cd4 central memory t cells miltenyi biotech  (Miltenyi Biotec)

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Schematic of the experimental workflow in CyTOF experiment. The placentas were obtained from individuals with normal pregnancy (NP, n=9), preeclampsia (PE, n=8), gestational diabetes mellitus (GDM, n=8) or GDM&PE (n=7). (B) t-SNE maps showing 7×10 5 CD45 + cells from the placenta overlaid with color-coded clusters and the distributions of B cells, CD4 + T cells, CD8 + T cells, DC, γδT cells, monocytes, granulocytes, MDSC, and NK cells. (C) Percentages of each cell type of CD45 + cells in placentas. (D) t-SNE maps showing the expression of CD3, CD8, CD4, γδTCR, CD14, CD15, CD56. (E) Heatmap showing the expression levels of markers in CD45 + cell subsets. Data were compared between NP and PE, NP and GDM, NP and GDM&PE using the Shapiro-wilk test and represented as mean±SEM (*P < 0.05, ** P < 0.01, *** P < 0.001; NS, not significant).

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Expressing

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Distribution of the CD4 + T cells analyzed using t-SNE. (B) Scatter dot plots showing the frequencies of cluster 8 of CD4 + T cells in the placentas of individuals with NP, PE, GDM and GDM&PE. (C) Expression of IL-17A and Foxp3 in CD45RO + CCR7 + CD4 + T cells in placentas of individuals with NP and PE using flow cytometry. (IL-17A: n=10 in NP group, n=15 in PE group; Foxp3: n=7 in NP group, n=9 in PE group). (D) Immunofluorescence co-staining of CD4 (red), CD45RO (green) and DAPI (blue) in frozen placental sections. The right panels show the enlargement of the dotted box in the left panel. Scale bar, 20 µm. (E) Scatter dot plots showing significantly altered markers of in cluster 8 of CD4 + T cells. (F) Distribution of the CD8 + T cells analyzed using t-SNE. (G) Scatter dot plots showing the frequencies of cluster 2 of CD8 + T cells in the placentas of individuals with NP, PE, GDM and GDM&PE. (H) Scatter dot plots showing significantly altered markers in cluster 2 of CD8 + T cells. (I) Distribution of the γδT cells analyzed using t-SNE. (J) Scatter dot plots showing the frequencies of cluster 15 of γδT cells in the placentas of individuals with NP, PE, GDM and GDM&PE. (K) Scatter dot plots showing significantly altered markers in cluster 15 of γδT cells. Data were compared between NP and PE, NP and GDM, NP and GDM&PE using the Shapiro-wilk test and represented as mean±SEM (*P < 0.05, ** P < 0.01, *** P < 0.001; NS, not significant).

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Expressing, Flow Cytometry, Immunofluorescence, Staining

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Heatmap showing the expression levels of markers in the CD4 + T subsets. (B) Heatmap showing the expression levels of markers in the CD8 + T cells. (C) Heatmap showing the expression levels of markers in the γδT cells.

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Expressing

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Schematic of the mice model adoptively transferred CD45 + F4/80 + CD206 - pro-inflam Macs or CD45 + F4/80 + CD206 + anti-inflam Macs. (B) Principal component analysis (PCA) reflected the differences between the two groups of macrophages (n=3). (C) The volcano map shows a comparison of the content and P value of gene expression between pro-inflam Macs and anti-inflam Macs. Differential expression genes were screened out when P < 0.05. Red dots indicate genes with increased expression in pro-inflam Macs. Blue dots indicate genes with decreased expression. (D) The volcano map shows differential expression genes between pro-inflam Macs and anti-inflam Macs. (E) Representative pathways enriched in the identified genes as determined by GSEA (p value<0.05). (F) Embryo abortion rate of the pregnant mice, body weight and crown-rump length of pups measured on day 18.5 of gestation. Black represents mice treated with DMSO (n=8); gray represents mice treated with PLX3397 (n=8); blue represents mice injected with CD45 + F4/80 + CD206 + anti-inflammatory macrophages (n=8); red represents mice injected with CD45 + F4/80 + CD206 - pro-inflammatory macrophages (n=8). (G) SBP and UACR of pregnant mice in the four groups. (H) Frequencies of CD44 + CD4 + IL-17A + cells, CD44 + CD8 + T cells and CD11b + Ly6G + granulocytes analyzed by flow cytometry.

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Comparison, Expressing, Injection, Flow Cytometry

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Embryo abortion rate of the pregnant mice, body weight and crown-rump length of pups measured on day 18.5 of gestation. Black represents mice treated with control liposomes (n=6); gray represents mice treated with clodronate liposomes (n=6); blue represents mice injected with CD45 + F4/80 + CD206 + anti-inflam Macs (n=6); red represents mice injected with CD45 + F4/80 + CD206 - pro-inflam Macs (n=6). (B) SBP and UACR of pregnant mice in the four groups. (C) Frequencies of CD44 + CD4 + IL-17A + cells, CD44 + CD8 + T cells and CD11b + Gr1 + granulocytes analyzed by flow cytometry.

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Liposomes, Injection, Flow Cytometry

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) UMAP maps showing the 12 clusters of mouse T/NK cells was listed in the up panel. Bar graph showing the frequencies of clusters of T/NK cells in the two groups of mice was listed in the down panel. (B) Heatmap showing clustering analysis for markers distinguished 12 different clusters of T/NK cells. (C) UMAP maps showing the distribution of specific markers of cluster 0, 1 and 2. (D) Dot plot depicting GO enrichment terms that were significantly enriched in the differentially expressed genes in cluster 0 from the pro-inflam Macs group and the control group. (E) Violin plot of specific differential gene expression in cluster 0 between the pro-inflam Macs group and the control group. (F) Frequencies of CD4 + CD44 + T cells and the percentages of IL-17A + cells in CD4 + CD44 + T cells at the maternal-fetal interface in Sham and RUPP group analyzed by flow cytometry. (G) The embryo abortion rate of pregnant mice, body weight, and crown-rump length of pups measured on day 18.5 of gestation in mice injected PBS, Sham mouse-derived or RUPP mouse-derived CD4 + CD44 + T cells. Black represents mice injected with PBS (n=6); blue represents mice injected with Sham mouse-derived CD4 + CD44 + T cells (n=6); red represents mice injected with RUPP mouse-derived CD4 + CD44 + T cells (n=6). (H) SBP and UACR of pregnant mice injected with PBS, Sham mouse-derived or RUPP mouse-derived CD4 + CD44 + T cells.

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Expressing, Flow Cytometry, Injection, Derivative Assay

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Experimental design of mice model with PE by reducing uterine perfusion pressure (RUPP) on day 12.5 of gestation. Mice with sham operation were considered as controls. Systolic blood pressure (SBP) and urine albumin creatine ratio (UACR) were measured on day 12.5 and 16.5 of gestation respectively. Mice were sacrificed on day 18.5 of gestation. (B) The embryo abortion rate of pregnant mice, body weight, and crown-rump length of pups measured on day 18.5 of gestation in the Sham and RUPP group. Embryo abortion rate = number of absorbed embryos/total number of embryos. Blue represents mice in the Sham group (n=9); Red represents mice in the RUPP group (n=9). (C) SBP and UACR of pregnant mice in Sham and RUPP group. (D) Embryo abortion rate of pregnant mice, body weight and crown-rump length of pups measured on day 18.5 of gestation. Black represents mice with previous normal pregnancy (n=9); gray represents mice with previous pregnancy with PE (n=9). (E) SBP and UACR of second pregnant mice with a history of PE or NP in the first pregnancy measured on day 16.5 of gestation. (F) Frequencies of CD4 + CD44 + T cells and the levels of IL-17A in CD4 + CD44 + T cells in mice with a pregnancy history with NP and PE analyzed by flow cytometry. Data were compared using the Student’s t-test and represented as mean±SEM (*P < 0.05, ** P < 0.01, *** P < 0.001; NS, not significant).

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: Flow Cytometry

Journal: bioRxiv

Article Title: Preeclampsia-specific immune cell network in placenta revealed by Cytometry by time of flight and single-cell RNA-seq

doi: 10.1101/2024.06.05.597577

Figure Lengend Snippet: (A) Signaling modules indicated by ligand-receptor pairing between macrophages and other types of immune cells at the maternal-fetal interface using CellPhoneDB. (B) Frequencies of IGF1 + CD14 + cells and IGFIR + CD4 + cells in placentas of individuals with NP and PE (n=12 in NP group, n=4 in PE group). (C) Schematic of the experimental workflow to induce memory-like T cells in vitro. Macrophages, after incubating with PBS, NP-EVs or PE-EVs, were co-cultured with CD4 + naïve T cells treated with DMSO or BMS-754807. Cells were isolated from human peripheral blood. (D) Frequencies of CD45RO + CCR7 + Th17 cells. Black represents CD4 + naïve T cells treated with DMSO; gray represents CD4 + naïve T cells cocultured with NP-EVs-treated macrophages; blue represents CD4 + naïve T cells co-cultured with PE-EVs-treated macrophages; red represents CD4 + naïve T cells treated with BMS-754807 before co-cultured with PE-EVs-treated macrophages (n=10 in each group). (E) Schematic of mice transferred CD4 + T cells treated with BMS-754807 or PBS. Anti-CD4 antibody was used to deplete CD4 + T cells in mice on day 10.5 of gestation. CD4 + T cells were transferred into mice on day 11.5 of gestation. 20 µg/kg lipopolysaccharide (LPS) was intraperitoneally injected on day 12.5 and 15.5 of gestation to induce a PE-like pregnant mice model. Mice were sacrificed on day 18.5 of gestation. (F) Embryo abortion rate of pregnant mice, body weight and crown-rump length of pups were measured on day 18.5 of gestation. Black represents the control group mice (n=6); gray represents mice treated with LPS (20μg/kg) to construct an animal model of PE (n=6); blue represents anti-CD4 antibody treated PE mice (n=6); red represents anti-CD4 antibody treated PE mice injected with CD4 + T cells with DMSO treatment (n=7); orange represents anti-CD4 antibody treated PE mice injected with CD4 + T cells with BMS754807 treatment (n=7). (G) SBP and UACR of pregnant mice in the five groups. (H) The frequencies of CD4 + CD44 + IL-17A + cells analyzed by flow cytometry. The results were compared using one-way ANOVA and represented as mean±SEM (*P < 0.05, ** P < 0.01, *** P < 0.001; NS, not significant).

Article Snippet: Human CD4 + memory T cells were isolated using a Human Central and Effector Memory CD4 + T Cell Isolation Kit (17865, STEMCELL, Canada).

Techniques: In Vitro, Cell Culture, Isolation, Injection, Construct, Animal Model, Flow Cytometry