J Korean Med Sci.  2022 Dec;37(48):e338. 10.3346/jkms.2022.37.e338.

Increased Pro-Inflammatory T Cells, Senescent T Cells, and Immune-Check Point Molecules in the Placentas of Patients With Gestational Diabetes Mellitus

• Kang YE
• Yi HS
• Yeo MK
• Kim JT
• Park D
• Jung Y
• Kim OS
• Lee SE
• Kim JM
• Joung KH
• Lee JH
• Ku BJ
• Lee M
• Kim HJ

• KMID: 2536732
• DOI: http://doi.org/10.3346/jkms.2022.37.e338

Abstract

Background
Gestational diabetes mellitus (GDM) is the most common metabolic complication of pregnancy. To define the altered pathway in GDM placenta, we investigated the transcriptomic profiles from human placenta between GDM and controls.
Methods
Clinical parameters and postpartum complications were reviewed in all participants. Differentially expressed canonical pathways were analyzed between the GDM and control groups based on transcriptomic analysis. CD4 ⁺ T, CD8 ⁺ T, and senescent T cell subsets were determined by flow cytometry based on staining for specific intracellular cytokines.
Results
Gene ontology analysis revealed that the placenta of GDM revealed upregulation of diverse mitochondria or DNA replication related pathways and downregulation of T-cell immunity related pathways. The maternal placenta of the GDM group had a higher proportion of CD4 ⁺ T and CD8 ⁺ T cells than the control group. Interestingly, senescent CD4 ⁺ T cells tended to increase and CD8 ⁺ T cells were significantly increased in GDM compared to controls, along with increased programmed cell death-1 (CD274 ⁺ ) expression. Programmed death-ligand 1 expression in syncytotrophoblasts was also significantly increased in patients with GDM.
Conclusion
This study demonstrated increased proinflammatory T cells, senescent T cells and immune-check point molecules in GDM placentas, suggesting that changes in senescent T cells and immune-escape signaling might be related to the pathophysiology of GDM.

Keyword

Gestational Diabetes; Placenta; RNA-Sequencing; T Cell Immunity; Senescent T Cells; Immune-Check Point Molecules

Figure

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  Fig. 1 Clinical information and differentially expressed canonical pathways in comparison between GDM patients and controls. (A) Clinical information including age, BMI, glycosylated hemoglobin (HbA1c), BMI, birth weight of infants, and weight gain. (B) DEGs in comparison between GDM patients and controls by volcano plot analysis. (C) Heatmap analysis of DEGs. (D) Upregulated differentially expressed canonical pathways in comparison between GDM patients and controls. (E) Downregulated differentially expressed canonical pathways in comparison between GDM patients and controls.GDM = gestational diabetes mellitus, BMI = body mass index, HbA1c = hemoglobin A1c, DEG = differentially expressed gene.

• 

  Fig. 2 Immunophenotype of CD4+ T cells of the maternal placenta in pregnant women with and without GDM. (A) Representative flow cytometry plots are presented for CD45RA and CD45RO expression by CD4+ T cells in patients with GDM (n = 6) and normal controls (n = 8). Statistical analysis of the population of CD45RA+CD45RO− (naïve) or CD45RA−CD45RO+ (memory) T cells in CD4+ T cells in the two groups. (B, C) The frequency of IFN-γ- and IL-17A-secreting cells in the population of CD4+ T cells was compared between the two groups. Data are expressed as mean ± standard error of the mean. Flow cytometry plots are representative of all independent experiments.GDM = gestational diabetes mellitus, IFN = interferon, IL = interleukin, FSC = forward scatter.*P < 0.05 compared with the corresponding controls.

• 

  Fig. 3 Immunophenotype of CD8+ T cells of the maternal placenta in pregnant women with and without GDM. (A) Representative flow cytometry plots are presented for CD45RA and CD45RO expression by CD8+ T cells in patients with GDM (n = 6) and normal controls (n = 8). Statistical analysis of the population of CD45RA+CD45RO− (naïve) or CD45RA−CD45RO+ (memory) T cells in CD8+ T cells in the two groups. (B) The frequency of IFN-γ-secreting cells in the population of CD8+ T cells was compared between the two groups. Data are expressed as mean ± standard error of the mean. Flow cytometry plots are representative of all independent experiments.GDM = gestational diabetes mellitus, IFN = interferon, IL = interleukin, FSC = forward scatter.*P < 0.05 compared with the corresponding controls.

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  Fig. 4 Population size of CD28−CD57+ senescent T cells from normoglycemic subjects and patients with GDM. (A, B) Representative flow cytometry plots are presented for CD57 and CD28 expression by CD4+ or CD8+ T cells in patients with GDM (n = 6) and normal controls (n = 8). Statistical analysis of the population of CD28− and CD57+ T cells in CD4+ and CD8+ T cells in the two groups. (C, D) CD279 (PD-1) expression of placental CD4+ and CD8+ T cell subsets from patients with GDM and normal controls. Data are expressed as mean ± standard error of the mean. Flow cytometry plots are representative of all independent experiments.GDM = gestational diabetes mellitus, PD-1 = programmed cell death-1.*P < 0.05 compared with the corresponding controls.

• 

  Fig. 5 Dysregulated immune-escape signaling in placenta from GDM participants. (A) Representative images of H&E statin and PD-L1 expression between control and GDM. PD-L1 were scored as weak, moderate, and marked. Scattered PD-L1 positive lymphocytes in implantation site of decidua. (B) Representative images of H&E statin and PD-1 expression between control and GDM. PD-1 were scored as negative, weak, and moderate. The number of PD-1 positive lymphocytes were counted per HPF. (C) mRNA expressions of genes related with PD-1, PD-L1, PD-L2, and CTLA-4 between two groups (Control vs. GDM). (D) Representative images of Western blot analysis about p-STAT1, STAT1, p-STAT3, STAT3, p-PI3K, PI3K. Comparison of densitometry of blot images between two groups (Control vs. GDM).H&E = hematoxylin and eosin, PD-L1 = programmed death-ligand 1, GDM = gestational diabetes mellitus, PD-1 = programmed cell death-1, HPF = high-power field, CTLA-4 = cytotoxic T cell antigen-4.*P  < 0.05 compared with the corresponding controls.

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