Journal: Frontiers in Molecular Biosciences

Article Title: Identification of ATRNL1 and WNT9A as novel key genes and drug candidates in hypertrophic cardiomyopathy: integrative bioinformatics and experimental validation

doi: 10.3389/fmolb.2024.1458434

Figure Lengend Snippet: Validation of key gene expression in cell models. (A) Phalloidin immunofluorescence staining of the HCM cell model established by ISO. (B) The integrated optical density (IOD) of phalloidin is significantly increased at 48 h. (C) ATRNL1 (mRNA) expression is significantly increased at 48 h. (D) WNT9A is significantly increased at 24 h and 48 h.

Article Snippet: The sections were incubated with rabbit anti- ATRNL1 (bs-11504R, Bioss, 1/200) or rabbit anti- WNT9A (bs-1933R, Bioss, 1/200) overnight at 4°C.

Techniques: Expressing, Immunofluorescence, Staining

Journal: Frontiers in Molecular Biosciences

Article Title: Identification of ATRNL1 and WNT9A as novel key genes and drug candidates in hypertrophic cardiomyopathy: integrative bioinformatics and experimental validation

doi: 10.3389/fmolb.2024.1458434

Figure Lengend Snippet: Pathological morphology and ATRNL1/WNT9A IHC staining in different groups. (A) HE stains of four groups: model, model + miR-1469, model + propylthiouracil, and model + triptolide. The model + miR-1469 and model + propylthiouracil groups showed a slight improvement of abnormalities in myocardial tissues. (B) IHC staining of ATRNL1 . (C) IHC staining of WNT9A . (D) Only propylthiouracil, but not miR-1469 or triptolide, significantly inhibits the expression of ATRNL1 in the HCM model. (E) All of the three drugs suppress WNT9A expression. * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The sections were incubated with rabbit anti- ATRNL1 (bs-11504R, Bioss, 1/200) or rabbit anti- WNT9A (bs-1933R, Bioss, 1/200) overnight at 4°C.

Techniques: Immunohistochemistry, Expressing

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 1. Wnt9a is induced specifically in renal tubular epithelium in human CKD. (A) Representative micrographs show the expression and localization of Wnt9a in various types of human CKD. Nontumor kidney tissue from the patients who had renal cell carcinoma and underwent nephrectomy was used as normal controls. Arrows indicate Wnt9a-positive tubules. Scale bar, 50 mm. (B) Representative mi- crographs show Wnt9a localized in those tubules with nuclei negative for expression of Ki67. Sequential paraffin-embedded kidney sections from patients with IgAN were immunostained for Wnt9a and Ki67. Yellow arrows indicate Wnt9a-positive tubules lack Ki67 nuclear expression. Scale bar, 50 mm. (C) Colocalization of Wnt9a and p16INK4A in tubules of patients with IgAN. Sequential paraffin-embedded kidney sections were immunostained for Wnt9a and p16INK4A. Colocalization of Wnt9a and p16INK4A in tubules is indicated by arrows. Scale bar, 50 mm. (D and E) Scatter plots with linear regression show significant correlation between Wnt9a expression levels and (D) p16INK4A

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Expressing

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 2. Wnt9a is commonly induced and accompanied by increasing expression of gH2AX and loss of Klotho expression in various animal models of CKD. (A) Representative western blots showing the renal expression of Wnt9a, gH2AX, and Klotho in sham and injured kidneys 1 or 3 days after IRI. (B and C) Graphical representations of (B) Wnt9a, gH2AX, and (C) Klotho expression levels in three groups, as indicated. *P,0.05 versus sham controls (n=5–6). (D) Representative staining showing induction of Wnt9a protein expression in tubules in the UUO group. Scale bar, 50 mm. (E) Representative western blots showing the renal expression of Wnt9a, gH2AX, and Klotho in sham and obstructed kidneys 7 days after UUO. (F and G) Graphical representations of (F) Wnt9a, gH2AX, and (G) Klotho protein expression levels in two groups, as indicated. *P,0.05 versus sham controls (n=5–6). (H) Representative western blots showing renal Wnt9a, gH2AX, and Klotho expression in ADR nephropathy 3 weeks after ADR injection. CTL, control. (I and J) Graphical representations of (I) Wnt9a and gH2AX and (J) Klotho protein expression levels in two groups, as indicated. *P,0.05 versus normal controls (n=5–6). (K) Colocalization of Wnt9a and p16INK4A in ADR nephropathy tubules. Sequential paraffin-embedded kidney sections were immunostained for Wnt9a and p16INK4A. Colocalization of Wnt9a and p16INK4A in tubules is indicated by arrows. Scale bar, 50 mm.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Expressing, Western Blot, Staining, Injection, Control

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 3. Overexpression of Wnt9a promotes tubular injury and impairs renal function in IRI. (A) Experimental design. Green arrow indicates the injection of pcDNA3 or pFlag-Wnt9a plasmid. Red arrows indicate the timing of renal IRI surgery. (B) Representative western blots showing renal expression of Wnt9a in three groups, as indicated. (C) Graphical representation of Wnt9a protein ex- pression levels in three groups. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (D) Representative micro- graphs show kidney morphology after IRI injury in different groups of mice, as indicated. Images of PAS staining are shown. Boxed areas are enlarged in the bottom panels. Arrows indicate injured tubules. Scale bar, 50 mm. (E) Quantitative analyses of injured tubules in three groups, as indicated. Kidney sections were subjected to PAS staining. At least 20 randomly selected fields were evaluated under 3400 magnification and results were averaged for each kidney. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (F) Urinary NAG level, expressed as international units per gram creatinine, in different groups. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (G and H) Ectopic expression of Wnt9a increased Scr and Ualb levels in IRI mice. Quan- titative analysis of (G) Scr and (H) Ualb levels in three groups, as indicated. Scr was expressed as milligrams per deciliter, and Ualb was expressed as micrograms per milligram urinary creatinine. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). Ucr, urinary creatinine; UNx, unilateral nephrectomy.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Over Expression, Injection, Plasmid Preparation, Western Blot, Expressing, Staining

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 4. Ectopic expression of Wnt9a aggravates renal fibrosis in IRI mice. (A) Representative micrographs show that Wnt9a exac- erbated renal interstitial fibrosis. Kidney sections were subjected to Masson trichrome staining. Arrow indicates positive staining. Scale bar, 50 mm. (B) Graphical representation of kidney fibrotic lesion area in different groups after quantitative determination. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (C) Representative western blots show renal expression of fibronectin and a-SMA in three groups, as indicated. (D and E) Graphical representations of (D) fibronectin and (E) a-SMA expression levels in different groups. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (F) Graphical representation of the relative abundance of collagen I mRNA in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (G) Representative immunostaining micrographs show fibronectin, collagen I, and Fsp-1 expression in different groups. Boxed areas are enlarged and presented in the right column. Arrows indicate positive staining. Scale bar, 50 mm.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Expressing, Staining, Western Blot, Immunostaining

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 5. In vivo expression of exogenous Wnt9a promotes the expressions of b-catenin and targeted genes. (A) Immunohisto- chemical staining showing the renal induction of b-catenin and MMP-7 after injection of Wnt9a expression plasmid. Arrows indicate positive staining. (B and C) Graphical representations of the relative abundance of (B) MMP-7 and (C) PAI-1 mRNA in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (D) Representative western blots show renal expression of b-catenin, MMP-7, PAI-1, and Snail 1 in three groups, as indicated. (E–H) Graphical representations show expression of (E) b-catenin, (F) MMP-7, (G) PAI-1, and (H) Snail 1 in three groups. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6).

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: In Vivo, Expressing, Staining, Injection, Plasmid Preparation, Western Blot

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 6. Overexpression of Wnt9a after IRI accelerates cellular senescence in renal tubules. (A) Expression of p16INK4A mRNA in different groups was assessed by real-time PCR. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (B) Rep- resentative western blots show that Klotho expression was further downregulated after injection of Wnt9a expression plasmid. (C) Graphical representations of Klotho protein expression levels in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (D) Representative staining micrographs show p16INK4A and TGF-b1 expression and SA–b-gal activity in different groups. Paraffin sections were immunostained with antibodies against p16INK4A and TGF-b1. Frozen kidney sections were stained for SA–b-gal activity, which appears as bright-blue granular staining in the cytoplasm of tubular epithelial cells. Boxed areas are enlarged and presented in the right column. Arrows indicate positive staining. Scale bar, 50 mm.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Over Expression, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Injection, Plasmid Preparation, Staining, Activity Assay

