Journal: International Journal of Biological Sciences

Article Title: METTL3 mediated m6A modification of HKDC1 promotes renal injury and inflammation in lead nephropathy

doi: 10.7150/ijbs.112463

Figure Lengend Snippet: METTL3 cKO protects against renal dysfunction and inflammation induced by lead acetate. (A) Schematic representation of the genetic approach used to generate METTL3 conditional knockout (cKO) mice. (B) METTL3 deficiency was confirmed using genomic DNA analysis. (C) Dot blot assay of m6A methylation modification abundance in lead nephropathy with or without METTL3 cKO. (D) Western blot analysis of METTL3 abundance in METTL3 cKO mice compared to METTL3 Flox/Flox mice. (E) Real-time PCR analysis of METTL3 mRNA expression in METTL3 cKO mice compared to METTL3 Flox/Flox mice (n = 6 biological replicates of mice, two tailed unpaired Student's t test). (F and H) Representative PAS staining picture and quantification analysis of kidneys from METTL3 Flox/Flox and METTL3 cKO mice treated with lead acetate. Scale bars = 100 μm. (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (G) Representative immunofluorescence staining of KIM-1 and proximal tubular markers (LTL). Scale bars = 100 μm. (I and J) Serum creatinine and BUN levels in METTL3 Flox/Flox and METTL3 cKO mice exposed to Pb (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (K) Western blot analysis of KIM-1 and NGAL levels in METTL3 Flox/Flox and METTL3 cKO mice exposed to Pb. (L) IHC staining for F4/80+ macrophage infiltration in METTL3 Flox/Flox and METTL3 cKO mice with lead nephropathy. (M) Real-time PCR analysis of inflammatory cytokines (TNF-α, MCP-1 and IL-1β) in lead acetate-induced lead nephropathy (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (N) Western blot analysis of p65 NF-κB phosphorylation in lead nephropathy. Data represents the mean ± S.E.M. ** P < 0.01, *** P < 0.001. M3FF : METTL3 Flox/Flox , M3cKO : METTL3 cKO.

Article Snippet: The levels of creatinine and BUN in the blood samples were measured using a Creatinine and BUN Assay Kit (Nanjing Jiancheng, China) according to the manufacturer's instructions.

Techniques: Knock-Out, Dot Blot, Methylation, Modification, Western Blot, Real-time Polymerase Chain Reaction, Expressing, Two Tailed Test, Staining, Immunofluorescence, Immunohistochemistry, Phospho-proteomics

Journal: International Journal of Biological Sciences

Article Title: METTL3 mediated m6A modification of HKDC1 promotes renal injury and inflammation in lead nephropathy

doi: 10.7150/ijbs.112463

Figure Lengend Snippet: METTL3 knockdown reduces renal injury and inflammatory response in lead acetate-induced mouse. (A) Schematic diagram of AAV9 mediated METTL3 silencing by tail vein injection. (B) The expression of METTL3 in mice. (C and D) Serum creatinine and BUN levels in Pb-induced mice with or without METTL3 knockdown (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (E) Real-time PCR analysis of KIM-1 expression in lead acetate nephropathy with and without METTL3 knockdown (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (F and G) PAS staining and scores for lead acetate-induced lead nephropathy in METTL3 knockdown mice. Scale bars = 100 μm. (H) Western blot and quantitative analysis of NGAL and KIM-1 in lead nephropathy with or without METTL3 knockdown. (I) Real-time PCR analysis of MCP-1 and IL-6 mRNA expression in lead acetate-treated METTL3 knockdown mice (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test).(J) Western blotting and quantitative analysis of p65 NF-κB phosphorylation in lead nephropathy with or without METTL3 knockdown. (K) IHC staining of F4/80+ macrophage infiltration in mice with lead nephropathy with or without METTL3 knockdown. Scale bars = 100 μm. Data represents the mean ± S.E.M. ***P < 0.001.

Article Snippet: The levels of creatinine and BUN in the blood samples were measured using a Creatinine and BUN Assay Kit (Nanjing Jiancheng, China) according to the manufacturer's instructions.

