cbs  (Proteintech)

Journal: Frontiers in Pharmacology

Article Title: Shensu IV maintains the integrity of the glomerular filtration barrier and exerts renal protective effects by regulating endogenous hydrogen sulfide levels

doi: 10.3389/fphar.2024.1447249

Figure Lengend Snippet: FIGURE 6 Shensu IV regulates the PI3K/AKT signaling pathway through H2S. (A) The effects of Shensu IV and NaHS on the mRNA expression of CD2AP, nephrin, CBS, CSE, NOX4, PI3K, and AKT in renal tissue of PAN rats were analyzed by RT-qPCR. (B) Western blot analysis of the effects of Shensu IV and NaHS on the protein levels of CD2AP, nephrin, CBS, CSE, NOX4, PI3K, p-PI3K,AKT,p-AKT in renal tissue of PAN rats. *P< 0.05, **P< 0.01, ***P< 0.001. Abbreviations: CD2AP, CD2-associated protein; CBS, Cystathionine β-synthase; CSE, Cystathionine γ-lyase; PI3K, Phosphoinositide 3-Kinase; AKT, Protein Kinase B.

Article Snippet: Membranes were blocked with 5% skim milk (Solarbio) to prevent nonspecific binding and then incubated with primary antibodies against CD2AP (1:2000; A01756-2, BOSTER, Wuhan, China), nephrin (1:2000; A01756-2, BOSTER), CBS (1: 10,000; 14787-1-AP, Proteintech, Wuhan, China), CSE (1: 4,000; 12217-1-AP, Proteintech), PI3K (1:2000; 60225-1-Ig, Proteintech), p-PI3K (1:2000; bs-3332R, BOSTER), AKT (1:10,000; 60203-2-Ig, Proteintech), p-AKT (1: 10,000; 66,444-1-lg, Proteintech), NOX4 (1: 8,000; 14347-1-AP, Proteintech), and GAPDH (1:40,000; 60004-1- Ig, Proteintech).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Frontiers in Pharmacology

Article Title: Shensu IV maintains the integrity of the glomerular filtration barrier and exerts renal protective effects by regulating endogenous hydrogen sulfide levels

doi: 10.3389/fphar.2024.1447249

Figure Lengend Snippet: FIGURE 8 Shensu IV regulates the PI3K/AKT signaling pathway through H2S in podocytes. (A) The effects of Shensu IV and NaHS on the mRNA expression of CD2AP, nephrin, CBS, CSE, NOX4, PI3K, and AKT in podocytes were analyzed by RT-qPCR. (B) Western blot analysis of the effects of Shensu IV and NaHS on the protein levels of CD2AP, nephrin, CBS, CSE, NOX4, PI3K, p-PI3K,AKT,p-AKT in PAN-induced podocyocytes. *P< 0.05, **P< 0.01, ***P< 0.001. Abbreviations: CD2AP, CD2-associated protein; CBS, Cystathionine β-synthase; CSE, Cystathionine γ-lyase; PI3K, Phosphoinositide 3-Kinase; AKT, Protein Kinase B.

Article Snippet: Membranes were blocked with 5% skim milk (Solarbio) to prevent nonspecific binding and then incubated with primary antibodies against CD2AP (1:2000; A01756-2, BOSTER, Wuhan, China), nephrin (1:2000; A01756-2, BOSTER), CBS (1: 10,000; 14787-1-AP, Proteintech, Wuhan, China), CSE (1: 4,000; 12217-1-AP, Proteintech), PI3K (1:2000; 60225-1-Ig, Proteintech), p-PI3K (1:2000; bs-3332R, BOSTER), AKT (1:10,000; 60203-2-Ig, Proteintech), p-AKT (1: 10,000; 66,444-1-lg, Proteintech), NOX4 (1: 8,000; 14347-1-AP, Proteintech), and GAPDH (1:40,000; 60004-1- Ig, Proteintech).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Cardiac complications occur in MsrB2 KO DM. A . Glucose tolerance test (GTT) in WT and MsrB2 KO under nonDM and DM conditions. B . Body weight in WT and MsrB2 KO DM mice during HFD feeding. The body weight was weighed every week. C . Electron microscopy (EM) images in WT and MsrB2 KO DM mice. The white arrow indicates the mitochondria (autophagosome and autolysosome). The black triangle indicates the membrane structure. A white triangle indicates an abnormal Z-line. D . H&E and trichrome staining of histological sections of the heart of WT and MsrB2 KO mice under nonDM and DM MsrB2 conditions. E . Fibrotic areas were quantified using the ImageJ (FIJI) analyzer on histological sections. The two-way ANOVA analysis was performed for p values

