Effect of locking galectin-receptor interaction on the immunopathology of small intestine of Toxoplasma gondii-infected mice

doi:10.12140/j.issn.1000-7423.2023.03.003

Abstract

Abstract:

Objective To investigate the regulatory effect of galectin-receptor interaction on the small intestine immunopathology of Toxoplasma gondii-infected mice. Methods Eighteen female BALB/c mice were randomly divided into 4 groups: 4 mice in uninfected group (naive group), 4 mice in lactose group (lactose group), 5 mice in T. gondii infection group (Tg group) and 5 mice in T. gondii infection + lactose group (Tg+lactose group). Each mouse in the Tg group and the Tg+lactose group was intraperitoneally (i.p.) injected with 1 000 tachyzoites of T. gondii RH strain, while the naive group and lactose group were i.p. injected with 0.2 ml PBS. Starting from day 0 post infection, each mouse in the Tg+lactose group and the lactose group was i.p. injected with 0.2 ml 0.2 mol/L of lactose, while each mouse in the naive group and the Tg group was i.p. injected with an equal volume of PBS, once in the morning and once in the evening for 7 consecutive days. After infection with T. gondii, the mice survival time in each group was recorded. The mice were euthanized on the 7th day after infection to collect middle segment of jejunum from each mouse for prepareing paraffin sections, which were stained with hematoxylin and eosin (HE) to observe the pathological changes; from the lower segment of jejunum of each mouse, total RNA was extracted and reverse-transcribed, and used in quantitative real-time reverse transcription PCR (qRT-PCR) with β-actin as an internal reference gene to detect the relative mRNA expression level of surface antigen 1 (SAG1), galectin-3, galectin-9, T cell immunoglobulin mucin 3 (Tim-3), leukocyte differentiation antigen 137 (CD137), interleukin 12 (IL-12), interferon-γ (IFN-γ), IL-10, IL-4, transforming growth factor β (TGF-β), chemokine receptor 2 (CCR2) and chitinase 3 like molecule 3 (Ym1). Results After infection with T. gondii, there was no mice died in the naive group and the lactose group. The survival time of the mice in the Tg group was 182-188 h, and the survival time of the mice in the Tg+lactose group was 180-182 h; the difference of the survival time between the two groups was statistically significant (χ² = 19.52, P < 0.05). HE staining showed no inflammation in the mice jejunal tissue in the naive group and lactose group. Shortened intestinal villus, shallower intestinal crypts, necrosis of epithelial cells at the top of villi and inflammatory cell infiltration in the intestinal mucosa were observed in the mice small intestine from the Tg group and Tg+lactose group. Compared with the Tg group, the pathological change of the mice small intestine in the Tg+lactose group was more severe. The qRT-PCR results showed that the relative mRNA expression of SAG1 in the mice small intestine of the Tg+lactose group was 9.17 ± 1.65, which was higher than that in the Tg group (1.00 ± 0.84, t = 4.40, P < 0.05). The relative mRNA expression levels of galectin-3 in the mce small intestine of the naive group, lactose group, Tg group, and Tg+lactose group were 1.00 ± 0.28, 1.71 ± 0.31, 2.46 ± 1.11, and 7.10 ± 1.57, respectively (F = 10.15, P < 0.01). The mRNA expression levels of galectin-9 in the 4 groups were 1.00 ± 0.31, 1.44 ± 0.26, 3.21 ± 1.01, and 7.00 ± 1.08, respectively (F = 14.53, P < 0.01). The mRNA expression levels of Tim-3 in the 4 groups were 1.00 ± 0.12, 0.88 ± 0.28, 1.64 ± 0.31, and 4.89 ± 0.69, respectively (F = 19.15, P < 0.01). The mRNA expression levels of CD137 in the 4 groups were 1.00 ± 0.42, 1.03 ± 0.30, 0.89 ± 0.11, and 3.84 ± 0.77, respectively (F = 8.46, P < 0.01). The mRNA expression levels of IL-12 in the 4 groups were 1.00 ± 0.35, 1.14 ± 0.56, 12.37 ± 4.43, and 18.42 ± 3.89, respectively (F = 10.18, P < 0.01). The mRNA expression levels of IFN-γ in the 4 groups were 1.00 ± 0.56, 1.65 ± 0.53, 5.57 ± 1.84, and 21.26 ± 6.48, respectively (F = 10.38, P < 0.01). The mRNA expression levels of IL-10 in the 4 groups were 1.00 ± 0.20, 1.10 ± 0.25, 8.65 ± 2.52, and 21.98 ± 3.96, respectively (F = 20.84, P < 0.01). The mRNA expression levels of IL-4, TGF-β, and CCR2 in the mice small intestine among the naive group, lactose group, Tg group, and Tg+lactose group had no statistically significant differences (F = 1.09, 4.74, and 2.03, P > 0.05). Ym1 mRNA expression was not detected in the naive group and lactose group, and Ym1 mRNA expression levels between the Tg group and the Tg+lactose group had no statistically significant difference (t = 0.24, P > 0.05). Conclusion Blockage of galectins-receptor interaction in mice infected with T. gondii leads to increased parasite burden in small intesting tissues, and aggravated pathological impairment, as well as upregulated expression of galectin-3, galectin-9, Tim-3, CD137, IL-10 and IFN-γ.

