Journal: Acta Biochimica et Biophysica Sinica

Article Title: G protein-coupled estrogen receptor 1 mediates proliferation and adipogenic differentiation of goat adipose-derived stem cells through ERK1/2-NF-κB signaling pathway

doi: 10.3724/abbs.2022031

Figure Lengend Snippet: GPER1 mediates 17β-estradiol-induced proliferation of gADSCs Expression of GPER1 was determined by immunofluorescence staining (A) and western blot analysis (B). Cell counting (C) and CCK-8 assay (D) were performed to detect cell proliferation of gADSCs treated with 17β-estradiol (E2; 0.1 μM), GPER1 agonist G1 (0.1 μM), or 17β-estradiol+GPER1 antagonist G15 (1 μM) for 4 days. (E) The protein level of GPER1 was determined by western blot analysis. After GPER1 silencing, cell counting (F) and CCK-8 assay (G) were performed to detect cell proliferation of gADSCs. (H,I) The mRNA levels of cyclin D1 and PCNA in gADSCs subjected to various treatments were measured by RT-qPCR. P3, P6, P9, and P12 represent the 3, 6, 9, and 12 generation of gADSCs respectively. The nuclei were stained with DAPI. GAPDH serves as the internal control. Scale bar=50 μm. *P<0.05, **P<0.01 (vs control); †P<0.05, ††P<0.01, †††P<0.001 (vs E2); and #P<0.05, ##P<0.01; ###P<0.001 (vs shRNA).

Article Snippet: During adipogenic differentiation, gADSCs were treated with 17β-estradiol (E2, 0.1 μM; Sigma), GPER1 agonist G1 (0.1 μM; Cayman, Ann Arbor, USA), GPER1 antagonist G15 (1 μM; Cayman), or mitogen-activated protein kinase kinase (MEK) inhibitor U0126 (10 μM; Selleck, Houston, USA) alone or in combination as specified.

Techniques: Expressing, Immunofluorescence, Staining, Western Blot, Cell Counting, CCK-8 Assay, Quantitative RT-PCR, shRNA

Journal: Acta Biochimica et Biophysica Sinica

Article Title: G protein-coupled estrogen receptor 1 mediates proliferation and adipogenic differentiation of goat adipose-derived stem cells through ERK1/2-NF-κB signaling pathway

doi: 10.3724/abbs.2022031

Figure Lengend Snippet: Activation of GPER1 promotes adipogenic differentiation of gADSCs (A) Representative images of intracellular lipid accumulation of gADSCs incubated with 17β-estradiol (E2; 0.1 μM), G1 (0.1 μM), or 17β-estradiol+G15 (1 μM) for 14 days. Lipid accumulation was detected by Oil red O staining. (B) Representative images of intracellular lipid accumulation of gADSCs with GPER1 silencing. Lipid accumulation was determined by Oil red O staining. (C,D) The mRNA levels of PPARγ, ACC, and FABP4 in gADSCs subjected to various treatments were measured by RT-qPCR. GAPDH serves as the internal control. Scale bar=100 μm. *P<0.05, **P<0.01 (vs control); †P<0.05, ††P<0.01 (vs E2); and #P<0.05, ##P<0.01 (vs shRNA).

Article Snippet: During adipogenic differentiation, gADSCs were treated with 17β-estradiol (E2, 0.1 μM; Sigma), GPER1 agonist G1 (0.1 μM; Cayman, Ann Arbor, USA), GPER1 antagonist G15 (1 μM; Cayman), or mitogen-activated protein kinase kinase (MEK) inhibitor U0126 (10 μM; Selleck, Houston, USA) alone or in combination as specified.