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques: Electron Microscopy, Membrane, Staining

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Decreased autophagy and SERCA2a-PLN function in MsrB2 KO DM mice. A . A Western blot analysis of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL, PNL, and GAPDH of nonDM (WT #1–3, MsrB2 KO #1–3) and DM mouse (WT #1–3, MsrB2 KO #1–6) hearts. GAPDH served as a loading control. B – D . Quantification of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL/PNL, and GAPDH signal intensity. E . Quantitative RT-PCR analysis MsrB2 and SERCA2a transcript levels in nonDM (WT #1–5, MsrB2 KO #1–5) and DM (WT #1–3, MsrB2 KO #1–6) mouse hearts. The two-way ANOVA analysis was performed for p values

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques: Western Blot, Control, Quantitative RT-PCR

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) and mitochondrial dysfunction in the heart of diabetic mice. A . Blood Glucose in nonDM and DM mice. B . Western blot analysis of MetO and MsrB2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. C . Quantification of MetO and MsrB2 signal intensity. D . Western blot analysis of Parkin and LC3II in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. E . Quantification of Parkin and LC3II signal intensity. F . Western blot analysis of MsrB2 and LC3I/II in nonDM (#1–3), DM mouse hearts (DM #1–3), and DM with NAC treatment (#1–5). GAPDH served as a loading control. G . Quantification of MsrB2 and LC3II signal intensity. H . Electron microscopy (EM) in nonDM mice. The white body indicates a mitophagy structure. I . EM in DM mice. The White body indicates mitophagy (autophagosome and autolysosome) structure. J . Quantification of mitochondria size in nonDM and DM mice. K . Western blot analysis of DRP1 and DRP1 and Mfn2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. L . Quantification of DRP1signal intensity. The nonparametric t -test was performed for p values

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques: Western Blot, Control, Electron Microscopy

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Electrophysiological dysfunctions occurred in MsrB2 KO DM mice. A ~ I . ECG recording results in nonDM (WT #1–4, MsrB2 KO #1–5) and DM (WT #1–11, MsrB2. KO #1–9) mouse hearts

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques:

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: MsrB2 induces autophagy by LC3 activation. A . Western blot analysis of MsrB2 and LC3I/II in H9C2 after MsrB2-GFP transfection. MsrB2 induced LC3II in a concentration-dependent manner. GAPDH served as a loading control. B . Quantification of MsrB2 and LC3II signal intensity. C . Western blot analysis of MsrB2, SERCA2a, and bMHC in WT and MsrB2 KO NMCM. After 4 h of starvation, 5.5 or 25 mM glucose was treated for 48 h. GAPDH served as a loading control. D . Quantification SERCA2a and bMHC signal intensity. E . Western blot analysis of MsrB2, LC3I/II, pDRP1, and DRP1 in NMCM after Ad-MsrB2-GFP infection. Ad-MsrB2-GFP 50 m.o.i infected into NMCM. Then, 5.5 or 25 mM glucose was treated for 48 h after 4 h starvation. GAPDH served as a loading control. F . Quantification of LC3II and pDRP1 signal intensity. The one-way ANOVA analysis was performed for p values

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques: Activation Assay, Western Blot, Transfection, Concentration Assay, Control, Infection

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) in human diabetic hearts. A . Western blot analysis of MetO in normal (NH #1–3) and diabetic human heart tissue(DH #1–6). GAPDH served as a loading control. B . Quantification of MetO signal intensity. C . Western blot analysis of methionine sulfoxide reductase A and B2 (MsrA and B2) in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as loading a loading control. D . Quantification of MsrA and MsrB2 signal intensity. E . Western blot analysis of LC3l/II and p62 in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as a loading control. F . Quantification of LC3l/II and p62 signal intensity. G . Western blot analysis of DRP1 and OPA1 in normal (NH #1–3) and diabetic heart tissues (DH #1–6). GAPDH served as a loading control. H . Quantification of MetO intensity

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques: Western Blot, Control

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Summary. Increased MsrB2 expression in diabetic mouse hearts suppressed cardiac complications. These results were confirmed by inducing diabetes in mice with suppressed MsrB2 expression (MsrB2 KO). In MsrB2 KO DM, the expression of SERCA2a, which regulates myocardial contractility, was reduced, cardiac fibrosis increased, and the number of mitochondrial structural abnormalities increased

Article Snippet: According to the manufacturer’s protocol, MsrB2-GFP, RFP-Pakin, and Cherry LC3 were transfected into H9C2 with Lipofectamine 3000 (Invitrogen, USA).

Techniques: Expressing

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Cardiac complications occur in MsrB2 KO DM. A . Glucose tolerance test (GTT) in WT and MsrB2 KO under nonDM and DM conditions. B . Body weight in WT and MsrB2 KO DM mice during HFD feeding. The body weight was weighed every week. C . Electron microscopy (EM) images in WT and MsrB2 KO DM mice. The white arrow indicates the mitochondria (autophagosome and autolysosome). The black triangle indicates the membrane structure. A white triangle indicates an abnormal Z-line. D . H&E and trichrome staining of histological sections of the heart of WT and MsrB2 KO mice under nonDM and DM MsrB2 conditions. E . Fibrotic areas were quantified using the ImageJ (FIJI) analyzer on histological sections. The two-way ANOVA analysis was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Electron Microscopy, Membrane, Staining

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Decreased autophagy and SERCA2a-PLN function in MsrB2 KO DM mice. A . A Western blot analysis of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL, PNL, and GAPDH of nonDM (WT #1–3, MsrB2 KO #1–3) and DM mouse (WT #1–3, MsrB2 KO #1–6) hearts. GAPDH served as a loading control. B – D . Quantification of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL/PNL, and GAPDH signal intensity. E . Quantitative RT-PCR analysis MsrB2 and SERCA2a transcript levels in nonDM (WT #1–5, MsrB2 KO #1–5) and DM (WT #1–3, MsrB2 KO #1–6) mouse hearts. The two-way ANOVA analysis was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Western Blot, Control, Quantitative RT-PCR

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) and mitochondrial dysfunction in the heart of diabetic mice. A . Blood Glucose in nonDM and DM mice. B . Western blot analysis of MetO and MsrB2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. C . Quantification of MetO and MsrB2 signal intensity. D . Western blot analysis of Parkin and LC3II in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. E . Quantification of Parkin and LC3II signal intensity. F . Western blot analysis of MsrB2 and LC3I/II in nonDM (#1–3), DM mouse hearts (DM #1–3), and DM with NAC treatment (#1–5). GAPDH served as a loading control. G . Quantification of MsrB2 and LC3II signal intensity. H . Electron microscopy (EM) in nonDM mice. The white body indicates a mitophagy structure. I . EM in DM mice. The White body indicates mitophagy (autophagosome and autolysosome) structure. J . Quantification of mitochondria size in nonDM and DM mice. K . Western blot analysis of DRP1 and DRP1 and Mfn2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. L . Quantification of DRP1signal intensity. The nonparametric t -test was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Western Blot, Control, Electron Microscopy

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Electrophysiological dysfunctions occurred in MsrB2 KO DM mice. A ~ I . ECG recording results in nonDM (WT #1–4, MsrB2 KO #1–5) and DM (WT #1–11, MsrB2. KO #1–9) mouse hearts

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques:

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: MsrB2 induces autophagy by LC3 activation. A . Western blot analysis of MsrB2 and LC3I/II in H9C2 after MsrB2-GFP transfection. MsrB2 induced LC3II in a concentration-dependent manner. GAPDH served as a loading control. B . Quantification of MsrB2 and LC3II signal intensity. C . Western blot analysis of MsrB2, SERCA2a, and bMHC in WT and MsrB2 KO NMCM. After 4 h of starvation, 5.5 or 25 mM glucose was treated for 48 h. GAPDH served as a loading control. D . Quantification SERCA2a and bMHC signal intensity. E . Western blot analysis of MsrB2, LC3I/II, pDRP1, and DRP1 in NMCM after Ad-MsrB2-GFP infection. Ad-MsrB2-GFP 50 m.o.i infected into NMCM. Then, 5.5 or 25 mM glucose was treated for 48 h after 4 h starvation. GAPDH served as a loading control. F . Quantification of LC3II and pDRP1 signal intensity. The one-way ANOVA analysis was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Activation Assay, Western Blot, Transfection, Concentration Assay, Control, Infection

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) in human diabetic hearts. A . Western blot analysis of MetO in normal (NH #1–3) and diabetic human heart tissue(DH #1–6). GAPDH served as a loading control. B . Quantification of MetO signal intensity. C . Western blot analysis of methionine sulfoxide reductase A and B2 (MsrA and B2) in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as loading a loading control. D . Quantification of MsrA and MsrB2 signal intensity. E . Western blot analysis of LC3l/II and p62 in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as a loading control. F . Quantification of LC3l/II and p62 signal intensity. G . Western blot analysis of DRP1 and OPA1 in normal (NH #1–3) and diabetic heart tissues (DH #1–6). GAPDH served as a loading control. H . Quantification of MetO intensity

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Western Blot, Control

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Summary. Increased MsrB2 expression in diabetic mouse hearts suppressed cardiac complications. These results were confirmed by inducing diabetes in mice with suppressed MsrB2 expression (MsrB2 KO). In MsrB2 KO DM, the expression of SERCA2a, which regulates myocardial contractility, was reduced, cardiac fibrosis increased, and the number of mitochondrial structural abnormalities increased

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Expressing

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Cardiac complications occur in MsrB2 KO DM. A . Glucose tolerance test (GTT) in WT and MsrB2 KO under nonDM and DM conditions. B . Body weight in WT and MsrB2 KO DM mice during HFD feeding. The body weight was weighed every week. C . Electron microscopy (EM) images in WT and MsrB2 KO DM mice. The white arrow indicates the mitochondria (autophagosome and autolysosome). The black triangle indicates the membrane structure. A white triangle indicates an abnormal Z-line. D . H&E and trichrome staining of histological sections of the heart of WT and MsrB2 KO mice under nonDM and DM MsrB2 conditions. E . Fibrotic areas were quantified using the ImageJ (FIJI) analyzer on histological sections. The two-way ANOVA analysis was performed for p values

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques: Electron Microscopy, Membrane, Staining

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Decreased autophagy and SERCA2a-PLN function in MsrB2 KO DM mice. A . A Western blot analysis of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL, PNL, and GAPDH of nonDM (WT #1–3, MsrB2 KO #1–3) and DM mouse (WT #1–3, MsrB2 KO #1–6) hearts. GAPDH served as a loading control. B – D . Quantification of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL/PNL, and GAPDH signal intensity. E . Quantitative RT-PCR analysis MsrB2 and SERCA2a transcript levels in nonDM (WT #1–5, MsrB2 KO #1–5) and DM (WT #1–3, MsrB2 KO #1–6) mouse hearts. The two-way ANOVA analysis was performed for p values

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques: Western Blot, Control, Quantitative RT-PCR

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) and mitochondrial dysfunction in the heart of diabetic mice. A . Blood Glucose in nonDM and DM mice. B . Western blot analysis of MetO and MsrB2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. C . Quantification of MetO and MsrB2 signal intensity. D . Western blot analysis of Parkin and LC3II in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. E . Quantification of Parkin and LC3II signal intensity. F . Western blot analysis of MsrB2 and LC3I/II in nonDM (#1–3), DM mouse hearts (DM #1–3), and DM with NAC treatment (#1–5). GAPDH served as a loading control. G . Quantification of MsrB2 and LC3II signal intensity. H . Electron microscopy (EM) in nonDM mice. The white body indicates a mitophagy structure. I . EM in DM mice. The White body indicates mitophagy (autophagosome and autolysosome) structure. J . Quantification of mitochondria size in nonDM and DM mice. K . Western blot analysis of DRP1 and DRP1 and Mfn2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. L . Quantification of DRP1signal intensity. The nonparametric t -test was performed for p values

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques: Western Blot, Control, Electron Microscopy

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Electrophysiological dysfunctions occurred in MsrB2 KO DM mice. A ~ I . ECG recording results in nonDM (WT #1–4, MsrB2 KO #1–5) and DM (WT #1–11, MsrB2. KO #1–9) mouse hearts

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques:

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: MsrB2 induces autophagy by LC3 activation. A . Western blot analysis of MsrB2 and LC3I/II in H9C2 after MsrB2-GFP transfection. MsrB2 induced LC3II in a concentration-dependent manner. GAPDH served as a loading control. B . Quantification of MsrB2 and LC3II signal intensity. C . Western blot analysis of MsrB2, SERCA2a, and bMHC in WT and MsrB2 KO NMCM. After 4 h of starvation, 5.5 or 25 mM glucose was treated for 48 h. GAPDH served as a loading control. D . Quantification SERCA2a and bMHC signal intensity. E . Western blot analysis of MsrB2, LC3I/II, pDRP1, and DRP1 in NMCM after Ad-MsrB2-GFP infection. Ad-MsrB2-GFP 50 m.o.i infected into NMCM. Then, 5.5 or 25 mM glucose was treated for 48 h after 4 h starvation. GAPDH served as a loading control. F . Quantification of LC3II and pDRP1 signal intensity. The one-way ANOVA analysis was performed for p values

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques: Activation Assay, Western Blot, Transfection, Concentration Assay, Control, Infection

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) in human diabetic hearts. A . Western blot analysis of MetO in normal (NH #1–3) and diabetic human heart tissue(DH #1–6). GAPDH served as a loading control. B . Quantification of MetO signal intensity. C . Western blot analysis of methionine sulfoxide reductase A and B2 (MsrA and B2) in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as loading a loading control. D . Quantification of MsrA and MsrB2 signal intensity. E . Western blot analysis of LC3l/II and p62 in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as a loading control. F . Quantification of LC3l/II and p62 signal intensity. G . Western blot analysis of DRP1 and OPA1 in normal (NH #1–3) and diabetic heart tissues (DH #1–6). GAPDH served as a loading control. H . Quantification of MetO intensity

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques: Western Blot, Control

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Summary. Increased MsrB2 expression in diabetic mouse hearts suppressed cardiac complications. These results were confirmed by inducing diabetes in mice with suppressed MsrB2 expression (MsrB2 KO). In MsrB2 KO DM, the expression of SERCA2a, which regulates myocardial contractility, was reduced, cardiac fibrosis increased, and the number of mitochondrial structural abnormalities increased

Article Snippet: We purchased Parkin and MsrB2 ORF clones from OriGene (USA) and subcloned them into RFP- and GFP-tagged vectors, respectively.

Techniques: Expressing

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Cardiac complications occur in MsrB2 KO DM. A . Glucose tolerance test (GTT) in WT and MsrB2 KO under nonDM and DM conditions. B . Body weight in WT and MsrB2 KO DM mice during HFD feeding. The body weight was weighed every week. C . Electron microscopy (EM) images in WT and MsrB2 KO DM mice. The white arrow indicates the mitochondria (autophagosome and autolysosome). The black triangle indicates the membrane structure. A white triangle indicates an abnormal Z-line. D . H&E and trichrome staining of histological sections of the heart of WT and MsrB2 KO mice under nonDM and DM MsrB2 conditions. E . Fibrotic areas were quantified using the ImageJ (FIJI) analyzer on histological sections. The two-way ANOVA analysis was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Electron Microscopy, Membrane, Staining

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Decreased autophagy and SERCA2a-PLN function in MsrB2 KO DM mice. A . A Western blot analysis of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL, PNL, and GAPDH of nonDM (WT #1–3, MsrB2 KO #1–3) and DM mouse (WT #1–3, MsrB2 KO #1–6) hearts. GAPDH served as a loading control. B – D . Quantification of MsrB2, LC3I/II, Parkin, SERCA2a, pPNL/PNL, and GAPDH signal intensity. E . Quantitative RT-PCR analysis MsrB2 and SERCA2a transcript levels in nonDM (WT #1–5, MsrB2 KO #1–5) and DM (WT #1–3, MsrB2 KO #1–6) mouse hearts. The two-way ANOVA analysis was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Western Blot, Control, Quantitative RT-PCR

