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Cell Signaling Technology Inc phospho nf κb
Phospho Nf κb, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 11398 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/phospho+nf+%CE%BAb/beta-Actin+Rabbit+mAb/pmc13049640-314-16-17
Average 99 stars, based on 11398 article reviews

phospho nf κb - by Bioz Stars, 2026-10

99/100 stars

Images

Incubation:

Article Title: Cannabinoid type 2 receptor regulates skeletal muscle regeneration by NLRP3-GSDMD mediated macrophage pyroptosis after injury.
Article Snippet: .. Sections were incubated overnight at 4 °C with primary antibodies against caspase‐1 (1:200, sc56036, Santa Cruz), IL‐1β (1:200, YT5201, Immunoway), and phospho‐NF‐κB (1:200, 4060, CST). .. Sections were incubated overnight at 4 °C with primary antibodies against caspase‐1 (1:200, sc56036, Santa Cruz), IL‐1β (1:200, YT5201, Immunoway), and phospho‐NF‐κB (1:200, 4060, CST).

Article Title: Cannabinoid type 2 receptor regulates skeletal muscle regeneration by NLRP3-GSDMD mediated macrophage pyroptosis after injury.
Article Snippet: .. After blocking with 5% BSA or 8% skim milk, membranes were incubated overnight at 4 °C with primary antibodies against: NLRP3 (1:1000, 1510S, CST), ASC (1:1000, sc514414, Santa Cruz), Pro‐caspase‐1 (1:1000, ab1872, Abcam), caspase‐1 (1:1000, sc56036, Santa Cruz), GSDMD (1:1000, ab209548, Abcam), GSDMD‐NT (1:1000, sc393656, Santa Cruz), IL‐18 (1:1000, ab191860, Abcam), IL‐1β (1:1000, abs120224, Absin), phospho‐PI3K (1:1000, 341468, ZENBIO), PI3K (1:1000, 251221, ZENBIO), phospho‐AKT (1:1000, 4060, CST), AKT (1:1000, 4691, CST), phospho‐NF‐κB (1:1000, 3033, CST), NF‐κB (1:1000, 10745, Proteintech), AR TI CL E IN P RE SS and GAPDH (1:3000, 60004, Proteintech). .. After blocking with 5% BSA or 8% skim milk, membranes were incubated overnight at 4 °C with primary antibodies against: NLRP3 (1:1000, 1510S, CST), ASC (1:1000, sc514414, Santa Cruz), Pro‐caspase‐1 (1:1000, ab1872, Abcam), caspase‐1 (1:1000, sc56036, Santa Cruz), GSDMD (1:1000, ab209548, Abcam), GSDMD‐NT (1:1000, sc393656, Santa Cruz), IL‐18 (1:1000, ab191860, Abcam), IL‐1β (1:1000, abs120224, Absin), phospho‐PI3K (1:1000, 341468, ZENBIO), PI3K (1:1000, 251221, ZENBIO), phospho‐AKT (1:1000, 4060, CST), AKT (1:1000, 4691, CST), phospho‐NF‐κB (1:1000, 3033, CST), NF‐κB (1:1000, 10745, Proteintech), AR TI CL E IN P RE SS and GAPDH (1:3000, 60004, Proteintech).

Article Title: Skin Barrier-Improving and Anti-Inflammatory Effects of Exosomes Derived from the Fructobacillus fructosus NSH-1 Strain Isolated from the Campsis grandiflora Flower.
Article Snippet: .. The membranes were then incubated overnight at 4oC with primary antibodies against iNOS (13120S, 1:500, Cell Signaling, USA), COX-2 (4842S, 1:500, Cell Signaling), phospho-ERK (9101S, 1:500, Cell Signaling), ERK (9102S, 1:500, Cell Signaling), phospho-JNK (9251S, 1:500, Cell Signaling), JNK (9252S, 1:500, Cell Signaling), phospho-p38 (9211S, 1:500, Cell Signaling), p38 (9212S, 1:500, Cell Signaling), phospho-NF-κB (#3033, 1:500, Cell Signaling), phospho-IκBα (#2859S, 1:500, Cell Signaling), filaggrin (sc-66192, 1:500, Santa Cruz Biotechnology, USA), loricrin (LS-C31863-100, 1:500, LSBio, USA), involucrin (sc-21748, 1:500, Santa Cruz Biotechnology), and β-actin (#3700, 1:5,000, Cell Signaling). ..

Blocking Assay:

Article Title: Cannabinoid type 2 receptor regulates skeletal muscle regeneration by NLRP3-GSDMD mediated macrophage pyroptosis after injury.
Article Snippet: .. After blocking with 5% BSA or 8% skim milk, membranes were incubated overnight at 4 °C with primary antibodies against: NLRP3 (1:1000, 1510S, CST), ASC (1:1000, sc514414, Santa Cruz), Pro‐caspase‐1 (1:1000, ab1872, Abcam), caspase‐1 (1:1000, sc56036, Santa Cruz), GSDMD (1:1000, ab209548, Abcam), GSDMD‐NT (1:1000, sc393656, Santa Cruz), IL‐18 (1:1000, ab191860, Abcam), IL‐1β (1:1000, abs120224, Absin), phospho‐PI3K (1:1000, 341468, ZENBIO), PI3K (1:1000, 251221, ZENBIO), phospho‐AKT (1:1000, 4060, CST), AKT (1:1000, 4691, CST), phospho‐NF‐κB (1:1000, 3033, CST), NF‐κB (1:1000, 10745, Proteintech), AR TI CL E IN P RE SS and GAPDH (1:3000, 60004, Proteintech). .. After blocking with 5% BSA or 8% skim milk, membranes were incubated overnight at 4 °C with primary antibodies against: NLRP3 (1:1000, 1510S, CST), ASC (1:1000, sc514414, Santa Cruz), Pro‐caspase‐1 (1:1000, ab1872, Abcam), caspase‐1 (1:1000, sc56036, Santa Cruz), GSDMD (1:1000, ab209548, Abcam), GSDMD‐NT (1:1000, sc393656, Santa Cruz), IL‐18 (1:1000, ab191860, Abcam), IL‐1β (1:1000, abs120224, Absin), phospho‐PI3K (1:1000, 341468, ZENBIO), PI3K (1:1000, 251221, ZENBIO), phospho‐AKT (1:1000, 4060, CST), AKT (1:1000, 4691, CST), phospho‐NF‐κB (1:1000, 3033, CST), NF‐κB (1:1000, 10745, Proteintech), AR TI CL E IN P RE SS and GAPDH (1:3000, 60004, Proteintech).