Techniques: Activation Assay, Incubation, Staining, Quantitative RT-PCR, shRNA

Journal: Acta Biochimica et Biophysica Sinica

Article Title: G protein-coupled estrogen receptor 1 mediates proliferation and adipogenic differentiation of goat adipose-derived stem cells through ERK1/2-NF-κB signaling pathway

doi: 10.3724/abbs.2022031

Figure Lengend Snippet: The cross-talk between GPER1 and NF-κB signaling (A) gADSCs were incubated with 17β-estradiol (E2; 0.1 μM), G1 (0.1 μM), or 17β-estradiol+G15 (1 μM) for 4 days. The expressions of nuclear p-p65 (A) and the total and phosphorylated protein level of ERK1/2 (C) were tested by western blot analysis. After GPER1 silencing, the nuclear protein level of p-p65 (B) and the total and phosphorylated protein level of ERK1/2 (D) were detected by western blot analysis. (E) gADSCs were incubated with 17β-estradiol (0.1 μM) or G1 (0.1 μM) with or without MEK inhibitor U0126 (10 μM) for 4 days, and the nuclear protein level of p-p65 was detected by western blot analysis. GAPDH serves as the internal control. *P<0.05, **P<0.01 (vs control); †P<0.05, ††P<0.01 (vs E2); #P<0.05, ##P<0.01 (vs shRNA); and ¶P<0.05 (vs G1).

Article Snippet: During adipogenic differentiation, gADSCs were treated with 17β-estradiol (E2, 0.1 μM; Sigma), GPER1 agonist G1 (0.1 μM; Cayman, Ann Arbor, USA), GPER1 antagonist G15 (1 μM; Cayman), or mitogen-activated protein kinase kinase (MEK) inhibitor U0126 (10 μM; Selleck, Houston, USA) alone or in combination as specified.

Techniques: Incubation, Western Blot, shRNA

Journal: Acta Biochimica et Biophysica Sinica

Article Title: G protein-coupled estrogen receptor 1 mediates proliferation and adipogenic differentiation of goat adipose-derived stem cells through ERK1/2-NF-κB signaling pathway

doi: 10.3724/abbs.2022031

Figure Lengend Snippet: Effect of 17β-estradiol targeting GPER1 on cell proliferation and adipogenic differentiation is through ERK1/2-NF-κB signaling pathway gADSCs were cultured with 17β-estradiol (E2; 0.1 μM) or G1 (0.1 μM) with or without U0126 (10 μM) for 4 days. Cell counting (A) and CCK-8 assay (B) were performed to detect cell proliferation. (C) The mRNA levels of cyclin D1 and PCNA of gADSCs were measured by RT-qPCR. gADSCs were cultured with 17β-estradiol (0.1 μM) or G1 (0.1 μM) with or without U0126 (10 μM) for 14 days. (D) Representative images of intracellular lipid accumulation of gADSCs detected by Oil red O staining. (E) The mRNA levels of PPARγ, ACC, and FABP4 of gADSCs were measured by RT-qPCR. GAPDH serves as the internal control. Scale bar=100 μm. †P<0.05, ††P<0.01 (vs E2); and¶P< 0.05, ¶¶P<0.01, ¶¶¶P<0.001 (vs G1).

Article Snippet: During adipogenic differentiation, gADSCs were treated with 17β-estradiol (E2, 0.1 μM; Sigma), GPER1 agonist G1 (0.1 μM; Cayman, Ann Arbor, USA), GPER1 antagonist G15 (1 μM; Cayman), or mitogen-activated protein kinase kinase (MEK) inhibitor U0126 (10 μM; Selleck, Houston, USA) alone or in combination as specified.

Techniques: Cell Culture, Cell Counting, CCK-8 Assay, Quantitative RT-PCR, Staining

Journal: Acta Biochimica et Biophysica Sinica

Article Title: G protein-coupled estrogen receptor 1 mediates proliferation and adipogenic differentiation of goat adipose-derived stem cells through ERK1/2-NF-κB signaling pathway

doi: 10.3724/abbs.2022031

Figure Lengend Snippet: Working model for the effect of estrogen binding to GPER1 on cell proliferation and adipogenic differentiation of goat adipose-derived stem cells Estrogenic GPER1 promotes cell proliferation and adipogenic differentiation of goat adipose-derived stem cells through activating the ERK1/2-NF-κB signaling pathway.