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 7. Knockdown of Wnt9a ameliorates renal injury after IRI. (A) Experimental design. Green arrow indicates the injection of pLVX-shRNA (Ctrl-shR) or pLVX-shWnt9a (Wnt9a-shR) plasmids. Red arrows indicate the timing of renal IRI surgery. (B) Repre- sentative western blots show expression of Wnt9a was nearly completely abolished after injection with Wnt9a-shR plasmid. (C) Graphical representation of Wnt9a protein expression levels in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (D and E) Knockdown of Wnt9a significantly decreased Scr and secretion of Ualb. Quantification of (D) Scr and (E) Ualb levels in three groups, as indicated. Ualb level was expressed as micrograms per milligram urinary creatinine. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (F) Representative immunofluo- rescence micrographs show collagen I and fibronectin expression in three groups. Arrows indicate positive staining. Scale bar, 50 mm. (G) Representative western blots show fibronectin and a-SMA expression in three groups, as indicated. (H and I) Graphical representations of (H) fibronectin and (I) a-SMA protein expression levels in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). (J) Representative micrographs show collagen deposition in different groups, as indicated. Paraffin sections were used for Masson trichrome staining. Arrow indicates positive staining. Scale bar, 50 mm. (K) Graphical representation of the extent of fibrosis. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6). Ucr, urinary creatinine; UNx, unilateral nephrectomy.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Knockdown, Injection, shRNA, Western Blot, Expressing, Plasmid Preparation, Staining

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 8. Knockdown of Wnt9a inhibits tubular cell senescence in IRI model. (A) Representative micrographs show staining for p16INK4A

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Knockdown, Staining

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 9. Knockdown of p16INK4A inhibits Wnt9a-induced renal fibrosis in IRI mice. (A) Experimental design. Red arrow indicates the injection of pcDNA3, pFlag-Wnt9a, or pFlag-Wnt9a with pLVX-shp16INK4A (p16-shR) plasmid. Black arrows indicate the timing of renal IRI surgery. (B) Representative western blots show expression of p16INK4A was largely abolished after injection with p16-shR plasmid. (C) Graphical representation of p16INK4A protein expression levels in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6); #P,0.05 versus IRI injected with pFlag-Wnt9a (n=5–6). (D) Knockdown of p16INK4A pre- vented the increase in Scr levels induced by Wnt9a in IRI mice. Quantitative assessment of Scr level (mg/dl) in four groups, as in- dicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6); #P,0.05 versus IRI injected with pFlag-Wnt9a (n=5–6). (E) Representative micrographs show collagen deposition in different groups, as indicated. Paraffin sections were used for Masson trichrome staining. Arrow indicates positive staining. Scale bar, 50 mm. (F) Graphical representation of the extent of fibrosis after quantitative determination. (G) Representative western blots show fibronectin and a-SMA expression in four groups, as in- dicated. (H and I) Graphical representations of (H) fibronectin and (I) a-SMA protein expression levels in different groups, as indicated. *P,0.05 versus sham controls (n=5–6); †P,0.05 versus IRI alone (n=5–6); #P,0.05 versus IRI injected with pFlag-Wnt9a (n=5–6). (J) Representative immunostaining micrographs show fibronectin expression in different groups. Arrow indicates positive staining. Scale bar, 50 mm. UNx, unilateral nephrectomy.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Knockdown, Injection, Plasmid Preparation, Western Blot, Expressing, Staining, Immunostaining

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 10. Wnt9a induces cellular senescence in cultured proximal tubular epithelial cells. (A) Expression of p16INK4A mRNA in dif- ferent groups was assessed by real-time PCR in HK-2 cells transfected with empty vector (pcDNA3) or Wnt9a expression plasmid (pFlag-Wnt9a) for 24 hours. *P,0.05 versus pcDNA3 group (n=3). (B) Representative micrographs of HK-2 cells immunostained for gH2AX (red) and counterstained with DAPI (blue) after transfection with pcDNA3 or pFlag-Wnt9a. Arrow indicates positive staining for gH2AX. Scale bar, 10 mm. (C) Representative western blots show Wnt9a transfection decreases the phosphorylation of Rb and pro- motes expression of p14ARF, p53, and p21. (D–G) Graphical representations of the levels of phosphorylated (D) Rb and (E) p14ARF, (F) p53, and (G) p21 protein expression in two groups, as indicated. *P,0.05 versus pcDNA3 group (n=3). (H) Representative western blots show incubation with recombinant Wnt9a for 24 hours dose-dependently promotes secretion of TGF-b1. (I) Graphical representations of TGF-b1 protein expression levels in four groups, as indicated. *P,0.05 versus medium alone (n=3). (J) Representative western blots show ICG-001inhibits Wnt9a-induced p53 and TGF-b1 expression. HK-2 cells were preincubated with ICG-001 (5 mM) or neutralizing antibody against type II receptor of TGF-b1 (5 mg/ml), then treated with Wnt9a (50 ng/ml) for 24 hours. (K and L) Graphical repre- sentations of (K) p53 and (L) TGF-b1 protein expression levels in four groups, as indicated. *P,0.05 versus medium alone (n=3); †P,0.05 versus Wnt9a alone (n=3). (M) Representative western blots show interference of p16INK4A blocks Wnt9a-induced increase in a-SMA and TGF-b1 expression. HK-2 cells were transfected with control (CTL) or p16INK4A-specific siRNA, then treated with Wnt9a (50 ng/ml) for 24 hours. (N–P) Graphical representations of (N) p16INK4A, (O) a-SMA, and (P) TGF-b1 protein expression levels in three groups, as indicated. *P,0.05 versus CTL siRNA alone (n=3); †P,0.05 versus Wnt9a alone (n=3). siRNA, small interfering RNA.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Cell Culture, Expressing, Real-time Polymerase Chain Reaction, Transfection, Plasmid Preparation, Staining, Western Blot, Phospho-proteomics, Incubation, Recombinant, Control, Small Interfering RNA

