖核酸酶A家族,成员11 Eosinophil-associated, ribonuclease A family, member 11
UBD	1.93	6.82E-04	泛素D Ubiquitin D
CCL17	1.57	7.62E-04	C-C基序趋化配体17 C-C motif chemokine ligand 17
HSD3B4	-4.66	1.46E-05	3-和甾体德尔塔-异构酶4 3 beta-and steroid delta-isomerase 4
SULT5A1	-3.55	2.34E-04	磺基转移酶家族5A,成员1 Sulfotransferase family 5A, member 1
EGFR	-3.06	1.66E-04	表皮生长因子受体 Epidermal growth factor receptor
A1BG	-2.97	3.78E-04	alpha-1-B糖蛋白 Alpha-1-B glycoprotein
CYP17A1	-2.64	7.22E-05	细胞色素P450家族17亚家族A成员1 Cytochrome P450 family 17 subfamily A member 1
AHSG	-2.27	1.14E-04	α2-HS糖蛋白 Alpha 2-HS glycoprotein
SERPINA1D	-2.25	2.64E-04	丝氨酸（或半胱氨酸）肽酶抑制剂,分支A,成员1D Serine (or cysteine) peptidase inhibitor, clade A, member 1D
APOE	-2.23	1.88E-04	载脂蛋白e Apolipoprotein E
ALB1	-1.80	6.16E-04	阿宾娜1 ALBINA 1
NSG1	-1.79	7.12E-04	神经元小泡转运相关1 Neuronal vesicle trafficking associated 1
2310005P05RIK	-1.62	6.78E-04	内切酶/外切酶/磷酸酶家族含域蛋白1 Endonuclease/exonuclease/phosphatase family domain containing 1
FBP1	-1.54	8.26E-04	果糖二磷酸酶1 Fructose-bisphosphatase 1

Table 2

Fig. 1

Fig. 2

Fig. 3

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基因名称Gene name	引物序列（5′ →3′） Primer sequence (5′ →3′)
CCL8	F：AACATGAAGGTCTCCGCTGG
	R：GCAGCAGGTGATTGGGGTAG
CHI3L1	F：GTTTTCCCAGTCACGAC
	R：CAGGAAACAGCTATGAC
β-actin	F：TTAGTTGCGTTACACCCTTTC
	R：ACCTTCACCGTTCCAGTTT