Journal: Neural regeneration research

Article Title: CHCHD2 Thr61Ile mutation impairs F1F0-ATPase assembly in in vitro and in vivo models of Parkinson's disease.

doi: 10.4103/1673-5374.378010

Figure Lengend Snippet: Figure 2 ｜Cellular ATP levels and F1F0-ATPase activity in isolated mitochondria in a in SH-SY5Y cell model of Parkinson’s disease. (A, B) Flag-vector: LV-Flag-vector (Ubi-MCS-3FLAG-SV40-puromycin vector)-transfected SH-SY5Y cells cultured in MEM-F12 medium; Flag-vector + MPP+: LV-Flag-vector- transfected SH-SY5Y cells treated with MPP+ (500 μM for 24 hours); CHCHD2-Flag + MPP+: LV-CHCHD2-Flag-transfected SH-SY5Y cells treated with MPP+ (500 μM for 24 hours); CHCHD2-T61I + MPP+: LV-CHCHD2-T61I-Flag-transfected SH-SY5Y cells treated with MPP+ (500 μM for 24 hours). (A) Detection of ATP levels in control or MPP+-treated SH-SY5Y cells stably transfected with empty vector or constructs encoding WT or T61I-mutant CHCHD2. (B) Effect of WT or T61I-mutant CHCHD2 on ATP synthase specific activity in control or MPP+-treated cells. (C) Effect of WT or T61I-mutant CHCHD2 on ATP synthase specific activity in AVV-transfected mice with or without MPTP treatment. CHCHD2-Flag + MPTP: AVV-CHCHD2-Flag–transfected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks); CHCHD2-T61I + MPTP: AVV-CHCHD2-T61I–transfected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks); Flag-vector: AVV-Flag (CMV- betaGlobin-MCS-3Flag-SV40 polyA (GV411) vector)-transfected mice treated with normal saline (twice a week, for 5 weeks); Flag-vector + MPTP: AVV-Flag-transfected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks). Data are expressed as the mean ± SD (n = 3 independent experiments). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (one-way analysis of variance followed by Tukey’s post hoc test). ATP: Adenosine triphosphate; MPP+: 1-methyl-4-phenylpyridinium; MPTP: 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine.

Article Snippet: Detection of mitochondrial permeability transition pore opening Opening of the mitochondrial permeability transition pore (mPTP) was detected by the calcein-AM/cobalt assay following the manufacturer’s instructions (C2009S, Beyotime Biotechnology).

Techniques: Activity Assay, Isolation, Plasmid Preparation, Transfection, Cell Culture, Control, Stable Transfection, Construct, Mutagenesis, Saline

Journal: Neural regeneration research

Article Title: CHCHD2 Thr61Ile mutation impairs F1F0-ATPase assembly in in vitro and in vivo models of Parkinson's disease.

doi: 10.4103/1673-5374.378010

Figure Lengend Snippet: Figure 3 ｜CHCHD2 T61I mutation promotes mitochondrial permeability transition pore (mPTP) opening in SH-SY5Y cells. CHCHD2-Flag: LV-CHCHD2-Flag-transfected SH-SY5Y cells; CHCHD2-T61I: LV-CHCHD2-T61I-Flag-transfected SH-SY5Y cells; Flag-vector: LV-Flag-vector (Ubi-MCS-3FLAG-SV40-puromycin vector)-transfected SH-SY5Y cells. (A) Calcein-AM and CoCl2 were administered to live cells expressing WT or T61I-mutant CHCHD2, and fluorescence intensity was detected by flow cytometry. (B) Effect of WT or the T61I-mutant CHCHD2 on mPTP opening as determined by detecting mitochondrial membrane potential using JC-1 fluorescent probes (white squares) in control or MPP+-treated SH-SY5Y cells. The fluorescence intensity of intracellular JC-1 aggregates (red) in the MPP+ + CHCHD2-Flag group was greater than that in the MPP+ + Flag-vector group. The CHCHD2-T61I group showed an increased amount of JC-1 monomers (green). Scale bars: 10 μm. (C) The mean fluorescence intensity of JC-1 aggregates or monomers in control or MPP+-treated SH-SY5Y cells expressing WT or T61I-mutant CHCHD2. Data are expressed as the mean ± SD (n = 3 independent experiments). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (one-way analysis of variance followed by Tukey’s post hoc test). Calcein-AM: Calcein acetoxymethyl ester; MPP+: 1-methyl-4-phenylpyridinium; mPTP: mitochondrial permeability transition pore; WT: wild type.