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 11. Wnt9a induces cellular senescence in primary cultured tubular cells. (A) Representative micrographs show freshly isolated tubules and cultured primary tubular cells. Renal tubules were isolated from mouse kidneys and cultivated for use as primary tubular cells. (B) Representative micrographs show staining of SA–b-gal activity induced by Wnt9a. The primary cultured tubular cells were treated with Wnt9a (50 ng/ml) for 8 days before staining. Arrow indicates positive staining. (C) Representative micrographs of cells immunostained for gH2AX (red) and E-cadherin (green) and counterstained with DAPI (blue). Primary tubular cells were treated with Wnt9a or medium alone for 8 days. Arrows indicate positive staining. Scale bar, 10 mm. (D) Representative western blots show p16INK4A, p19ARF, phosphorylated Rb, and PCNA expression levels in two groups, as indicated. (E and F) Graphical representation of (E) p16INK4A and p19ARF, and (F) phosphorylated Rb and PCNA expression levels in two groups, as indicated. The primary cultured tubular cells were treated with Wnt9a (50 ng/ml) for 4 days before assessment. *P,0.05 versus medium alone (n=3). Ctrl, control.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Cell Culture, Isolation, Staining, Activity Assay, Western Blot, Expressing, Control

Journal: Journal of the American Society of Nephrology

Article Title: Wnt9a Promotes Renal Fibrosis by Accelerating Cellular Senescence in Tubular Epithelial Cells

doi: 10.1681/asn.2017050574

Figure Lengend Snippet: Figure 12. Wnt9a induces communication between senescent tubules and activated fibroblasts. (A) Representative western blots showing expression of PCNA and cyclin D1 after NRK-49F cells were treated with or without Wnt9a recombinant protein (50 ng/ml) for 24 hours. (B and C) Graphical representation of (B) PCNA and (C) cyclin D1 protein expression levels in two groups, as indicated. *P,0.05 versus medium alone (n=3). (D) Graphical representation shows that Wnt9a promoted NRK-49F cell proliferation as assessed by a colorimetric MTT assay. *P,0.05 versus medium alone (n=3). (E) Flow chart shows the experimental design and procedures. HK-2 cells were transfected with the Wnt9a expression plasmid for 24 hours to induce cellular senescence or with empty vector as controls. The supernatant was collected (Wnt9a-TCM or CTL-TCM) and used to stimulate NRK-49F cells. (F) Representative micrographs of NRK- 49F cells immunostained for fibronectin (red) and counterstained with DAPI (blue). NRK-49F cells were plated on coverslips and preincubated with neutralizing antibody against TGF-b1 receptor II (5 mg/ml) for 1 hour, then treated with Wnt9a-TCM or CTL-TCM for 24 hours before immunostaining. Scale bar, 20 mm. (G) Representative western blots show PCNA and a-SMA expression in three groups, as indicated. (H and I) Graphical representations of (H) PCNA and (I) a-SMA protein expression levels in different groups, as indicated. *P,0.05 versus CTL-TCM group (n=4); †P,0.05 versus Wnt9a-TCM alone (n=4). (J) Representative western blots show TGF- b1 promoted Wnt9a expression in cultured fibroblasts. NRK-49F cells were stimulated with TGF-b1 (5 ng/ml) for 24 hours. Whole-cell kidney lysates were analyzed for Wnt9a. (K) Graphical representation of Wnt9a levels in (J). *P,0.05 versus control (n=3). (L) Schematic presentation of reciprocal activation loop between senescent tubular cells and activated fibroblasts. In pathologic conditions, Wnt9a is dramatically upregulated. Wnt9a induces tubular cell senescence and promotes fibroblast activation. Tubular cells exhibiting the SASP secrete TGF-b1, which aggravates fibroblast proliferation and facilitates matrix protein production and deposition. The activated fibroblasts also produce Wnt9a, which further promotes tubular cell senescence. All of these lead to the pathogenesis of renal fibrotic foci. CTL-TCM, the supernatant collected from tubular cells with empty vector transfection; ECM, excellular matrix.

