26196 1 ap  (Proteintech)

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Reduced expression of GLP1R and GIPR in PDLSCs under multiple stress conditions. a Western blot assay show the expression of GLP1R and GIPR and PKA activity in PDLSCs respectively treated with cigarette, E-cig, hypoxia, LPS, alcohol, HG as well as MOF stimulation. Bubble plot indicates the quantitative analysis of the grayscale values of each band normalized to the β-actin (All western blot assays in this study were performed three duplications). Green bubbles indicate a significant decrease, whereas red bubbles indicate a significant increase compared to NC (gray bubble) ( P < 0.05) determined by Student’s t -test. Gray bubbles in other groups indicate no statistical significance compared with NC. b–d FRET-based biosensors detect the intracellular levels of cAMP ( b ), Ca 2+ ( c ), and PKA activity ( d ). CFP and YFP ratiometric images are shown as a pseudocolored green (CFP) and magenta (YFP) overlay (top). Quantifications of CFP/YFP emission ratio in the panels of sensor in different groups ( n = 20 in each group) (bottom). A ratio value more than 1 represents increased cAMP level, decreased Ca 2+ level, or inactivated PKA. e Western blot assay showing the expression of GLP1R and GIPR in PDLSCs derived from five healthy donors and five patients with periodontitis. The interpretation of bubble plot is illustrated in Fig. 1a. f , g Immunofluorescence show green fluorescence expression of the classical marker STRO-1, which is used to locate PDLSCs, and simultaneously demonstrate the co-localized red fluorescence expression of GLP1R ( f ) and GIPR ( g ) in NC (top) and PD (bottom) in vivo. H&E staining (right panel) is first performed to guide the identification of periodontal ligament tissue at second molar of NC and PD in serial sections, followed by immunofluorescence detection. Scale bar, 20 μm. Histogram show the differential positive expression rate of GLP1R or GIPR in STRO-1 positive cells compared between NC and PD. Green column indicate a significant decrease compared with NC (gray column) ( P < 0.05) determined by Student’s t -test (five fields for each group). NC, normal PDLSCs as negative control; Cig, cigarette; E-cig, E-cigarette; LPS, lipopolysaccharide; HG, high glucose; MOF, mild orthodontic force; FRET, Förster resonance energy transfer; HC, healthy control; PD, periodontitis

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: Expressing, Western Blot, Activity Assay, Derivative Assay, Immunofluorescence, Fluorescence, Marker, In Vivo, Staining, Negative Control, Förster Resonance Energy Transfer, Control

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Cell growth of PDLSCs enhanced by GIPRA. a Western blot assay validate the expression of GLP1R and GIPR in PDLSCs treated with siRNA-mediated knockdown, ectopic overexpression, and agonists of GLP1R and GIPR. The interpretation of bubble plot is illustrated in Fig. a. b CCK-8 assay indicating the cell growth enhanced by GIPR agonist, but suppressed by GIPR knockdown ( n = 5 in each group). The interpretation of line colors in the line chart is the same in Fig. a. c Colony formation assay showing the crystal violet spots in different groups of PDLSCs (top). Histogram shows the statistical analysis of colony counts by Student’s t -test (bottom) ( n = 5 in each group). The interpretation of column colors is the same as in Fig. a. d Stacked column chart show the proportion of cells at different stages of cell cycle. * Significant increase of G2/M phase in GIPR KD and DKD groups as well as decrease of G2/M phase in GIPRA, 2GRA, and BGM0504 ( P < 0.05) compared with NC determined by Student’s t -test ( n = 5 in each group). e Venn diagram show the intersection of differentially expressed genes (|log 2 FC > 1|, P < 0.05) among the GIPR KD, GIPRA, and 2GRA groups compared with NC group. f The bubble plot displaying the top ten functions of cell proliferation (GO biological process) associated with the 1151 differentially expressed genes in e . g GSEA identifies the activated enrichment of the MAPK/ERK signaling pathway in response to GIPRA treatment compared with NC group. NC, normal PDLSCs as negative control; KD, knockdown; OE, overexpression; DKD, GLP1R and GIPR double knockdown; DOE, GLP1R and GIPR double overexpression; GLP1RA, GLP1R agonist; GIPRA, GIPR agonist; 2GRA, GLP1R/GIPR agonist-1; BGM0504, BGM0504 injection of GLP-1 and GIP dual receptor agonist (developed by BrightGene)

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: Western Blot, Expressing, Knockdown, Over Expression, CCK-8 Assay, Colony Assay, Negative Control, Injection

