Mechanical Ventilation-Related High Stretch Mainly Induces Endoplasmic Reticulum Stress and Thus Mediates Inflammation Response in Cultured Human Primary Airway Smooth Muscle Cells

Abstract

Ventilator-induced lung injury (VILI) occurs in mechanically ventilated patients of respiratory disease and is typically characterized by airway inflammation. However, recent studies increasingly indicate that a major cause of VILI may be the excessive mechanical loading such as high stretch (>10% strain) on airway smooth muscle cells (ASMCs) due to mechanical ventilation (MV). Although ASMCs are the primary mechanosensitive cells in airways and contribute to various airway inflammation diseases, it is still unclear how they respond to high stretch and what mediates such a response. Therefore, we used whole genome-wide mRNA-sequencing (mRNA-Seq), bioinformatics, and functional identification to systematically analyze the mRNA expression profiles and signaling pathway enrichment of cultured human ASMCs exposed to high stretch (13% strain), aiming to screen the susceptible signaling pathway through which cells respond to high stretch. The data revealed that in response to high stretch, 111 mRNAs with count ≥100 in ASMCs were significantly differentially expressed (defined as DE-mRNAs). These DE-mRNAs are mainly enriched in endoplasmic reticulum (ER) stress-related signaling pathways. ER stress inhibitor (TUDCA) abolished high-stretch-enhanced mRNA expression of genes associated with ER stress, downstream inflammation signaling, and major inflammatory cytokines. These results demonstrate in a data-driven approach that in ASMCs, high stretch mainly induced ER stress and activated ER stress-related signaling and downstream inflammation response. Therefore, it suggests that ER stress and related signaling pathways in ASMCs may be potential targets for timely diagnosis and intervention of MV-related pulmonary airway diseases such as VILI.

Association between volume of lung damage and endoplasmic reticulum stress expression among severe COVID-19 ICU patients

“…The association found between GRP78 plasma level and lung damage volume in our cohort combined with a non-correlation between ERS and systemic in ammation (IL-6) strengthen their hypothesis and describe for the rst time this potential link among infected humans. This hypothesis is also comforted by recent studies in mouse models showing that mechanical volume induced lung injury was the predominant trigger of ERS, preceding and contributing to lung in ammation (35,36). As it is known that ERS expression after tissue injury last longer than systemic in ammation in humans, and given its association with damaged lung volume, it would be interesting to study the link between ERS expression and long-term prognosis among COVID-19 patients ( brosis, bronchiectasis, chronic dyspnea,…) (33).…”

High Stretch Associated with Mechanical Ventilation Promotes Piezo1-Mediated Migration of Airway Smooth Muscle Cells

“…Cell line mouse ASMCs (mASMCs, #BNCC359490, BeNa Culture Collection, Beijing, China) were purchased from BeNa Culture Collection (Beijing, China) and cultured in Dulbecco's modified Eagle's medium (DMEM, #11885092, Thermo Fisher) supplemented with 10% fetal bovine serum (FBS, #16000-044, Thermo Fisher), 2 mM L-glutamine, 100 units/mL penicillin, and 100 µg/mL streptomycin in an incubator containing 5% CO 2 humidified at 37 • C according to the method described previously [20,21,35]. mASMCs were identified with immunostaining antibodies against α-SMA (#BM0002).…”

“…For the high-stretch experiment, 2 × 10 4 cells/cm 2 mASMCs in exponential proliferation were plated on type I collagen-coated Bioflex 6-well plates and were serum-deprived for 24 h. Subsequently, we exposed the cells grown on Bioflex plates to a 13% cyclic strain at 0.5 Hz for 72 h (a sinusoidal wave, 1 s of deformation alternating with 1 s of relaxation) using the Flexcell FX-5000 system (Flexcell International, Hillsborough, NC, USA) to simulate MV-associated high stretch on ASMCs as previously described [4,20,21,48]. The mASMCs grown on Bioflex plates in static condition for 72 h were used as controls.…”

“…It is well known that cell migration involves integrins because the interaction between integrins and ligands in the extracellular matrix determines the adhesion and, thus, migration speed of the cell [19]. Moreover, cell migration has been characterized by two distinct modes, namely, slow integrin-dependent or fast integrin-independent cell migration, and the transition between these two modes was predominantly determined by the level of cell adhesion and expression of various members of the integrin family, especially integrin αV and β1 in ASMCs [20,21]. For example, reduced expression of integrin α2β1 and weakened cell adhesion are associated with cell migration transition towards the integrin-independent mode in human fibrosarcoma and non-small lung carcinoma cells [22].…”

High Stretch Modulates cAMP/ATP Level in Association with Purine Metabolism via miRNA–mRNA Interactions in Cultured Human Airway Smooth Muscle Cells

“…Primary human ASMCs (#BNCC339826) were purchased from the BeNa Culture Collection (Beijing, China) and used in passages 3-10 which maintained normal "hill and valley" morphology and cellular functions, including physiological response to contractile/relaxant agonist and α-smooth muscle actin (SMA) expression (#ab119952, Abcam, Waltham, MA, USA) [26,[35][36][37]. These cells were cultured in DMEM supplemented with 10% FBS, 2 mM L-glutamine, 100 units/mL penicillin, and 100 µg/mL streptomycin in an incubator containing 5% CO 2 humidified at 37 • C according to the method described previously.…”