Article Snippet: During adipogenic differentiation, gADSCs were treated with 17β-estradiol (E2, 0.1 μM; Sigma), GPER1 agonist G1 (0.1 μM; Cayman, Ann Arbor, USA), GPER1 antagonist G15 (1 μM; Cayman), or mitogen-activated protein kinase kinase (MEK) inhibitor U0126 (10 μM; Selleck, Houston, USA) alone or in combination as specified.

Techniques: Binding Assay, Derivative Assay

Journal: Pharmacological research

Article Title: Breaking the synergism of iron overload and miR-122 to rescue lipid accumulation and peroxidation in MASLD.

doi: 10.1016/j.phrs.2025.107728

Figure Lengend Snippet: Fig. 5. Liver-specifically restoring miR-122 inhibited lipid peroxidation and liver injury induced by iron overload and miR-122 knockout. (A) miR-122 expression and (B) iron content in the liver of miR-122+/+ and miR-122-/- mice with iron overload (IO) and tail-vein injection of recombinant rAAV8 carrying miR-122 (AAV8). (C) Liver fat content and (D) PUFAs proportion of miR-122+/+ and miR-122-/- mice with iron overload and AAV8 treatment. (E) Liver MDA content and (F) liver LPO content of miR-122+/+ and miR-122-/- mice with iron overload and AAV8 treatment. (G) Serum ALT content and (H) serum AST content of miR-122+/+ and miR- 122-/- mice with iron overload and AAV8 treatment. (I) Prussian blue staining, H&E staining, oil red staining, Sirius red staining, and α-SMA staining of liver slices. Scale bar: 100 μm. (J-Q) mRNA expressions of Aacs, Fads2, Agpat1, Mogat2, Slc27a1, Cyp2b9, Cyp2a22, and Cyp3a11 in the liver of miR-122+/+ and miR-122-/- mice with iron overload (IO) and tail-vein injection of recombinant rAAV8 carrying miR-122 (AAV8). (R) Protein expressions of Aacs, Fads2, Agpat1, Mogat2, and Slc27a1 in the liver of miR-122+/+ and miR-122-/- mice with iron overload and AAV8 treatment. * : p < 0.05; **: p < 0.01; *** : p < 0.001, tested by one-way ANOVA with post-hoc Tukey’s multiple comparisons, sample size was presented in figure.

Article Snippet: Antibodies used in this study were β-actin (1:5000, Cat. 20536–1-AP, Proteintech, China), Aacs (1:500, Cat. 13815–1-AP, Proteintech, China), Fads2 (1:500, Cat. 28034–1-AP, Proteintech, China), Agpat1 (1:500, Cat. 10601–1-AP, Proteintech, China), Mogat2 (1:500, Cat. 19514–1-AP, Proteintech, China), Slc27a1 (1:500, Cat. DF7716, Affinity Biosciences, China).

Techniques: Knock-Out, Expressing, Injection, Recombinant, Staining

Journal: Pharmacological research

Article Title: Breaking the synergism of iron overload and miR-122 to rescue lipid accumulation and peroxidation in MASLD.