other:

Article Title: Hemoglobin inhibits fibroblast-to-cardiomyocyte reprogramming via TLR2/TLR4-dependent chromatin compaction
Article Snippet: Band intensities were analyzed with either ImageJ or Syngene software.

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Image Search Results

Cytokines Activated Their Downstream Pathways in HK-2 Cells. The phosphorylation of ERK, JNK, p38, and NF-κB in HK-2 cells treated with TNF-α (A) or IL-1β (B) was analyzed over a time course ranging from 15 min to 4 h. The phosphorylation of Smad1/5/9 and Smad2/3 in HK-2 cells treated with TGF-β1 (C) or BMP-2 (D) was analyzed over a time course ranging from 15 min to 4 h. GAPDH was served as the loading control.

Journal: Frontiers in Cell and Developmental Biology

Article Title: Inflammation-induced osteogenic signaling promotes calcium phosphate crystal formation in kidneys via MAPK, NF-κB, and smad pathways

doi: 10.3389/fcell.2026.1831072

Figure Lengend Snippet: Cytokines Activated Their Downstream Pathways in HK-2 Cells. The phosphorylation of ERK, JNK, p38, and NF-κB in HK-2 cells treated with TNF-α (A) or IL-1β (B) was analyzed over a time course ranging from 15 min to 4 h. The phosphorylation of Smad1/5/9 and Smad2/3 in HK-2 cells treated with TGF-β1 (C) or BMP-2 (D) was analyzed over a time course ranging from 15 min to 4 h. GAPDH was served as the loading control.

Article Snippet: Primary antibodies for immunoblotting such as OPN (A19092), ALP (A0514), OPG (A2100), Runx2 (A2851), p-JNK1/2/3 (AP0631), JNK1/2/3 (A4867), p-NF-κB (AP0944), NF-κB (A19653), p-p38 (AP0057), p38 (A14401), p-ERK (AP0485), ERK (A4782), p-Smad1/5/9 (AP0850), Smad1 (A21734), p-Smad2/3 (AP0548), Smad2/3 (A7536), TNF-α (A11534), IL-1β (A11534), TGF-β1 (A16640), BMP-2 (A0231), β-actin (AC028), α-Tubulin (A6830), and GAPDH (A19056) were from ABclonal (Woburn, MA, United States).

Techniques: Phospho-proteomics, Control

Pro-inflammatory Cytokines Upregulated Osteogenesis-related Proteins via the MAPK/NF-κB and Smad Signaling Pathway. HK-2 cells were pretreated with 10 µM ERK inhibitor (U0126), JNK inhibitor (SP600125), p38 inhibitor (SB203580), or NF-κB inhibitor (BAY 11–7,082) for 2 h, and then treated with 20 ng/mL TNF-α (A) or IL-1β (B) for 72 h (C) HK-2 cells were pretreated with 5 µM Smad2/3 inhibitor (SB-525334) for 2 h and then treated with 200 ng/mL TGF-β1 for 72 h (D) HK-2 cells were pretreated with 10 µM Smad1/5/8 inhibitor (LDN-193189) for 2 h and then treated with 100 ng/mL BMP-2 for 72 h. The expression of osteoblast markers (ALP, OPN, and OPG) was examined by Western blotting. The quantitative results showed the fold change of the protein expression compared to the control group. GAPDH served as the loading control. Data are presented as individual data points (scatter plot) with the mean ± SEM; n = 3; * P < 0.05, ** P < 0.01, *** P < 0.001 versus the control group; # P < 0.05, ## P < 0.01, ### P < 0.001 versus cytokine group (One-way ANOVA).

Journal: Frontiers in Cell and Developmental Biology

Article Title: Inflammation-induced osteogenic signaling promotes calcium phosphate crystal formation in kidneys via MAPK, NF-κB, and smad pathways

doi: 10.3389/fcell.2026.1831072

Figure Lengend Snippet: Pro-inflammatory Cytokines Upregulated Osteogenesis-related Proteins via the MAPK/NF-κB and Smad Signaling Pathway. HK-2 cells were pretreated with 10 µM ERK inhibitor (U0126), JNK inhibitor (SP600125), p38 inhibitor (SB203580), or NF-κB inhibitor (BAY 11–7,082) for 2 h, and then treated with 20 ng/mL TNF-α (A) or IL-1β (B) for 72 h (C) HK-2 cells were pretreated with 5 µM Smad2/3 inhibitor (SB-525334) for 2 h and then treated with 200 ng/mL TGF-β1 for 72 h (D) HK-2 cells were pretreated with 10 µM Smad1/5/8 inhibitor (LDN-193189) for 2 h and then treated with 100 ng/mL BMP-2 for 72 h. The expression of osteoblast markers (ALP, OPN, and OPG) was examined by Western blotting. The quantitative results showed the fold change of the protein expression compared to the control group. GAPDH served as the loading control. Data are presented as individual data points (scatter plot) with the mean ± SEM; n = 3; * P < 0.05, ** P < 0.01, *** P < 0.001 versus the control group; # P < 0.05, ## P < 0.01, ### P < 0.001 versus cytokine group (One-way ANOVA).

Techniques: Expressing, Western Blot, Control

Hypothesized Model of Randall’s Plaque Formation. a, In a high-calcium renal environment, activated macrophages and inflammasomes release pro-inflammatory cytokines, activating signaling pathways such as MAPK, NF-κB, and Smad in renal tubular cells. b, These signaling pathways upregulate proteins involved in osteogenesis, such as Runx2, ALP, OPG, and OPN, and result in an osteogenic transition. c, Calcium-enriched and osteogenesis-related protein-enriched vesicles are released from the basal side of tubule epithelial cells. Osteogenesis-related proteins directly promote the nucleation, growth and aggregation of CaP crystals in these vesicles. d, CaP aggregates-enriched vesicles move to the renal interstitium, calcifying the interstitial collagen, and eventually leading to the formation of Randall’s plaque in the renal papillae. This figure was created by BioRender.com .

Journal: Frontiers in Cell and Developmental Biology

Article Title: Inflammation-induced osteogenic signaling promotes calcium phosphate crystal formation in kidneys via MAPK, NF-κB, and smad pathways

doi: 10.3389/fcell.2026.1831072

Figure Lengend Snippet: Hypothesized Model of Randall’s Plaque Formation. a, In a high-calcium renal environment, activated macrophages and inflammasomes release pro-inflammatory cytokines, activating signaling pathways such as MAPK, NF-κB, and Smad in renal tubular cells. b, These signaling pathways upregulate proteins involved in osteogenesis, such as Runx2, ALP, OPG, and OPN, and result in an osteogenic transition. c, Calcium-enriched and osteogenesis-related protein-enriched vesicles are released from the basal side of tubule epithelial cells. Osteogenesis-related proteins directly promote the nucleation, growth and aggregation of CaP crystals in these vesicles. d, CaP aggregates-enriched vesicles move to the renal interstitium, calcifying the interstitial collagen, and eventually leading to the formation of Randall’s plaque in the renal papillae. This figure was created by BioRender.com .

Techniques: Protein-Protein interactions

Journal: Materials Today Bio

Article Title: Exosomal lncRNA OTUD6B-AS1 as a pathogenic nanocarrier promotes inflammatory macrophage polarization in endometritis via a targetable ceRNA circuit

doi: 10.1016/j.mtbio.2026.103027

Figure Lengend Snippet: Exosomes from LPS-stimulated EECs induce pro-inflammatory macrophage activation. (A) Schematic diagram of the experimental setup for exosome uptake. (B) Fluorescence microscopy images showing the uptake of PKH67-labeled exosomes (green) by macrophages. Cytoskeleton was stained with Phalloidin (red), and nuclei were stained with DAPI (blue). (C) Western blotting analysis of phosphorylated NF-κB p65 (p-p65) in macrophages treated with Control-exo or LPS-exo. (D, E) Representative immunofluorescence (IF) staining images (D) and quantitative analysis (E) of iNOS (greed) in macrophages. (F, G) Representative IF staining images (F) and quantitative analysis (G) of Arg1 (red) in macrophages. (H) Schematic diagram of co-culture experiments. (I, J) Relative mRNA expression levels of iNOS (I) and Arg1 (J) in macrophages after co-culture with EECs. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The antibodies used include TSG101 (Genuin Biotech, Cat # 51942), CD81 (Santa Cruz Biotechnology, Cat # sc-18877), AKT (abcam, Cat # AB81283), Phospho-AKT (abcam, Cat # AB179463 ), NF-κB p65 (abcam, Cat # AB32536), Phospho-NF-κB p65 (Cell Signaling Technology, Cat # 3033S), Notch2 (Santa Cruz Biotechnology, Cat # sc-51869), RBP-Jκ (Santa Cruz Biotechnology, Cat # sc-271128), and β-actin (Bioss, Cat # bs-0061R).

Techniques: Activation Assay, Fluorescence, Microscopy, Labeling, Staining, Western Blot, Control, Immunofluorescence, Co-Culture Assay, Expressing

Transcriptomic profiling reveals significant enrichment of lncRNA OTUD6B-AS1 in exosomes derived from LPS-stimulated EECs. (A, B) LPS-exo was treated with RNase A alone or in combination with Triton X-100 for 4 h, and then co-incubated with macrophages. Relative expression levels of iNOS (A) and Arg1 (B) in macrophages. (C) Schematic overview of the RNA sequencing and analysis workflow. (D) Volcano plot showing differentially expressed lncRNAs in LPS-exo compared to Control-exo. (E, F) Gene Ontology (GO) biological process enrichment analysis (E) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis (F) of the differentially expressed lncRNAs. (G) qPCR validation of the 6 upregulated lncRNAs in Control-exo and LPS-exo. (H) LPS-exo was treated with RNase A alone or in combination with Triton X-100 for 4 h, and then co-incubated with macrophages. Relative mRNA expression level of lncRNA OTUD6B-AS1 in macrophages. (I) A proposed competing endogenous RNA (ceRNA) network involving lncRNA OTUD6B-AS1, miR-128, and Notch2. (J) Relative mRNA expression level of lncRNA OTUD6B-AS1 in control and LPS-stimulated EECs. (K, L) RNA fluorescence in situ hybridization (RNA-FISH) showing the subcellular localization of lncRNA OTUD6B-AS1 (red) in EECs (K) and its quantitative cytoplasmic/nuclear distribution (L). Nuclei were stained with DAPI (blue). (M – P) Relative mRNA expression levels of lncRNA OTUD6B-AS1 (M − O) and miR-128 (P) in endometrial tissues from healthy cows and cows with endometritis, as determined by qPCR (O, P) and RNA-FISH (M) with quantification (N). (Q) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 protein levels in endometrial tissues. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Journal: Materials Today Bio

Article Title: Exosomal lncRNA OTUD6B-AS1 as a pathogenic nanocarrier promotes inflammatory macrophage polarization in endometritis via a targetable ceRNA circuit

doi: 10.1016/j.mtbio.2026.103027

Figure Lengend Snippet: Transcriptomic profiling reveals significant enrichment of lncRNA OTUD6B-AS1 in exosomes derived from LPS-stimulated EECs. (A, B) LPS-exo was treated with RNase A alone or in combination with Triton X-100 for 4 h, and then co-incubated with macrophages. Relative expression levels of iNOS (A) and Arg1 (B) in macrophages. (C) Schematic overview of the RNA sequencing and analysis workflow. (D) Volcano plot showing differentially expressed lncRNAs in LPS-exo compared to Control-exo. (E, F) Gene Ontology (GO) biological process enrichment analysis (E) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis (F) of the differentially expressed lncRNAs. (G) qPCR validation of the 6 upregulated lncRNAs in Control-exo and LPS-exo. (H) LPS-exo was treated with RNase A alone or in combination with Triton X-100 for 4 h, and then co-incubated with macrophages. Relative mRNA expression level of lncRNA OTUD6B-AS1 in macrophages. (I) A proposed competing endogenous RNA (ceRNA) network involving lncRNA OTUD6B-AS1, miR-128, and Notch2. (J) Relative mRNA expression level of lncRNA OTUD6B-AS1 in control and LPS-stimulated EECs. (K, L) RNA fluorescence in situ hybridization (RNA-FISH) showing the subcellular localization of lncRNA OTUD6B-AS1 (red) in EECs (K) and its quantitative cytoplasmic/nuclear distribution (L). Nuclei were stained with DAPI (blue). (M – P) Relative mRNA expression levels of lncRNA OTUD6B-AS1 (M − O) and miR-128 (P) in endometrial tissues from healthy cows and cows with endometritis, as determined by qPCR (O, P) and RNA-FISH (M) with quantification (N). (Q) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 protein levels in endometrial tissues. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Techniques: Derivative Assay, Incubation, Expressing, RNA Sequencing, Control, Biomarker Discovery, Fluorescence, In Situ Hybridization, Staining, Western Blot

EECs-derived exosomes induce pro-inflammatory macrophage activation via delivery of lncRNA OTUD6B-AS1. (A) Relative mRNA expression level of lncRNA OTUD6B-AS1 in macrophages treated with Control-exo or LPS-exo. (B, C) RNA-FISH images (B) and quantitative analysis (C) showing lncRNA OTUD6B-AS1 (red) transfer to macrophages after co-culture with Control-exo or LPS-exo. Nuclei were stained with DAPI (blue). (D) Relative mRNA expression level of lncRNA OTUD6B-AS1 in macrophages after transfection with lncRNA OTUD6B-AS1 overexpression plasmids (OE-lncRNA) or control plasmids (OE-NC). ( E – I) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (E), along with immunofluorescence (IF) quantitative analysis of iNOS (F, G) and Arg1 (H, I) protein levels in macrophages after transfection with OE-lncRNA or OE-NC. (J – N) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (J), along with IF quantitative analysis of iNOS (K, L) and Arg1 (M, N) protein levels in macrophages after lncRNA OTUD6B-AS1 knockdown (si-lncRNA) or control treatment (si-NC). (O) Relative mRNA expression level of lncRNA OTUD6B-AS1 in exosomes isolated from lncRNA OTUD6B-AS1-knockdown LPS-stimulated EECs (si-lncRNA-LPS-exo) or exosomes from siRNA NC-transfected LPS-stimulated EECs (si-NC-LPS-exo). (P – S) IF quantitative analysis of iNOS (P, Q) and Arg1 (R, S) protein levels in macrophages treated with si-lncRNA-LPS-exo or si-NC-LPS-exo. (T) Relative mRNA expression levels of iNOS and Arg1 in macrophages treated with exosomes isolated from control EECs overexpressing lncRNA OTUD6B-AS1 (OE-lncRNA-Control-exo) or exosomes from control plasmids-transfected EECs (OE-NC-Control-exo). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Journal: Materials Today Bio

Article Title: Exosomal lncRNA OTUD6B-AS1 as a pathogenic nanocarrier promotes inflammatory macrophage polarization in endometritis via a targetable ceRNA circuit

doi: 10.1016/j.mtbio.2026.103027

Figure Lengend Snippet: EECs-derived exosomes induce pro-inflammatory macrophage activation via delivery of lncRNA OTUD6B-AS1. (A) Relative mRNA expression level of lncRNA OTUD6B-AS1 in macrophages treated with Control-exo or LPS-exo. (B, C) RNA-FISH images (B) and quantitative analysis (C) showing lncRNA OTUD6B-AS1 (red) transfer to macrophages after co-culture with Control-exo or LPS-exo. Nuclei were stained with DAPI (blue). (D) Relative mRNA expression level of lncRNA OTUD6B-AS1 in macrophages after transfection with lncRNA OTUD6B-AS1 overexpression plasmids (OE-lncRNA) or control plasmids (OE-NC). ( E – I) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (E), along with immunofluorescence (IF) quantitative analysis of iNOS (F, G) and Arg1 (H, I) protein levels in macrophages after transfection with OE-lncRNA or OE-NC. (J – N) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (J), along with IF quantitative analysis of iNOS (K, L) and Arg1 (M, N) protein levels in macrophages after lncRNA OTUD6B-AS1 knockdown (si-lncRNA) or control treatment (si-NC). (O) Relative mRNA expression level of lncRNA OTUD6B-AS1 in exosomes isolated from lncRNA OTUD6B-AS1-knockdown LPS-stimulated EECs (si-lncRNA-LPS-exo) or exosomes from siRNA NC-transfected LPS-stimulated EECs (si-NC-LPS-exo). (P – S) IF quantitative analysis of iNOS (P, Q) and Arg1 (R, S) protein levels in macrophages treated with si-lncRNA-LPS-exo or si-NC-LPS-exo. (T) Relative mRNA expression levels of iNOS and Arg1 in macrophages treated with exosomes isolated from control EECs overexpressing lncRNA OTUD6B-AS1 (OE-lncRNA-Control-exo) or exosomes from control plasmids-transfected EECs (OE-NC-Control-exo). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Techniques: Derivative Assay, Activation Assay, Expressing, Control, Co-Culture Assay, Staining, Transfection, Over Expression, Western Blot, Immunofluorescence, Knockdown, Isolation

lncRNA OTUD6B-AS1 acts as a ceRNA by sponging miR-128 to facilitate pro-inflammatory macrophage activation. (A) Relative mRNA expression level of miR-128 in macrophages treated with Control-exo or LPS-exo. (B) Luciferase reporter assay in HEK293T cells co-transfected with wild-type (WT) or mutant (MUT) lncRNA OTUD6B-AS1 reporter plasmids and miR-128 mimic or mimic NC. (C) RNA pull-down detection of the enrichment of miR-128 to lncRNA OTUD6B-AS1. (D) Ago2 RIP assay analysis of the enrichment of lncRNA OTUD6B-AS1 pulled-down from the Ago2 protein. (E) Relative mRNA expression level of miR-128 in macrophages transfected with OE-NC or OE-lncRNA. (F – J) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (F), along with immunofluorescence (IF) quantitative analysis of iNOS (G, H) and Arg1 (I, J) protein levels in macrophages co-transfected with OE-lncRNA and miR-128 mimic or mimic NC. (K, L) Relative mRNA expression levels of IL-1β (K) and IL-6 (L) in macrophages co-transfected with OE-lncRNA and miR-128 mimic or mimic NC. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

Journal: Materials Today Bio

Article Title: Exosomal lncRNA OTUD6B-AS1 as a pathogenic nanocarrier promotes inflammatory macrophage polarization in endometritis via a targetable ceRNA circuit

doi: 10.1016/j.mtbio.2026.103027

Figure Lengend Snippet: lncRNA OTUD6B-AS1 acts as a ceRNA by sponging miR-128 to facilitate pro-inflammatory macrophage activation. (A) Relative mRNA expression level of miR-128 in macrophages treated with Control-exo or LPS-exo. (B) Luciferase reporter assay in HEK293T cells co-transfected with wild-type (WT) or mutant (MUT) lncRNA OTUD6B-AS1 reporter plasmids and miR-128 mimic or mimic NC. (C) RNA pull-down detection of the enrichment of miR-128 to lncRNA OTUD6B-AS1. (D) Ago2 RIP assay analysis of the enrichment of lncRNA OTUD6B-AS1 pulled-down from the Ago2 protein. (E) Relative mRNA expression level of miR-128 in macrophages transfected with OE-NC or OE-lncRNA. (F – J) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (F), along with immunofluorescence (IF) quantitative analysis of iNOS (G, H) and Arg1 (I, J) protein levels in macrophages co-transfected with OE-lncRNA and miR-128 mimic or mimic NC. (K, L) Relative mRNA expression levels of IL-1β (K) and IL-6 (L) in macrophages co-transfected with OE-lncRNA and miR-128 mimic or mimic NC. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

Techniques: Activation Assay, Expressing, Control, Luciferase, Reporter Assay, Transfection, Mutagenesis, Western Blot, Immunofluorescence

Notch2 mediates the regulatory effect of the lncRNA OTUD6B-AS1/miR-128 axis on macrophage activation. (A) Predictive analysis of miR-128 targets using multiple databases. (B) Western blotting analysis of Notch2 protein levels in macrophages treated with Control-exo or LPS-exo. (C) Western blotting analysis of Notch2 protein levels in macrophages transfected with OE-NC or OE-lncRNA. (D – H) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (D), along with immunofluorescence (IF) quantitative analysis of iNOS (E, F) and Arg1 (G, H) protein levels in macrophages treated with OE-NC or OE-lncRNA and the Notch2 inhibitor DAPT. (I) Luciferase reporter assay in HEK293T cells co-transfected with WT or MUT Notch2 3′UTR reporter plasmids and miR-128 mimic or mimic NC. (J, K) Relative protein (J) and mRNA (K) expression levels of Notch2 in macrophages transfected with miR-128 mimic or mimic NC. (L – P) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (L), along with IF quantitative analysis of iNOS (M, N) and Arg1 (O, P) protein levels in macrophages co-treated with miR-128 inhibitor or inhibitor NC and DAPT. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

Journal: Materials Today Bio

Article Title: Exosomal lncRNA OTUD6B-AS1 as a pathogenic nanocarrier promotes inflammatory macrophage polarization in endometritis via a targetable ceRNA circuit

doi: 10.1016/j.mtbio.2026.103027

Figure Lengend Snippet: Notch2 mediates the regulatory effect of the lncRNA OTUD6B-AS1/miR-128 axis on macrophage activation. (A) Predictive analysis of miR-128 targets using multiple databases. (B) Western blotting analysis of Notch2 protein levels in macrophages treated with Control-exo or LPS-exo. (C) Western blotting analysis of Notch2 protein levels in macrophages transfected with OE-NC or OE-lncRNA. (D – H) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (D), along with immunofluorescence (IF) quantitative analysis of iNOS (E, F) and Arg1 (G, H) protein levels in macrophages treated with OE-NC or OE-lncRNA and the Notch2 inhibitor DAPT. (I) Luciferase reporter assay in HEK293T cells co-transfected with WT or MUT Notch2 3′UTR reporter plasmids and miR-128 mimic or mimic NC. (J, K) Relative protein (J) and mRNA (K) expression levels of Notch2 in macrophages transfected with miR-128 mimic or mimic NC. (L – P) Western blotting analysis of Notch2, RBP-Jκ, and p-p65 (L), along with IF quantitative analysis of iNOS (M, N) and Arg1 (O, P) protein levels in macrophages co-treated with miR-128 inhibitor or inhibitor NC and DAPT. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

Techniques: Activation Assay, Western Blot, Control, Transfection, Immunofluorescence, Luciferase, Reporter Assay, Expressing

A proposed model illustrating the exosome-mediated lncRNA OTUD6B-AS1/miR-128/Notch2 axis in aggravating endometritis. Upon LPS-induced damage, endometrial epithelial cells (EECs) release increased exosomes carrying elevated levels of lncRNA OTUD6B-AS1. These exosomes are taken up by endometrial macrophages. The transferred lncRNA OTUD6B-AS1 acts as a molecular sponge to sequester miR-128, leading to the derepression and upregulation of its target gene, Notch2. The enhanced Notch2 signaling subsequently promotes macrophage polarization towards a pro-inflammatory M1 phenotype, characterized by increased NF-κB activation and iNOS expression, thereby exacerbating endometrial inflammation and tissue damage.

Journal: Materials Today Bio

Article Title: Exosomal lncRNA OTUD6B-AS1 as a pathogenic nanocarrier promotes inflammatory macrophage polarization in endometritis via a targetable ceRNA circuit

doi: 10.1016/j.mtbio.2026.103027

Figure Lengend Snippet: A proposed model illustrating the exosome-mediated lncRNA OTUD6B-AS1/miR-128/Notch2 axis in aggravating endometritis. Upon LPS-induced damage, endometrial epithelial cells (EECs) release increased exosomes carrying elevated levels of lncRNA OTUD6B-AS1. These exosomes are taken up by endometrial macrophages. The transferred lncRNA OTUD6B-AS1 acts as a molecular sponge to sequester miR-128, leading to the derepression and upregulation of its target gene, Notch2. The enhanced Notch2 signaling subsequently promotes macrophage polarization towards a pro-inflammatory M1 phenotype, characterized by increased NF-κB activation and iNOS expression, thereby exacerbating endometrial inflammation and tissue damage.

Techniques: Activation Assay, Expressing

$PIEZO1-dependent epithelial signaling regulates MMT via PKA/NF-κB P65 pathway. A Western blotting was performed to detect the phosphorylation level of AKT, ERK1/2, CREB, PKA and NF-κB P65 in THP-1 macrophages with CM for 24h (n = 5). P values were calculated using Student’s t-test. B Immunofluorescence showing the expression of CD68 + p-PKA + cells or CD68 + p-NF-κB P65 + cells in colonic mucosa between healthy controls and CD patients (n = 4). P values were calculated using Student’s t-test. C Endogenous interaction between PKA and NF-κB P65 in THP-1 macrophages. Co-immunoprecipitation (Co-IP) analysis demonstrating the physical association between PKA and P65. THP-1 macrophages were stimulated with CM from Caco-2 transfected with either shCTL or shPIEZO1. Lysates were immunoprecipitated (IP) with anti-PKA antibodies or an IgG isotype control, followed by immunoblotting (IB) with anti-PKA and anti-P65 antibodies. The presence of P65 in the PKA-IP fraction confirms the formation of the PKA/P65 complex. GAPDH was used as the loading control for Input lysates. D THP-1 macrophages were treated with CM derived from Caco-2-shCTL cells at 50% or 100% confluence (denoted as 50 and 100). For the 100 condition, cells were further transfected with siNC or siP65 as indicated. The protein expression of α-SMA, COL1A2 and NF-κB P65 were analyzed by Western blotting (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. E THP-1 macrophages were treated with CM derived from Caco-2-shCTL cells at 50% or 100% confluence (denoted as 50 and 100). For the 100 condition, cells were further treated with the PKA inhibitor H-89 or NF-κB activator 1(activator-1), either alone or in combination, as indicated. Western blotting detected the expression of α-SMA, COL1A2 and the phosphorylation level of PKA and NF-κB P65 (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. Data are represented as mean ± SD, ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05. Scale bar =50 μm. CM conditions are defined as in Fig.$

Journal: Cell Communication and Signaling : CCS

Article Title: Intestinal epithelial PIEZO1 regulates macrophage-to-myofibroblast transition via epithelial-derived TGF-α signaling in Crohn’s disease

doi: 10.1186/s12964-026-02941-w

Figure Lengend Snippet: PIEZO1-dependent epithelial signaling regulates MMT via PKA/NF-κB P65 pathway. A Western blotting was performed to detect the phosphorylation level of AKT, ERK1/2, CREB, PKA and NF-κB P65 in THP-1 macrophages with CM for 24h (n = 5). P values were calculated using Student’s t-test. B Immunofluorescence showing the expression of CD68 + p-PKA + cells or CD68 + p-NF-κB P65 + cells in colonic mucosa between healthy controls and CD patients (n = 4). P values were calculated using Student’s t-test. C Endogenous interaction between PKA and NF-κB P65 in THP-1 macrophages. Co-immunoprecipitation (Co-IP) analysis demonstrating the physical association between PKA and P65. THP-1 macrophages were stimulated with CM from Caco-2 transfected with either shCTL or shPIEZO1. Lysates were immunoprecipitated (IP) with anti-PKA antibodies or an IgG isotype control, followed by immunoblotting (IB) with anti-PKA and anti-P65 antibodies. The presence of P65 in the PKA-IP fraction confirms the formation of the PKA/P65 complex. GAPDH was used as the loading control for Input lysates. D THP-1 macrophages were treated with CM derived from Caco-2-shCTL cells at 50% or 100% confluence (denoted as 50 and 100). For the 100 condition, cells were further transfected with siNC or siP65 as indicated. The protein expression of α-SMA, COL1A2 and NF-κB P65 were analyzed by Western blotting (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. E THP-1 macrophages were treated with CM derived from Caco-2-shCTL cells at 50% or 100% confluence (denoted as 50 and 100). For the 100 condition, cells were further treated with the PKA inhibitor H-89 or NF-κB activator 1(activator-1), either alone or in combination, as indicated. Western blotting detected the expression of α-SMA, COL1A2 and the phosphorylation level of PKA and NF-κB P65 (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. Data are represented as mean ± SD, ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05. Scale bar =50 μm. CM conditions are defined as in Fig.

Article Snippet: The antibodies used for Immunofluorescent staining listed as followed: rabbit anti-α-SMA (Proteintech, China; Cat. #14395-1-AP; 1:100), rabbit anti-COL1A2 (Solarbio, China; Cat. #K004211P; 1:50), rabbit anti-phospho-PKA (ImmunoWay, China; Cat. #YP0226; 1:50), rabbit anti-PKA (ImmunoWay, China; Cat. #YT3749; 1:50), rabbit anti-phospho-NF-κB P65 (ABclonal, China; Cat. #AP0124; 1:50), rabbit anti-NF-κB P65 (HuaBio, China; Cat. #ET1603-12; 1:50), mouse anti-CD68 (Immunoway, USA; Cat. #YM3050; 1:50), rabbit anti-TGF-α (BD-PT4626, Biodragon, China; Cat. #BD-PT4626; 1:50), Goat anti-rabbit IgG H&L (Alexa Fluor ® 647) (Abcam, UK; Cat. #ab150079; 1:100) and goat anti-mouse IgG H&L (Alexa Fluor ® 488) (Abcam, UK; Cat. #ab150113; 1:100).

Techniques: Western Blot, Phospho-proteomics, Immunofluorescence, Expressing, Immunoprecipitation, Co-Immunoprecipitation Assay, Transfection, Control, Derivative Assay

Intestinal epithelial PIEZO1 regulates MMT via TGF-α signaling. A The production of TGF-β1 in the CM derived from Caco-2-shCTL or Caco-2-shPIEZO1 for 24h was assessed by ELISA (n = 15). P values were calculated using Student’s t-test. B Volcano plot of differentially expressed genes. C The production of TGF-α in the CM was assessed by ELISA (n = 21). P values were calculated using Student’s t-test. D Pharmacological modulation of PIEZO1 regulates TGF-α secretion. Caco-2-shCTL cells were cultured to 50% or 100% confluence and treated with the PIEZO1 agonist Yoda1 or inhibitor GsMTx4, as indicated. TGF-α levels in supernatant were measured by ELISA (n = 10). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. E Rescue experiment demonstrating that TGF-α mediates PIEZO1-dependent macrophage activation. THP-1 macrophages were treated with CM derived from Caco-2-shCTL or Caco-2-shPIEZO1 cells at 50% or 100% confluence, as indicated. Recombinant TGF-α was added to KD-100-treated macrophages to restore signaling activity. Western blotting was performed to detect the COL1A2, α-SMA, p-NF-κB P65 and p-PKA (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. F THP-1 macrophages were treated with CM derived from Caco-2-shCTL cells at 50% or 100% confluence (denoted as 50 and 100). For neutralization experiments, CM collected at 100% confluence was pre-incubated with an anti–TGF-α neutralizing antibody or isotype IgG control prior to macrophage stimulation. Protein expression of ECM-associated markers and downstream signaling molecules was analyzed by Western blot (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. G Western blotting was performed to detect the COL1A2, α-SMA, p-NF-κB P65 and p-PKA in RAW264.7 with TGF-α (n = 5). P values were calculated using one-way ANOVA and Dunnett’s multiple comparison test. H Western blotting was performed to detect COL1A2, α-SMA, p-NF-κB P65 and p-PKA in BMDMs with TGF-α (10 ng/mL) (n = 4). This concentration was selected based on dose–response experiments. P values were calculated using Student’s t-test. I Immunofluorescence showing the expression of TGF-α in colonic mucosa between healthy controls and CD patients. (n = 4). P values were calculated using Student’s t-test. Data are represented as mean ± SD, ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05. Scale bar = 50μm. CM conditions are defined as in Fig.

Journal: Cell Communication and Signaling : CCS

Article Title: Intestinal epithelial PIEZO1 regulates macrophage-to-myofibroblast transition via epithelial-derived TGF-α signaling in Crohn’s disease

doi: 10.1186/s12964-026-02941-w

Figure Lengend Snippet: Intestinal epithelial PIEZO1 regulates MMT via TGF-α signaling. A The production of TGF-β1 in the CM derived from Caco-2-shCTL or Caco-2-shPIEZO1 for 24h was assessed by ELISA (n = 15). P values were calculated using Student’s t-test. B Volcano plot of differentially expressed genes. C The production of TGF-α in the CM was assessed by ELISA (n = 21). P values were calculated using Student’s t-test. D Pharmacological modulation of PIEZO1 regulates TGF-α secretion. Caco-2-shCTL cells were cultured to 50% or 100% confluence and treated with the PIEZO1 agonist Yoda1 or inhibitor GsMTx4, as indicated. TGF-α levels in supernatant were measured by ELISA (n = 10). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. E Rescue experiment demonstrating that TGF-α mediates PIEZO1-dependent macrophage activation. THP-1 macrophages were treated with CM derived from Caco-2-shCTL or Caco-2-shPIEZO1 cells at 50% or 100% confluence, as indicated. Recombinant TGF-α was added to KD-100-treated macrophages to restore signaling activity. Western blotting was performed to detect the COL1A2, α-SMA, p-NF-κB P65 and p-PKA (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. F THP-1 macrophages were treated with CM derived from Caco-2-shCTL cells at 50% or 100% confluence (denoted as 50 and 100). For neutralization experiments, CM collected at 100% confluence was pre-incubated with an anti–TGF-α neutralizing antibody or isotype IgG control prior to macrophage stimulation. Protein expression of ECM-associated markers and downstream signaling molecules was analyzed by Western blot (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. G Western blotting was performed to detect the COL1A2, α-SMA, p-NF-κB P65 and p-PKA in RAW264.7 with TGF-α (n = 5). P values were calculated using one-way ANOVA and Dunnett’s multiple comparison test. H Western blotting was performed to detect COL1A2, α-SMA, p-NF-κB P65 and p-PKA in BMDMs with TGF-α (10 ng/mL) (n = 4). This concentration was selected based on dose–response experiments. P values were calculated using Student’s t-test. I Immunofluorescence showing the expression of TGF-α in colonic mucosa between healthy controls and CD patients. (n = 4). P values were calculated using Student’s t-test. Data are represented as mean ± SD, ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05. Scale bar = 50μm. CM conditions are defined as in Fig.

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay, Cell Culture, Activation Assay, Recombinant, Activity Assay, Western Blot, Neutralization, Incubation, Control, Expressing, Comparison, Concentration Assay, Immunofluorescence

Intestinal epithelial PIEZO1 deficiency attenuates colonic fibrosis in a chronic DSS-induced colitis–associated fibrosis model. A Schematic diagram of the chronic DSS-induced colitis-associated fibrosis model. B Body weight changes during DSS treatment (n = 6 mice per group). Statistical significance was analyzed using two-way ANOVA followed by Tukey’s multiple comparisons test. C Disease activity index (DAI) scores in control and CKO mice during the experimental period (n = 6 mice per group). Statistical significance was analyzed using two-way ANOVA followed by Tukey’s multiple comparisons test. Data are presented as mean ± SD. D Representative colon length changes in the indicated group (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. E Serum TGF-α levels measured by ELISA (n = 4). P values were calculated using Student’s t-test. F Immunofluorescence showed the expression of TGF-α in colonic tissues of colonic fibrosis in a mouse model (n = 6). P values were calculated using Student’s t-test. G Representative images of Masson’s trichrome-stained sections in the indicated group and the quantified area of ECM deposition per high-powered field (n = 4). P values were calculated using Student’s t-test. H Immunofluorescence showing the expression of CD68 + COL1A2 + cells and CD68 + α-SMA + cells in colonic tissues in the indicated group (n = 4). P values were calculated using Student’s t-test. I Immunofluorescence showing the expression of CD68 + p-PKA + cells and CD68 + p-NF-κB P65 + cells in colonic tissues in the indicated group (n = 4). P values were calculated using Student’s t-test. DAPI (blue), CD68 (green), and target proteins (red). Data are represented as mean ± SD, ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05. Scale bar = 50 μm

Journal: Cell Communication and Signaling : CCS

Article Title: Intestinal epithelial PIEZO1 regulates macrophage-to-myofibroblast transition via epithelial-derived TGF-α signaling in Crohn’s disease

doi: 10.1186/s12964-026-02941-w

Figure Lengend Snippet: Intestinal epithelial PIEZO1 deficiency attenuates colonic fibrosis in a chronic DSS-induced colitis–associated fibrosis model. A Schematic diagram of the chronic DSS-induced colitis-associated fibrosis model. B Body weight changes during DSS treatment (n = 6 mice per group). Statistical significance was analyzed using two-way ANOVA followed by Tukey’s multiple comparisons test. C Disease activity index (DAI) scores in control and CKO mice during the experimental period (n = 6 mice per group). Statistical significance was analyzed using two-way ANOVA followed by Tukey’s multiple comparisons test. Data are presented as mean ± SD. D Representative colon length changes in the indicated group (n = 5). P values were calculated using one-way ANOVA and Tukey’s multiple comparisons test. E Serum TGF-α levels measured by ELISA (n = 4). P values were calculated using Student’s t-test. F Immunofluorescence showed the expression of TGF-α in colonic tissues of colonic fibrosis in a mouse model (n = 6). P values were calculated using Student’s t-test. G Representative images of Masson’s trichrome-stained sections in the indicated group and the quantified area of ECM deposition per high-powered field (n = 4). P values were calculated using Student’s t-test. H Immunofluorescence showing the expression of CD68 + COL1A2 + cells and CD68 + α-SMA + cells in colonic tissues in the indicated group (n = 4). P values were calculated using Student’s t-test. I Immunofluorescence showing the expression of CD68 + p-PKA + cells and CD68 + p-NF-κB P65 + cells in colonic tissues in the indicated group (n = 4). P values were calculated using Student’s t-test. DAPI (blue), CD68 (green), and target proteins (red). Data are represented as mean ± SD, ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05. Scale bar = 50 μm

Techniques: Activity Assay, Control, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Expressing, Staining

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