Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: Discovery and validation of STAT5B as a key regulatory factor. ( A ) DoRothEA analysis identifies 32 TFs with significantly altered activity in oligodendrocytes in the PD group (12 activated, 20 downregulated). ( B ) Differential ranking of TFs reveals significant downregulation of STAT5B mRNA in snRNA-seq results (****, p < 0.0001). ( C ) Violin plot confirms significant decrease in STAT5B expression in oligodendrocytes from the PD group compared to the control group. ( D , E ) LFB staining shows decreased myelin density correlating with STAT5B mRNA (r = 0.83, p < 0.00074) and protein (r = 0.76, p < 0.0039) expression in the SN of mice.

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Biomarker Discovery, Activity Assay, Expressing, Control, Staining

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: The expression of STAT5B was significantly reduced in the PD model. ( A ) Representative image of immunofluorescence staining in the SN of the MPTP-induced mouse model (scale bars = 100 μm). ( B ) Quantification of co-localization using Pearson’s correlation coefficient (PCC). The y-axis represents the PCC, which quantifies the degree of spatial overlap between the STAT5B and Olig2 signals. PCC values range from −1 to +1, where +1 indicates perfect co-localization, 0 indicates no correlation (random distribution), and -1 indicates perfect segregation. Each data point represents the PCC calculated from individual image fields. The data are presented as the means ± SEMs of 3 independent experiments. Data are presented from male (n = 3) mice, aged 8 weeks. ( C ) qRT-PCR analysis of STAT5B mRNA expression in MO3.13 cells treated with 200 and 500 μM MPP + for 24 h. The data are presented as the means ± SEMs of 3 independent experiments. ( D , E ) Western blot analysis of STAT5B protein expression in MO3.13 cells treated with 500 μM MPP + for 24 h. The data are presented as the means ± SEMs of 6 independent experiments (*, p < 0.05; **, p < 0.01).

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Expressing, Immunofluorescence, Staining, Quantitative RT-PCR, Western Blot

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: LFB staining of myelin in co-cultures of differentiated MO3.13 cells and SH-SY5Y neuronally differentiated cells. ( A , B ) LFB staining of STAT5B overexpression in MPP + -treated MO3.13 cells. ( C , D ) LFB staining of STAT5B knockdown in MO3.13 cells. The results are expressed as the mean ± SD (n = 3; 10× scale bars = 200 μm; **, p < 0.01; ***, p < 0.001).

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Staining, Over Expression, Knockdown

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: Overexpression of oligodendrocyte STAT5B improved myelin damage in MPTP-induced mice. ( A , B ) LFB staining showed that overexpression of oligodendrocyte STAT5B increased myelin density in MPTP-induced mice. The results are expressed as the mean ± SD (10× scale bars = 200 μm). Data are presented from male (n = 6) mice, aged 8 weeks. ( C , D ) TEM analysis revealed that oligodendrocyte STAT5B overexpression increased axonal myelin thickness and significantly reduced g-ratio in MPTP-treated mice. The results are expressed as the mean ± SD (5 randomly selected myelinated axons per SN field; 20.0 k× scale bars = 1 μm). ( E ) Scatter plot illustrating the individual g -ratio values and the distribution of axonal sizes. Data are presented from male (n = 3) mice, aged 8 weeks (*, p < 0.05; **, p < 0.01; ***, p < 0.01).

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Over Expression, Staining

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: Overexpression of oligodendrocyte STAT5B ameliorated dopamine neuronal damage in MPTP-treated mice. ( A ) qRT-PCR analysis of TH mRNA expression in the mouse SN. ( B ) Western blot analysis and ( C ) quantification of TH protein levels in the mouse SN. ( D , E ) Immunohistochemical staining of TH protein in the mouse SN. ( F , G ) Immunohistochemical staining of NfL protein in the mouse SN. The results are expressed as the mean ± SD (10× scale bars = 200 μm; 40× scale bars = 50 μm; *, p < 0.05; **, p < 0.01; ***, p < 0.001). Data are presented from male (n = 6) mice, aged 8 weeks.

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Over Expression, Quantitative RT-PCR, Expressing, Western Blot, Immunohistochemical staining, Staining

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: Overexpression of oligodendrocyte STAT5B improved motor function in MPTP-treated mice. ( A , B ) Pole test results showing total time and turn time. ( C ) Rotarod test results showing fall latency. ( D – F ) Gait analysis results showing movement speed, stride length, and support time. NS: no support, SLS: single-leg support, CLS: contralateral limb support, HLS: homologous limb support, ILS: ipsilateral limb support, TLS: three-limb support, FLS: four-limb support. The results are expressed as the mean ± SD (*, p < 0.05; **, p < 0.01; ***, p < 0.001). Data are presented from male (n = 14) mice, aged 8 weeks.

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Over Expression

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: STAT5B overexpression promoted MBP expression and reduced myelin injury. ( A , B ) Luciferase reporter assay showing the binding efficiency of STAT5B to the MBP promoter region (n = 6). ( C – H ) In MO3.13 cells overexpressing STAT5B : ( C ) qRT-PCR and ( D , E ) Western blot analysis of MBP expression (n = 4). ( F – H ) Immunofluorescently stained STAT5B and MBP expression (n = 3, scale bars = 50 μm). The results are expressed as the mean ± SD (**, p < 0.01; ***, p < 0.001).

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Over Expression, Expressing, Luciferase, Reporter Assay, Binding Assay, Quantitative RT-PCR, Western Blot, Staining

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: Knockdown of STAT5B decreases MBP expression and aggravates myelin damage. ( A – F ) In STAT5B -knockdown MO3.13 cells: ( A ) qRT-PCR analysis of MBP mRNA expression (n = 4), ( B , C ) Western blot analysis of MBP protein expression (n = 4), and ( D – F ) immunofluorescently stained STAT5B and MBP expression (n = 3, scale bars = 50 μm). ( G ) qRT-PCR analysis of MBP mRNA expression in the mouse SN. ( H , I ) Western blot analysis and quantification of MBP protein levels in the mouse SN. The results are expressed as the mean ± SD (*, p < 0.05; **, p < 0.01; ***, p < 0.001). Data are presented from male (n = 6) mice, aged 8 weeks.

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Knockdown, Expressing, Quantitative RT-PCR, Western Blot, Staining

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: STAT5B mRNA stability and promoter methylation study. ( A ) Actinomycin D transcription inhibition assay to detect STAT5B mRNA stability. Line chart showing STAT5B mRNA expression at different time points after treatment with actinomycin D. The results are expressed as the mean ± SD (n = 6). ( B ) The MethPrimer website predicts that there are extensive CpG islands in the promoter region of STAT5B (light blue areas are CpG island regions). ( C , D ) Top 10 CpG sites with significantly increased methylation in the PD group compared to the control group, as determined by MethylTarget sequencing: ( C ) MethylTarget sequencing scatter plot—Y-axis: “CG methyl level”, “CG density”; X-axis: methylation site positions. ( D ) MethylTarget sequencing heatmap—Y-axis: methylation sites; X-axis: groups.

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Methylation, Inhibition, Expressing, Control, Sequencing

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: Effects of DNMT3A on methylation levels in the STAT5B promoter region. ( A ) mRNA expression of DNA methylation-related genes assessed by qRT-PCR (n = 6, **, p < 0.01; ***, p < 0.001). ( B , C ) Methylation-specific PCR (MSP) analysis of the STAT5B promoter region at site 2202 in DNMT3A -knockdown and MPP + -induced MO3.13 cells. ( D , E ) MSP analysis of the STAT5B promoter region at site 2202 in DNMT3A -overexpression MO3.13 cells.

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Methylation, Expressing, DNA Methylation Assay, Quantitative RT-PCR, Knockdown, Over Expression

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: DNMT3A knockdown and overexpression affect STAT5B expression. ( A – D ) In DNMT3A -knockdown MO3.13 cells: ( A – D ) qRT-PCR and ( C , D ) Western blot analysis of STAT5B expression (n = 6). ( E – H ) In MO3.13 cells with DNMT3A overexpression or STAT5B co-overexpression with DNMT3A : ( E , F ) qRT-PCR analysis of DNMT3A and STAT5B mRNA and ( G , H ) Western blot analysis of STAT5B protein expression (n = 6). The results are expressed as the mean ± SD (*, p < 0.05; **, p < 0.01; ***, p < 0.001).

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Knockdown, Over Expression, Expressing, Quantitative RT-PCR, Western Blot

Journal: Cells

Article Title: Oligodendrocyte-Specific STAT5B Overexpression Ameliorates Myelin Impairment in Experimental Models of Parkinson’s Disease

doi: 10.3390/cells14151145

Figure Lengend Snippet: DNMT3A-mediated downregulation of MBP via STAT5B in oligodendrocytes. ( A – E ) In DNMT3A -knockdown MO3.13 cells: ( A – C ) qRT-PCR and Western blot analysis of MBP expression (n = 6). ( D , E ) LFB staining indicates improved myelin integrity in SH-SY5Y neuronally differentiated cells co-cultured with differentiated DNMT3A -knockdown MO3.13 cells (n = 3, scale bars = 200 μm). The results are expressed as the mean ± SD (*, p < 0.05; **, p < 0.01; ***, p < 0.001). ( F – J ) In MO3.13 cells with DNMT3A and STAT5B co-overexpression: ( F – H ) qRT-PCR and Western blot analysis of MBP expression (n = 6). ( I , J ) LFB staining shows improved myelin integrity in SH-SY5Y neuronally differentiated cells co-cultured with MO3.13 cells co-overexpressing STAT5B and DNMT3A (n = 3). The results are expressed as the mean ± SD (*, p < 0.05; **, p < 0.01; ***, p < 0.001).