doi: 10.1016/j.phrs.2025.107728

Figure Lengend Snippet: Fig. 6. Systematically restoring miR-122 inhibited lipid peroxidation and liver injury induced by iron overload and miR-122 knockout. (A) miR-122 expression and (B) iron content in the liver with miR-122 knockout (KO), iron overload (IO), and tail-vein injection of miR-122 agomir. (C) Fat content and (D) PUFAs proportion of the liver with miR-122 KO, IO, and miR-122 agomir treatments. (E) MDA content and (F) LPO content of the liver with miR-122 KO, IO, and miR-122 agomir treatments. (G) Serum ALT content and (H) serum AST content of mice with miR-122 KO, IO, and miR-122 agomir treatments (I) Prussian blue staining, H&E staining, oil red staining, Sirius red staining, and α-SMA staining of liver slices. Scale bar: 100 μm. (J) mRNA expressions of Aacs, Fads2, Agpat1, Mogat2, Slc27a1, Cyp2b9, Cyp2a22, and Cyp3a11 in the liver with miR-122 knockout (KO), iron overload (IO), and tail-vein injection of miR-122 agomir. (K) Protein expressions of Aacs, Fads2, Agpat1, Mogat2, and Slc27a1 in the liver with miR-122 KO, IO, and miR-122 agomir treatments. * : p < 0.05; **: p < 0.01; *** : p < 0.001, tested by one-way ANOVA with post-hoc Tukey’s multiple comparisons, sample size was presented in figure.

Article Snippet: Antibodies used in this study were β-actin (1:5000, Cat. 20536–1-AP, Proteintech, China), Aacs (1:500, Cat. 13815–1-AP, Proteintech, China), Fads2 (1:500, Cat. 28034–1-AP, Proteintech, China), Agpat1 (1:500, Cat. 10601–1-AP, Proteintech, China), Mogat2 (1:500, Cat. 19514–1-AP, Proteintech, China), Slc27a1 (1:500, Cat. DF7716, Affinity Biosciences, China).

Techniques: Knock-Out, Expressing, Injection, Staining

Journal: Pharmacological research

Article Title: Breaking the synergism of iron overload and miR-122 to rescue lipid accumulation and peroxidation in MASLD.

doi: 10.1016/j.phrs.2025.107728

Figure Lengend Snippet: Fig. 7. A high-throughput screening identified dihydroberberine to upregulate miR-122 expression and prevent iron overload-induced MASLD. (A) The high- throughput screening results. Luminescence of the luciferase reporter were detected to represent miR-122 expression. The relative luminescence intensity was calculated by luminescence intensity of compound / luminescence intensity of blank control. Data were from two biological replicates. (B) The scheme of dihy droberberine (DHB) intervention in high-iron diet induced MASLD model. Dihydroberberine was i.p. administrated at 10 mg/kg, twice per week for 4 weeks. (C) The liver iron content. (D) The liver miR-122 expression level. (E) The liver TG content and (F) the liver total lipid content. (G) Oil red staining, Sirius red staining, and α-SMA staining of liver slices. Scale bar: 50 μm. (H) The liver weight / body weight ratio of mice. (I) Hepatic MDA content and (J) hepatic LPO content. (K) Total PUFAs proportion and two specific PUFA proportion, (L) C20:3n3 and (M) C22:6n3, in the liver. (N) mRNA expressions of Aacs, Fads2, Agpat1, Mogat2, Slc27a1, Cyp2a22, and Cyp3a11 in the liver. (O) Protein expressions of Aacs, Fads2, Agpat1, Mogat2, and Slc27a1 in the liver. * : p < 0.05; * *: p < 0.01; * ** : p < 0.001, tested by one-way ANOVA with post-hoc Tukey’s multiple comparisons, n = 6.

Article Snippet: Antibodies used in this study were β-actin (1:5000, Cat. 20536–1-AP, Proteintech, China), Aacs (1:500, Cat. 13815–1-AP, Proteintech, China), Fads2 (1:500, Cat. 28034–1-AP, Proteintech, China), Agpat1 (1:500, Cat. 10601–1-AP, Proteintech, China), Mogat2 (1:500, Cat. 19514–1-AP, Proteintech, China), Slc27a1 (1:500, Cat. DF7716, Affinity Biosciences, China).

