Journal: Cell death discovery

Article Title: YAP is required for prostate development, regeneration, and prostate stem cell function.

doi: 10.1038/s41420-023-01637-1

Figure Lengend Snippet: Fig. 8 YAP regulates prostate stem cells through Notch signaling and Hedgehog signaling. A The morphology of shYAP and control prostate spheres cultured in Matrigel. B The number of prostate spheres were measured from 3 pairs of sphere samples. The data is presented as mean ± SEM (**P < 0.01). C The diameter of prostate spheres was measured in at least five prostate spheres from three pairs of sphere samples. The data are presented as mean ± SEM (**P < 0.01). D The expression of YAP, CTGF and Cyr61 in prostate spheres. E Notch and Hedgehog signaling pathways were compared by qPCR between shYAP and control prostate spheres. F Notch and Hedgehog signaling pathways in early prostate development were compared via qPCR between scYAPKO and WT in 2-week-old mouse prostates. G Prostate spheres treated 10 days with recombinant Shh or control media.

Article Snippet: In several experiments, 0.25ug/ml recombinant mouse Sonic Hedgehog (MedChemExpress) was used to treat prostate spheres.

Techniques: Control, Cell Culture, Expressing, Protein-Protein interactions, Recombinant

Journal: Theranostics

Article Title: Nintedanib enhances the efficacy of PD-L1 blockade by upregulating MHC-I and PD-L1 expression in tumor cells

doi: 10.7150/thno.65828

Figure Lengend Snippet: Analysis of mechanisms related to immune activation. (A) Heat map of the expression of signature genes of interferon-gamma (IFN-γ) response (B) quantified by signature score. (C) Heat map of the expression of signature genes of IFN-α response (D) quantified by signature score. Gene Set Enrichment Analysis (GSEA) plot and gene sets for (E) RESPONSE TO INTERFERON-GAMMA and (F) ANTIGEN PROCESSING AND PRESENTATION. Flow cytometry analysis of (G) major histocompatibility complex class I (MHC-I) and (H) PD-L1 expression after different treatments in MC38 cells. The MHC-I expression levels were evaluated by MFI in (I) MC38, (J) LLC, (K) 4T1, and (O) B16F10 cells. The PD-L1 expression levels were evaluated by MFI in (L) MC38, (M) LLC, (N) 4T1, and (P) B16F10 cells. (Q) Representative western blot images of p-STAT3/STAT3, transporter associated with antigen processing 1 (TAP1), PD-L1 and beta-2-microglobulin (β2M).

Article Snippet: Recombinant mouse interferon-gamma (rMuIFN-γ; HY-P7071) and recombinant human interferon-gamma (rHuIFN-γ; HY-P7025) were purchased from MedChem Express (Monmouth Junction, NJ, USA).

Techniques: Activation Assay, Expressing, Flow Cytometry, Western Blot

Journal: Stem Cell Reviews and Reports

Article Title: HucMSCs-derived Exosomes Promote Lung Development in Premature Birth via Wnt5a/ROCK1 Axis

doi: 10.1007/s12015-024-10824-1

Figure Lengend Snippet: Proteomics analysis reveals differentially expressed proteins in term-Exos and preterm Exos. A : Hierarchical clustering of exosome proteomes in two groups ( n =3). Blue to red scale: heatmap representing the log2 fold changes in protein pool levels relative to term-Exos and preterm-Exos (14ba: term-Exos group, 15aa: preterm-Exos group). B : The GO enrichments of differentially expressed proteins in two groups. The Y-axis represents the GO function names, the X-axis values are the significance P -values transformed by -Log10, the different colors of the circles indicates the number of differential proteins contained in the function, and the size of the circles represents the enrichment ratio (E-ration) of the differential proteins contained in the function. C : The bubble diagram was used to show the information of the top 20 enriched pathways in the KEGG enrichment analysis. The Y-axis represents the KEGG pathway name, the X-axis represents the RichFactor value, the color of the circle represents the enriched significance P -value after Log10 transformation, with the size of the circles indicating the E-ration of the differential proteins, and the different colors represent the number of differentially expressed proteins contained in the pathway. D : Volcano plot, the differential protein screening results are presented in this form. In the graph, the points represent proteins, with blue indicating downregulated proteins, red indicating upregulated proteins, and grayish-black indicating no difference, when compared term-Exos with preterm-Exos. E : The top 9 proteins in expression of upregulated proteins in term-Exos group compared with preterm-Exos, of which Wnt5a was included. F , G : WB was used to further verify the difference of Wnt5a between the two groups ( n =3, ** P<0.01 )

Article Snippet: Mix equal volumes and concentrations of recombinant human Wnt5a-GST protein (MCE, USA) and recombinant human ROCK1-His protein (proteintech, CHN), and divide them into three groups: GST-Wnt5a + ROCK1-His (Input) group, IgG group, and GST pull-down group.

Techniques: Transformation Assay, Expressing

Journal: Stem Cell Reviews and Reports

Article Title: HucMSCs-derived Exosomes Promote Lung Development in Premature Birth via Wnt5a/ROCK1 Axis

doi: 10.1007/s12015-024-10824-1

Figure Lengend Snippet: Wnt5a could directly interact with ROCK1. A : To further research the probably mechanism of Wnt5a in promoting lung development, the h-AT2 cells were transfected with Wnt5a-Flag to make Wnt5a be overexpressed. After 72h, proteins were collected to conduct Co-IP experiment via flag beads. Then, proteins of IgG and IP groups were conducted proteomic analysis, and then common functional annotations (GO classification, subcellular localization classification, COG/KOG classification, and KEGG pathway classification analysis) were made for the identified proteins. B : Venn analysis of the identified peptides, of which existing in IP-group only were ranked from highest to lowest and taken the top ten proteins. C : AlphaFold Server was used to predict interactions between Wnt5a and ROCK1. The results indicated a potential interaction between these two proteins, with the blue area representing the possible functional domains that are closely associated. D : In Co-IP experiment, Wnt5a and ROCK1 proteins were positively expressed in both Input group and IP group, suggesting the interaction between Wnt5a and ROCK1. In the pull-down experiment, human recombinant protein WNT5A-GST was first mixed with human recombinant protein Rock1-His in equal volume and concentration, and then the protein mixture was purified by incubation with GST magnetic beads, and the expression of His label was verified by WB to further verify the direct interaction between Wnt5a and ROCK1

Article Snippet: Mix equal volumes and concentrations of recombinant human Wnt5a-GST protein (MCE, USA) and recombinant human ROCK1-His protein (proteintech, CHN), and divide them into three groups: GST-Wnt5a + ROCK1-His (Input) group, IgG group, and GST pull-down group.

Techniques: Transfection, Co-Immunoprecipitation Assay, Functional Assay, Recombinant, Concentration Assay, Purification, Incubation, Magnetic Beads, Expressing

Journal: Stem Cell Reviews and Reports

Article Title: HucMSCs-derived Exosomes Promote Lung Development in Premature Birth via Wnt5a/ROCK1 Axis

doi: 10.1007/s12015-024-10824-1

Figure Lengend Snippet: Term-Exos promoted the phosphorylation of ROCK1 and accelerated autophagy via Wnt5a. A : To evaluate whether term-Exos could mediate the phosphorylation of ROCK1 and be related with autophagy of lung epithelial cells, we examined the expressions of ROCK1, p-ROCK1, Wnt5a and LC3B in lung tissues by WB. B : Compared with Sham groups, the expression of LC3B in term-Exos groups were accelerated both in E13.5 and E18.5 stages, which in preterm-Exos group was only increased in E18.5 ( n =3, *P<0.05, **P<0.01 ). C : Compared with Sham groups, the expressions of Wnt5a were increased both in E13.5 and E18.5 stages of term-Exos groups, but just promoted in E18.5 stage of preterm-Exos group, which was obviously lower than term-Exos group ( n =3 *P<0.05,**P<0.01 ). D : The expressions of ROCK1 of term-Exos groups were obviously increased in E13.5 and E18.5 stages, while in preterm-Exos groups was obviously increased in E18.5 stages ( n =3, *P<0.05,**P<0.01 ). E: The phosphorylation of ROCK1 was calculated by the ratio of p-ROCK1 to ROCK1 protein expression levels. Compared with Sham groups and term-Exos groups, the ratio of preterm-Exos group was decreased in E13.5 and E18.5 ( n =3, *P<0.05,**P<0.01 ). F : The expressions of Atg5 in E13.5 and E18.5 groups were examined via WB. G : In E13.5 stage, the expressions of Atg5 in preterm group was the highest while in E13.5 stage it decreased to the lowest ( n =3 , *P<0.05, **P<0.01 )

Article Snippet: Mix equal volumes and concentrations of recombinant human Wnt5a-GST protein (MCE, USA) and recombinant human ROCK1-His protein (proteintech, CHN), and divide them into three groups: GST-Wnt5a + ROCK1-His (Input) group, IgG group, and GST pull-down group.

Techniques: Expressing

Journal: Stem Cell Reviews and Reports

Article Title: HucMSCs-derived Exosomes Promote Lung Development in Premature Birth via Wnt5a/ROCK1 Axis

doi: 10.1007/s12015-024-10824-1

Figure Lengend Snippet: A schematic model showing the proposed mechanism for the role of exosomal Wnt5a in lung development. Term-Exos: Term infant umbilical cord mesenchymal stem cells-derived exosomes, Ror: Receptor tyrosine kinase-like orphan receptor; FZD 6 : Frizzled receptor 6; ROCK1: Rho-associated protein kinase 1; RhoA: Ras homolog gene family, member A; LC3B: Microtubule-associated proteins 1A/1B light chain 3B; P: Phosphorylation; α-SMA: alpha smooth muscle actin

Article Snippet: Mix equal volumes and concentrations of recombinant human Wnt5a-GST protein (MCE, USA) and recombinant human ROCK1-His protein (proteintech, CHN), and divide them into three groups: GST-Wnt5a + ROCK1-His (Input) group, IgG group, and GST pull-down group.

Techniques: Derivative Assay