Journal: Advanced Science

Article Title: Metabolic Interplay in Acute Lung Injury: PARK7 Integrates FADS1/2‐Dependent PUFA Metabolism and H3K14 Lactylation to Attenuate Endothelial Ferroptosis and Dysfunction

doi: 10.1002/advs.202508725

Figure Lengend Snippet: PARK7 regulates FADS1/2 expression by upregulating bone morphogenetic protein (BMP)‐SMAD1/5/9 signaling. A) Quantitative real‐time PCR (qPCR) analysis of the expression of FADS1/2 precursor mRNA (n = 3). B) Gene set enrichment analysis (GSEA) of signaling pathways from the Pathway Interaction Database (PID). C) GSEA analysis of BMP signaling. D) Heatmap of expression levels for key BMP signaling molecules in human umbilical vein endothelial cells (HUVECs). E,F) qPCR (E) and Western blot (WB) (F) analysis of BMP2, BMP4, BMPR1A, and BMPR1B expression changes after PARK7 knockdown in HUVECs (n = 3). G) Immunofluorescence detection of phosphorylated SMAD1/5/9 (p‐SMAD1/5/9) expression in HUVECs with PARK7 overexpression or knockdown. H) WB analysis of expression levels of FADS1 and FADS2, the phosphorylation levels of SMAD1/5/9 in whole‐cell lysates, and the content of p‐SMAD1/5/9 in nuclear proteins were detected by WB in HUVECs overexpressing PARK7 , knocking down PARK7 , and treated with the BMP agonist sb4. I) WB quantification of the p‐SMAD1/5/9 to total SMAD5 ratio in HUVEC whole cell lysates (n = 4). J) WB quantification of nuclear p‐SMAD1/5/9 in HUVECs (n = 4). K) WB quantification of FADS1 and FADS2 in whole cell lysates of HUVECs (n = 4). L) ChIP‐qPCR validation of p‐SMAD1/5/9 binding to FADS1/2 gene promoters (n = 4). M) qPCR analysis of the expression of BMP2/4 and BMPR1A/B precursor mRNA (n = 4). N) WB analysis of NRF2 and KEAP1 after PARK7 knockdown. O) CoIP analysis of NRF2‐ubiquitin conjugation status following PARK7 knockdown. P) qPCR analysis of the expression of BMP2/4 after PARK7 knockdown and NEF2L2 overexpression (n = 4). Q) ChIP‐qPCR validation of NRF2 binding to BMP2/4 gene promoters (n = 3). R) Changes in BMPR1A/B mRNA levels over time after cycloheximide (CHX) administration (n = 3). S) qPCR analysis of the expression of BMPR1A/B after PARK7 knockdown and CMLD‐2 administration (n = 4). T) Western blot analysis of HuR expression changes following tBHP stimulation or PARK7 knockout, and the restorative effect of NAC. U) RIP assay validating changes in the binding capacity of HuR to BMPR1A/B mRNA (n = 5). V) Schematic diagram illustrating the dual pathways through which PARK7 regulates BMP/BMPR expression. CHX, cycloheximide; NAC, N‐Acetyl‐cysteine; tBHP, t‐Butyl hydroperoxide; ChIP, chromatin immunoprecipitation; CoIP, Co‐Immunoprecipitation; RIP, RNA immunoprecipitation; OE, overexpression; NC, negative control; Mock, transfection with vector plasmids and addition of corresponding solvents. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: Main reagents used in this study included LPS (L9143; Sigma–Aldrich, USA), nigericin (HY‐127019; MCE, China), ferrostatin‐1 (T6500; TargetMol, China), SC26196 ( T12856 ; TargetMol, China), DHA (T5369; TargetMol, China), erastin (HY‐15763; MCE, China), ML385 (T4360; TargetMol, China), ALA (T3P2904; TargetMol, China), CMLD‐2 ( T36493 ; TargetMol, China), sodium lactate (HY‐B2227B; MCE, China), 2‐DG(HY‐13966; MCE, China), BMP agonist sb4 (HY‐12469; MCE, China), medium chain triglycerides (MCT, S25953 ; Yuanye, China), puromycin (HY‐K1057; MCE, China), cycloheximide (HY‐12320; MCE, China) and ROS assay reagent DCFH‐DA (HY‐D0940; MCE, China).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Protein-Protein interactions, Western Blot, Knockdown, Immunofluorescence, Over Expression, Phospho-proteomics, ChIP-qPCR, Biomarker Discovery, Binding Assay, Ubiquitin Proteomics, Conjugation Assay, Knock-Out, Chromatin Immunoprecipitation, Immunoprecipitation, RNA Immunoprecipitation, Negative Control, Transfection, Plasmid Preparation

Journal: International Journal of Molecular Sciences

Article Title: Therapeutic Advances in Diabetes, Autoimmune, and Neurological Diseases

doi: 10.3390/ijms22062805

Figure Lengend Snippet: Approved and investigational biologics and small molecules for RA.

Article Snippet: SM , Decernotinib , Aclaris Therapeutics , Phase II , n/a , Oral.

Techniques:

Journal: Frontiers in Neurology

Article Title: Selective Inhibition of Janus Kinase 3 Has No Impact on Infarct Size or Neurobehavioral Outcomes in Permanent Ischemic Stroke in Mice

doi: 10.3389/fneur.2017.00363

Figure Lengend Snippet: (A) Dose–response curves measuring infarct volume in vehicle-treated and 0.03, 0.1, 0.3, 3, and 10 mg/kg of decernotinib-treated mice. There was no significant difference in infarct size across groups (vehicle, n = 8; 0.03 mg/kg, n = 6; 0.1 mg/kg, n = 8; 0.3 mg/kg, n = 8; 3 mg/kg, n = 10; and 10 mg/kg, n = 8). Data were analyzed using a one-way ANOVA with a Dunnett’s multiple comparison post-test. (B) Representative 2,3,5-triphenyl-tetrazolium chloride (TTC) images of vehicle- and 10 mg/kg decernotinib-treated mice to show extent of cortical injury in our model of permanent middle cerebral artery occlusion (MCAO). (C) Representative immunoblots of brain tissue obtained from the ipsilateral (CXI) and contralateral (CXC) cerebral cortex depicting reduced phosphorylation of Janus kinase 3 (JAK3) in the 10 mg/kg decernotinib group compared to vehicle. There is no effect of decernotinib on stroke-induced increase in total JAK3 levels. (D,E) Graphical summary of immunoblot data (* P < 0.05 compared to vehicle ipsilateral; vehicle, n = 9; 10 mg/kg decernotinib, n = 5). Data were analyzed using a two-way ANOVA. (F) Latency to remove the sticky tape from the contralateral paw, measuring sensorimotor deficits, in vehicle and decernotinib groups. Stroke induced a significant increase in the latency to remove the sticker at 24 h in the vehicle-treated group. Comparing both treatments at 24 h, there was a trend toward a better performance in the adhesive removal test in mice given decernotinib, but this effect did not reach a statistical significance (two-way ANOVA with Bonferroni post-test; vehicle n = 9; decernotinib at a dose of 10 mg/kg, n = 10).

Article Snippet: Animals were treated with vehicle or 0.03, 0.1, 0.3, 1.0, 3.0, or 10 mg/kg decernotinib (Cat # CT-DECE; ChemieTek, Indianapolis, IN, USA), a highly selective JAK3 inhibitor.

Techniques: Comparison, Western Blot, Phospho-proteomics, Adhesive

Journal: Frontiers in Neurology

Article Title: Selective Inhibition of Janus Kinase 3 Has No Impact on Infarct Size or Neurobehavioral Outcomes in Permanent Ischemic Stroke in Mice

doi: 10.3389/fneur.2017.00363

Figure Lengend Snippet: (A) IL-21 is increased in the ipsilateral cortex compared to the contralateral cortex (** P < 0.01) (B) IL-4 does not appreciably change between treatment groups or hemisphere. (C) There is a trend toward increased IL-7 in the ipsilateral cortex. (D) There is no change in IL-9 between groups. (E) There is no change in IL-15 between groups. (F) There is no significant change in IL-2 across groups. (G) Ccl2 is significantly increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (*** P < 0.01). (H) Cxcl10 is significantly increased in the ipsilateral cortex in both groups (*** P < 0.01; ** P < 0.01). (I) Cxcl1 is increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (*** P < 0.01). (J) IL-1 β is increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (*** P < 0.01). (K) IL-6 is increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (** P < 0.01; *** P < 0.01). (L) Tnfα is increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (*** P < 0.01). (M) Ptgs2 is increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (* P < 0.05; *** P < 0.01). (N) Mmp-9 is increased in the ipsilateral cortex in vehicle- and 10 mg/kg decernotinib-treated groups (** P < 0.01) Statistical analyses performed with a two-way ANOVA and Bonferroni post-test; vehicle n = 5, 10 mg/kg decernotinib, n = 7.

Article Snippet: Animals were treated with vehicle or 0.03, 0.1, 0.3, 1.0, 3.0, or 10 mg/kg decernotinib (Cat # CT-DECE; ChemieTek, Indianapolis, IN, USA), a highly selective JAK3 inhibitor.

Techniques:

Journal: Frontiers in Neurology

Article Title: Selective Inhibition of Janus Kinase 3 Has No Impact on Infarct Size or Neurobehavioral Outcomes in Permanent Ischemic Stroke in Mice

doi: 10.3389/fneur.2017.00363

Figure Lengend Snippet: (A) Representative immunoblot of mouse brain tissue. (B) Graphical summary of phosphorylated STAT3 immunoblot data normalized to actin (* P < 0.05; ** P < 0.01, vehicle n = 9; 10 mg/kg decernotinib, n = 5). Phosphorylated STAT3 is increased in the ipsilateral cortex compared to the contralateral. Phosphorylated STAT3 is decreased in the ipsilateral cortex with 10 mg/kg decernotinib compared to the vehicle-treated group. (C) Graphical summary of total STAT3 immunoblot data normalized to actin (vehicle, n = 9; 10 mg/kg decernotinib, n = 5). Total STAT3 does not vary between groups.

Article Snippet: Animals were treated with vehicle or 0.03, 0.1, 0.3, 1.0, 3.0, or 10 mg/kg decernotinib (Cat # CT-DECE; ChemieTek, Indianapolis, IN, USA), a highly selective JAK3 inhibitor.

Techniques: Western Blot