Budesonide with surfactant decreases systemic responses in mechanically ventilated preterm lambs exposed to fetal intra-amniotic lipopolysaccharide

Ren

et al. 2022

Preprint

Objective To investigate the anti-inflammatory role of H2 in LPS-induced BPD via regulating TNF-α/NF-κB signaling pathway in placenta. Methods We induced a neonatal rat model of BPD by injecting lipopolysaccharide (LPS, 1ug) into the amniotic fluid at embryonic day 16.5(E16.5). Treatment of 30% hydrogen gas for 4 hours/day with continuously 5days. We primarily analyzed the neonatal outcomes and then compared inflammatory levels from Control group (CON), LPS group (LPS) and LPS with H2 inhalation group (LPS + H2). TUNEL and Hematoxylin-Eosin (HE) staining were performed to evaluate inflammatory and apoptotic levels. We further used RNA sequencing and ELISA assay to examine differentially expressed proteins and mRNA levels of tumor necrosis factor-α (TNF-α), nuclear factor kappa-B (NF-κB) (p65), interleukin (IL)-6, IL-18, IL-1β, C-C motif chemokine ligand 2(CCL2) and C-X-C motif chemokine ligand 1(CXCL1). Bioinformatics analysis (GO and KEEG) of RNA-seq and correlation analysis were applied to clarify the mechanisms of H2 anti-inflammatory effect on LPS-induced BPD. Results We found the H2 inhalation decreased production of inflammatory cytokines/chemokines (IL-6, IL-18, IL-1β, CCL2, CXCL1) in LPS-induced placenta to rescue from the BPD. Upon administration of H2, infiltration degree of LPS-induced placenta was reduced and infiltrating significantly narrowed down. Hydrogen normalized LPS-induced perturbed lung development, reduced lung apoptotic index, death ratio of fetus and neonate. Meanwhile, H2 also upregulated the survival ratio. RNA-seq and Elisa demonstrated that both mRNA and protein levels of TNF-α/NF-κB signaling pathway were activated by LPS, and H2 relieved the pro-inflammatory function of LPS on TNF-α/NF-κB-stimulated placenta. Correlation analysis showed a positive association of TNF-α vs both NF-κB and inflammatory cytokines/chemokines. Conclusion H2 inhalation alleviated LPS-induced BPD by inhibiting excessive pro-inflammatory cytokines and inflammatory chemokines via the TNF-α/NF-κB signaling pathway in placenta and may be a potential therapeutic strategy for BPD.

Section: Discussionmentioning

confidence: 71%

Hydrogen gas ameliorates the LPS-induced BPD via inhibiting the activation of TNF-α/NF-κB inflammatory signaling pathway in placenta

Ren

et al. 2022

Preprint

“…However, the model still has drawbacks such as complex experimental operations, significant trauma, and susceptibility to infection. CAM can produce excessive inflammatory factors in the fetus, which affect the maturation of the fetal lung, cause fetal lung structural remodeling, and affect the contractile function of pulmonary vessels, thus leading to the occurrence of BPD [ 19 , 20 ]. Our results indicated that the LPS group exhibited a disorder of intrauterine development: intrauterine fetal death (IUFD) rates were high in the LPS group.…”

Section: Discussionmentioning

confidence: 99%

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

Zhang,

Ren,

Zhang

et al. 2024

Eur J Med Res

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-1β, 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.

“…In vitro and in vivo studies have demonstrated the compatibility of Poractant alfa with the standard intratracheal budesonide dose of 0.25 mg/kg, showing a good stability of budesonide within the phospholipid suspension for 24 h, no detrimental effects in the surface tension reducing properties of Poractant alfa, and positive long-term effects on lung performance [48][49][50]. Moreover, the intratracheal delivery of Poractant alfa and budesonide reduces the concentration of proinflammatory cytokines in the bronchoalveolar lavage fluid and the expression of inflammatory markers in the lung, liver, and brain of preterm lambs exposed to various prenatal and postnatal insults, implying both a local and systemic effect of intratracheally delivered budesonide [10,[51][52][53]. To complete the preclinical profiling of the combination of Poractant alfa and budesonide, we applied MSI to visualize the pulmonary distribution of budesonide delivered with or without Poractant alfa.…”

Section: Discussionmentioning

confidence: 99%

Surfactant-Assisted Distal Pulmonary Distribution of Budesonide Revealed by Mass Spectrometry Imaging

et al. 2021

Self Cite

Direct lung administration of budesonide in combination with surfactant reduces the incidence of bronchopulmonary dysplasia. Although the therapy is currently undergoing clinical development, the lung distribution of budesonide throughout the premature neonatal lung has not yet been investigated. Here, we applied mass spectrometry imaging (MSI) to investigate the surfactant-assisted distal lung distribution of budesonide. Unlabeled budesonide was either delivered using saline as a vehicle (n = 5) or in combination with a standard dose of the porcine surfactant Poractant alfa (n = 5). These lambs were ventilated for one minute, and then the lungs were extracted for MSI analysis. Another group of lambs (n = 5) received the combination of budesonide and Poractant alfa, followed by two hours of mechanical ventilation. MSI enabled the label-free detection and visualization of both budesonide and the essential constituent of Poractant alfa, the porcine surfactant protein C (SP-C). 2D ion intensity images revealed a non-uniform distribution of budesonide with saline, which appeared clustered in clumps. In contrast, the combination therapy showed a more homogeneous distribution of budesonide throughout the sample, with more budesonide distributed towards the lung periphery. We found similar distribution patterns for the SP-C and budesonide in consecutive lung tissue sections, indicating that budesonide was transported across the lungs associated with the exogenous surfactant. After two hours of mechanical ventilation, the budesonide intensity signal in the 2D ion intensity maps dropped dramatically, suggesting a rapid lung clearance and highlighting the relevance of achieving a uniform surfactant-assisted lung distribution of budesonide early after delivery to maximize the anti-inflammatory and maturational effects throughout the lung.