Article Snippet: Detection of mitochondrial permeability transition pore opening Opening of the mitochondrial permeability transition pore (mPTP) was detected by the calcein-AM/cobalt assay following the manufacturer’s instructions (C2009S, Beyotime Biotechnology).

Techniques: Mutagenesis, Permeability, Transfection, Plasmid Preparation, Expressing, Fluorescence, Flow Cytometry, Membrane, Control

Journal: Neural regeneration research

Article Title: CHCHD2 Thr61Ile mutation impairs F1F0-ATPase assembly in in vitro and in vivo models of Parkinson's disease.

doi: 10.4103/1673-5374.378010

Figure Lengend Snippet: Figure 5 ｜CHCHD2 T61I mutation aggravates movement deficits and nigral DA neuron function in a mouse model of PD. (A) Experimental design. AAV-CHCHD2, AAV-CHCHD2 T61I, or AAV-Vector was stereotaxically injected into the substantia nigra pars compacta, and 4 weeks later mice were injected intraperitoneally with normal saline or MPTP for 5 weeks. One day after the last MPTP/normal saline injection, behavioral tests were performed, and then the mice were sacrificed. Flag-vector: AVV-Flag(CMV-betaGlobin-MCS-3Flag-SV40 polyA (GV411) vector)-transfected mice treated with normal saline (twice a week, for 5 weeks); Flag-vector+MPTP: AVV-Flag– transfected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks); CHCHD2-Flag+MPTP: AVV-CHCHD2-Flag–infected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks); CHCHD2-T61I+MPTP: AVV-CHCHD2-T61I–transfected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks). (B) The grasping test was used to examine grip strength. (C) The pole-climbing test was used to examine bradykinesia. (D) The rotarod test was used to examine motor coordination. (E) Representative immunohistochemical staining of TH-positive neurons in the substantia nigra (SN). There were fewer TH-positive neurons in the SN in the AAV-CHCHD2-T61I + MPTP group than in the AAV-Flag-vector + MPTP and AAV-CHCHD2-Flag + MPTP groups. Scale bars: 800 μm. (F) Quantification of the TH-positive cells shown in E. (G) Representative images of double-immunofluorescent staining for CHCHD2-Flag (green, Alexa Fluor 488) and TH (red, Alexa Fluor 555) (white squares) in the SN. There were significantly fewer TH-positive neurons in the SN in the AAV-CHCHD2-T61I + MPTP group than in the AAV-Flag-vector + MPTP and AAV-CHCHD2-Flag + MPTP groups. Scale bars: 50 μm. (H) Quantification of TH mean fluorescence in the SN as shown in G. (I) Representative immunoblot for TH in the SN. (J) TH expression in the SN. Results are expressed as the mean ± SD (n ≥ 6). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (one-way analysis of variance followed by Tukey’s post hoc test). ACTB: β-Actin; AAV: adeno-associated virus; DA: dopaminergic; DAPI: 4′,6-diamidino-2-phenylindole; MPTP: 1-methyl-4-phenyl- 1,2,3,6-tetrahydropyridine; TH: tyrosine hydroxylase; SN: substantia nigra.

Article Snippet: Detection of mitochondrial permeability transition pore opening Opening of the mitochondrial permeability transition pore (mPTP) was detected by the calcein-AM/cobalt assay following the manufacturer’s instructions (C2009S, Beyotime Biotechnology).

