Hydrogen gas inhalation ameliorates LPS-induced BPD by inhibiting inflammation via regulating the TLR4-NFκB-IL6/NLRP3 signaling pathway in the placenta.

INTRODUCTION: Hydrogen (H2) is regarded as a novel therapeutic agent against several diseases owing to its inherent biosafety. Bronchopulmonary dysplasia (BPD) has been widely considered among adverse pregnancy outcomes, without effective treatment. Placenta plays a role in defense, synthesis, and immunity, which provides a new perspective for the treatment of BPD. This study aimed to investigate if H2 reduced the placental inflammation to protect the neonatal rat against BPD damage and potential mechanisms. METHODS: We induced neonatal BPD model by injecting lipopolysaccharide (LPS, 1 µg) into the amniotic fluid at embryonic day 16.5 as LPS group. LPS + H2 group inhaled 42% H2 gas (4 h/day) until the samples were collected. We primarily analyzed the neonatal outcomes and then compared inflammatory levels from the control group (CON), LPS group and LPS + H2 group. HE staining was performed to evaluate inflammatory levels. RNA sequencing revealed dominant differentially expressed genes. Bioinformatics analysis (GO and KEGG) of RNA-seq was applied to mine the signaling pathways involved in protective effect of H2 on the development of LPS-induced BPD. We further used qRT-PCR, Western blot and ELISA methods to verify differential expression of mRNA and proteins. Moreover, we verified the correlation between the upstream signaling pathways and the downstream targets in LPS-induced BPD model. RESULTS: Upon administration of H2, the inflammatory infiltration degree of the LPS-induced placenta was reduced, and infiltration significantly narrowed. Hydrogen normalized LPS-induced perturbed lung development and reduced the death ratio of the fetus and neonate. RNA-seq results revealed the importance of inflammatory response biological processes and Toll-like receptor signaling pathway in protective effect of hydrogen on BPD. The over-activated upstream signals [Toll-like receptor 4 (TLR4), nuclear factor kappa-B p65 (NF-κB p65), Caspase1 (Casp1) and NLR family pyrin domain containing 3 (NLRP3) inflammasome] in LPS placenta were attenuated by H2 inhalation. The downstream targets, inflammatory cytokines/chemokines [interleukin (IL)-6, IL-18, IL-1ß, C-C motif chemokine ligand 2 (CCL2) and C-X-C motif chemokine ligand 1 (CXCL1)], were decreased both in mRNA and protein levels by H2 inhalation in LPS-induced placentas to rescue them from BPD. Correlation analysis displayed a positive association of TLR4-mediated signaling pathway both proinflammatory cytokines and chemokines in placenta. CONCLUSION: H2 inhalation ameliorates LPS-induced BPD by inhibiting excessive inflammatory cytokines and chemokines via the TLR4-NFκB-IL6/NLRP3 signaling pathway in placenta and may be a potential therapeutic strategy for BPD.

Tubeimoside­1 induces apoptosis in human glioma U251 cells by suppressing PI3K/Akt­mediated signaling pathways.

Tubeimoside-1 (TBMS1), a traditional Chinese herb extracted from Bolbostemma paniculatum (Maxim.), induces apoptosis in a number of human cancer cell lines. TBMS1 has been reported to induce apoptosis in human glioma cells, however the mechanism remains to be elucidated. The present study explored TBMS1­induced PI3K/Akt­related pathways in human glioma cells. The human glioma U251 and the human astrocyte (HA) cell lines were treated with various concentrations of TBMS1. MTT assays were conducted to analyze cell viability. Cell cycle distribution and the rate of apoptosis were assessed using flow cytometry. BrdU incorporation and Hoechst 33342 staining were performed to analyze the cell cycle and apoptosis, respectively. Western blotting was performed to investigate protein expression levels. The results demonstrated that TBMS1 reduced cell viability in human glioma cells U251 by suppressing Akt phosphorylation. Subsequently, TBMS1 inhibited DNA synthesis and induced G2/M phase arrest by targeting the PI3K/Akt/p21 and the cyclin­dependent kinase 1/cyclin B1 signaling cascades. In addition, TBMS1 triggered apoptosis via the PI3K/Akt­mediated Bcl­2 signaling pathway. These results demonstrated that TBMS1 prevented the progression of gliomas via the PI3K/Akt­dependent pathway, which provided a theoretical basis for in vivo studies to use TBMS1 as potential therapy for the prevention of cancer.