proteintech 29906 1 ap tsc2  (Proteintech)

Journal: Scientific Reports

Article Title: Macrophages lacking TSC2 have mTORC1-dependent increased GPNMB and ameliorate ventricular dysfunction/remodeling after ischemia-reperfusion

doi: 10.1038/s41598-025-24699-w

Figure Lengend Snippet: Constitutive rapamycin sensitive activation of mTORC1 in MΦ TSC2−/− BMDMs favors pro-inflammatory polarization responses in vitro. A , Immunoblot of TSC2 protein expression in BMDMs from MΦ TSC2+/+ versus MΦ TSC2−/− mice ( n = 5 per group, unpaired t-test). B , Upper : Immunoblot for P70S6K and 4E-BP1 phosphorylation and total protein in BMDMs from same two models incubated with vehicle or rapamycin. ( n = 4 per group); Lower : phospho/total protein. C , Left : Immunoblot of Akt activation from same experiment ( n = 4 per group), Right : Summary data; all analyzed by one-way ANOVA, Dunnett multiple correction test. D , Proliferation of BMDMs from both groups with vehicle (Con), LPS/IFN-γ or IL-4 stimulation. Upper : cell numbers ( n = 20/group), Lower : cell size ( n = 60/group). E , Response of BMDMs isolated from each model to IFN-γ + LPS stimulation ± rapamycin ( n = 6/group), F BMDM responses to IL-4 ± rapamycin ( n = 6/group). Data and mean ± SEM displayed, data normalized to MΦ-TSC2 +/+ results for each analysis, using two-way ANOVA with Šidák multiple comparisons test. * P < 1e-20 vs. MΦTSC2 +/+ , † P < 1e-10, § P < 0.05 vs. vehicle within genotype. 4E-BP1, eukaryotic translation initiation factor 4E-binding protein 1; P70S6K, ribosomal protein S6 kinase; Arg1, arginase 1; LPS, lipopolysaccharide; Mrc1, mannose receptor C-type 1; Nos2, nitric oxide synthase 2; Retnla, resistin-like alpha; TNFα, tumor necrosis factor alpha, other abbreviations in text.

Article Snippet: The I/R protocol was performed in three different cohorts: (1) control and MΦ TSC2−/− mice; (2) the same groups pre-administered 400 μg anti-Ly6G antibody (clone 1A8, BioXCell) i.p. starting 24 h prior to I/R and dosed 2x/week for 4 weeks (neutrophil depletion group) following established protocol ; and (3) control and MΦ TSC2−/− randomized to receive vehicle (3.3 mL/kg of 0.9% NaCl, 5% PEG400, 5% Tween 20, and 3% ethanol) or rapamycin (Target Mol Cat#53123-88-9, 2 mg/kg in vehicle) daily by intraperitoneal injection.

Techniques: Activation Assay, In Vitro, Western Blot, Expressing, Phospho-proteomics, Incubation, Isolation, Binding Assay

Journal: Scientific Reports

Article Title: Macrophages lacking TSC2 have mTORC1-dependent increased GPNMB and ameliorate ventricular dysfunction/remodeling after ischemia-reperfusion

doi: 10.1038/s41598-025-24699-w

Figure Lengend Snippet: MΦTSC2 −/− mice are protected against post-I/R cardiac dysfunction. A , Representative echocardiographic images at baseline and 4 weeks post– I/R in MΦ TSC2+/+ and MΦ TSC2−/− mice. B , Quantification of left ventricular ejection fraction, end-diastolic short axis diameter, stroke diameter (end-diastolic – end-systolic diameter), and percent anterior wall thickening ( n = 6 per group). C , Lung wet-to-dry weight ratios ( n = 6 per group). D , Masson’s trichrome stained cross section with percent fibrosis area ( n = 5 per group, unpaired t-test). E , Gene expression for hypertrophic and matrix remodeling genes, data normalized to MΦ TSC2−/− (WT) Sham. A-type natriuretic peptide, Nppa , b-myosin heavy chain, Myh7 , collagen type 2a1, Col2a1 , metalloproteinase 2, Mmp2 ( n = 6/group). All data shown with mean ± SD, panels B , C , and E analyzed with two-way ANOVA with Šidák multiple comparisons test.

Article Snippet: The I/R protocol was performed in three different cohorts: (1) control and MΦ TSC2−/− mice; (2) the same groups pre-administered 400 μg anti-Ly6G antibody (clone 1A8, BioXCell) i.p. starting 24 h prior to I/R and dosed 2x/week for 4 weeks (neutrophil depletion group) following established protocol ; and (3) control and MΦ TSC2−/− randomized to receive vehicle (3.3 mL/kg of 0.9% NaCl, 5% PEG400, 5% Tween 20, and 3% ethanol) or rapamycin (Target Mol Cat#53123-88-9, 2 mg/kg in vehicle) daily by intraperitoneal injection.

Techniques: Staining, Gene Expression

Journal: Scientific Reports

Article Title: Macrophages lacking TSC2 have mTORC1-dependent increased GPNMB and ameliorate ventricular dysfunction/remodeling after ischemia-reperfusion

doi: 10.1038/s41598-025-24699-w

Figure Lengend Snippet: Rapamycin prevents protection against cardiac I/R injury in MΦ TSC2–/– mice. A , B Example M-mode echocardiograms and summary data at 4 weeks post sham or I/R intervention in MΦ TSC2+/+ and MΦ TSC2–/– mice treated with either vehicle or rapamycin ( n = 5/group). Same parameters and analysis as shown in Fig. . C Lung wet-to-dry weight ratio, ( n = 5/group). D Representative heart sections and fibrotic area quantitation ( n = 7/group, unpaired t-test). E Gene expression of hypertrophy and matrix remodeling genes in same hearts ( n = 6/group). Analysis in panels B , C , and E are by two-way ANOVA with Šidák multiple comparison test. All data are shown along with mean ± SD.

Article Snippet: The I/R protocol was performed in three different cohorts: (1) control and MΦ TSC2−/− mice; (2) the same groups pre-administered 400 μg anti-Ly6G antibody (clone 1A8, BioXCell) i.p. starting 24 h prior to I/R and dosed 2x/week for 4 weeks (neutrophil depletion group) following established protocol ; and (3) control and MΦ TSC2−/− randomized to receive vehicle (3.3 mL/kg of 0.9% NaCl, 5% PEG400, 5% Tween 20, and 3% ethanol) or rapamycin (Target Mol Cat#53123-88-9, 2 mg/kg in vehicle) daily by intraperitoneal injection.

Techniques: Quantitation Assay, Gene Expression, Comparison

Journal: Scientific Reports

Article Title: Macrophages lacking TSC2 have mTORC1-dependent increased GPNMB and ameliorate ventricular dysfunction/remodeling after ischemia-reperfusion

doi: 10.1038/s41598-025-24699-w

Figure Lengend Snippet: Reduced myocardial pro-inflammatory immune cell infiltration 5 days post-I/R in MΦ TSC2−/− mice. A Number of CD45 + per mg tissue between MΦTSC2 +/+ and MΦTSC2 –/– hearts ( n = 5/group). B Number of CD64 + macrophages per mg in hearts from both models ( n = 5/group). C , Representative flow cytometry density plots for CCR2 + /CD64 + macrophages ( n = 5/group). D , Number of Ly6C + monocytes per mg tissue in both groups, ( n = 5/group). E Representative flow cytometry analysis of macrophage subsets ( n = 5/group). F Ly6G + (neutrophils), CD8 + and CD4 + (T cells), and CD19 + (B cells) each normalized to mg tissue for both groups ( n = 5). Analysis used two-tailed unpaired t-tests. Data are mean ± SEM. CD45, leukocyte common antigen; CCR2, C-C chemokine receptor type 2; CD64, Fc gamma receptor 1; Ly6C, lymphocyte antigen 6 complex locus C; Ly6G, lymphocyte antigen 6 complex locus G; MHCII, major histocompatibility complex class II; SEM, standard error of the mean; TSC2, tuberous sclerosis complex 2.

Article Snippet: The I/R protocol was performed in three different cohorts: (1) control and MΦ TSC2−/− mice; (2) the same groups pre-administered 400 μg anti-Ly6G antibody (clone 1A8, BioXCell) i.p. starting 24 h prior to I/R and dosed 2x/week for 4 weeks (neutrophil depletion group) following established protocol ; and (3) control and MΦ TSC2−/− randomized to receive vehicle (3.3 mL/kg of 0.9% NaCl, 5% PEG400, 5% Tween 20, and 3% ethanol) or rapamycin (Target Mol Cat#53123-88-9, 2 mg/kg in vehicle) daily by intraperitoneal injection.

Techniques: Flow Cytometry, Two Tailed Test, Immunopeptidomics

Journal: Scientific Reports

Article Title: Macrophages lacking TSC2 have mTORC1-dependent increased GPNMB and ameliorate ventricular dysfunction/remodeling after ischemia-reperfusion

doi: 10.1038/s41598-025-24699-w

Figure Lengend Snippet: Enhanced GPNMB protein in MΦ TSC2–/– BMDM and post IR-myocardium. A Western blot and quantification show increased GPNMB protein levels in BMDMs isolated from MΦ TSC2–/– mice compared to MΦ TSC2+/+ controls measured under basal conditions ( n = 5 per group, unpaired two-tailed t-test). B GPNMB levels elevated in MΦ TSC2–/– BMDMs are significantly reduced after rapamycin treatment ( n = 4 per group). C Western blot of cardiac tissue lysates 4 weeks post–I/R shows increased GPNMB expression in MΦ TSC2–/– hearts that is absent in the cohort treated with rapamycin ( n = 4 per group, D ELISA quantification of GPNMB protein from the same cardiac tissues confirmed the elevation in MΦTSC2 –/– hearts and its suppression by rapamycin ( n = 6 per group). Analysis in panels B – D , one-way ANOVA with Dunnett multiple comparisons correction. Data are mean ± SD. Rapa – rapamycin.

Article Snippet: The I/R protocol was performed in three different cohorts: (1) control and MΦ TSC2−/− mice; (2) the same groups pre-administered 400 μg anti-Ly6G antibody (clone 1A8, BioXCell) i.p. starting 24 h prior to I/R and dosed 2x/week for 4 weeks (neutrophil depletion group) following established protocol ; and (3) control and MΦ TSC2−/− randomized to receive vehicle (3.3 mL/kg of 0.9% NaCl, 5% PEG400, 5% Tween 20, and 3% ethanol) or rapamycin (Target Mol Cat#53123-88-9, 2 mg/kg in vehicle) daily by intraperitoneal injection.

Techniques: Western Blot, Isolation, Two Tailed Test, Expressing, Enzyme-linked Immunosorbent Assay