Techniques: Mutagenesis, Plasmid Preparation, Injection, Saline, Transfection, Infection, Immunohistochemical staining, Staining, Fluorescence, Western Blot, Expressing, Virus

Journal: Neural regeneration research

Article Title: CHCHD2 Thr61Ile mutation impairs F1F0-ATPase assembly in in vitro and in vivo models of Parkinson's disease.

doi: 10.4103/1673-5374.378010

Figure Lengend Snippet: Figure 8 ｜CHCHD2 modulates OSCP expression in MPP+-/MPTP-induced PD models. (A, B) Flag-vector: LV-Flag-vector (Ubi-MCS-3FLAG-SV40-puromycin vector)-transfected SH-SY5Y cells cultured in MEM-F12 medium; Flag-vector+MPP+: LV-Flag-vector–transfected SH- SY5Y cells treated with MPP+ (500 μM for 24 hours); CHCHD2-Flag+MPP+: LV-CHCHD2-Flag–transfected SH-SY5Y cells treated with MPP+ (500 μM for 24 hours); CHCHD2-T61I + MPP+: LV-CHCHD2-T61I-Flag–transfected SH-SY5Y cells treated with MPP+ (500 μM for 24 hours). (A) Representative immunoblot of OSCP, CHCHD2, and ACTB in MPP+-treated SH-SY5Y cells. (B) Quantification of relative OSCP expression shown in A. (C, D) Flag-vector: AVV-Flag(CMV-betaGlobin-MCS-3Flag-SV40 polyA (GV411) vector)-transfected mice treated with normal saline (twice a week, for 5 weeks); Flag-vector + MPTP: AVV-Flag-infected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks); CHCHD2-Flag + MPTP: AVV-CHCHD2-Flag- infected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks); CHCHD2-T61I + MPTP: AVV-CHCHD2-T61I–transfected mice treated with MPTP (25 mg/kg, twice a week, for 5 weeks). (C) Representative immunoblot of OSCP, CHCHD2, and ACTB in control or MPTP-treated mice. (D) Quantification of relative OSCP expression shown in C. Data normalized to SH-SY5Y cells overexpressing Flag-vector or C57BL/6J mice transfected with AAV-Flag are expressed as the mean ± SD (n = 3 independent experiments for cells, n = 6 for animals). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (one-way analysis of variance followed by Tukey’s post hoc test). ACTB: β-Actin; MPP+: 1-methyl-4-phenylpyridinium; MPTP: 1-methyl- 4-phenyl-1,2,3,6-tetrahydropyridine; OSCP: oligomycin sensitivity conferral protein.

Article Snippet: Detection of mitochondrial permeability transition pore opening Opening of the mitochondrial permeability transition pore (mPTP) was detected by the calcein-AM/cobalt assay following the manufacturer’s instructions (C2009S, Beyotime Biotechnology).

Techniques: Expressing, Plasmid Preparation, Transfection, Cell Culture, Western Blot, Saline, Infection, Control

Journal: Frontiers in Immunology

Article Title: Bactericidal/Permeability-Increasing Protein Is an Enhancer of Bacterial Lipoprotein Recognition

doi: 10.3389/fimmu.2018.02768

Figure Lengend Snippet: Competition of bLPs with lipopolysaccharide for binding to BPI. BPI binding assays with LPS biotin (A–C,F–H) or Pam 2 CSK 4 biotin-coated plates (D,E) . “unc.” shows binding of rBPI in uncoated wells treated otherwise identically (A,B) . rBPI was pre-incubated with increasing concentrations of LPS EC (A,D) , the racemate Pam 3 CSK 4 (B) , ( R )-Pam 3 CSK 4 (C,E) and ( S )-Pam 3 CSK 4 (C) or peptidoglycan of S. aureus (PGN SA; F ). Furthermore, pre-incubations of rBPI with different heat-inactivated bacterial lysates are shown (G) . Preparations of rBPI and neutrophil BPI of two different sources [BPI N(W) and BPI N(A) ] were pre-incubated with ( R )-Pam 3 CSK 4 (H) . Absorbance measured at 450 nm for wells with BPI alone was set to 100% to ensure comparability between the different ligands. All results are shown as means ± SD of three biological replicates.

Article Snippet: Bactericidal/permeability-increasing protein (BPI) extracted from neutrophils was purchased from Athens Research and Technology [Athens, GA, USA; BPI N(A) ] and Wieslab AB [Malmö, Sweden; BPI N(W) ].

Techniques: Binding Assay, Incubation

Journal: BMC pregnancy and childbirth

Article Title: Expression of insulin-like growth factor binding protein-3 in HELLP syndrome.

doi: 10.1186/s12884-023-06074-7

Figure Lengend Snippet: Fig. 1 Levels of IGFBP-3/IGF-1/TGF-β1/VEGF in maternal blood of three groups (*P < 0.05, **P < 0.01)

Article Snippet: The reagents comprised: Human IGFBP-3 ELISA kit (R&B, DY675, USA), IGF-1 ELISA kit (R&B, DY291-05, USA), TGF-β1 ELISA kit (R&B, DY240-5,USA), Human VEGF ELISA Kit (R&B, DY293B-5,USA), goat serum (Proteintech Inc., B900780, USA), Tween-20 (Sigma Inc., T2700, USA), substrate solution (R&B,DY999, USA), bovine serum albumin (Sigma, B2064-50,USA), Annexin V-FITC/PI apoptosis assay kit (Novizan, A21101, China), endothelial cell culture medium (ScienCell, 1001, USA), recombinant human IGFBP-3 protein (YiqiaoShenzhou Technology Co., Ltd., 10430-H07H, China).

Techniques:

Journal: BMC pregnancy and childbirth

Article Title: Expression of insulin-like growth factor binding protein-3 in HELLP syndrome.

doi: 10.1186/s12884-023-06074-7

Figure Lengend Snippet: Fig. 2 Levels of IGFBP-3/IGF-1/TGF-β1/VEGF in umbilical blood of three groups (*P < 0.05, **P < 0.01)

Article Snippet: The reagents comprised: Human IGFBP-3 ELISA kit (R&B, DY675, USA), IGF-1 ELISA kit (R&B, DY291-05, USA), TGF-β1 ELISA kit (R&B, DY240-5,USA), Human VEGF ELISA Kit (R&B, DY293B-5,USA), goat serum (Proteintech Inc., B900780, USA), Tween-20 (Sigma Inc., T2700, USA), substrate solution (R&B,DY999, USA), bovine serum albumin (Sigma, B2064-50,USA), Annexin V-FITC/PI apoptosis assay kit (Novizan, A21101, China), endothelial cell culture medium (ScienCell, 1001, USA), recombinant human IGFBP-3 protein (YiqiaoShenzhou Technology Co., Ltd., 10430-H07H, China).

Techniques:

Journal: BMC pregnancy and childbirth

Article Title: Expression of insulin-like growth factor binding protein-3 in HELLP syndrome.

doi: 10.1186/s12884-023-06074-7

Figure Lengend Snippet: Fig. 3 Correlation analysis between IGFBP-3 and IGF-1/TGF-β1/VEGF (**P < 0.01, Line 1: results of correlation analysis in maternal blood; Line 2: results of correlation analysis in umbilical blood. The correlation coefficient (r) and regression equation are shown in the figure

Article Snippet: The reagents comprised: Human IGFBP-3 ELISA kit (R&B, DY675, USA), IGF-1 ELISA kit (R&B, DY291-05, USA), TGF-β1 ELISA kit (R&B, DY240-5,USA), Human VEGF ELISA Kit (R&B, DY293B-5,USA), goat serum (Proteintech Inc., B900780, USA), Tween-20 (Sigma Inc., T2700, USA), substrate solution (R&B,DY999, USA), bovine serum albumin (Sigma, B2064-50,USA), Annexin V-FITC/PI apoptosis assay kit (Novizan, A21101, China), endothelial cell culture medium (ScienCell, 1001, USA), recombinant human IGFBP-3 protein (YiqiaoShenzhou Technology Co., Ltd., 10430-H07H, China).

Techniques:

Journal: Respiratory Research

Article Title: Sodium propionate alleviates bronchopulmonary dysplasia by inhibiting ferroptosis through the SLC7A11/GPX4 pathway in pulmonary endothelial cells

doi: 10.1186/s12931-026-03535-3

Figure Lengend Snippet: Ferroptosis and vascular arrest are present in the rat model of BPD induced by hyperoxia. A Comparison of body size and lung volume of rats after euthanasia; B Weight plot of rats in CON and BPD groups; C HE staining of rat lung tissue, scale bar: 50 μm; D Determination of mRNA contents of PTGS2, SLC7A11 and GPX4 in rat lung tissues, GAPDH was used as an internal control; E Quantitative analysis of radial alveolar count in rat lung tissues by HE staining; F - G . Protein expression and gray value quantitative analysis of PTGS2, SLC7A11 and GPX4 in lung tissue of rats, GAPDH was used as a whole protein internal control; The levels of LPO, MDA, SOD and GSH in serum of ( H - K ). L Determination of VEGFA and CD31 mRNA content in rat lung tissue, GAPDH was used as internal control; Protein expression and gray value quantification of VEGFA and CD31 in lung tissues of ( M - N ). GAPDH was used as a whole protein internal control. N = 6, p < 0.05 for statistically significant (* p < 0.05, * * p < 0.01, * * * p < 0.001, * * * * p < 0.0001)

Article Snippet: The GAPDH and CD31 antibodies were procured from Proteintech (Cat No.60004-1-Ig, Cat No.11265-1-AP, Wuhan, China), while the PTGS2, SLC7A11, GPX4, and VEGFA antibodies were acquired from MedChemExpress (HY- P80091 , HY- P80935 , HY- P80450 , HY- P80929 , MCE, USA).

Techniques: Comparison, Staining, Control, Expressing

Journal: Respiratory Research

Article Title: Sodium propionate alleviates bronchopulmonary dysplasia by inhibiting ferroptosis through the SLC7A11/GPX4 pathway in pulmonary endothelial cells

doi: 10.1186/s12931-026-03535-3

Figure Lengend Snippet: Effect of SLC7A11 knockdown on SP intervention. A - C SLC7A11 siRNA knockdown efficiency was detected by RT-qPCR and Western blot; D - E PTGS2, CD31 and VEGFA protein detection and gray value quantification of HUVECs, GAPDH was used as an internal control; F - H , J Fluorescence plots and quantification of LPO and ROS of HUVECs, scale scale =100 μm, This fluorescent probe can emit red fluorescence under normal circumstances, and the fluorescence will change from red to green as lipid peroxidation occurs. The formation of lipid peroxides can be detected with high sensitivity through the fluorescence intensities of red and green.; I GSH measurement of HUVEC. Two-sided unpaired t test was used for comparison between the two groups. N =6, p <0.05 was considered statistically significant (* p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001, ns: no difference)

Article Snippet: The GAPDH and CD31 antibodies were procured from Proteintech (Cat No.60004-1-Ig, Cat No.11265-1-AP, Wuhan, China), while the PTGS2, SLC7A11, GPX4, and VEGFA antibodies were acquired from MedChemExpress (HY- P80091 , HY- P80935 , HY- P80450 , HY- P80929 , MCE, USA).

Techniques: Knockdown, Quantitative RT-PCR, Western Blot, Control, Fluorescence, Comparison

Journal: Respiratory Research

Article Title: Sodium propionate alleviates bronchopulmonary dysplasia by inhibiting ferroptosis through the SLC7A11/GPX4 pathway in pulmonary endothelial cells

doi: 10.1186/s12931-026-03535-3

Figure Lengend Snippet: Effect of GPX4 knockdown on SP intervention. A - C GPX4 siRNA knockdown efficiency was detected by RT-PCR and western blot; D - E PTGS2, CD31 and VEGFA protein detection and gray value quantification of HUVECs, GAPDH was used as an internal control; F - H , J Fluorescence plots and quantification of LPO and ROS of HUVEC, scale scale =100 μm, This fluorescent probe can emit red fluorescence under normal circumstances, and the fluorescence will change from red to green as lipid peroxidation occurs. The formation of lipid peroxides can be detected with high sensitivity through the fluorescence intensities of red and green.; I GSH measurement of HUVECs. Two-sided unpaired t test was used for comparison between the two groups. N =6, p <0.05 was considered statistically significant (* p <0.05, ** p <0.01, *** p <0.001, ns: no difference)

Article Snippet: The GAPDH and CD31 antibodies were procured from Proteintech (Cat No.60004-1-Ig, Cat No.11265-1-AP, Wuhan, China), while the PTGS2, SLC7A11, GPX4, and VEGFA antibodies were acquired from MedChemExpress (HY- P80091 , HY- P80935 , HY- P80450 , HY- P80929 , MCE, USA).

Techniques: Knockdown, Reverse Transcription Polymerase Chain Reaction, Western Blot, Control, Fluorescence, Comparison

Journal: Respiratory Research

Article Title: Sodium propionate alleviates bronchopulmonary dysplasia by inhibiting ferroptosis through the SLC7A11/GPX4 pathway in pulmonary endothelial cells

doi: 10.1186/s12931-026-03535-3

Figure Lengend Snippet: A proposed mechanistic model illustrating how SP alleviates BPD. In HUVECs, hyperoxia exposure suppresses the SLC7A11/GPX4 antioxidant axis, leading to increased lipid peroxidation (LPO) and reactive oxygen species (ROS), thereby triggering ferroptosis. Concurrently, the expression of pro-angiogenic factors (VEGFA, CD31) is downregulated, resulting in impaired vascular development and alveolar simplification—the hallmark pathological features of BPD. SP treatment counteracts hyperoxia-induced damage by upregulating the SLC7A11/GPX4 pathway. The activation of this axis inhibits ferroptosis (reducing LPO and ROS) and restores the expression of VEGFA and CD31, thereby promoting angiogenesis and preserving alveolar structure. This model summarizes the core finding: SP exerts protective effects against BPD by modulating ferroptosis and vascular development through the SLC7A11/GPX4 signaling axis

Article Snippet: The GAPDH and CD31 antibodies were procured from Proteintech (Cat No.60004-1-Ig, Cat No.11265-1-AP, Wuhan, China), while the PTGS2, SLC7A11, GPX4, and VEGFA antibodies were acquired from MedChemExpress (HY- P80091 , HY- P80935 , HY- P80450 , HY- P80929 , MCE, USA).

Techniques: Expressing, Activation Assay, Preserving

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: The enhanced efficacy of MMCs following catalpol pretreatment may be associated with VEGF-A secretion. ( A ) A total of 183 differentially expressed genes were identified between the ordinary MMC group and the catalpol-pretreated MMC group. ( B ) RNA-seq volcano plot illustrating the genes that were differentially expressed between the cisplatin treatment group and ordinary TCMK-1 cells ( n = 4), with red indicating upregulated gsenes, blue representing downregulated genes, and gray denoting nondifferentially expressed genes. ( C ) GO enrichment analysis revealed the top 20 biological processes associated with the identified pathways. ( D , E ) Results from the GSEA of the GO terms ranked by P value and Q value, indicating the activation of retinol metabolism and VEGF production-related pathways in catalpol-pretreated MMCs. ( F ) ELISA results demonstrating that the content of VEGF-A in the conditioned medium from the catalpol-pretreated MMC group was significantly greater than that in the control group ( n = 3). con-CM: conventional MMC supernatant group; cata-CM: catalpol-pretreated MMC supernatant group. ** P < 0.01

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: RNA Sequencing, Activation Assay, Enzyme-linked Immunosorbent Assay, Control

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: Knockdown of VEGF-A in MMCs Abrogates the Protective Effect of Catalpol-Pretreated MMCs against AKI. ( A , B ) Western blot showing the expression of VEGF-A in MMCs transfected with different siRNAs and treated with catalpol for 24 h ( n = 3). ( C , D ) Serum levels of SCr and BUN in the different groups of mice ( n = 6). ( E - I ) Semiquantitative analysis of histopathological scores ( n = 6) and representative images of PAS-stained kidney tissue sections from different groups of mice (400×). ( J , K ) Western blot showing the expression level of the injury marker KIM-1 and the inflammatory markers p-p65 and TNF-α in the kidneys of mice from different groups and statistical analysis ( n = 6). (L–N) Levels of the oxidative stress markers T-SOD, GSH, and MDA in the kidney tissues of mice from different groups ( n = 6). ( O , P ) Western blot showing the expression of the necroptosis markers RIP, RIP3, and MLKL, as well as their phosphorylated forms, in the kidneys of mice from different groups and statistical analysis ( n = 6). ( Q - T ) Immunohistochemical staining images showing the expression of the necroptosis marker MLKL (400×). Groups in panels A-B: MMC-cata: catalpol-pretreated MMCs; NC: MMCs transfected with the empty vector; siRNA1: MMCs transfected with siRNA1; siRNA2: MMCs transfected with siRNA2; siRNA3: MMCs transfected with siRNA3. Groups in panels C-T: Ctrl: Control group; CP: cisplatin model group; MMC-cata: catalpol-pretreated MMC treatment group; siRNA3: siRNA3-transfected MMC treatment group. *** P < 0.001, ** P < 0.01, * P < 0.05

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: Knockdown, Western Blot, Expressing, Transfection, Staining, Marker, Immunohistochemical staining, Plasmid Preparation, Control

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: Neutralizing VEGF-A or Blocking VEGFR2 Abrogates the Protective Effect of Catalpol-Pretreated MMCs in vitro. ( A , B) Immunohistochemical staining images showing the expression of VEGFR2 in the kidneys of mice from different groups (400×). ( C , D ) Immunofluorescence staining images showing the expression of VEGFR2 in TCMK-1 cells from different groups (400×). ( E , F ) CCK-8 and LDH assay results showing the proliferative activity and cytotoxicity of TCMK-1 cells in different groups ( n = 6). ( G , H ) Western blot showing the expression of the inflammatory markers p-p65 and TNF-α in cells from different groups and statistical analysis ( n = 3). ( I - P ) Levels of the oxidative stress indicators ROS and T-SOD and ROS staining in TCMK-1 cells ( n = 3). ( Q - R ) Western blot showing the expression of the necroptosis markers RIP, RIP3, and MLKL, as well as their phosphorylated forms, in cells from different groups and statistical analysis ( n = 3). CON : control group; CP : cisplatin injury group; cata-CM : catalpol-pretreated MMC supernatant group; VEGF-ab : catalpol-pretreated MMC supernatant + VEGF-A neutralizing antibody group; Ki8751: catalpol-pretreated MMC supernatant + VEGFR2 inhibitor Ki8751 group; MAZ51: catalpol-pretreated MMC supernatant + VEGFR3 inhibitor MAZ51 group. *** P < 0.001, ** P < 0.01, * P < 0.05

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: Blocking Assay, In Vitro, Immunohistochemical staining, Staining, Expressing, Immunofluorescence, CCK-8 Assay, Lactate Dehydrogenase Assay, Activity Assay, Western Blot, Control

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: Application of VEGF-A Alone Alleviates Cisplatin-Induced Renal Tubular Epithelial Cell Injury. ( A , B ) CCK-8 and LDH assay results showing the proliferative activity and cytotoxicity of TCMK-1 cells in different groups ( n = 6). ( C , D ) Western blot showing the expression of the inflammatory markers p-p65 and TNF-α in cells from different groups and statistical analysis ( n = 3). ( E - J ) The levels of the oxidative stress indicators ROS and T-SOD and ROS staining in TCMK-1 cells ( n = 3). ( K , L ) Western blot showing the expression of the necroptosis markers RIP, RIP3, and MLKL, as well as their phosphorylated forms, in cells from different groups and statistical analysis ( n = 3). CON control group; CP cisplatin injury group; VEGF-A VEGF-A treatment group; Ki8751: VEGF-A + VEGFR2 inhibitor Ki8751 group. *** P < 0.001, ** P < 0.01, * P < 0.05

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: CCK-8 Assay, Lactate Dehydrogenase Assay, Activity Assay, Western Blot, Expressing, Staining, Control

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: Pretreatment of MMCs with catalpol may suppress the STAT3 pathway and activate the p38 pathway. ( A , B ) Western blot showing the expression of STAT3, p38, and their phosphorylated proteins in kidney tissues from mice in different groups ( n = 6). Ctrl: control group; CP: cisplatin model group; MMC-cata: catalpol-pretreated MMC treatment group; siRNA3: siRNA3-transfected MMC treatment group. ( C , D ) Western blot showing the expression of STAT3, p38, and their phosphorylated proteins in cells from different groups ( n = 3). CON: control group; CP: cisplatin injury group; cata-CM: catalpol-pretreated MMC supernatant group; VEGF-ab: catalpol-pretreated MMC supernatant + VEGF-A neutralizing antibody group; Ki8751: catalpol-pretreated MMC supernatant + VEGFR2 inhibitor Ki8751 group; MAZ51: catalpol-pretreated MMC supernatant + VEGFR3 inhibitor MAZ51 group. ( E – F ) Western blot showing the expression of STAT3, p38, and their phosphorylated proteins in cells from different groups ( n = 3). CON: control group; CP: cisplatin injury group; VEGF-A: VEGF-A treatment group; and Ki8751: VEGF-A + VEGFR2 inhibitor Ki8751 group. *** P < 0.001, ** P < 0.01, * P < 0.05

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: Western Blot, Expressing, Control, Transfection

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: Catalpol binds to Wnt3A, activating the canonical Wnt pathway in MMCs and promoting VEGF-A secretion. ( A , B ) Molecular docking images of catalpol and Wnt3A. ( C ) Docking score of the interaction between catalpol and Wnt3A. ( D , E ) Interaction sites of catalpol and Wnt3A. ( F ) RMSDs of the molecular dynamics simulations of catalpol with the Wnt3A protein. ( G , H) RMSFs of the molecular dynamics simulations of catalpol and the Wnt3A protein; the numerical sequence represents different binding sites. ( I ) Free energy decomposition of different binding sites of catalpol to the Wnt3A protein. ( J ) Surface plasmon resonance was used to precisely measure the binding affinity and binding mode of catalpol to Wnt3A. ( K , L ) Western blot showing the expression levels of VEGF-A and Wnt3A, β-catenin, and TCF-4 in MMCs treated with different drugs ( n = 3). (M) VEGF-A content in the conditioned medium of MMCs under different drug treatments ( n = 3). Groups in panels J-K: MMC-con: normal MMCs; MMC-cata: catalpol pretreated MMCs; MSAB: catalpol- + canonical Wnt pathway blocker MSAB-pretreated MMCs. Groups in panel L: Con-CM: normal MMC-conditioned medium; cata-CM: catalpol-pretreated MMC-conditioned medium; MSAB: catalpol- + canonical Wnt pathway blocker MSAB-pretreated MMC-conditioned medium. *** P < 0.001, ** P < 0.01, * P < 0.05

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: Sequencing, Binding Assay, SPR Assay, Western Blot, Expressing

Journal: Stem Cell Research & Therapy

Article Title: Pretreatment of metanephric mesenchymal cells with catalpol mitigates acute kidney injury through VEGF-A secretion via multiple mechanisms

doi: 10.1186/s13287-026-04914-9

Figure Lengend Snippet: Catalpol-pretreated MMCs alleviate cisplatin-induced acute kidney injury through multiple pathways by promoting VEGF-A secretion. Catalpol-pretreated MMCs activate the canonical Wnt pathway, increasing the synthesis and secretion of VEGF-A. VEGF-A acts on VEGFR2 in renal tubular epithelial cells, mitigating inflammation, oxidative stress, and necroptosis by activating the p38 signaling pathway and suppressing STAT3 signaling, thereby alleviating cisplatin-induced acute kidney injury

Article Snippet: Ultimately, 5 ng/ml VEGF-A (HY- P72280 , MCE, USA), 0.1 μg/ml VEGF-A neutralizing antibody (sc-7249, Santa Cruz, USA), 10 nM VEGFR2 blocker Ki8751 (HY-12038, MCE, USA), and 5 μM VEGFR3 blocker MAZ51 (HY-116624, MCE, USA) were used.

Techniques: