Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: mRNA-seq of rat tissues. To explore the related molecular mechanism of bronchiolitis obliterans, high-throughput analysis was conducted using the lung tissues of rats. (A) DEGs in DA group rats relative to the control are shown using a volcano plot. DEGs were defined as an absolute value of log2FC >1.5 and adjusted-P<0.001. (B) Quantitative PCR was carried out to verify the results of mRNA-seq. Four DEGs were randomly selected for verification. (C) Gene Ontology analysis revealed that apoptosis, inflammation, fibrosis, EMT and epithelium-associated items were enriched by DEGs. (D) Venn diagram shows that 47 DEGs commonly existed in the six datasets. (E) Heatmap of 47 DEGs demonstrated that most genes were functional proteins. (F) CIP2A was located on chromosome 11 and was increased in DA-treated rats. n=6. CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; DA, diacetyl; DEGs, differentially expressed genes; EMT, epithelial-mesenchymal transition; FC, fold change; mRNA-seq, mRNA-sequencing; SLC1A6, solute carrier family 1 member 6; ERN2, endoplasmic reticulum to nucleus signaling 2; RNASE2, ribonuclease A family member 2; TPSB2, tryptase β2.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: High Throughput Screening Assay, Control, Real-time Polymerase Chain Reaction, Functional Assay, Sequencing

Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: CIP2A expression is increased in DA group rats. (A) Quantitative PCR and western blotting, and (B) immunohistochemistry (scale bar: 50 µm) were used to detect the increased expression of CIP2A in samples from the DA group, which was initially determined by mRNA-sequencing. n=6. CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; DA, diacetyl.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Immunohistochemistry, Sequencing

Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: Inhibiting CIP2A attenuates the injuries induced by DA in the bronchi and lungs of rats. To evaluate the effect of CIP2A inhibition on bronchiolitis obliterans, Eth, a CIP2A inhibitor, was injected subcutaneously into DA group rats. (A) Animal experimental timeline. Representative (B) H&E and (C) Masson's trichrome-stained lung slices (scale bar: 200 µm). Quantitative analysis of severity is shown. (D) Bronchoalveolar lavage fluid total cell count, and macrophage, neutrophil, lymphocyte and eosinophil counts in the control rats, DA-exposed rats and Eth-treated DA-exposed rats. (E) Western blotting and (F) immunohistochemistry were used to measure the protein expression levels of CIP2A in lung tissues. n=6. CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; DA, diacetyl; Eth, ethoxysanguinarine; H&E, hematoxylin and eosin.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: Inhibition, Injection, Staining, Cell Counting, Control, Western Blot, Immunohistochemistry, Expressing

Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: Inhibiting cell proliferation regulating inhibitor of protein phosphatase 2A weakens the epithelial-mesenchymal transition and inflammation induced by DA in the bronchi and lungs of rats. (A) E-cadherin and (B) Vimentin levels were detected by immunofluorescence (scale bar: 50 µm). (C) Western blotting was used to measure the protein expression levels of α-SMA, fibronectin, Snail and iNOS. (D) Enzyme-linked immunosorbent assay kits were used to analyze the levels of IL-1β, IL-6 and TNF-α. n=6. α-SMA, α-smooth muscle actin; CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; DA, diacetyl; IL, interleukin; iNOS, inducible NO synthase; TNF-α, tumor necrosis factor-α.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: Immunofluorescence, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: Inhibiting CIP2A reduces damage caused by DA in human primary bronchial epithelial cells. To explore the cellular mechanisms, an in vitro cellular model of bronchiolitis obliterans was employed. (A) Eth (6 µM) showed the best inhibitory effect on CIP2A expression. (B) Western blotting was used to measure the protein expression levels of α-SMA, fibronectin, Snail and iNOS. (C) Enzyme-linked immunosorbent assay kits were used to analyze the levels of IL-1β, IL-6 and TNF-α. (D) E-cadherin and (E) Vimentin levels were detected by immunofluorescence (scale bar: 50 µm). n=3. α-SMA, α-smooth muscle actin; CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; DA, diacetyl; Eth, ethoxysanguinarine; IL, interleukin; iNOS, inducible NO synthase; TNF-α, tumor necrosis factor-α.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: In Vitro, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Immunofluorescence

Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: Inhibiting CIP2A blocks the activation of NF-κB signaling in HPBECs. The expression of CIP2A, and phosphorylation of IKBα and expression of p65 were assessed by western blotting. n=3. CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; DA, diacetyl; HPBECs, human primary bronchial epithelial cells; IκBα, inhibitor of NF-κB α; NF-κB, nuclear factor-κB; p-, phosphorylated.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: Activation Assay, Expressing, Western Blot

Journal: Molecular Medicine Reports

Article Title: CIP2A promotes bronchiolitis obliterans by activating the NF‑κB pathway

doi: 10.3892/mmr.2025.13473

Figure Lengend Snippet: CIP2A promotes BO by activating the NF-κB pathway. BO is caused by DA and mediated by CIP2A. CIP2A deficiency inhibits fibrosis, inflammation and EMT in BO models through suppressing the NF-κB pathway. Solid and dashed arrows both indicate promotion. The solid arrow reveals the molecular mechanism through which the dashed arrow operates. BO, bronchiolitis obliterans; DA, diacetyl; CIP2A, cell proliferation regulating inhibitor of protein phosphatase 2A; EMT, epithelial-mesenchymal transition; NF-κB, nuclear factor-κB.

Article Snippet: After blocking with 1% BSA (Sangon Biotech Co., Ltd.) for 15 min at 4°C, the slices were incubated with CIP2A antibody (1:100; cat. no. bs-5948R; BIOSS) overnight at 4°C and with Goat Anti-Rabbit IgG/HRP (1:100; cat. no. SE134; Beijing Solarbio Science & Technology Co., Ltd.) for 45 min. Diaminobenzidine was used as a chromogenic substrate and the nuclei were counterstained using hematoxylin at room temperature for 3 min.

Techniques: