Journal: Frontiers in Molecular Biosciences

Article Title: JAK-centric explainable few-shot gene-expression diagnosis framework for alopecia via MultiPLIER priors and relation-style set-to-set comparison

doi: 10.3389/fmolb.2025.1753206

Figure Lengend Snippet: RT-qPCR validation of the IL2RB/IL2RG–EOMES–GZMA cytotoxic/JAK axis in alopecia areata (AA). Relative mRNA levels of EOMES, GZMA, IL2RB, and IL2RG are significantly elevated in AA lesional scalp compared with healthy controls, whereas androgenetic alopecia (AGA) samples are comparable to controls, indicating that this cytotoxic/JAK module is transcriptionally active in AA but largely quiescent in AGA. Data are shown as mean ± SD. Statistical annotations: ** p < 0.01 vs. control; n.s., not significant (AGA vs. control).

Article Snippet: Equal amounts of protein were resolved by SDS–PAGE and transferred to PVDF membranes at 200 mA for 2 h. Membranes were blocked in 5% non-fat milk at room temperature for 2 h and incubated overnight at 4 ° C with primary antibodies against β -actin (1:4000; 20536-1-AP, Proteintech, China), EOMES (1:5000; 83945-5-RR, Proteintech, China), GZMA (1:500; 11288-1-AP, Proteintech, China), IL2RB (1:5000; 13602-1-AP, Proteintech, China), and IL2RG (1:500; 11409-1-AP, Proteintech, China).

Techniques: Quantitative RT-PCR, Biomarker Discovery, Control

Journal: Frontiers in Molecular Biosciences

Article Title: JAK-centric explainable few-shot gene-expression diagnosis framework for alopecia via MultiPLIER priors and relation-style set-to-set comparison

doi: 10.3389/fmolb.2025.1753206

Figure Lengend Snippet: GSEA plots for key genes. (a) EOMES. (b) GZMA. (c) IL2RB. (d) IL2RG.

Article Snippet: Equal amounts of protein were resolved by SDS–PAGE and transferred to PVDF membranes at 200 mA for 2 h. Membranes were blocked in 5% non-fat milk at room temperature for 2 h and incubated overnight at 4 ° C with primary antibodies against β -actin (1:4000; 20536-1-AP, Proteintech, China), EOMES (1:5000; 83945-5-RR, Proteintech, China), GZMA (1:500; 11288-1-AP, Proteintech, China), IL2RB (1:5000; 13602-1-AP, Proteintech, China), and IL2RG (1:500; 11409-1-AP, Proteintech, China).

Techniques:

Journal: Frontiers in Molecular Biosciences

Article Title: JAK-centric explainable few-shot gene-expression diagnosis framework for alopecia via MultiPLIER priors and relation-style set-to-set comparison

doi: 10.3389/fmolb.2025.1753206

Figure Lengend Snippet: Western blot validation of the IL2RB/IL2RG–EOMES–GZMA cytotoxic/JAK axis at the protein level. Protein expression of EOMES, GZMA, IL2RB, and IL2RG is markedly upregulated in AA lesional scalp compared with healthy controls, concordant with the RT-qPCR results, whereas AGA-affected scalp shows no significant difference relative to controls. Representative immunoblots and densitometric quantification (normalized to β -actin) are shown. Data are presented as mean ± SD. Statistical annotations: ** p < 0.01 vs. control; n.s., not significant (AGA vs. control).

Article Snippet: Equal amounts of protein were resolved by SDS–PAGE and transferred to PVDF membranes at 200 mA for 2 h. Membranes were blocked in 5% non-fat milk at room temperature for 2 h and incubated overnight at 4 ° C with primary antibodies against β -actin (1:4000; 20536-1-AP, Proteintech, China), EOMES (1:5000; 83945-5-RR, Proteintech, China), GZMA (1:500; 11288-1-AP, Proteintech, China), IL2RB (1:5000; 13602-1-AP, Proteintech, China), and IL2RG (1:500; 11409-1-AP, Proteintech, China).

Techniques: Western Blot, Biomarker Discovery, Expressing, Quantitative RT-PCR, Control

Journal: Frontiers in Immunology

Article Title: In Vivo Priming of Peritoneal Tumor-Reactive Lymphocytes With a Potent Oncolytic Virus for Adoptive Cell Therapy

doi: 10.3389/fimmu.2021.610042

Figure Lengend Snippet: Vaccinia virus expressing IL15/IL15Rα (vvDD-IL15/Rα) treatment leads to enrichment of memory CD8 + T cells with an activated phenotype. Immunocompetent B6 mice were inoculated IP with 5.0x10 5 MC38-luc cells (day −9), randomized and treated with intraperitoneal (IP) injection of PBS, vvDD or vvDD-IL15/Rα at a dose of 5.0x107 pfu/mouse (day 0). Ten days following i.p. oncolytic virotherapy, peritoneal CD8 + T cells were isolated via peritoneal lavage and characterized via flow cytometry (in addition to day 3, 6, 8, and 14 for figure A). (A) Regional delivery of vvDD-IL15/Rα greatly enhanced and prolonged peritoneal CD8 + T cell infiltration on day 6 to 14 following virotherapy. (B) The amount of peritoneal central memory (Tcm, CD44 + CD62L + ) and effector memory (Tem, CD44 + CD62L - ) T cells is increased 10 days following vvDD-IL15/Rα injection. (C) Representative flow cytometry charts of memory CD8 + T cell subsets. vvDD-IL15Rα treatment resulted in increased Ki67 (D) , CXCR3 (E) , CD122 (F) , Tim3 - PD1 int (G) , CD103 (H) , and CD44 + PD-1 + (I) staining compared to vvDD or PBS 10 days following injection. PD-1 high (J) , TIGIT + (K) , and Tim-3 + (L) staining were not increased after vvDD-IL15/Rα treatment compared to vvDD or PBS treatment. Representative flow cytometry charts of PD-1 expression by treatment group are depicted in . N=8–12. All values presented as mean ± SEM. *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. Data are combined from two independent experiments.

Article Snippet: FITC conjugated anti-human CD122 (clone REA167) was purchased from Miltenyi Biotec.

Techniques: Virus, Expressing, Injection, Isolation, Flow Cytometry, Staining

Journal: Frontiers in Immunology

Article Title: In Vivo Priming of Peritoneal Tumor-Reactive Lymphocytes With a Potent Oncolytic Virus for Adoptive Cell Therapy

doi: 10.3389/fimmu.2021.610042

Figure Lengend Snippet: Characterization of human peritoneal CD8 + and CD4 + T cells derived from peritoneal metastasized tumors. Peritoneal fluid was collected from 14 patients undergoing peritoneal surgery, and peritoneal lymphocytes were extracted and characterized. (A) Flowchart of flow cytometry gating to identify peritoneal CD4 and CD8 T cells. Single cells gating, dead cells exclusion via Zombie aqua staining. Subsequently CD3 + cells were separated into CD4 + or CD8 + cells. (B) Distribution of naïve T cells (Tn, CD45RO - CCR7 + ), central memory T cells (Tcm, CD45RO + CCR7 + ), effector memory T cells (Tem, CD45RO + CCR7 - ) and highly differentiated effector T cells (Temra, CD45RO - CCR7 - ) gated from CD8 + T cells. (C) Representative flow cytometry chart of peritoneal memory CD8 + T cell subsets. (D) Levels of 4-1BB and Ox40 on CD8 + T cells were analyzed. (E–H) Analysis of PD-1 and Tim-3 demonstrated the majority of CD8 + cells were PD-1 high and an average of 20% expressed Tim-3. Representative flow charts of PD-1 and Tim-3 expression are shown in (F, H) respectively. (I, J) (representative flow chart): The shared beta-subunit of the IL-2- and IL15-receptor (CD122) showed a mean expression in peritoneal CD8 + T cells of 29.1%. (K–M) Peritoneal CD8 + T cells from 7 patients were tested for secretion of cytotoxic IFN-γ by ELISA (K) and 4-1BB surface expression by flow (L, M) (representative images) after overnight coculture with correlating tumor digest. Monocytes were used as negative controls, CD3/CD28 dynabeads as a positive control. All values presented as mean ± SEM. *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001.

Article Snippet: FITC conjugated anti-human CD122 (clone REA167) was purchased from Miltenyi Biotec.

Techniques: Derivative Assay, Flow Cytometry, Staining, Expressing, Enzyme-linked Immunosorbent Assay, Positive Control

Journal: Molecular Medicine Reports

Article Title: Expression, immunogenicity and clinical significance analysis of thyroid-stimulating hormone receptor fusion proteins

doi: 10.3892/mmr.2025.13639

Figure Lengend Snippet: Changes of specific regulatory T cell subsets in mice after immunization. (A) Flow cytometry analysis of changes in CD4 + CD25 + T cells in the hTSHR289, hTSHR290 and hTSHR410 groups after immunization. (B) Flow cytometry analysis of changes in CD8 + CD122 + T cells in the hTSHR289, hTSHR290 and hTSHR410 groups after immunization. *P<0.05 (One-way ANOVA with Tukey's post hoc test. hTSHR, human thyroid-stimulating hormone receptor.

Article Snippet: The analyte detectors were as follows: CD3 + antibody (cat. no. 565643; Becton, Dickinson and Company), CD4 + antibody (cat. no. F21004A02; Multi Sciences Biotech), CD8 + antibody (cat. no. F2100801; Multi Sciences Biotech), CD25 antibody (cat. no. E-AB-F1102C; Wuhan Elabscience Biotechnology Co., Ltd.) and CD122 antibody (cat. no. E-AB-F1029D; Wuhan Elabscience Biotechnology Co., Ltd.).

Techniques: Flow Cytometry

Journal: Cells

Article Title: Tolerance Induced by Antigen-Loaded PLG Nanoparticles Affects the Phenotype and Trafficking of Transgenic CD4 + and CD8 + T Cells

doi: 10.3390/cells10123445

Figure Lengend Snippet: Antigen-encapsulating PLG/GP33 nanoparticles expand CD8 + Tregs in transgenic P14 mice. 3–4 female P14 mice were injected intravenously with either PLG/GP33 or a control PLG (PLG/OVA323 or PLG). Spleens and/or lymph nodes (pooled axillary, brachial, and inguinal) were harvested and counted 3 days later. Viable T cells were analyzed by flow cytometry. Statistical significance was determined by Student’s t -test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ( A ) Expression of FoxP3, CD28, or CD122 was determined by geometric mean fluorescence intensity (MFI). ( B ) Sub-populations were determined by percentage of viable CD8 + cells that were double positive for CD122 and PD-1, Ly49, or CD38. ( C ) Female P14 mice ( n = 3) at various ages (young 4 weeks of age, adult 8 WOA, old 12 WOA) were injected intravenously with either PLG/GP33 or PLG/OVA323. Expansion of Tregs was determined by percentage of all viable CD8 + T cells that were double positive for CD122 and PD-1 by flow cytometry. Statistical significance between the groups was determined by considering three ages as three replicates.

Article Snippet: CD122 + cells were positively selected and removed from whole spleens by first staining with biotin conjugated anti-CD122 monoclonal antibody and then removing the stained cells with a biotin positive selection kit (STEMCELL Technologies, Vancouver, BC, Canada) according to the manufacturer’s instructions.

Techniques: Transgenic Assay, Injection, Control, Flow Cytometry, Expressing, Fluorescence

Journal: Cells

Article Title: Tolerance Induced by Antigen-Loaded PLG Nanoparticles Affects the Phenotype and Trafficking of Transgenic CD4 + and CD8 + T Cells

doi: 10.3390/cells10123445

Figure Lengend Snippet: Expanded CD8 + CD122 + Tregs reduce viability of CD4 + T cells. For each experiment, spleen cells from female P14 mice were harvested 3 days after intravenous injection with PLG/GP33 or control PLG. For each mouse, 5 separate ex vivo cultures were set up. CD4 + T cell viability was determined by flow cytometry. Statistical significance was determined by Student’s t -test. ns not significant, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ( A ) Whole spleen cultures from PLG/GP33-treated P14 or untreated wild-type mice were incubated at 37 °C for 3 or 24 h with GP33 (1 μg/mL), anti-CD3 (1 μg/mL), or both. ( B ) Whole spleen cultures from P14 mice treated with either PLG/GP33 or control PLG (PLG/OVA323 or PLG) were incubated at 37 °C for 3 h with 1 μg/mL anti-CD3. ( C ) CD122 + cells from mice treated with PLG/GP33 were depleted by magnetic cell separation and incubated at 37 °C for 3 h with 1 μg/mL anti-CD3. Results are from three separate pooled experiments.

Article Snippet: CD122 + cells were positively selected and removed from whole spleens by first staining with biotin conjugated anti-CD122 monoclonal antibody and then removing the stained cells with a biotin positive selection kit (STEMCELL Technologies, Vancouver, BC, Canada) according to the manufacturer’s instructions.

Techniques: Injection, Control, Ex Vivo, Flow Cytometry, Incubation, Magnetic Cell Separation