Article Snippet: Primary antibody incubations (4 °C overnight) included the following: Olig2 mouse monoclonal (1:300, sc-515947, Santa Cruz Biotechnology, Shanghai, China), STAT5B rabbit (1:200, CSB-PA022815LA01HU, CUSABIO, Wuhan, China) and mouse (1:200, 66427-1-Ig, Proteintech, Wuhan, China) monoclonals, and MBP rabbit monoclonal (1:200, 10458-1-AP, Proteintech, Wuhan, China).

Techniques: Knockdown, Quantitative RT-PCR, Western Blot, Expressing, Staining, Cell Culture, Over Expression

Journal: International Journal of Molecular Sciences

Article Title: Chr23-miR-200s and Dmrt1 Control Sexually Dimorphic Trade-Off Between Reproduction and Growth in Zebrafish

doi: 10.3390/ijms26041785

Figure Lengend Snippet: Chr23-miR-200s regulate growth of female zebrafish through targeting stat5b 3′UTR. ( A ) QRT-PCR analysis of gh expression levels in pituitary of WT and chr23-miR-200s-KO (miR-200s −/− ) female zebrafish. ( B ) ELISA analysis of plasma GH in miR-200s −/− and WT female zebrafish. ( C ) Stat5b mRNA expression levels in liver of female fish. ( D ) Western blot analysis of Stat5b expression in liver. Representative images (top) and the quantitation (bottom; n = 3 per assay). ( E ) MiR-200b/200a/429a seed region and predicted target sites within 3′UTR of stat5b , and the mutated 3′UTR sequence of stat5b (3′UTR-Mut). The seed sequence pairing regions are marked in red, while other pairing base sequences are marked in blue, and the green bases indicate the mutated pairing region in 3′UTR of stat5b . ( F ) Validation of potential target of miR-200s by a dual-luciferase reporter assay in HEK-293T cells. NC stands for the negative control. Data are represented as mean ± SD ( p < 0.05 *, p < 0.01 **; p < 0.001 ***). ( G ) Fold enrichment of stat5b by using qRT-PCR in the sample pulled down by biotinylated miR-200s (miR-200b/-200a/-429a) and negative control (NC). ( H , I ) Statistics of body length ( H ) and body weight ( I ) of female self-bred offspring of chr23-miR-200s-KO and stat5b double heterozygotes at 3 mpf. The groups with different letters indicate that they are statistically significant ( p < 0.05).

Article Snippet: After blocking, the membranes were incubated overnight at 4 °C with primary antibodies, rabbit polyclonal anti-Stat5b antibody (1:1000, #A21613; ABclonal, Wuhan, China), or rabbit monoclonal anti-β-actin antibody (1:100,000, #AC026; ABclonal, Wuhan, China).

Techniques: Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay, Clinical Proteomics, Western Blot, Quantitation Assay, Sequencing, Biomarker Discovery, Luciferase, Reporter Assay, Negative Control

Journal: International Journal of Molecular Sciences

Article Title: Chr23-miR-200s and Dmrt1 Control Sexually Dimorphic Trade-Off Between Reproduction and Growth in Zebrafish

doi: 10.3390/ijms26041785

Figure Lengend Snippet: Dmrt1 regulates somatic growth in male zebrafish by mediating stat5b expression. ( A , B ) Gh mRNA levels in pituitary ( A ) and plasma GH levels ( B ) of WT and dmrt1 −/− male zebrafish at 5 mpf. ( C , D ) Relative expression of stat5b mRNA ( C ) and Stat5b protein ( D ) in liver. Representative images of Western blot (top) and the quantitation (bottom; n = 3 per assay). ( E ) Binding site of transcription factor Dmrt1 in the stat5b promoter region predicted by JASPAR. ( F ) Schematic representation of the normal and mutational stat5b promoter luciferase constructs and their activities in dual-luciferase assay. Dmrt1 overexpression inhibited the luciferase activity of stat5b promoter in HEK-293T cells. The empty vector pcDNA3.1 + and pGL3-basic were transfected as the control. ( G , H ) The body length and body weight of male self-bred offspring of dmrt1 and stat5b double heterozygotes at 3 mpf. The groups with different letters indicate that they are statistically significant ( p < 0.05). ( I ) The body length and body weight of male dmrt1 transgenic line at 3 mpf ( n = 20). ( J ) Relative expression of stat5b mRNA in liver of dmrt1 transgenic male fish. Data are represented as mean ± SD ( p < 0.05 *, p < 0.01 **; p < 0.001 ***; ns, no significant).

Article Snippet: After blocking, the membranes were incubated overnight at 4 °C with primary antibodies, rabbit polyclonal anti-Stat5b antibody (1:1000, #A21613; ABclonal, Wuhan, China), or rabbit monoclonal anti-β-actin antibody (1:100,000, #AC026; ABclonal, Wuhan, China).

Techniques: Expressing, Clinical Proteomics, Western Blot, Quantitation Assay, Binding Assay, Luciferase, Construct, Over Expression, Activity Assay, Plasmid Preparation, Transfection, Control, Transgenic Assay