Techniques: Knockdown, Injection, Expressing, Real-time Polymerase Chain Reaction, Staining, Western Blot, Phospho-proteomics, Immunohistochemistry

Journal: International Journal of Biological Sciences

Article Title: METTL3 mediated m6A modification of HKDC1 promotes renal injury and inflammation in lead nephropathy

doi: 10.7150/ijbs.112463

Figure Lengend Snippet: The METTL3 inhibitor STM2457 prevents renal injury and inflammation induced by lead acetate in vitro and in vivo. (A) Effect of different concentrations of STM2457 on the viability of HK2 cells by MTT assay. (B) Real-time PCR analysis of KIM-1 in STM24574 -treated HK2 cells in response to lead acetate treatment (n = 3 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (C) Western blot analysis of KIM-1 in STM24574 -treated HK2 cells in response to lead acetate treatment. (D) Western blot analysis of p65 NF-κB phosphorylation. in STM24574 -treated HK2 cells following lead acetate treatment. (E) Representative immunofluorescence staining of KIM-1 in lead acetate-treated HK2 cells with and without STM24574 . Scale bars = 100 μm. (F) Representative immunofluorescence staining of pp65 in lead acetate-treated HK2 cells with and without STM24574 . Scale bars = 100 μm. (G) Dot blot assay of m6A methylation in lead acetate-treated HK2 cells with and without STM24574 . (H) MeRIP-qPCR analysis of alterations in m6A modifications of HKDC1 in lead acetate-stimulated HK2 cells with or without STM24574 (n = 4 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (I) Real-time PCR analysis of HKDC1 expression in lead acetate-treated HK2 cells with and without STM24574 (n = 4 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (J) Schematic diagram of STM2457 administration. (K and L) Serum creatinine and BUN levels in lead acetate-induced lead nephropathy mouse models with and without STM2457 treatment (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (M) PAS staining in a lead acetate-induced lead nephropathy mouse model with and without STM2457 treatment. Scale bars = 100 μm. (N) Real-time PCR analysis of MCP-1, TNF-α and IL-1β mRNA expression in lead acetate-treated mice with STM24574 (n = 6 biological replicates, one-way ANOVA with Tukey's multiple comparisons test). (O) Western blot analysis of KIM-1and NGAL levels, and p65 NF-κB phosphorylation in STM2457-treated mice following lead acetate treatment. Data represents the mean ± S.E.M. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The levels of creatinine and BUN in the blood samples were measured using a Creatinine and BUN Assay Kit (Nanjing Jiancheng, China) according to the manufacturer's instructions.

Techniques: In Vitro, In Vivo, MTT Assay, Real-time Polymerase Chain Reaction, Western Blot, Phospho-proteomics, Immunofluorescence, Staining, Dot Blot, Methylation, Expressing

Journal: Nutrients

Article Title: Dietary Probiotic Pediococcus acidilactici GKA4, Dead Probiotic GKA4, and Postbiotic GKA4 Improves Cisplatin-Induced AKI by Autophagy and Endoplasmic Reticulum Stress and Organic Ion Transporters

doi: 10.3390/nu16203532

Figure Lengend Snippet: The experimental framework ( A ) and the renoprotective efficacy of GKA4, dead probiotic GKA4, and postbiotic GKA4 alleviates neurotoxicity induced by cisplatin treatment. Oral administration of GKA4, dead probiotic GKA4, and postbiotic GKA4 at 250 mg/kg was carried out daily for 10 consecutive days, with cisplatin being administered one hour after the seventh dose. The sacrifice of the mice occurred on the eleventh day. The levels of BUN ( B ) and CRE in the serum ( C ), the renal sections stained with H&E (400×) ( D ), and the renal injury scale ( E ) were assessed. Means ± S.E.M ( n = 5) are shown in the presentation of the data. Statistical significance ( p < 0.001) is indicated by ### when compared with the control group sample. Statistical significance at *** p < 0.001 was evident in contrast to the cisplatin group.

Article Snippet: Detection kits for creatinine (CRE) and blood urea nitrogen (BUN) were obtained from HUMAN Diagnostics Worldwide (Wiesbaden, Germany).

Techniques: Staining, Control

Journal: Frontiers in Pharmacology

Article Title: Delineation of renal protein profiles in aristolochic acid I-induced nephrotoxicity in mice by label-free quantitative proteomics

doi: 10.3389/fphar.2024.1341854

Figure Lengend Snippet: Effects of AAI on renal function in mice. (A) Serum creatinine levels (n = 6). (B) Blood urea nitrogen levels (n = 6). Data are expressed as mean ± SEM (* p < 0.05 and ***p < 0.001 as compared to the control group).

Article Snippet: The levels of serum creatinine (SCR) and blood urea nitrogen (BUN) were measured using commercially available kits (SKT-217 and SKT-213, StressMarq Biosciences, Victoria, British Columbia, Canada), according to the manufacturer’s protocols.

Techniques: Control