Article Snippet: HK-2 cells or NRK-49F cells were treated with human recombinant Wnt9a (R & D Systems, Minneapolis, MN) at the indicated concentration or transfected with Wnt9a expression plasmid (pFlag-Wnt9a).

Techniques: Western Blot, Expressing, Recombinant, MTT Assay, Transfection, Plasmid Preparation, Immunostaining, Cell Culture, Control, Activation Assay

Journal: Scientific Reports

Article Title: Enhancement of angiotensin II type 1 receptor-associated protein suppresses kidney inflammation in a mouse model of aristolochic acid nephropathy

doi: 10.1038/s41598-025-08642-7

Figure Lengend Snippet: Effects of AA treatment on renal expression of cellular senescence-related genes in ATRAP transgenic (Tg19) and littermate control (LC) mice. Relative renal mRNA expression of ( a ) p16, ( b ) Wnt9a and ( c ) Klotho, and ( d ) protein expression of SIRT1 in LC and Tg19 mice. Values are expressed as mean ± SE (n = 8 per group) and were analyzed using two-way ANOVA with Bonferroni’s post-hoc test. * P < 0.05 vs. vehicle; *** P < 0.001 vs. vehicle; † P < 0.05 vs. LC mice. Wnt9a, Wnt family member 9A; SIRT1, Sirtuin1; AA, aristolochic acid; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The TaqMan probes used for PCR were collagen 1α1 Mm00801666_g1; collagen 3α1, Mm01254476_m1; TGF-β, Mm01178820_m1; F4/80, Mm00802529_m1; MCP-1, Mm00441242_m1; TNF-α, Mm00443258_m1; IL-1β, Mm00434228_m1; Kim-1, Mm00506686_m1; IL-6, 00446190_m1; IFN-γ, Mm01168134_m1; Nox2, Mm00627_m1; Hmox-1, Mm00516005_m1; p16, Mm494449_m1; Wnt9a, Mm00460518_m1; Klotho, Mm00502002_m1; Angiotensinogen, Mm00599662_m1.

Techniques: Expressing, Transgenic Assay, Control

Journal: Molecular Metabolism

Article Title: Global mRNA sequencing of human skeletal muscle: Search for novel exercise-regulated myokines

doi: 10.1016/j.molmet.2017.01.007

Figure Lengend Snippet: Transcripts up-regulated 2 h after 45 min acute exercise of 70% of VO 2 max.

Article Snippet: The following pre-designed primers and probe sets were used (TaqMan assays; ThermoFisher, Foster City, California, United States) RPLP0 (Hs99999902_m1), MTRNR2L4 (Hs04276154_s1), SMPDL3A (Hs00378308_m1), FAM20C (Hs00398243_m1), WNT9A (Hs00243321_m1), TEK (Hs00945155_m1), FLT1 (Hs01052961_m1), CSF1 (Hs00174164_m1), C8G (Hs01113922_g1), CHSY1 (Hs00208704_m1), IL4R (Hs00166237_m1), LCN10 (Hs01596612_g1), STC2 (Hs00175027_m1), THBS4 (Hs00170261_m1), IGFBP2 (Hs01040719_m1), KAZALD1 (Hs00368867_g1), TNFRSF25 (Hs00600930_g1), and SCT (Hs00360814_g1).

Techniques: Binding Assay, Sequencing, Cell Differentiation, Immunopeptidomics

Journal: Regenerative Biomaterials

Article Title: Embedding MSCs in Si-HPMC hydrogel decreased MSC-directed host immune response and increased the regenerative potential of macrophages

doi: 10.1093/rb/rbac022

Figure Lengend Snippet: Gene and Taqman gene expression assay reference

Article Snippet: wnt9a , Wnt family member 9A , Rn01496604_m1.

Techniques: Gene Expression