Journal: Journal of Translational Medicine

Article Title: Secretion of miRNA-326-3p by senescent adipose exacerbates myocardial metabolism in diabetic mice

doi: 10.1186/s12967-022-03484-7

Figure Lengend Snippet: Removal of epididymal adipose tissue alleviates diastolic dysfunction in STZ mice. A Representative images of senescence associated beta-galactosidase (SA-β-gal) staining of EAT. B Detection of SA-β-gal positive cells from Con- or STZ-EAT (n = 3 per group). C Relative gene expression of senescence associated genes in EAT from Con- or STZ-mice (n = 3 per group). D Experimental design of EAT removal surgery. E Representative of doppler and tissue doppler echocardiography and evaluation of diastolic function (n = 6 per group). F Sarcomere length tracing of isolated murine cardiomyocytes using an Ionoptix HTS system. Evaluation of G myocardial contraction, H fraction shortening and I calcium handling in Langendorff-isolated adult mouse ventricular myocytes (AMVMs) (n = 5 per group). J Representative micrographs of heart tissue sections examined by transmission electron micrographs. Data are presented as the mean ± SEM; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 compared to Con group; # P < 0.05, ## P < 0.01, ### P < 0.001 compared to STZ + EATr group

Article Snippet: Briefly, adipose tissue chunks were collected in PBS, fixed with fixative solution (Solarbio) for 15 min. Then, the adipose tissue chunks were washed 3 times in PBS and placed in SA-β-gal activity solution (Solarbio).

Techniques: Staining, Expressing, Isolation, Transmission Assay

Journal: Journal of Translational Medicine

Article Title: Secretion of miRNA-326-3p by senescent adipose exacerbates myocardial metabolism in diabetic mice

doi: 10.1186/s12967-022-03484-7

Figure Lengend Snippet: LEVs isolated from senescent EAT in diabetic mice drives contractile and mitochondrial dysfunction in NMVMs. A Representative transmission electron micrograph of isolated LEVs. B Representative micrograph of EAT derived LEVs (PKH67-labeled, green) co-cultured with NMVMs. C Contractile velocity of untreated (UT) or NMVMs treated with Con-EAT LEVs or STZ-EAT LEVs (n = 6 per group). D Representative micrograph and E quantification of untreated (UT) and NMVMs treated with Con-EAT LEVs or STZ-EAT LEVs stained for mitochondrial membrane potential (TMRM), mitochondrial superoxide (MitoSox), and DAPI (n = 5 per group). F Real-time oxygen consumption rates (OCR) were evaluated for untreated (UT) or NMVMs treated with Con-EAT LEVs or STZ-EAT LEVs; basal and maximal respiration rates are shown (n = 8–9 per group). G Percentage of SA-β-gal positive cells in STZ-EAT cultured ex vivo in presence of Seno or Veh (n = 3 per group). H Relative gene expression of senescence associated genes in STZ-EAT cultured ex vivo in presence of Seno or Veh. (n = 3 per group). I Contractile velocity, J mitochondrial membrane potential and mitochondrial superoxide relative intensity fluorescence of NMVMs treated with LEVs from STZ-EAT cultured ex vivo in presence of Seno or Veh (n = 5–6 per group). (K) Real-time oxygen consumption rates (OCR) were evaluated for NMVMs treated with LEVs from STZ-EAT cultured ex vivo in presence of Seno or Veh, basal and maximal respiration rates are shown (n = 8–9 per group). Data are presented as the mean ± SEM; *P < 0.05, ***P < 0.001, ****P < 0.0001 compared to UT group; ## P < 0.01, ### P < 0.001, #### P < 0.0001 compared to Con-EAT LEVs group; $ P < 0.05, $$ P < 0.01, $$$ P < 0.001 compared to Veh-STZ-EAT LEVs group

Article Snippet: Briefly, adipose tissue chunks were collected in PBS, fixed with fixative solution (Solarbio) for 15 min. Then, the adipose tissue chunks were washed 3 times in PBS and placed in SA-β-gal activity solution (Solarbio).

Techniques: Isolation, Transmission Assay, Derivative Assay, Labeling, Cell Culture, Staining, Ex Vivo, Expressing, Fluorescence

Journal: Journal of Translational Medicine

Article Title: Secretion of miRNA-326-3p by senescent adipose exacerbates myocardial metabolism in diabetic mice

doi: 10.1186/s12967-022-03484-7

Figure Lengend Snippet: Senolytic treatment alleviates cardiac function in STZ mice. A Experiment design for senolytic treatment in vivo. B Representative images and C quantification of (SA-β-gal) staining of EAT isolated from Veh or Seno treated STZ-mice (n = 3 per group). D Relative gene expression of senescence associated genes in Veh or Seno treated STZ-EAT (n = 3 per group). E Representative of doppler and tissue doppler echocardiography and evaluation of diastolic function (n = 6 per group). F Contractile function and G calcium handling of AMVMs isolated from Veh or Seno treated STZ-mice were evaluated (n = 5 per group). H Representative transmission electron micrographs of Veh or Seno treated STZ-mice hearts. Relative miRNA-326-3p expression in I blood and J AMVMs from Veh or Seno treated STZ-mice (n = 4–5 per group). K Protein levels of Rictor, p-AKT, AKT and Gapdh in AMVMs isolated from Veh or Seno treated STZ-mice evaluated by immunoblotting (n = 3). Data are presented as the mean ± SEM; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 compared to Veh group

Article Snippet: Briefly, adipose tissue chunks were collected in PBS, fixed with fixative solution (Solarbio) for 15 min. Then, the adipose tissue chunks were washed 3 times in PBS and placed in SA-β-gal activity solution (Solarbio).

Techniques: In Vivo, Staining, Isolation, Expressing, Transmission Assay, Western Blot

Journal: Gastroenterology

Article Title: The epigenetic reader, BRD2, mediates cholangiocyte senescence via interaction with ETS1

doi: 10.1053/j.gastro.2023.03.235

Figure Lengend Snippet: A - D . Control NHC (no LPS), NHCsen (LPS), and PSCDCs (PSC) were stained for SA-β-gal and immunoblots performed to assess senescence (p21) and BET protein expression. Approximately 40% of the NHCsen (A) and PSCDCs (C) exhibited SA-β-gal positivity. Following treatment with BET protein inhibitors (JQ1, dBET1, and GSK046), SA-β-gal positivity was observed in less than 10% of the cholangiocytes. n = 5 independent experiments. NHCsen exhibited increased BRD2, BRD4, and p21, while the BET protein inhibitors reduced their expression. (B) . PSCDCs exhibited increased BRD2 and p21 compared to NHC control and the BET inhibitors reduced expression of both (D) . Western blots shown are representative from 3 separate experiments. E . Non-cholestatic liver disease patient cholangiocytes were isolated from bile and grown as organoids, induced to senescence (LPS), and cultured in the presence/absence of BET inhibitors for 48 hours. Each BET inhibitor decreased SA-β-gal positivity as measured by fluorescent intensity, n = 3 independent experiments. F . Immunoblots reveal that induced senescent cholangiocyte organoids exhibited increased BRD2, BRD4, and p21 protein expression compared to control cholangiocyte organoids. BET inhibitors reduced expression of each of these. G. Cholangiocytes isolated from PSC patient bile (PSC) were grown as organoids and exhibited increased SA-β-gal positivity compared to non-cholestatic liver disease patient cholangiocyte organoids (Normal); each BET inhibitor decreased PSC organoid SA-β-gal positivity, n = 3 independent experiments. Representative immunoblots from n = 3 independent experiments.

Article Snippet: Cells grown in 12-well plates were washed, fixed, and stained with the SA-β-gal (Cell Biolabs).

Techniques: Control, Staining, Western Blot, Expressing, Isolation, Cell Culture