Journal: International Dental Journal

Article Title: Endothelial Nitric Oxide Synthase Restores Diabetic Dentin Regeneration via AKT/Endothelial Nitric Oxide Synthase Axis

doi: 10.1016/j.identj.2026.109511

Figure Lengend Snippet: Both p-AKT/AKT and eNOS expression are downregulated in dental pulp from diabetic patients. (A and B) Immunohistochemistry staining results of eNOS in dental pulp tissue in NT group and DM group. (C-E) Western Blot confirmed the phosphorylation degree of AKT and eNOS in NT group and DM group. Scale bar: 100 μm. Data presented as mean ± SD.

Article Snippet: Short hairpin RNA (shRNA) lentiviral vectors for NOS3 knockdown, NOS3 overexpression vectors, and control lentiviral vectors were constructed by GeneChem.

Techniques: Expressing, Immunohistochemistry, Staining, Western Blot, Phospho-proteomics

Journal: International Dental Journal

Article Title: Endothelial Nitric Oxide Synthase Restores Diabetic Dentin Regeneration via AKT/Endothelial Nitric Oxide Synthase Axis

doi: 10.1016/j.identj.2026.109511

Figure Lengend Snippet: High glucose affects the proliferation and differentiation of hDPSCs and inhibits the expression of eNOS in hDPSCs via the AKT pathway. (A) Analysis of CCK-8 assay in DPSCs cultured in media with different glucose concentrations. (B) Quantification of ALP activity in DPSCs cultured in media with different glucose concentrations. (C) Quantification of ALP activity in DPSCs from blank, mannitol and HG groups. (D-F) Western Blot confirmed the proteins level of p-AKT, AKT and eNOS in blank, HG and SC79 group. Data are presented as mean ± SD.

Article Snippet: Short hairpin RNA (shRNA) lentiviral vectors for NOS3 knockdown, NOS3 overexpression vectors, and control lentiviral vectors were constructed by GeneChem.

Techniques: Expressing, CCK-8 Assay, Cell Culture, Activity Assay, Western Blot

Journal: International Dental Journal

Article Title: Endothelial Nitric Oxide Synthase Restores Diabetic Dentin Regeneration via AKT/Endothelial Nitric Oxide Synthase Axis

doi: 10.1016/j.identj.2026.109511

Figure Lengend Snippet: Knockdown of eNOS inhibits the osteo/odontogenic differentiation capacity of hDPSCs. (A and B) ALP staining and quantification of ALP activity results in vector and eNOSsh group. (C and D) Alizarin red staining and Ca 2+ ion measurement results in vector and eNOSsh group. (E-G) Western Blot confirmed the proteins level of DSPP and DMP1 in vector and eNOSsh group. (H-K) RT-qPCR results showed the downregulation of RUNX2(H), OCN(I), BSP(J), and DSPP(K) mRNA expression levels in eNOSsh group. Data are presented as mean ± SD.

Article Snippet: Short hairpin RNA (shRNA) lentiviral vectors for NOS3 knockdown, NOS3 overexpression vectors, and control lentiviral vectors were constructed by GeneChem.

Techniques: Knockdown, Staining, Activity Assay, Plasmid Preparation, Western Blot, Quantitative RT-PCR, Expressing

Journal: International Dental Journal

Article Title: Endothelial Nitric Oxide Synthase Restores Diabetic Dentin Regeneration via AKT/Endothelial Nitric Oxide Synthase Axis

doi: 10.1016/j.identj.2026.109511

Figure Lengend Snippet: Overexpression of eNOS could rescue the impairment of osteo/odontogenic capability of hDPSCs in high-glucose conditions. (A and B) ALP staining and quantification of ALP activity results. (C and D) Alizarin red staining and Ca 2+ ion measurement results. (E-G) Western Blot confirmed the proteins level of DSPP and DMP1. (H-K) RT-qPCR results showed the upregulation of mRNA expression levels of BSP(H), RUNX2(I), DSPP(J), and DMP1(K) in eNOS group. Data are expressed as mean ± SD.

Article Snippet: Short hairpin RNA (shRNA) lentiviral vectors for NOS3 knockdown, NOS3 overexpression vectors, and control lentiviral vectors were constructed by GeneChem.

Techniques: Over Expression, Staining, Activity Assay, Western Blot, Quantitative RT-PCR, Expressing

Journal: International Dental Journal

Article Title: Endothelial Nitric Oxide Synthase Restores Diabetic Dentin Regeneration via AKT/Endothelial Nitric Oxide Synthase Axis

doi: 10.1016/j.identj.2026.109511

Figure Lengend Snippet: Overexpression of eNOS in DPSCs promotes dentin regeneration in the extraction sockets of rabbit jaws. (A and B) Micro-CT analysis. (C) Observation of cross-section in the lower 1/3 segment of the extraction socket. (D) Gross observation of regeneration tissue in rabbit extraction socket. Data presented as mean ± SD.

Article Snippet: Short hairpin RNA (shRNA) lentiviral vectors for NOS3 knockdown, NOS3 overexpression vectors, and control lentiviral vectors were constructed by GeneChem.

Techniques: Over Expression, Extraction, Micro-CT

Journal: International Dental Journal

Article Title: Endothelial Nitric Oxide Synthase Restores Diabetic Dentin Regeneration via AKT/Endothelial Nitric Oxide Synthase Axis

doi: 10.1016/j.identj.2026.109511

Figure Lengend Snippet: Histological examination of regenerated tissues in the extraction sockets of rabbit jaws. (A and B) HE staining results. (C and D) SEM examination of regeneration tissues in rabbit extraction socket. (E-G) Immunohistochemistry staining results of DSPP in regenerated dentin-like tissue in the vector group and eNOS group. (H-J) Immunohistochemistry staining results of DMP1 in regenerated dentin-like tissue in the vector group and eNOS group. Scale bar: 200 μm (A), 100 μm (B, E, H), 50 μm (F, I), 5 μm (C), 2 μm (D). Data presented as mean ± SD. AB: alveolar bone; rD: regenerated dentin-like tissue; Od and black arrow: odontoblast-like cells.

Article Snippet: Short hairpin RNA (shRNA) lentiviral vectors for NOS3 knockdown, NOS3 overexpression vectors, and control lentiviral vectors were constructed by GeneChem.

Techniques: Extraction, Staining, Immunohistochemistry, Plasmid Preparation

Journal: International Journal of Molecular Medicine

Article Title: Exosomal circ0000549 promotes MNNG-induced gastric cancer through miR-15b-5p/KIF1B

doi: 10.3892/ijmm.2026.5785

Figure Lengend Snippet: Characterization and expression of circ0000549 in TGES-1 cells and exosomes. (A) Coincidence was observed for the differentially expressed circular RNAs in the GEO database. (B) Heat map illustrating the differential expression of circRNAs, with circ0000549 standing out as highly expressed. (C) Diagram of the cyclization mechanism of circ0000549. (D) Verification through agarose gel electrophoresis showing the specificity of circ0000549 amplification from cDNA. (E) RT-qPCR results demonstrating the expression levels of circ0000549 before and after RNase R treatment, confirming its resistance to RNase R and stability compared to linear RNA. (F) The levels of circ0000549 and its parental gene SRSF5 were assessed after treatment with actinomycin D. (G) The expression of circ0000549 in TGES-1 cells and TGES-1-EX by RT-qPCR. (H) Localization of circ0000549 determined by FISH, original magnification, ×60 (oil immersion). (I) The level of circ0000549 in GES-1 cells after TGES-1-EX treatment by RT-qPCR. (J) Serum levels of N-nitrosoproline were determined by LC-MS/MS. (K) The mRNA expression of circ0000549 in serum exosomes, n=26. (L) Diagnostic efficacy of serum exosome circ0000549, n=40. (M) Protein levels of EMT and stemness markers in GES-1 cells after treatment with serum exosomes. (N) The migration ability of GES-1 cells after treatment with serum exosomes, scale bar, 200 μ m. (O) The mRNA expression of EMT and stemness markers in GES-1 cells after treatment with serum exosomes. ** P<0.01, *** P<0.001, **** P<0.0001. GEO, Gene Expression Omnibus; RT-qPCR, reverse transcription-quantitative PCR; EMT, epithelial-mesenchymal transition; GC, gastric cancer; FISH, fluorescence in situ hybridization; Serum ex, serum derived exosomes from patients with gastric cancer; DAPI, 4',6-diamidino-2-phenylindole.

Article Snippet: Analysis of tumor tissues revealed that circ0000549 expression was markedly decreased in the circ0000549 knockdown exosome group ( ).

Techniques: Expressing, Quantitative Proteomics, Agarose Gel Electrophoresis, Amplification, Quantitative RT-PCR, Liquid Chromatography with Mass Spectroscopy, Diagnostic Assay, Migration, Gene Expression, Reverse Transcription, Real-time Polymerase Chain Reaction, Fluorescence, In Situ Hybridization, Derivative Assay

Journal: International Journal of Molecular Medicine

Article Title: Exosomal circ0000549 promotes MNNG-induced gastric cancer through miR-15b-5p/KIF1B

doi: 10.3892/ijmm.2026.5785

Figure Lengend Snippet: Circ0000549 promoted migration, proliferation, invasion and EMT in GES-1 cells. (A) The overexpression of circ0000549 in TGES-1 cells transfection with si1-circ0000549 and si2-circ0000549 was detected by RT-qPCR. (B) The effect of knocking down circ0000549 on the proliferation of TGES-1 cells was determined through a CCK-8 assay. (C) The effect of knocking down circ0000549 on the proliferative capacity of GES-1 cells was observed. (D) The abilities of TGES-1 cells to migrate and invade were evaluated, scale bar, 200 μ m. (E) The protein levels of EMT-related and stemness were analyzed after knocking down circ0000549. (F) The mRNAs expression of EMT-related and stemness markers. (G) The level of circ0000549 in TGES-1 cells was measured 48 h post-transfection with a lentivirus designed to overexpress circ0000549. (H) The expression of circ0000549 after puromycin screening. (I) Assessed the proliferative capacity of TGES-1 cells following overexpression of circ0000549. (J) Changes in the clonogenic ability of TGES-1 cells after overexpression of circ0000549. (K) The effect of overexpression of circ0000549 on the migration and invasion abilities in TGES-1 cells, scale bar, 200 μ m. (L) Changes in the levels of EMT-related and stemness protein after overexpression of circ0000549. (M) The expression of EMT-related and stemness mRNA in TGES-1 cells after overexpression of circ0000549. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001. EMT, epithelial-mesenchymal transition; RT-qPCR, reverse transcription-quantitative PCR; si1, siRNA1; si2, siRNA2; siNC, siRNA negative control; LV-OE-NC, lentivirus overexpression negative control; LV-OE, lentivirus overexpression; MOI, multiplicity of infection.

Article Snippet: Analysis of tumor tissues revealed that circ0000549 expression was markedly decreased in the circ0000549 knockdown exosome group ( ).

Techniques: Migration, Over Expression, Transfection, Quantitative RT-PCR, CCK-8 Assay, Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control, Infection

Journal: International Journal of Molecular Medicine

Article Title: Exosomal circ0000549 promotes MNNG-induced gastric cancer through miR-15b-5p/KIF1B

doi: 10.3892/ijmm.2026.5785

Figure Lengend Snippet: Exosomal circ0000549 promotes the proliferation, migration, invasion, EMT and stemness in GES-1 cells. (A) Measure the level of circ0000549 in TGES-1 cells 48 h post-transfection with a lentivirus designed for circ0000549 knockdown. (B) The level of circ0000549 after puromycin screening. (C) The knockdown/overexpression of exosomes was detected by TEM, scale bar, 100 nm. (D) The size of exosomes was determined using NTA. (E) The protein levels of surface markers of exosomes were detected. (F) The levels of circ0000549 in TGES-1-EX with knockdown/overexpression of circ0000549. (G) The level of circ0000549 in GES-1 cells after SH-circ0000549-EX/OE-circ0000549-EX treatment. (H) Assessment of GES-1 cell proliferation post-treatment with SH-circ0000549-EX/OE-circ0000549-EX. (I) Evaluation of the clonality of GES-1 cells after SH-circ0000549-EX/OE-circ0000549-EX treatment. (J) Examination of the migratory and invasive capabilities of GES-1 cells following SH-circ0000549-EX/OE-circ0000549-EX treatment, scale bar, 200 μ m. (K) Determination of EMT and stemness protein expression levels in GES-1 cells treated SH-circ0000549-EX/OE-circ0000549-EX. (L) Quantification of EMT and stemness mRNA levels in GES-1 cells after SH-circ0000549-EX/OE-circ0000549-EX treatment using RT-qPCR. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001. TEM, transmission electron microscopy; NTA, nanoparticle tracking analysis; RT-qPCR, reverse transcription-quantitative PCR; NC, negative control; EX, exosome; OE-circ549-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus overexpression; OE-NC-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus overexpression negative control; SH-circ0000549-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus knockdown; SH-NC-EX; exosomes from TGES-1 cells treated with circ0000549 lentivirus knockdown negative control; MOI, multiplicity of infection.

Article Snippet: Analysis of tumor tissues revealed that circ0000549 expression was markedly decreased in the circ0000549 knockdown exosome group ( ).

Techniques: Migration, Transfection, Knockdown, Over Expression, Expressing, Quantitative RT-PCR, Transmission Assay, Electron Microscopy, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control, Infection

Journal: International Journal of Molecular Medicine

Article Title: Exosomal circ0000549 promotes MNNG-induced gastric cancer through miR-15b-5p/KIF1B

doi: 10.3892/ijmm.2026.5785

Figure Lengend Snippet: miR-15b-5p is the target of circ0000549 and weakens the protumor effect of circ0000549. (A) circInteractome database showed that AGO2 had binding ability with the flanking region of circ0000549. (B) The online database predicted the intersection of miRNA bound by circ0000549. (C) The expression of miRNA after knocking down circ0000549 by RT-qPCR. (D) The level of miRNA after overexpression of circ0000549 by RT-qPCR. (E) The expression of miR-15b-5p was assessed. (F) A dual luciferase reporter gene assay was conducted to investigate the interaction between circ0000549 and miR-15b-5p. (G) The level of miR-15b-5p expression in TGES-1 cells was assessed following transfection with miR-15b-5p mimics or an inhibitor. (H) The level of circ0000549 was assessed following transfection with miR-15b-5p mimics or an inhibitor. (I) The proliferation of cells was evaluated after co-transfecting them with OE-circ0000549 and miR-15b-5p mimics, or with si-circ0000549 and miR-15b-5p inhibitor. (J) Evaluation of the clonality of TGES-1 cells after co-transfection of OE-circ0000549 and miR-15b-5p mimics/si-circ0000549 and miR-15b-5p inhibitor. (K) The migration and invasion capabilities of TGES-1 cells were assessed following co-transfection with OE-circ0000549 and miR-15b-5p mimics, or with si-circ0000549 and miR-15b-5p inhibitor, scale bar, 200 μ m. (L) Level of miR-15b-5p following SH-circ0000549-EX/OE-circ0000549-EX treatment by RT-qPCR. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001. RT-qPCR, reverse transcription-quantitative PCR; si1, siRNA1; si2, siRNA2; siNC, siRNA negative control; LV-OE-NC, lentivirus overexpression negative control; LV-OE, lentivirus overexpression; WT, Wild Type; MUT, Mutant; miR-NC, miR-15b-5p mimics negative control; inhibitor Ctrl, miR-15b-5p inhibitor control; OE-circ, lentivirus overexpression of circ0000549; miR mimics, miR-15b-5p mimics; miR-inhibitor, miR-15b-5p inhibitor.

Article Snippet: Analysis of tumor tissues revealed that circ0000549 expression was markedly decreased in the circ0000549 knockdown exosome group ( ).

Techniques: Binding Assay, Expressing, Quantitative RT-PCR, Over Expression, Luciferase, Reporter Gene Assay, Transfection, Cotransfection, Migration, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control, Mutagenesis, Control

Journal: International Journal of Molecular Medicine

Article Title: Exosomal circ0000549 promotes MNNG-induced gastric cancer through miR-15b-5p/KIF1B

doi: 10.3892/ijmm.2026.5785

Figure Lengend Snippet: Circ0000549 in exosomes aids in promoting the aggressive evolution of GES-1 cells, mainly through miR-15b-5p/KIF1B/PI3K/AKT axis. (A) Internet databases were used to predict which genes are regulated by miR-15b-5p. (B) The levels of candidate target genes after knockdown/overexpression of circ0000549 or transfection of miR-15b-5p mimics/inhibitor by RT-qPCR. (C) mRNA level of KIF1B was detected. (D) The effect of co transfection with OE circ0000549 and miR-15b-5p mimic/si circ00000549 and miR-15b-5p inhibitor on the expression of KIF1B. (E) A dual luciferase reporter gene assay was employed to elucidate the interaction between KIF1B and miR-15b-5p. (F) GeneCards database was used to predict KIF1B downstream pathways online. (G) The pathway activity of the PI3K/AKT in TGES-1 cells after knocking down/overexpressing circ0000549. (H) Changes in expression levels of KIF1B and PI3K/AKT pathway proteins of GES-1 cells after SH-circ0000549-EX/OE-circ0000549-EX treatment. (I) The effect on KIF1B level in TGES-1 cells subsequent to KIF1B knockdown. (J) The level of KIF1B in TGES-1 cells which was knocked down KIF1B after treatment with OE-circ0000549 exosomes by RT-qPCR. (K) Detection of changes in expression levels of KIF1B and PI3K/AKT pathway proteins in TGES-1 cells which was knocked down KIF1B after treatment with OE-circ0000549 exosomes. (L) Detection of changes in the proliferation ability of TGES-1 cells. (M) Clonality of TGES-1 cells which was knocked down KIF1B after treatment with OE-circ0000549 exosomes. (N) Alterations in the migratory and invasive capabilities of TGES-1 cells treated with OE-circ0000549 exosomes and KIF1B knockdown in combination. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001. miR, microRNA; RT-qPCR, reverse transcription-quantitative PCR; NC, negative control; LV-OE-NC, lentivirus overexpression negative control; LV-OE, lentivirus overexpression; LV-SH-circ0000549, lentivirus knockdown of circ0000549; LV-SH-NC; lentivirus knockdown of circ0000549 negative control; miR-NC, miR-15b-5p mimics negative control; inhibitor Ctrl, miR-15b-5p inhibitor control; OE-circ549, lentivirus overexpression of circ0000549; OE-circ549-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus overexpression; siKIF1B, siRNA of KIF1B; siNC, siRNA negative control; SH-circ549-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus knockdown; SH-NC-EX; exosomes from TGES-1 cells treated with circ0000549 lentivirus knockdown negative control.

Article Snippet: Analysis of tumor tissues revealed that circ0000549 expression was markedly decreased in the circ0000549 knockdown exosome group ( ).

Techniques: Knockdown, Over Expression, Transfection, Quantitative RT-PCR, Cotransfection, Expressing, Luciferase, Reporter Gene Assay, Activity Assay, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control, Control

Journal: International Journal of Molecular Medicine

Article Title: Exosomal circ0000549 promotes MNNG-induced gastric cancer through miR-15b-5p/KIF1B

doi: 10.3892/ijmm.2026.5785

Figure Lengend Snippet: Exosomal circ0000549 tumorigenicity, EMT, proliferation and stemness. (A) The downregulation of circ0000549 impairs the tumorigenic capacity of TGES-1 cells. (B) The effect of SH-circ0000549-EX treatment on tumor weight. (C) The effect of SH-circ0000549-EX treatment on tumor volume. (D) The level of circ0000549 in tumor tissues after SH-circ0000549-EX treatment by RT-qPCR. (E) The levels of stemness, EMT and proliferation proteins in tumor tissues after SH-circ0000549-EX treatment. (F) The mRNA expression of stemness, EMT and proliferation makers mRNA in tumor tissues after SH-circ0000549-EX treatment. (G) Immunohistochemical analysis of tumor tissues treated with SH-circ0000549-EX revealed alterations in the levels of EMT and stemness markers, magnification, ×400. (H) The effect of OE-circ0000549-EX alone or in combination with miR-15b-5p mimic on the tumorigenicity of TGES-1 cells. (I) The effect of OE-circ0000549-EX alone or in combination with miR-15b-5p mimic on tumor weight and volume. (J) The levels of stemness, EMT and proliferation-related markers in tumor tissues following treatment with OE-circ0000549-EX alone or in combination with miR-15b-5p mimic. (K) The expression of stemness, EMT and proliferation mRNA in tumor tissues after treated with OE-circ0000549-EX alone or in combination with miR-15b-5p mimic. (L) The levels of vimentin and Nanog in tumor tissues after SH-circ0000549-EX treatment by Immunohistochemistry, magnification, ×200. * P<0.05, ** P<0.01, *** P<0.001, # P<0.05, ## P<0.01. EMT, epithelial-mesenchymal transition; RT-qPCR, reverse transcription-quantitative PCR; NC, negative control; SH-circ549-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus knockdown; SH-NC-EX; exosomes from TGES-1 cells treated with circ0000549 lentivirus knockdown negative control; OE-circ549-EX, exosomes from TGES-1 cells treated with circ0000549 lentivirus overexpression.