Key words: Toxoplasma gondii, Infected mice, Small intestine, Galectin

CLC Number:

R531.8

OU Yangran, LIU Xingzhuo, HUANG Shiguang, LYU Fangli. Effect of locking galectin-receptor interaction on the immunopathology of small intestine of Toxoplasma gondii-infected mice[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2023, 41(3): 279-285.

Figures/Tables 5

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References 27

[1]	Wu YP, Ji TK, Bai SY, et al. Progress in diagnosis of toxoplasmosis[J]. Chin J Am Infect Dis, 2021, 1-17. (in Chinese)
	(吴云萍, 祭天锴, 白邵缘, 等. 弓形虫病诊断研究进展[J]. 中国动物传染病学报, 2021, 1-17.)
[2]	Jones JL, Kruszon-Moran D, Sanders-Lewis K, et al. Toxoplasma gondii infection in the United States, 1999—2004, decline from the prior decade[J]. Am J Trop Med Hyg, 2007, 77(3): 405-410. doi: 10.4269/ajtmh.2007.77.405
[3]	Snyder LM, Denkers EY. From initiators to effectors: roadmap through the intestine during encounter of Toxoplasma gondii with the mucosal immune system[J]. Front Cell Infect Microbiol, 2020, 10: 614701. doi: 10.3389/fcimb.2020.614701
[4]	Gazzinelli RT, Hieny S, Wynn TA, et al. Interleukin 12 is required for the T-lymphocyte-independent induction of interferon gamma by an intracellular parasite and induces resistance in T-cell-deficient hosts[J]. Proc Natl Acad Sci USA, 1993, 90(13): 6115-6119. pmid: 8100999
[5]	Hunter CA, Subauste CS, Van Cleave VH, et al. Production of gamma interferon by natural killer cells from Toxoplasma gondii-infected SCID mice: regulation by interleukin-10, interleukin-12, and tumor necrosis factor alpha[J]. Infect Immun, 1994, 62(7): 2818-2824. doi: 10.1128/iai.62.7.2818-2824.1994 pmid: 7911785
[6]	Shi WK, Xue CY, Su XZ, et al. The roles of galectins in parasitic infections[J]. Acta Trop, 2018, 177: 97-104. doi: S0001-706X(16)30509-5 pmid: 28986248
[7]	Nio-Kobayashi J. Histological mapping and subtype-specific functions of galectins in health and disease[J]. Trends Glycosci Glycotechnol, 2018, 30(172): SJ47-SJ53. doi: 10.4052/tigg.1737.1SJ
[8]	Gao ZY, Liu ZN, Wang R, et al. Galectin-3 is a potential mediator for atherosclerosis[J]. J Immunol Res, 2020, 2020: 5284728.
[9]	Dong R, Zhang M, Hu QY, et al. Galectin-3 as a novel biomarker for disease diagnosis and a target for therapy (Review)[J]. Int J Mol Med, 2018, 41(2): 599-614. doi: 10.3892/ijmm.2017.3311 pmid: 29207027
[10]	Vasta GR. Roles of galectins in infection[J]. Nat Rev Microbiol, 2009, 7(6): 424-438. doi: 10.1038/nrmicro2146 pmid: 19444247
[11]	Wu YF, Lyu FL. Galectin-receptor interactions on the regulation of small intestinal pathology of Plasmodium berghei-infected mice[J]. J Trop Med, 2019, 19(5): 541-544. (in Chinese)
	(吴一凡, 吕芳丽. 半乳糖凝集素-受体相互作用对感染伯氏疟原虫小鼠小肠病理的调节[J]. 热带医学杂志, 2019, 19(5): 541-544.)
[12]	Yan JH, Lyu FL. The galectin-receptor interaction may regulate intestinal mucosal immunity of mice infected with Trichinella spiralis[J]. J Trop Med, 2021, 21(5): 540-543, 622. (in Chinese)
	(颜景海, 吕芳丽. 半乳糖凝集素-受体相互作用对旋毛虫感染小鼠肠道黏膜免疫的调节[J]. 热带医学杂志, 2021, 21(5): 540-543, 622.)
[13]	Lee JN, Kim J, Lee JH, et al. SIRT1 promotes host protective immunity against Toxoplasma gondii by controlling the FoxO-autophagy axis via the AMPK and PI3K/AKT signalling pathways[J]. Int J Mol Sci, 2022, 23(21): 13578. doi: 10.3390/ijms232113578
[14]	Zhang YX, He J, Zheng HQ, et al. Association of TREM-1, IL-1β, IL-33/ST2, and TLR expressions with the pathogenesis of ocular toxoplasmosis in mouse models on different genetic backgrounds[J]. Front Microbiol, 2019, 10: 2264. doi: 10.3389/fmicb.2019.02264 pmid: 31649630
[15]	Bernardes ES, Silva NM, Ruas LP, et al. Toxoplasma gondii infection reveals a novel regulatory role for galectin-3 in the interface of innate and adaptive immunity[J]. Am J Pathol, 2006, 168(6): 1910-1920. pmid: 16723706
[16]	Zhu C, Anderson AC, Schubart A, et al. The Tim-3 ligand galectin-9 negatively regulates T helper type 1 immunity[J]. Nat Immunol, 2005, 6(12): 1245-1252. doi: 10.1038/ni1271 pmid: 16286920
[17]	Bitra A, Doukov T, Wang J, et al. Crystal structure of murine 4-1BB and its interaction with 4-1BBL support a role for galectin-9 in 4-1BB signaling[J]. J Biol Chem, 2018, 293(4): 1317-1329. doi: 10.1074/jbc.M117.814905
[18]	Eriksen LL, Nielsen MA, Laursen TL, et al. Early loss of T lymphocyte 4-1BB receptor expression is associated with higher short-term mortality in alcoholic hepatitis[J]. PLoS One, 2021, 16(8): e0255574. doi: 10.1371/journal.pone.0255574
[19]	He J, Hou YH, Lu FL. Blockage of galectin-receptor interactions attenuates mouse hepatic pathology induced by Toxoplasma gondii infection[J]. Front Immunol, 2022, 13: 896744. doi: 10.3389/fimmu.2022.896744
[20]	Hu XH, Tang MX, Mor G, et al. Tim-3: Expression on immune cells and roles at the maternal-fetal interface[J]. J Reprod Immunol, 2016, 118: 92-99. doi: 10.1016/j.jri.2016.10.113
[21]	Wu C, Thalhamer T, Franca RF, et al. Galectin-9-CD44 interaction enhances stability and function of adaptive regulatory T cells[J]. Immunity, 2014, 41(2): 270-282. doi: 10.1016/j.immuni.2014.06.011 pmid: 25065622
[22]	Raetz M, Hwang SH, Wilhelm CL, et al. Parasite-induced TH1 cells and intestinal dysbiosis cooperate in IFN-γ-dependent elimination of Paneth cells[J]. Nat Immunol, 2013, 14(2): 136-142. doi: 10.1038/ni.2508 pmid: 23263554
[23]	Heimesaat MM, Bereswill S, Fischer A, et al. Gram-negative bacteria aggravate murine small intestinal Th1-type immunopathology following oral infection with Toxoplasma gondii[J]. J Immunol, 2006, 177(12): 8785-8795. doi: 10.4049/jimmunol.177.12.8785 pmid: 17142781
[24]	Wu B, Lyu FL. Progress of CD8⁺ T cell-mediated immune response to Toxoplasma gondii infection[J]. Chin J Parasitol Parasit Dis, 2014, 32(2): 143-147. (in Chinese)
	(吴斌, 吕芳丽. CD8⁺ T细胞免疫应答在刚地弓形虫感染免疫中的功能研究进展[J]. 中国寄生虫学与寄生虫病杂志, 2014, 32(2): 143-147.)
[25]	Lang C, Gross U, Lüder CGK. Subversion of innate and adaptive immune responses by Toxoplasma gondii[J]. Parasitol Res, 2007, 100(2): 191-203. doi: 10.1007/s00436-006-0306-9
[26]	Kobayashi M, Aosai F, Hata H, et al. Toxoplasma gondii: difference of invasion into tissue of digestive organs between susceptible and resistant strain and influence of IFN-gamma in mice inoculated with the cysts perorally[J]. J Parasitol, 1999, 85(5): 973-975. pmid: 10577740
[27]	Shin EH, Chun YS, Kim WH, et al. Immune responses of mice intraduodenally infected with Toxoplasma gondii KI-1 tachyzoites[J]. Korean J Parasitol, 2011, 49(2): 115-123. doi: 10.3347/kjp.2011.49.2.115

基因名称 Genes name	引物序列（5'→ 3'） Primer sequence(5'→ 3')	基因ID Gene ID
β⁃actin	F TGGAATCCTGTGGCATCCATGAAA	114610
β⁃actin	R TAAAACGCAGCTCAGTAACAGTCC	114610
SAG1	F CTGTAGCCTCGGAGGAAACACA	202150
SAG1	R GCCAGGTTTGTGTCCTCATGCT	202150
galectin⁃3	F AACACGAAGCAGGACAATAACTGG	168540
galectin⁃3	R GCAGTAGGTGAGCATCGTTGAC	168540
galectin⁃9	F CTGGAATCCCTCCTGTGGTGTA	168590
galectin⁃9	R CCTCGTAGCATCTGGCAAGACA	168590
Tim⁃3	F ACAGACACTGGTGACCCTCCAT	171285
Tim⁃3	R CAGCAGAGACTCCCACTCCAAT	171285
CD137	F CCAAGTACCTTCTCCAGCATAGG	219420
CD137	R GCGTTGTGGGTAGAGGAGCAAA	219420
IL⁃12	F ACGAGAGTTGCCTGGCTACTAG	161590
IL⁃12	R CCTCATAGATGCTACCAAGGCAC	161590
IFN⁃γ	F CAGCAACAGCAAGGCGAAAAAGG	159780
IFN⁃γ	R TTTCCGCTTCCTGAGGCTGGAT	159780
IL⁃10	F CGGGAAGACAATAACTGCACCC	161530
IL⁃10	R CGGTTAGCAGTATGTTGTCCAGC	161530
IL⁃4	F ATCATCGGCATTTTGAACGAGGTC	161890
IL⁃4	R ACCTTGGAAGCCCTACAGACGA	161890
TGF⁃β	F TGATACGCCTGAGTGGCTGTCT	218030
TGF⁃β	R CACAAGAGCAGTGAGCGCTGAA	218030
CCR2	F ATCCACGGCATACTATCAACATC	127720
CCR2	R CAAGGCTCACCATCATCGTAG	127720
Ym1	F CAGGTCTGGCAATTCTTCTGAA	126550
Ym1	R GTCTTGCTCATGTGTGTAAGTGA	126550