Techniques: High Throughput Screening Assay, Expressing, Luciferase, Control, Staining

Journal: Cell death discovery

Article Title: Neuronal RBM5 modulates cell signaling responses to traumatic and hypoxic-ischemic injury in a sex-dependent manner.

doi: 10.1038/s41420-023-01677-7

Figure Lengend Snippet: Fig. 4 Effect of RBM5 KO and sex on neuronal survival in stretch-injury. A Box plots show well peak pressure. B Purity of male and female floxed cultures from independent culture isolations in Fig. 4. C Box plots show post-injury LDH levels (n = 9/group) in control (empty vector) versus CRE (RBM5 KO) neurons. D Representative blots (n = 4/group) for RBM5, SBDPs, caspase-3, pAKT, total AKT, phosphorylated CREB (pCREB), and total CREB levels. E–H Densitometric quantification of RBM5, SBDP 145, pAKT and pCREB normalized to total protein stain (n = 8/ group). Box plots show median, max, min, and IQR. Data were significant at p < 0.05. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. N.S. not significant. RBM5 RNA-binding Motif 5, SBDPs spectrin breakdown products, AKT Protein Kinase B, CREB cAMP response element-binding protein.

Article Snippet: Primary Antibodies: α-Fodrin, Cat#BML-FG6090, Lot#08231207 (Enzo); pan-AKT, Cat#4691, Lot#28 (Cell Signaling Technologies); pan-AKT, Cat#2920 S, Lot#3 and #8 (Cell Signaling Technologies); phosho-AKT (Ser473), Cat#4060 S, Lot#23 and #27 (Cell Signaling Technologies); ERK, Cat#4696 S, Lot#22 (Cell Signaling Technologies); phospho-ERK, Cat#4377 S, Lot#10; PERK, Cat#LSC757915, Lot#143906 (LSBio); RBM5, Cat#19930-1-AP, Lot#00049598 (ProteinTech); Caspase-3, Cat#19677-1-AP Lot#00094596 and #00109139; GPR30, Cat#PA5-28647, Lot#YD3884109C (ThermoFisher); Estrogen Receptor α, Cat#ab76228, Lot#1016753-3 (Abcam).

Techniques: Control, Plasmid Preparation, Staining, RNA Binding Assay, Binding Assay

Journal: Cell death discovery

Article Title: Neuronal RBM5 modulates cell signaling responses to traumatic and hypoxic-ischemic injury in a sex-dependent manner.

doi: 10.1038/s41420-023-01677-7

Figure Lengend Snippet: Fig. 5 Effect of RBM5 KO and sex on neuronal survival in OGD. A Box plots show post-injury LDH levels in sex-separated WT Neurons (n = 8- 9/group). B Purity of WT male and female cultures from independent culture isolations in Fig. 5A. C Box plots show post-injury LDH levels (n = 9/group) in control (empty vector) versus CRE (RBM5 KO) neurons. D Purity of male and female floxed cultures from independent culture isolations in Fig. 5C–K. E Representative blots (n = 4/group) for RBM5, SBDPs, caspase-3, pAKT, total AKT, pCREB, and total CREB levels. F–K Densitometric quantification of RBM5, SBDP 145, SBDP 120, caspase-3, pAKT and pCREB normalized to total protein stain (n = 8/group). Box plots show median, max, min, and IQR. Data were significant at p < 0.05. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. RBM5 RNA- binding Motif 5, SBDPs spectrin breakdown products, AKT protein kinase B, CREB cAMP response element-binding protein.

Article Snippet: Primary Antibodies: α-Fodrin, Cat#BML-FG6090, Lot#08231207 (Enzo); pan-AKT, Cat#4691, Lot#28 (Cell Signaling Technologies); pan-AKT, Cat#2920 S, Lot#3 and #8 (Cell Signaling Technologies); phosho-AKT (Ser473), Cat#4060 S, Lot#23 and #27 (Cell Signaling Technologies); ERK, Cat#4696 S, Lot#22 (Cell Signaling Technologies); phospho-ERK, Cat#4377 S, Lot#10; PERK, Cat#LSC757915, Lot#143906 (LSBio); RBM5, Cat#19930-1-AP, Lot#00049598 (ProteinTech); Caspase-3, Cat#19677-1-AP Lot#00094596 and #00109139; GPR30, Cat#PA5-28647, Lot#YD3884109C (ThermoFisher); Estrogen Receptor α, Cat#ab76228, Lot#1016753-3 (Abcam).