Article Snippet: Analysis of tumor tissues revealed that circ0000549 expression was markedly decreased in the circ0000549 knockdown exosome group ( ).

Techniques: Quantitative RT-PCR, Expressing, Immunohistochemical staining, Immunohistochemistry, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control, Knockdown, Over Expression

Journal: iScience

Article Title: ATF6 ameliorates renal warm ischemia-reperfusion injury through FHL2-mediated NF-κB signaling pathway

doi: 10.1016/j.isci.2026.115173

Figure Lengend Snippet: Renal warm I/R injury in mice increases the expression of ATF6 (A) RT-qPCR analysis of ATF6 expression in mouse kidney tissues. (B–C) Representative Western blot images and quantification of ATF6 and activated ATF6 (cATF6) expression in mouse kidney tissues subjected to Sham and ischemia 1h/reperfusion 6h (I/R). (D–E) Representative IHC images and quantification of ATF6 expression in kidney tissues (Scale bars, 100 μm and 50 μm). (F–G) ATF6 expression in cultured HK-2 in response to normoxia and hypoxia/reoxygenation (H/R) treatment was determined by RT-qPCR and Western blot. (H) Quantitative statistical analysis of ATF6 protein. Data represent mean ± SDs, and statistical analysis was performed using t test. (∗ p < 0.05). T-vs-C means IR-vs-Sham.

Article Snippet: For PTECs-specific ATF6 knockdown, adeno-associated viruses (AAV)-based delivery system with pAAV9-MCS-miR30shRNA (Atf6)-WPRE or pAAV9-MCS-miR30shRNA (NC)-WPRE (0.5 × 1012 vg, OBiO Technology, Shanghai, China) were injected by tail vein injection to C57BL/6 mice.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Cell Culture

Journal: iScience

Article Title: ATF6 ameliorates renal warm ischemia-reperfusion injury through FHL2-mediated NF-κB signaling pathway

doi: 10.1016/j.isci.2026.115173

Figure Lengend Snippet: ATF6 plays a protective role in renal ischemia-reperfusion injury (A–B) The levels of serum Cr and BUN in mice. (C–D) The levels of serum IL-6 and TNF-α in mice. (E–F) The representative images and kidney injury assessment score of HE staining of the kidney. Ceapin-A7 (10 μM) is an ATF6 inhibitor, and AA147 (10 μM) is an ATF6 agonist. Data represent mean ± SDs, and statistical analysis was performed using one-way ANOVA. (∗ p < 0.05).

Article Snippet: For PTECs-specific ATF6 knockdown, adeno-associated viruses (AAV)-based delivery system with pAAV9-MCS-miR30shRNA (Atf6)-WPRE or pAAV9-MCS-miR30shRNA (NC)-WPRE (0.5 × 1012 vg, OBiO Technology, Shanghai, China) were injected by tail vein injection to C57BL/6 mice.

Techniques: Staining

Journal: iScience

Article Title: ATF6 ameliorates renal warm ischemia-reperfusion injury through FHL2-mediated NF-κB signaling pathway

doi: 10.1016/j.isci.2026.115173

Figure Lengend Snippet: ATF6 plays a protective role in renal ischemia-reperfusion injury (A–B) The levels of serum Cr and BUN in mice. (C–D) The levels of serum IL-6 and TNF-α in mice. (E–F) The representative images and kidney injury assessment score of HE staining of the kidney. (G–H) RT-qPCR analysis of TNF-α and IL-6 expression in cultured HK-2 in response to H/R treatment. (I–J) RT-qPCR analysis of IL-6 and TNF-α expression in cultured HK-2 in response to H/R using siATF6. Data represent mean ± SDs, and statistical analysis was performed using one-way ANOVA. (∗ p < 0.05).

Article Snippet: For PTECs-specific ATF6 knockdown, adeno-associated viruses (AAV)-based delivery system with pAAV9-MCS-miR30shRNA (Atf6)-WPRE or pAAV9-MCS-miR30shRNA (NC)-WPRE (0.5 × 1012 vg, OBiO Technology, Shanghai, China) were injected by tail vein injection to C57BL/6 mice.

Techniques: Staining, Quantitative RT-PCR, Expressing, Cell Culture

Journal: iScience

Article Title: ATF6 ameliorates renal warm ischemia-reperfusion injury through FHL2-mediated NF-κB signaling pathway

doi: 10.1016/j.isci.2026.115173

Figure Lengend Snippet: ATF6 inhibits the activation of the NF-κB pathway (A) Western blot detection of the expression of NF-κB pathway-related proteins in renal tissue; GAPDH was used as a loading control. (B) Quantitative analysis of protein expression, data represent mean ± SDs, and statistical analysis was performed using t test. (∗ p < 0.05). (C) After hypoxia/reoxygenation in the WT group and knockdown group, the expression of NF-κB pathway-related proteins was detected by Western blot, GAPDH was used as a loading control. (D) Quantitative analysis of protein expression. (E) After hypoxia/reoxygenation in the WT group and the ATF6 overexpression group, the expression of NF-κB pathway-related proteins was detected by Western blot, GAPDH was used as a loading control. (F) Quantitative analysis of protein expression. Data represent mean ± SDs, and statistical analysis was performed using one-way ANOVA. (∗ p < 0.05).

Article Snippet: For PTECs-specific ATF6 knockdown, adeno-associated viruses (AAV)-based delivery system with pAAV9-MCS-miR30shRNA (Atf6)-WPRE or pAAV9-MCS-miR30shRNA (NC)-WPRE (0.5 × 1012 vg, OBiO Technology, Shanghai, China) were injected by tail vein injection to C57BL/6 mice.

Techniques: Activation Assay, Western Blot, Expressing, Control, Knockdown, Over Expression

Journal: iScience

Article Title: ATF6 ameliorates renal warm ischemia-reperfusion injury through FHL2-mediated NF-κB signaling pathway

doi: 10.1016/j.isci.2026.115173

Figure Lengend Snippet: The expression of FHL2 is transcriptionally regulated by ATF6 (A) Expression of FHL2 in RNA-seq after using siATF6. (B) RT-qPCR analysis of FHL2 expression in mouse kidney tissues. (C) Representative Western blot images and quantification of FHL2 expression in mouse kidney tissues (GAPDH as the loading control). (D–E) Representative IHC images and quantification of FHL2 expression in kidney tissues (Scale bars, 100 μm). (F–G) FHL2 expression in cultured HK-2 in response to normoxia and hypoxia/reoxygenation (H/R) treatment was determined by Western blot. GAPDH was used as a loading control. (H–I) The expression of FHL2 and ATF6 in HK-2 cells after ATF6 knockdown was examined by Western blot. GAPDH was used as a loading control; (J-K) The expression of FHL2 and ATF6 in HK-2 cell lines after ATF6 overexpression was examined by Western blot. GAPDH was used as a loading control. (L) The input of ATF6 and IgG binding the FHL2 promoters in HK-2 after hypoxia/physoxia treatment by CHIP-qPCR. (M) Luciferase activity of transfected HEK-293T targeting FHL2 and its mutant after H/R using the dual-luciferase reporter assay. (N) The putative ATF6-binding sites in the FHL2 promoter and the nucleotide sequences representing the predicted binding sequences with the red capital letters indicating core binding elements; Data represent mean ± SDs, and statistical analysis was performed using one-way ANOVA. (∗ p < 0.05).

Article Snippet: For PTECs-specific ATF6 knockdown, adeno-associated viruses (AAV)-based delivery system with pAAV9-MCS-miR30shRNA (Atf6)-WPRE or pAAV9-MCS-miR30shRNA (NC)-WPRE (0.5 × 1012 vg, OBiO Technology, Shanghai, China) were injected by tail vein injection to C57BL/6 mice.

Techniques: Expressing, RNA Sequencing, Quantitative RT-PCR, Western Blot, Control, Cell Culture, Knockdown, Over Expression, Binding Assay, ChIP-qPCR, Luciferase, Activity Assay, Transfection, Mutagenesis, Reporter Assay