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) and mitochondrial dysfunction in the heart of diabetic mice. A . Blood Glucose in nonDM and DM mice. B . Western blot analysis of MetO and MsrB2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. C . Quantification of MetO and MsrB2 signal intensity. D . Western blot analysis of Parkin and LC3II in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. E . Quantification of Parkin and LC3II signal intensity. F . Western blot analysis of MsrB2 and LC3I/II in nonDM (#1–3), DM mouse hearts (DM #1–3), and DM with NAC treatment (#1–5). GAPDH served as a loading control. G . Quantification of MsrB2 and LC3II signal intensity. H . Electron microscopy (EM) in nonDM mice. The white body indicates a mitophagy structure. I . EM in DM mice. The White body indicates mitophagy (autophagosome and autolysosome) structure. J . Quantification of mitochondria size in nonDM and DM mice. K . Western blot analysis of DRP1 and DRP1 and Mfn2 in nonDM (#1–3) and DM mouse hearts (DM #1–3). GAPDH served as a loading control. L . Quantification of DRP1signal intensity. The nonparametric t -test was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Western Blot, Control, Electron Microscopy

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Electrophysiological dysfunctions occurred in MsrB2 KO DM mice. A ~ I . ECG recording results in nonDM (WT #1–4, MsrB2 KO #1–5) and DM (WT #1–11, MsrB2. KO #1–9) mouse hearts

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques:

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: MsrB2 induces autophagy by LC3 activation. A . Western blot analysis of MsrB2 and LC3I/II in H9C2 after MsrB2-GFP transfection. MsrB2 induced LC3II in a concentration-dependent manner. GAPDH served as a loading control. B . Quantification of MsrB2 and LC3II signal intensity. C . Western blot analysis of MsrB2, SERCA2a, and bMHC in WT and MsrB2 KO NMCM. After 4 h of starvation, 5.5 or 25 mM glucose was treated for 48 h. GAPDH served as a loading control. D . Quantification SERCA2a and bMHC signal intensity. E . Western blot analysis of MsrB2, LC3I/II, pDRP1, and DRP1 in NMCM after Ad-MsrB2-GFP infection. Ad-MsrB2-GFP 50 m.o.i infected into NMCM. Then, 5.5 or 25 mM glucose was treated for 48 h after 4 h starvation. GAPDH served as a loading control. F . Quantification of LC3II and pDRP1 signal intensity. The one-way ANOVA analysis was performed for p values

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Activation Assay, Western Blot, Transfection, Concentration Assay, Control, Infection

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: ROS induced methionine sulfoxidation (MetO) in human diabetic hearts. A . Western blot analysis of MetO in normal (NH #1–3) and diabetic human heart tissue(DH #1–6). GAPDH served as a loading control. B . Quantification of MetO signal intensity. C . Western blot analysis of methionine sulfoxide reductase A and B2 (MsrA and B2) in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as loading a loading control. D . Quantification of MsrA and MsrB2 signal intensity. E . Western blot analysis of LC3l/II and p62 in normal (NH #1–3) and diabetic heart tissue (DH #1–6). GAPDH served as a loading control. F . Quantification of LC3l/II and p62 signal intensity. G . Western blot analysis of DRP1 and OPA1 in normal (NH #1–3) and diabetic heart tissues (DH #1–6). GAPDH served as a loading control. H . Quantification of MetO intensity

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Western Blot, Control

Journal: Diabetology & Metabolic Syndrome

Article Title: Methionine sulfoxide reductase B2 protects against cardiac complications in diabetes mellitus

doi: 10.1186/s13098-024-01390-0

Figure Lengend Snippet: Summary. Increased MsrB2 expression in diabetic mouse hearts suppressed cardiac complications. These results were confirmed by inducing diabetes in mice with suppressed MsrB2 expression (MsrB2 KO). In MsrB2 KO DM, the expression of SERCA2a, which regulates myocardial contractility, was reduced, cardiac fibrosis increased, and the number of mitochondrial structural abnormalities increased

Article Snippet: Mouse heart tissue lysates and cell lysates (after transient transfection) were mixed with the specific target antibody [1 μg of LC3 anti-rabbit antibody (Abcam) and 2 μg of Parkin anti-goat antibody (Abcam) or 1.5 μg of Parkin anti-rabbit antibody (Abcam) for Parkin IP; 1 μg of MsrB2 anti-rabbit antibody (Yale) for MsrB2 IP; and GFP-Trap bead (Chromotek, USA), and the same species IgG control with HC] and incubated overnight at 4 °C.

Techniques: Expressing