Techniques: Control, Plasmid Preparation, Staining, RNA Binding Assay, Binding Assay

Journal: Cell death discovery

Article Title: Neuronal RBM5 modulates cell signaling responses to traumatic and hypoxic-ischemic injury in a sex-dependent manner.

doi: 10.1038/s41420-023-01677-7

Figure Lengend Snippet: Fig. 6 Effect of estradiol on neuronal survival in RBM5 KO neurons in OGD. A, B Box plots of post-injury LDH levels in male or female neurons treated with or without 1 µM 17β estradiol (E2) (n = 6/group). C Purity of male and female floxed cultures from independent culture isolations in Fig. 6. D, E Representative blots (n = 3/group) for SBDP levels in male and female neurons, respectively. F–I Densitometric quantification of SBDP 145 and SBDP 120 normalized to total protein stain (n = 6/group). Box plots show median, max, min, and IQR. Data were significant at p < 0.05. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. SBDPs spectrin breakdown products.

Article Snippet: Primary Antibodies: α-Fodrin, Cat#BML-FG6090, Lot#08231207 (Enzo); pan-AKT, Cat#4691, Lot#28 (Cell Signaling Technologies); pan-AKT, Cat#2920 S, Lot#3 and #8 (Cell Signaling Technologies); phosho-AKT (Ser473), Cat#4060 S, Lot#23 and #27 (Cell Signaling Technologies); ERK, Cat#4696 S, Lot#22 (Cell Signaling Technologies); phospho-ERK, Cat#4377 S, Lot#10; PERK, Cat#LSC757915, Lot#143906 (LSBio); RBM5, Cat#19930-1-AP, Lot#00049598 (ProteinTech); Caspase-3, Cat#19677-1-AP Lot#00094596 and #00109139; GPR30, Cat#PA5-28647, Lot#YD3884109C (ThermoFisher); Estrogen Receptor α, Cat#ab76228, Lot#1016753-3 (Abcam).

Techniques: Staining

Journal: Cell death discovery

Article Title: Neuronal RBM5 modulates cell signaling responses to traumatic and hypoxic-ischemic injury in a sex-dependent manner.

doi: 10.1038/s41420-023-01677-7

Figure Lengend Snippet: Fig. 7 Global proteomic analysis in male and female RBM5 KO neurons in OGD. A Dendrogram of 240 differentially expressed (DE) proteins across the 6 treatment groups from three independent culture isolations (n = 3/group). B Venn diagram on sex differences in protein targets altered 24 h after OGD in EV (control genotype) neurons. C Gene IDs of DE proteins after OGD in EV-neurons. D Volcano plot of DE proteins in male EV-injured versus male RBM5 KO-injured neurons. E Volcano plot of DE proteins in female EV-injured versus female RBM5 KO neurons.

Article Snippet: Primary Antibodies: α-Fodrin, Cat#BML-FG6090, Lot#08231207 (Enzo); pan-AKT, Cat#4691, Lot#28 (Cell Signaling Technologies); pan-AKT, Cat#2920 S, Lot#3 and #8 (Cell Signaling Technologies); phosho-AKT (Ser473), Cat#4060 S, Lot#23 and #27 (Cell Signaling Technologies); ERK, Cat#4696 S, Lot#22 (Cell Signaling Technologies); phospho-ERK, Cat#4377 S, Lot#10; PERK, Cat#LSC757915, Lot#143906 (LSBio); RBM5, Cat#19930-1-AP, Lot#00049598 (ProteinTech); Caspase-3, Cat#19677-1-AP Lot#00094596 and #00109139; GPR30, Cat#PA5-28647, Lot#YD3884109C (ThermoFisher); Estrogen Receptor α, Cat#ab76228, Lot#1016753-3 (Abcam).

Techniques: Control