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Cell differentiation of PDLSCs enhanced by GLP1RA. a Representative images of Alizarin Red S staining of mineralized nodules in PDLSCs by osteogenic induction and siRNA-mediated knockdown and ectopic overexpression of GLP1R or GIPR as well as agonists of GLP1R and GIPR. Calcium deposits are visualized as red-stained areas, indicating extracellular matrix mineralization. Quantification was performed by dye extraction and absorbance measurement at 562 nm. Scar bar, 100 μm. b Representative images of Oil Red O staining in PDLSCs cultured by adipogenic induction and siRNA-mediated knockdown and ectopic overexpression of GLP1R or GIPR as well as agonists of GLP1R and GIPR. Red staining indicate intracellular lipid droplet accumulation. Quantification was performed by dye extraction and absorbance measurement at 490 nm. Scale bar, 100 μm. c Representative images of Alcian Blue staining in PDLSCs cultured by chondrogenic induction and siRNA-mediated knockdown and ectopic overexpression of GLP1R or GIPR as well as agonists of GLP1R and GIPR. Blue coloration indicates the deposition of sulfated glycosaminoglycans, a hallmark of chondrogenic differentiation. Quantification was performed by dye extraction and absorbance measurement at 620 nm. Scale bar, 100 μm. d Double immunofluorescence staining show the co-localization of NeuN (Red) and GFAP (Green) representing neurons and astrocytes in PDLSCs by neurogenic induction and siRNA-mediated knockdown and ectopic overexpression of GLP1R or GIPR as well as agonists of GLP1R and GIPR. Scar bar, 50 μm. Histogram shows the differential percentages of NeuN or GFAP positive cells among these groups (five fields for per group) determined by one-way ANOVA. e Western blot assay show the expression of STRO-1, CD146, and vimentin as mesenchymal biomarkers of PDLSCs by osteogenic induction with siRNA-mediated knockdown and ectopic overexpression of GLP1R or GIPR as well as agonists of GLP1R and GIPR. In statistical analysis, the meanings of the red, green, and gray colors are illustrated in Fig. a ( n = 5 in each group). Interpretation of abbreviations can be found in the legend of Fig.

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: Cell Differentiation, Staining, Knockdown, Over Expression, Extraction, Cell Culture, Double Immunofluorescence Staining, Western Blot, Expressing

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Translation shut off by upregulated IFIT proteins. a Heatmap showing the differentially expressed genes (|log 2 FC|> 1, P < 0.05) compared between GLP1R knockdown, GLP1RA, 2GRA, and NC among the multiple lineage differentiation system of PDLSCs. b–e Dumbbell plots illustrating the changes in gene expression of IFIT1 , IFIT2 , IFIT3 , IRF9 , IFI44 , IFI6 , IFIH1 , OAS1 , OASL , RSAD2 , and BST2 between GLP1R knockdown, GLP1RA, 2GRA, and NC in osteogenesis ( b ), adipogenesis ( c ), chondrogenesis ( d ), and neurogenesis ( e ) from RNA sequencing data. f Western blot assay showing the expression of classic markers of osteogenesis COL1A1, BGLAP, and RUNX2 in PDLSCs for osteogenic differentiation system treated with GLP1R knockdown, GLP1RA, and 2GRA. g Western blot assay showing the expression of classic markers of adipogenesis CEBPA, FABP4, and PPARG in PDLSCs for adipogenic differentiation system treated with GLP1R knockdown, GLP1RA, and 2GRA. h Western blot assay show the expression of classic markers of chondrogenesis ACAN, COL2A1, and SOX9 in PDLSCs for chondrogenic differentiation system treated with GLP1R knockdown, GLP1RA, and 2GRA. i Western blot assay show the expression of classic markers of neurogenesis MAP2, NES, and TUBB3 in PDLSCs for adipogenic differentiation system treated with GLP1R knockdown, GLP1RA, and 2GRA. j Western blot assay showing the interaction of IFIT1 with IFIT2, IFIT3, eIF3C, and eIF3E within the immunoprecipitation of IFIT1 from PDLSCs under undifferentiated, osteogenic, GLP1R knockdown, GLP1RA, and 2GRA treated conditions. k Western blot assay show the interaction of eIF3C with IFIT1, IFIT2, IFIT3, eIF3E, and RPS3 within the immunoprecipitation of eIF3C from PDLSCs under undifferentiated, osteogenic, GLP1R knockdown, GLP1RA, and 2GRA treated conditions. Red boxes highlight the robust protein interaction in undifferentiated PDLSCs and osteogenic PDLSCs with GLP1R knockdown. KD, knockdown; NC, PDLSCs for regular differentiation accordingly; Un, undifferentiated PDLSCs; OS, osteogenesis; 2GRA, GLP1R/GIPR agonist-1

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: Knockdown, Gene Expression, RNA Sequencing, Western Blot, Expressing, Immunoprecipitation

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Bioinformatic analysis of transcriptome-wide occupancy of IFIT1 in osteogenic PDLSCs. a Volcano plot showing the total 2909 genes (1744 increased and 1165 decreased binding genes) corresponding to IFIT1 differentially bound (|log 2 FC|> 1, P < 0.05) transcripts in GLP1R knockdown versus NC PDLSCs of osteogenic differentiation system. b , c Pie charts showing the positional distribution of IFIT1 binding across transcript regions in 1744 increased ( b ) and 1165 decreased binding genes ( c ). d , e RNA binding motif at the 5′ UTR of IFIT1 ( d ) and eIF3C ( e ) in PDLSCs of NC (top) and GLP1R knockdown (bottom) with osteogenic differentiation. f, g Bubble chart showing the top functions of genes with elevated ( f ) and reduced ( g ) IFIT1 peaks at the 5′ UTR by Gene Ontology analysis (biological process) in GLP1R knockdown versus NC PDLSCs of osteogenic differentiation system. h – l IGV showing the classic examples of differentially binding peaks at 5′ UTR of IFIT1 and eIF3C on SPP1 ( h ), FGF18 ( i ), COL1A1 ( j ), MMP14 ( k ), and BMP2 ( l ) in GLP1R knockdown versus NC PDLSCs of osteogenic differentiation system. m Western blot assay validate the protein expression of SPP1, FGF18, COL1A1, MMP14, and BMP2 in GLP1R knockdown versus NC PDLSCs of osteogenic differentiation system. The meanings of the green and gray colors are illustrated in Fig. a. NC, regular osteogenic differentiation of PDLSCs

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: Binding Assay, Knockdown, RNA Binding Assay, Western Blot, Expressing

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: The protective effect of PDLSCs by GLP1RA and GIPRA from periodontitis in vivo. a Representative bidimensional views of the maxillary molars of PD mice model treated with GLP1RA and GIPRA by micro-CT. Scale bar, 0.5 mm. Measurement of the distance from CEJ to ABC around second molar. Histogram shows the differential distance among different groups. Red columns indicate a significant increase compared with NC (gray column), whereas yellow columns indicate a significant decrease compared with red ( P < 0.05) determined by Student’s t -test ( n = 5 in each group). b Representative images of TRAP staining of the maxillary molars of those groups from ( a ). Scale bar, 20 μm. Measurement of the osteoclast number around second molar. Histogram show the differential distance among different groups. c Representative bidimensional views of the maxillary molars of PD mice model treated with GLP1RA, GIPRA, and PDLSC transplantation by micro-CT ( n = 5 in each group). Scale bar, 0.5 mm. Measurement of the distance from CEJ to ABC around second molar. Histogram shows the differential distance among different groups. Green columns indicate a significant decrease compared with NC (gray columns). d Representative images of TRAP staining of the maxillary molars of those groups from c . Scale bar, 20 μm. Measurement of the osteoclast number around second molar. Histogram shows the differential distance among different groups. e – i Immunofluorescence shows mCherry fluorescence derived from exogenous PDLSCs ( n = 5 in each group). The fluorescence indicated the expression of genes such as CD146 (e), RUNX2 (f), CD29 ( g ), UCP1 ( h ), and IFIT1 ( i ), thereby enabling the evaluation of the proliferation and multilineage differentiation of the transplanted PDLSCs under different treatment conditions. Scale bar, 20 μm. WT, wild type; KO, GLP1R −/− /GIPR −/− ; PD, periodontitis; PDLSCs, PDLSCs transplantation; GLP1RA, GLP1R agonist; GIPRA, GIPR agonist; 2GRA, GLP1R/GIPR agonist-1; CEJ, cemento-enamel junction; ABC, alveolar bone crest

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: In Vivo, Micro-CT, Staining, Transplantation Assay, Immunofluorescence, Fluorescence, Derivative Assay, Expressing

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Clinical impact of GLP1R and GIPR agonists on periodontitis severity. a , b Pie charts illustrating the distribution of periodontitis stage ( a ) and grade ( b ) in patients treated with GLP1R/GIPR agonists ( n = 74) (left) and those not treated ( n = 76) (right). ( c , d ) Box plots show the relationship between the duration of GLP1R/GIPR agonist administration and the severity of periodontitis, represented by stage ( c ) and grade ( d ). Data suggest a potential association between prolonged agonist use and reduced periodontitis severity. The classification of periodontitis stage and grade was assessed by the clinical attachment loss, probing pocket depth, and radiographic bone loss on the basis of the 2017 AAP/EFP classification

Article Snippet: Cells were incubated with primary antibodies against NeuN (1: 500, cat. no. 26975-1-AP, Proteintech), GFAP (1: 1000, cat. no. 60190-1-lg, Proteintech), GLP1R (1: 1000, same with Western blot), GIPR (1: 500, same with Western blot), STRO-1 (1: 1000, same with Western blot), cAMP (1: 100, cat. no. ab134901, Abcam) diluted in blocking buffer overnight at 4 °C.

Techniques: