SENP1 knockdown-mediated CTCF SUMOylation enhanced its stability and alleviated lipopolysaccharide-evoked inflammatory injury in human lung fibroblasts via regulation of FOXA2 transcription.

BACKGROUND: Excessive inflammation is the main cause of treatment failure in neonatal pneumonia (NP). CCCTC-binding factor (CTCF) represents an important node in various inflammatory diseases. In the present study, we tried to clarify the function and underlying molecular mechanism of CTCF on an in vitro cellular model of NP, which was generated by simulating the human lung fibroblast cell line WI-38 with lipopolysaccharide (LPS). METHODS: The SUMOylation level and protein interaction were verified by Co-immunoprecipitation assay. Cell viability was measured by Cell Counting Kit-8 assay. Inflammatory factors were examined by Enzyme-linked immunosorbent assay. Cell apoptosis was evaluated by TUNEL assay. The binding activity of CTCF to target promoter was tested by chromatin immunoprecipitation and luciferase reporter assay. RESULTS: LPS treatment restrained cell viability, promoted the production of inflammatory factors, and enhanced cell apoptosis. CTCF overexpression played anti-inflammatory and anti-apoptotic roles. Furthermore, CTCF was modified by SUMOylation with small ubiquitin-like modifier protein 1 (SUMO1). Interfering with sumo-specific protease 1 (SENP1) facilitated CTCF SUMOylation and protein stability, thus suppressing LPS-evoked inflammatory and apoptotic injuries. Moreover, CTCF could bind to the forkhead box protein A2 (FOXA2) promoter region to promote FOXA2 expression. The anti-inflammatory and anti-apoptotic roles of CTCF are associated with FOXA2 activation. In addition, SENP1 knockdown increased FOXA2 expression by enhancing the abundance and binding ability of CTCF. CONCLUSIONS: SUMOylation of CTCF by SENP1 knockdown enhanced its protein stability and binding ability and it further alleviated LPS-evoked inflammatory injury in human lung fibroblasts by positively regulating FOXA2 transcription.

NRF1 knockdown alleviates lipopolysaccharide-induced pulmonary inflammatory injury by upregulating DKK3 and inhibiting the GSK-3ß/ß-catenin pathway.

Excessive inflammatory injury is the main cause of the incidence of severe neonatal pneumonia (NP) and associated deaths. Although dickkopf-3 (DKK3) exhibits anti-inflammatory activity in numerous pathological processes, its role in NP is still unknown. In this study, human embryonic lung WI-38 and MRC-5 cells were treated with lipopolysaccharide (LPS) to induce inflammatory injury of NP in vitro. The expression of DKK3 was downregulated in LPS-stimulated WI-38 and MRC-5 cells. DKK3 overexpression decreased LPS-induced inhibition of cell viability, and reduced LPS-induced apoptosis of WI-38 and MRC-5 cells. DKK3 overexpression also reduced LPS-induced production of pro-inflammatory factors such as ROS, IL-6, MCP-1, and TNF-α. Nuclear respiratory factors 1 (NRF1) knockdown was found to upregulate DKK3 and inactivate the GSK-3ß/ß-catenin pathway in LPS-injured WI-38 and MRC-5 cells. NRF1 knockdown also suppressed LPS-induced inhibition on cell viability, repressed LPS-induced apoptosis, and inhibited the accumulation of ROS, IL-6, MCP-1, and TNF-α in LPS-injured WI-38 and MRC-5 cells. DKK3 knockdown or re-activation of the GSK-3ß/ß-catenin pathway reversed the inhibitory effects of NRF1 knockdown on LPS-induced inflammatory injury. In conclusion, NRF1 knockdown can alleviate LPS-triggered inflammatory injury by regulating DKK3 and the GSK-3ß/ß-catenin pathway.

A Diastereoselective Multicomponent Reaction for Construction of Alkynylamide-Substituted α,ß-Diamino Acid Derivatives To Hunt Hits.

A highly diasetereoselective Mannich-type multicomponent reaction was developed to rapidly construct alkynylamide-substituted α,ß-diamino acid derivatives from simple starting materials under mild conditions in moderate to good yields for hit hunting. Most of the resulting products 4 exhibited good anticancer activity in HCT116, BEL7402, and SMMC7721 cells.

Peripheral blood MDSCs, IL-10 and IL-12 in children with asthma and their importance in asthma development.

OBJECTIVE: To investigate myeloid-derived suppressor cell (MDSC) accumulation and interleukin 10 (IL-10) and interleukin 12 (IL-12) levels during the onset of asthma in both pediatric patients and mouse models, as well as their possible roles in the development of asthma. METHODS: Peripheral blood samples were gathered from children with asthma attacks (attack group) and alleviated asthma (alleviated group), as well as two control groups, children with pneumonia and healthy children. The pathological characteristics of asthma in asthmatic mice, budesonide-treated asthmatic mice, and normal control mice were also evaluated by immunohistochemistry (IHC) and hematoxylin and eosin (H&E) staining. RESULTS: MDSC accumulation and serum IL-10 levels were significantly elevated in the children with asthma compared with the budesonide-treated alleviated group, normal healthy controls, and pneumonia controls (p<0.05), whereas those in the latter three groups showed no statistical differences (p>0.05). The level of serum IL-12 in the asthmatic children was drastically reduced compared to the budesonide-treated alleviated group, healthy controls, and pneumonia controls (p<0.05), whereas the latter three groups showed no significant differences in their serum IL-12 levels. The percentage of MDSCs in children with asthma was positively correlated with the level of serum IL-10 and negatively correlated with the level of serum IL-12. The levels of MDSCs and IL-10 in asthmatic mice were significantly higher than those in the normal control mice (both p<0.05) and were reduced after budesonide treatment (both p<0.05). IL-12 expression in the asthmatic mice was significantly lower than the control and was increased upon budesonide treatment (both p<0.05). CONCLUSION: During the onset of asthma, the accumulation of MDSCs and the level of serum IL-10 increase, while the level of IL-12 decreases. These fluctuations may play an important role in the development of asthma.

[Changes to CD4(+)CD25(high+)CD127(low) regulatory T cells in peripheral blood from children with bronchiolitis, and its clinical significance].

OBJECTIVE: To study changes to CD4(+)CD25(high+)CD127(low) regulatory T cells (Treg) in peripheral blood from children with bronchiolitis, and to explore its clinical significance. METHODS: Thirty-one children with bronchiolitis and aged under two years were randomly enrolled as the bronchiolitis group, and 25 under two-year-olds with bronchopneumonia were randomly enrolled as the bronchopneumonia group. A further twenty-five children with non-infectious diseases such as hernia and renal calculus served as the control group. The level of CD4(+)CD25(high+)CD127(low) Treg in peripheral blood was measured by multi-color detection and multi-parameter flow cytometry. RESULTS: The proportion of CD4(+)CD25(high+)CD127(low) Treg in peripheral blood in the bronchiolitis group (8.0%±2.1%) was significantly lower than in the bronchopneumonia (9.6%±2.6%; P<0.05) and control groups (11.3%±2.9%; P<0.05). CONCLUSIONS: CD4(+)CD25(high+)CD127(low) Treg level in peripheral blood may be an index of immunological function in infants. A decreased level of CD4(+)CD25(high+)CD127(low) Treg in peripheral blood suggests that Treg cells may be involved in the pathogenesis and development of bronchiolitis.