Journal: Aging Cell

Article Title: Environmental Enrofloxacin Exposure as a Modifiable Driver of Mitochondria‐Mediated Intestinal Aging and Barrier Dysfunction

doi: 10.1111/acel.70526

Figure Lengend Snippet: Effects of PQQ treatment on mitochondrial function, inflammation, and hypoxia levels in the intestine following ENR exposure. Experimental design diagram (A). Representative immunofluorescence images of mitochondrial function‐related proteins (Hsp60, Cox4, Tomm20, Grp75, Cox5a) and quantification (B–I). Representative immunofluorescence images of CD3 and quantification (J–K). Representative immunofluorescence images of hypoxia markers and quantification (L, M). Data are presented as the mean ± standard error of the mean. Statistical significance among the Control, ENR, and ENR + PQQ groups was assessed using one‐way ANOVA followed by Tukey's multiple‐comparisons test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Intestinal tissues were fixed (4% paraformaldehyde, 15 min), blocked (10% goat serum albumin, 0.4% Triton X‐100), and incubated with primary antibodies from Servicebio: Cdkn1a (GB11153, 1:300), Cdkn2a ( GB151143 , 1:300), Tomm20 ( GB151481 , 1:1000), Grp75 ( GB112273 , 1:650), Cox5a ( GB111676 , 1:500), Hsp60 ( GB112464 , 1:800), Cox4 (GB11250, 1:200), CD3 (GB13014, 1:100), Mucin‐2 (GB11344, 1:500), Occludin ( GB111401 , 1:750), Zo‐1 ( GB115686 , 1:1000), and Claudin‐1 ( GB112543 , 1:1000).

Techniques: Immunofluorescence, Control

Journal: Frontiers in Immunology

Article Title: Overexpression of mitochondrial STAT3 protein improves colonic inflammation and fibrosis in inflammatory bowel disease by enhancing mitochondrial function

doi: 10.3389/fimmu.2026.1728341

Figure Lengend Snippet: mtSTAT3 regulates mitochondrial function and fibrosis in human intestinal cells. Overexpression of mtSTAT3 reduced the fibrosis marker C18. (A) Lysates of C18 cells transfected with mock and mtSTAT3 overexpression vector were analyzed for mtSTAT3 protein in mitochondria by Western blotting. (B, C) mRNA levels of the OXPHOS complex genes COX4 and ATP5O and the fibrosis genes aSMA, fibronectin, and COL1A1 according to real-time PCR.

Article Snippet: The tissue sections were stained for COX4 (NB110–39115; Novus, St. Louis, MO, USA), p727-STAT3 (ab32143; Abcam, Cambridge, UK), and DAPI (D3571; Invitrogen, Waltham, MA, USA); washed with phosphate-buffered saline (PBS); fixed in 4% paraformaldehyde; washed again with PBS; and blocked for 30 min with 10% normal goat serum.

Techniques: Over Expression, Marker, Transfection, Plasmid Preparation, Western Blot, Real-time Polymerase Chain Reaction

Journal: bioRxiv

Article Title: The COX2-PGE2-PKA Axis Suppresses Antiviral Immunity by Inhibiting mtDNA-Dependent STING Activation

doi: 10.64898/2026.04.03.716411

Figure Lengend Snippet: (A) Proteomic workflow in HEK293 cells with doxycycline-inducible expression of PKA Cα wild-type (WT) or W197R mutant (MUT). (B) The top 8 significantly enriched KEGG pathways within the mitochondrial quality control category identified from the proteomic dataset. (C) Schematic of the mt-mKeima mitophagy reporter. (D) Representative Airyscan live-cell imaging of THP-1 macrophages expressing mt-mKeima sensor treated with 1 µM PGE 2 in the presence of either 1 µM EP4 inhibitor (EP4i) or 1 µM PKA inhibitor (PKAi) for 16 hours. Scale bar, 10 µm. (E) Quantification of mitolysosome numbers shown in (D) (n = 10). Data were quantified from one representative experiment of three. (F) Representative images of live-cell 4D lattice light sheet imaging on THP-1 macrophages treated as in (D) . Scale bar, 20 µm. (G-H) Quantification of net mitolysosome displacement (n = 10) (G) and average mitolysosome speed (n = 12) (H) from imaging in (F) . (I) THP-1 macrophages treated with PGE 2 at indicated concentrations in the presence or absence of 1 µM PKA inhibitor (PKAi) for 16 hours. Mitochondrial fractions were isolated and subjected to immunoblotting with indicated antibodies. Band intensities were quantified and normalized to COX4 expression for PINK1 (J) and pUB Ser65 (K) (n = 3). (L) Working model illustrating that PGE 2 induces mitophagy and mitochondrial biogenesis to enhance mitochondrial homeostasis in a EP4- and PKA-dependent manner. Data are presented as mean ± s.e.m. Statistical significance was determined by one-way ANOVA followed by Sidak’s multiple comparisons test. p -values are indicated.

Article Snippet: Antibodies for PKA substrates (#9624), PKA Cα (#5842), PKA RIα/β (#3927), GAPDH (#2118), STING (#13647), pTBK1 S172 (5483), TBK1 (#3504), pIRF3 S396 (#4947), IRF3 (#4302), TFAM (#8076), HSP90 (#4877), pUB S65 (#62802), COX4 (#4850), pPINK1 S228 (#89010), STOML2 (#89199), Vinculin (#13901) were purchased from Cell Signaling Technology.

Techniques: Expressing, Mutagenesis, Control, Live Cell Imaging, Imaging, Isolation, Western Blot

Journal: bioRxiv

Article Title: The COX2-PGE2-PKA Axis Suppresses Antiviral Immunity by Inhibiting mtDNA-Dependent STING Activation

doi: 10.64898/2026.04.03.716411

Figure Lengend Snippet: (A) Network of mitochondrial quality control proteins interacting with wild-type (WT) and mutant (MUT) PKA Cα with BFDR σ; 0.2. Interactors are colored by log 2 fold change (WT/MUT spectral counts) with WT-specific interactors in dark purple and MUT-specific interactors in dark green. Edges to the central bait (yellow) represent interactions detected in this study. (B) Whole cell lysates of THP-1 macrophages were subjected to pulldown using 8-AHA-cAMP (RIα), Rp-8-AHA-cAMPS (holoenzyme), or HaloLink resin (control), followed by immunoblotting with indicated antibodies. (C-D) THP-1 macrophages were stimulated with 1 µM PGE 2 for 6 hours, followed by incubation with 1 µM cycloheximide for the indicated times. Cell lysates were collected and subjected to immunoblotting with the indicated antibodies. Representative blots from three independent experiments are shown (C) . Band intensities for STOML2 were quantified and normalized to HSP90 intensity (n = 3) (D) . (E-G) THP-1 macrophages were transfected with either scramble siRNA (siCTRL) or STOML2 siRNA (siSTOML2), followed by treatment with PGE 2 at indicated concentrations for 16 hours. Mitochondrial fractions were isolated and subjected to immunoblotting with the indicated antibodies (E) . Band intensities for PINK1 (F) and pUB Ser65 (G) were quantified and normalized to COX4 intensity (n = 3). (H) THP-1 macrophages were transfected with either scramble siRNA (siCTRL) or STOML2 siRNA (siSTOML2), followed by mock or HSV-1 infection in the presence or absence of 1 µM PGE 2 . At 16 h.p.i, cytosol fractions were isolated and subjected to qPCR to assess the presence of mt-Dloop and mt-ND1 regions (n=3). (I) THP-1 macrophages were transfected with either scramble siRNA (siCTRL) or STOML2 siRNA (siSTOML2), followed by mock or HSV-1 infection in the presence or absence of 1 µM PGE 2 . At 16 h.p.i, whole cell lysates were collected and subjected to RT-qPCR to assess mRNA levels of IFNβ (n=3). (J) THP-1 macrophages were transfected with either scramble siRNA (siCTRL) or STOML2 siRNA (siSTOML2), followed by mock or HSV-1 infection in the presence or absence of 1 µM PGE 2 . At 16 h.p.i, whole cell lysates were collected and subjected to qPCR to assess HSV-1 UL30 genomic abundance (n=3). All experiments were performed with three independent biological replicates and repeated at least twice with reproducible results. Data are presented as mean ± s.e.m. Statistical significance was determined by one-way ANOVA followed by Sidak’s multiple comparisons test. p -values are indicated.

Article Snippet: Antibodies for PKA substrates (#9624), PKA Cα (#5842), PKA RIα/β (#3927), GAPDH (#2118), STING (#13647), pTBK1 S172 (5483), TBK1 (#3504), pIRF3 S396 (#4947), IRF3 (#4302), TFAM (#8076), HSP90 (#4877), pUB S65 (#62802), COX4 (#4850), pPINK1 S228 (#89010), STOML2 (#89199), Vinculin (#13901) were purchased from Cell Signaling Technology.

Techniques: Control, Mutagenesis, Western Blot, Incubation, Transfection, Isolation, Infection, Quantitative RT-PCR