Journal: Frontiers in Cell and Developmental Biology

Article Title: Integrating single-cell and bulk RNA sequencing data reveals RGS4 as a functional driver in a proliferative subgroup of SF-1 lineage PitNETs

doi: 10.3389/fcell.2026.1815682

Figure Lengend Snippet: Rgs4 promotes tumor proliferation in vitro . (A,E,I) CellTiter-Glo assessing cell viability following Rgs4 overexpression in AtT20 cells, GH3 cells (E) , and MMQ cells (I) . (B,F,J) RT–qPCR analysis confirming Rgs4 overexpression and quantifying downstream mRNA changes in AtT20 cells, GH3 cells (F) , and MMQ cells (J) . (C,G,K) CellTiter-Glo assays evaluating cell viability after Rgs4 knockdown in AtT20 cells, GH3 cells (G) , and MMQ cells (K) . (D,H,L) RT-qPCR validation of Rgs4 knockdown and associated transcriptional changes in AtT20 cells, GH3 cells (H) , and MMQ cells ( L ).

Article Snippet: The AtT20, GH3, and MMQ cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, United States).

Techniques: In Vitro, Over Expression, Quantitative RT-PCR, Knockdown, Biomarker Discovery

Journal: Frontiers in Cell and Developmental Biology

Article Title: Integrating single-cell and bulk RNA sequencing data reveals RGS4 as a functional driver in a proliferative subgroup of SF-1 lineage PitNETs

doi: 10.3389/fcell.2026.1815682

Figure Lengend Snippet: Rgs4 promotes tumor proliferation in vivo . ( A–C) Tumor growth curves from subcutaneous xenograft models generated by injecting AtT20 cells (A) , MMQ cells (B) , and GH3 cells (C) . ( D–F) Representative images of subcutaneous xenograft tumors formed after injection of AtT20 cells (D) , with corresponding images for MMQ (E) and GH3 (F) cell–derived tumors. ( G–I) Ki-67 expression and its H-score was detected and calculated by immunohistochemistry in AtT20 cells (G) , MMQ cells (H) , and GH3 cells (I) . P-value was calculated by student t test.

Article Snippet: The AtT20, GH3, and MMQ cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, United States).

Techniques: In Vivo, Generated, Injection, Derivative Assay, Expressing, Immunohistochemistry

Journal: Frontiers in Cell and Developmental Biology

Article Title: Integrating single-cell and bulk RNA sequencing data reveals RGS4 as a functional driver in a proliferative subgroup of SF-1 lineage PitNETs

doi: 10.3389/fcell.2026.1815682

Figure Lengend Snippet: RGS4 inhibition induces apoptosis in PitNETs cells. (A–C) Cell viability of three pituitary tumor cell lines following treatment with the RGS4 inhibitor at multiple concentrations and time points, measured using the CellTiter-Glo luminescence assay. (D) Flow cytometry analysis assessing apoptosis rates in the three cell lines after exposure to different doses of the RGS4 inhibitor. (E) RT-qPCR quantification of Bax and Bcl-2 mRNA levels in the three cell lines following Rgs4 overexpression or knockdown. (F) Apoptosis of primary PitNET cells following RGS4 inhibitor treatment was assessed by flow cytometry. (G) Cell viability of primary PitNET cells following RGS4 inhibitor treatment was measured using the CellTiter-Glo assay. (H) Western blot analysis evaluating protein expression of Bax and Bcl-2 in response to increasing concentrations of the RGS4 inhibitor in the three cell lines. (I) Transcriptomic profiling of AtT20 cells showing Bax and Bcl-2 expression changes after RGS4 inhibitor treatment (5 μM, 24 h).

Article Snippet: The AtT20, GH3, and MMQ cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, United States).

Techniques: Inhibition, Luminescence Assay, Flow Cytometry, Quantitative RT-PCR, Over Expression, Knockdown, Glo Assay, Western Blot, Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Integrating single-cell and bulk RNA sequencing data reveals RGS4 as a functional driver in a proliferative subgroup of SF-1 lineage PitNETs

doi: 10.3389/fcell.2026.1815682

Figure Lengend Snippet: RGS4 regulates p53 protein ubiquitination and stability. (A,B) KEGG pathway and HALLMARK gene set enrichment analyses of transcriptomic data from AtT20 cells treated with an RGS4 inhibitor. (C) ssGSEA-based quantification of p53 signaling pathway activity in AtT20 transcriptome datasets following RGS4 inhibition. (D) Trp53 mRNA expression from RNA-seq data in RGS4 inhibitor–treated AtT20 cells. (E) Western blot analysis of p53 and RGS4 protein levels after Rgs4 overexpression in four cell lines. (F) Western blot analysis of p53 and RGS4 expression in response to 24 h of exposure to increasing concentrations of the RGS4 inhibitor in four cell lines. (G) IP assays showing increased p53 ubiquitination in 293T cells co-transfected with HA-Ub, p53-3×FLAG, and RGS4 expression plasmid. (H) Co-IP assays demonstrating altered p53 ubiquitination in 293T cells co-transfected with HA-Ub and p53-3×FLAG following treatment with an RGS4 inhibitor. (I) CHX chase assays showing p53 protein stability in RGS4 -overexpressing cell lines at the indicated time points.

Article Snippet: The AtT20, GH3, and MMQ cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, United States).

Techniques: Ubiquitin Proteomics, Activity Assay, Inhibition, Expressing, RNA Sequencing, Western Blot, Over Expression, Transfection, Plasmid Preparation, Co-Immunoprecipitation Assay

Journal: Endocrinology

Article Title: miR-375 Regulation of SSTR2 Expression in Corticotroph Pituitary Cells: Somatostatin Receptor Ligands Effects

doi: 10.1210/endocr/bqaf107

Figure Lengend Snippet: (A) Expression of circulating miR-375 in sera of 17 normal subjects and 26 patients with CD. (B) Expression of miR-375 in 2 normal pituitaries and 6 corticotroph pituitary adenomas tissues. (C) Expression of miR-375 in GH3 and AtT20/D16 cell lines. (D) Expression of miR-375 in AtT20/D16 cell line under DEX 10 −8 M treatment after 24 hours, 48 hours and 6 days. (E) Sstr2 gene expression in GH3 and AtT20/D16 cell lines. (F) Sstr2 gene expression modulation by DEX 10 −8 M treatment after 24 hours, 48 hours, and 6 days in AtT20/D16 cell line. Data in the graphs performed in cell lines represent mean ± SEM of 3 independent experiments. * P < .05, ** P < .01, *** P < .001, **** P < .0001 vs control or among the groups.

Article Snippet: AtT20/D16 cell line (CRL-1795), a strain of the mouse corticotroph pituitary tumor model AtT20, and GH3 cell line (CCL-82.1), a rat somatotroph pituitary tumor model, were provided by American Type Culture Collection (ATCC) and cultured in DMEM media supplemented with 10% fetal bovine serum (FBS) (Gibco, code number 16140071), 200 mM L-glutamine (Gibco, code number 25030081) and 1 × 10 5 U/L penicillin and streptomycin (Gibco, code number 15140122).

Techniques: Expressing, Gene Expression, Control

Journal: Endocrinology

Article Title: miR-375 Regulation of SSTR2 Expression in Corticotroph Pituitary Cells: Somatostatin Receptor Ligands Effects

doi: 10.1210/endocr/bqaf107

Figure Lengend Snippet: (A) Scheme of seed match between miR-375 and Sstr2 in mouse and SSTR2 in human. (B) Protein expression and densitometry of SSTR2 in AtT20/D16 treated with miR-375 inhibitor and mimic, evaluated by WB. The data in the graphs are expressed as a percentage of control and represent the mean ± SEM of 8 independent experiments. (C) Protein expression and fluorescence intensity of SSTR2 in AtT20/D16 treated with miR-375 inhibitor and mimic and in 2 human corticotroph primary cultures and (D) treated with miR-375 inhibitor evaluated by IF. The data in the graphs are expressed as a percentage of control normalized to cell number and represent the mean ± SEM of 3 independent experiments in cell lines. * P < .05, ** P < .01, *** P < .001, **** P < .0001 vs control or among the groups.

Article Snippet: AtT20/D16 cell line (CRL-1795), a strain of the mouse corticotroph pituitary tumor model AtT20, and GH3 cell line (CCL-82.1), a rat somatotroph pituitary tumor model, were provided by American Type Culture Collection (ATCC) and cultured in DMEM media supplemented with 10% fetal bovine serum (FBS) (Gibco, code number 16140071), 200 mM L-glutamine (Gibco, code number 25030081) and 1 × 10 5 U/L penicillin and streptomycin (Gibco, code number 15140122).

Techniques: Expressing, Control, Fluorescence

Journal: Endocrinology

Article Title: miR-375 Regulation of SSTR2 Expression in Corticotroph Pituitary Cells: Somatostatin Receptor Ligands Effects

doi: 10.1210/endocr/bqaf107

Figure Lengend Snippet: Evaluation of cell proliferation in AtT20/D16 treated with (A) OCT 10 −8 M and 10 −7 M and (B) PAS 10 −8 M and 10 −7 M with and without miR-375 inhibitors after 3 days of treatment by DNA assay. The data in the graphs are expressed as a percentage of control and represent the mean ± SEM of 3 independent experiments. * P < .05, ** P < .01, *** P < .001, **** P < .0001 among the groups.

Article Snippet: AtT20/D16 cell line (CRL-1795), a strain of the mouse corticotroph pituitary tumor model AtT20, and GH3 cell line (CCL-82.1), a rat somatotroph pituitary tumor model, were provided by American Type Culture Collection (ATCC) and cultured in DMEM media supplemented with 10% fetal bovine serum (FBS) (Gibco, code number 16140071), 200 mM L-glutamine (Gibco, code number 25030081) and 1 × 10 5 U/L penicillin and streptomycin (Gibco, code number 15140122).

Techniques: Control

Journal: Endocrinology

Article Title: miR-375 Regulation of SSTR2 Expression in Corticotroph Pituitary Cells: Somatostatin Receptor Ligands Effects

doi: 10.1210/endocr/bqaf107

Figure Lengend Snippet: Cellular localization of SSTR2 protein expression in AtT20/D16 evaluated by IF in the presence of OCT 10 −7 M for 20 minutes, miR-375 inhibitor and their combination. Images were captured at 60× magnification. The images reported in the figure represent 1 of the 3 independent experiments.

Article Snippet: AtT20/D16 cell line (CRL-1795), a strain of the mouse corticotroph pituitary tumor model AtT20, and GH3 cell line (CCL-82.1), a rat somatotroph pituitary tumor model, were provided by American Type Culture Collection (ATCC) and cultured in DMEM media supplemented with 10% fetal bovine serum (FBS) (Gibco, code number 16140071), 200 mM L-glutamine (Gibco, code number 25030081) and 1 × 10 5 U/L penicillin and streptomycin (Gibco, code number 15140122).

Techniques: Expressing

Journal: Endocrinology

Article Title: miR-375 Regulation of SSTR2 Expression in Corticotroph Pituitary Cells: Somatostatin Receptor Ligands Effects

doi: 10.1210/endocr/bqaf107

Figure Lengend Snippet: Intracellular signaling modulation of proliferation markers, ERK1/2, and apoptotic markers, caspase 3, and PARP, in AtT20/D16 treated with OCT 10 −8 M and 10 −7 M, miR-375 inhibitor, and their combination, evaluated by WB.

Article Snippet: AtT20/D16 cell line (CRL-1795), a strain of the mouse corticotroph pituitary tumor model AtT20, and GH3 cell line (CCL-82.1), a rat somatotroph pituitary tumor model, were provided by American Type Culture Collection (ATCC) and cultured in DMEM media supplemented with 10% fetal bovine serum (FBS) (Gibco, code number 16140071), 200 mM L-glutamine (Gibco, code number 25030081) and 1 × 10 5 U/L penicillin and streptomycin (Gibco, code number 15140122).

Techniques: