Repair Of Acute Lung Injury Research Articles

Differential roles of regulatory T cells in acute respiratory infections.

Acute respiratory infections trigger an inflammatory immune response with the goal of pathogen clearance; however, overexuberant inflammation causes tissue damage and impairs pulmonary function. CD4+FOXP3+ regulatory T cells (Tregs) interact with cells of both the innate and the adaptive immune system to limit acute pulmonary inflammation and promote its resolution. Tregs also provide tissue protection and coordinate lung tissue repair, facilitating a return to homeostatic pulmonary function. Here, we review Treg-mediated modulation of the host response to respiratory pathogens, focusing on mechanisms underlying how Tregs promote resolution of inflammation and repair of acute lung injury. We also discuss potential strategies to harness and optimize Tregs as a cellular therapy for patients with severe acute respiratory infection and discuss open questions in the field.

Diosmetin alleviates acute lung injury caused by lipopolysaccharide by targeting barrier function.

Acute lung injury (ALI) is an acute and devastating disease caused by systemic inflammation e.g. patients infected with bacteria and viruses such as SARS-CoV-2 have an unacceptably high mortality rate. It has been well documented that endothelial cell damage and repair play a central role in the pathogenesis of ALI because of its barrier function. Nevertheless, the leading compounds that effectively accelerate endothelial cell repair and improve barrier dysfunction in ALI are largely unknown. In the present study, we found that diosmetin had promising characteristics to inhibit the inflammatory response and accelerate the repair of endothelial cells. Our results indicated that diosmetin accelerated wound healing and barrier repair by improving the expression of the barrier-related proteins, including zonula occludens-l (ZO-1) and occludin, in human umbilical vein endothelial cells (HUVECs) treated with lipopolysaccharide (LPS). Meanwhile, diosmetin administration significantly inhibited inflammatory response by decreasing the content of TNFα and IL-6 in the serum, alleviated lung injury by reducing lung wet/dry (W/D) ratio and histologic score, improved endothelial hyperpermeability by decreasing protein levels and neutrophil infiltration in the bronchoalveolar lavage fluid (BALF) and increasing ZO-1 and occludin expression in the lung tissues of LPS-treated mice. Mechanistically, diosmetin also mediated the expression of Rho A and ROCK1/2 in HUVECs treated with LPS, and fasudil, a Rho A inhibitor remarkably inhibited the role of diosmetin in ZO-1 and occludin proteins. All these findings of this study revealed that diosmetin can be an effective protector of lung injury and the Rho A/ROCK1/2 signal pathway plays a pivotal role in diosmetin accelerating barrier repair in ALI.

Mesenchymal stromal cells ameliorate acute lung injury induced by LPS mainly through stanniocalcin-2 mediating macrophage polarization.

BackgroundAcute lung injury (ALI) is a devastating syndrome with no effective pharmacological therapies in the clinic. Mesenchymal stromal cells (MSCs) have been demonstrated to promote inflammation resolution and tissue repair in ALI. However, the specific mechanisms of this have not been clearly elucidated. Stanniocalcin-2 (STC2) is a stress-responsive protein that has anti-oxidative properties. Our previous study found that STC2 is a highly expressed stanniocalcin in MSCs, which may be involved in immunomodulatory activities. However, the role of STC2 in MSCs to resolve ALI has never been elucidated.MethodsSpecific shRNA was used to downregulate STC2 in MSCs. We detected ROS, cell apoptosis, and paracrine factors changes in MSCs. STC2-associated antioxidant genes were also investigated by Co-immunoprecipitation (Co-IP) and immunofluorescence. Macrophage (THP1 cells) phenotype transitions were measured by flow cytometry after coculturing with MSCs in vitro. Then, we used MSCs to treat LPS-induced ALI in mice, and assessed injury scores inflammation, and antioxidant activities in the lungs of the mice. Alveolar macrophage (AM) phenotypes and CFSE-labeled MSC apoptosis in collected bronchoalveolar fluids (BALF) were also analyzed by flow cytometry.ResultsAfter the STC2 knockdown, MSCs increased ROS generation and cell apoptosis after PX12 pretreatment. The antioxidant protein Nrf2 was colocalized with STC2 in the nucleus. A lack of STC2 expression in MSCs produced less interleukin 10 (IL10) and blunted macrophage polarization in THP1 cells. Furthermore, in the murine LPS-induced ALI model, the STC2 knockdown counteracted the inflammatory resolution and antioxidative effect of MSCs in the lungs. MSCshSTC2-treated mice had a higher lung injury score than the controls, which may be attributed to diminished AM polarization and increased apoptosis of MSCs in vivo.ConclusionsCollectively, these results suggested that STC2 is essential to the anti-oxidative and anti-inflammation properties of MSCs and could prove to be crucial for stem cell therapies for ALI.

Deficiency of HIF-1α enhances influenza A virus replication by promoting autophagy in alveolar type II epithelial cells

ABSTRACT Infection of influenza A virus (IAV) can trigger exaggerated pulmonary inflammation and induce acute lung injury (ALI). Limiting IAV replication and alleviation of pulmonary inflammation are two important therapeutic strategies for influenza virus infection. Recent studies have shown that hypoxia inducible factor-1α (HIF-1α) is an essential factor for the development and repair of ALI; however, the role and the underlying mechanisms of HIF-1α in IAV-induced ALI remain elusive. Here, we demonstrated that lung epithelial cell-specific Hif1α knockout mice infected with IAV developed more lung IAV replication and severe lung inflammation, which led to increased mortality compared to IAV-infected control mice. Moreover, knockdown of HIF1A in A549 cells (human alveolar type II epithelial cell line) promoted IAV replication in vitro. Mechanistically, knockdown of HIF1A reduced glycolysis by regulating transcription of glycolysis-related enzymes, which subsequently activated the AMPKα-ULK1 signalling pathway. Interestingly, AMPKα-ULK1 signalling promoted autophagy and augmented IAV replication. Taken together, deficiency of HIF-1α in lung epithelial cells reduces glycolysis and enhances AMPKα-ULK1-mediated autophagy, which finally facilitates IAV replication. These findings have deepened our understanding of the role of HIF-1α in regulating IAV replication and provided us novel therapeutic targets for combating influenza infection.

Effects of Vascular Endothelial Growth Factor in Recovery Phase of Acute Lung Injury in Mice.

To test the hypothesis that exogenous administration of vascular endothelial growth factor (VEGF) promotes lung repair in acute lung injury (ALI). ALI was induced by intranasal lipopolysaccharide (LPS) administration in mice, followed by different treatment protocols for 7 days in 3 groups (n = 6, each) including the LPS, the VEGF and the anti-VEGF group. At day 7, peripheral blood and lungs were collected. Lung wet-to-dry (W/D) ratio and lung injury score were measured. Immunohistochemistry assay was employed to detect the number of pulmonary vessels. Circulating endothelial progenitor cells (EPCs) was detected using flow cytometric analysis, and the apoptosis of lung cells was determined by TUNEL staining. VEGF treatment reduced W/D ratio and pulmonary neutrophil infiltration in the VEGF group compared with the LPS group. The treatment of VEGF increased the number of pulmonary vessels, and significantly increased the number of circulating EPC cells. Moreover, administration of VEGF decreased the percentage of apoptotic cells in the VEGF group. Our results suggest that VEGF may contribute to vascular endothelial repair and function as a protective factor against ALI.

Metformin-stimulated AMPK-α1 promotes microvascular repair in acute lung injury

Acute lung injury secondary to sepsis is a leading cause of mortality in sepsis-related death. Present therapies are not effective in reversing endothelial cell dysfunction, which plays a key role in increased vascular permeability and compromised lung function. AMP-activated protein kinase (AMPK) is a molecular sensor important for detection and mediation of cellular adaptations to vascular disruptive stimuli. In this study, we sought to determine the role of AMPK in resolving increased endothelial permeability in the sepsis-injured lung. AMPK function was determined in vivo using a rat model of endotoxin-induced lung injury, ex vivo using the isolated lung, and in vitro using cultured rat pulmonary microvascular endothelial cells (PMVECs). AMPK stimulation using N1-(α-d-ribofuranosyl)-5-aminoimidizole-4-carboxamide or metformin decreased the LPS-induced increase in permeability, as determined by filtration coefficient (Kf) measurements, and resolved edema as indicated by decreased wet-to-dry ratios. The role of AMPK in the endothelial response to LPS was determined by shRNA designed to decrease expression of the AMPK-α1 isoform in capillary endothelial cells. Permeability, wounding, and barrier resistance assays using PMVECs identified AMPK-α1 as the molecule responsible for the beneficial effects of AMPK in the lung. Our findings provide novel evidence for AMPK-α1 as a vascular repair mechanism important in the pulmonary response to sepsis and identify a role for metformin treatment in the management of capillary injury.

Novel concepts of acute lung injury and alveolar-capillary barrier dysfunction

In this review we summarize recent major advances in our understanding on the molecular mechanisms, mediators, and biomarkers of acute lung injury (ALI) and alveolar-capillary barrier dysfunction, highlighting the role of immune cells, inflammatory and noninflammatory signaling events, mechanical noxae, and the affected cellular and molecular entities and functions. Furthermore, we address novel aspects of resolution and repair of ALI, as well as putative candidates for treatment of ALI, including pharmacological and cellular therapeutic means.

Repair of Lipopolysaccharide-Induced Acute Lung Injury in Mice by Endothelial Progenitor Cells, Alone and in Combination With Simvastatin

Endothelial progenitor cells (EPCs) are involved in endothelium repair of acute lung injury (ALI). Numerous studies have demonstrated that statins can promote EPC function in vitro and in vivo; therefore, the purpose of this study was to determine whether simvastatin enhances the function of EPCs participating in the repair of ALI. BALB/C mice were initially pretreated with simvastatin by intraperitoneal administration 24 h before, and again at the time of, intratracheal instillation of lipopolysaccharide (LPS) and subsequently treated with EPCs by i.v. transplantation 2 h later. The effects of capillary permeability, endothelium repair, and inflammatory cytokines were measured. This study revealed that both simvastatin administration and EPC transplantation can reduce the severity of LPS-induced ALI in mice, and the effect can be further improved by combining the two therapies. The administration of simvastatin and EPC transplantation can reduce the severity of LPS-induced ALI in mice, and improvement is moderately enhanced in some respects when EPC transplantation is combined with simvastatin administration. The beneficial role of simvastatin on EPCs may be a component of its pleiotropic effects. Although the exact mechanism remains unknown, the combined administration of simvastatin and EPC transplantation may be a potentially important, cell-based, inflammation-mediated therapy for patients with ALI/ARDS.

CD166pos Subpopulation From Differentiated Human ES and iPS Cells Support Repair of Acute Lung Injury

Previous efforts to derive lung progenitor cells from human embryonic stem (hES) cells using embryoid body formation or stromal feeder cocultures had been limited by low efficiencies. Here, we report a step-wise differentiation method to drive both hES and induced pluripotent stem (iPS) cells toward the lung lineage. Our data demonstrated a 30% efficiency in generating lung epithelial cells (LECs) that expresses various distal lung markers. Further enrichment of lung progenitor cells using a stem cell marker, CD166 before transplantation into bleomycin-injured NOD/SCID mice resulted in enhanced survivability of mice and improved lung pulmonary functions. Immunohistochemistry of lung sections from surviving mice further confirmed the specific engraftment of transplanted cells in the damaged lung. These cells were shown to express surfactant protein C, a specific marker for distal lung progenitor in the alveoli. Our study has therefore demonstrated the proof-of-concept of using iPS cells for the repair of acute lung injury, demonstrating the potential usefulness of using patient's own iPS cells to prevent immune rejection which arise from allogenic transplantation.

Intrapleural delivery of MSCs attenuates acute lung injury by paracrine/endocrine mechanism.

Two different repair mechanisms of mesenchymal stem cells (MSCs) are suggested to participate in the repair of acute lung injury (ALI): (i) Cell engraftment mechanism, (ii) Paracrine/endocrine mechanism. However, the exact roles they play in the repair remain unclear. The aim of the study was to evaluate the role of paracrine/endocrine mechanism using a novel intrapleural delivery method of MSCs. Either 1 × 106 MSCs in 300 μl of PBS or 300 μl PBS alone were intrapleurally injected into rats with endotoxin-induced ALI. On days 1, 3 or 7 after injections, samples of lung tissues and bronchoalveolar lavage fluid (BALF) were collected from each rat for assessment of lung injury, biochemical analysis and histology. The distribution of MSCs was also traced by labelling the cells with 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI). MSCs intrapleural injection significantly improved LPS-induced lung histopathology compared with PBS-treated group at day 3. There was also a significant decrease in total cell counts and protein concentration in BALF at day 7 in the MSCs -treated rats compared to PBS control group. Tracking the DAPI-marked MSCs showed that there were no exotic MSCs in the lung parenchyma. MSCs administration resulted in a down-regulation of pro-inflammatory response to endotoxin by reducing TNF-α both in the BALF and in the lung, while up-regulating the anti-inflammatory cytokine IL-10 in the lung. In conclusion, treatment with intrapleural MSCs administration markedly attenuates the severity of endotoxin-induced ALI. This role is mediated by paracrine/endocrine repair mechanism of MSCs rather than by the cell engraftment mechanism.

Hepatocyte growth factor levels in Legionella pneumonia: A retrospective study

BackgroundHepatocyte growth factor (HGF) is known to be involved in the resolution of pulmonary inflammation and repair of acute lung injury. Legionella pneumonia is sometimes complicated by acute lung injury. Our study aimed to determine the role of serum HGF levels in Legionella pneumonia.MethodsSera from patients with Legionella pneumonia (42 cases), other bacterial pneumonia (33 cases), pulmonary tuberculosis (19 cases), and normal controls (29 cases) were collected. The serum HGF levels for each serum sample were determined by sandwich ELISA. Clinical and laboratory data were collected by reviewing the medical charts.ResultsSerum HGF levels were higher in patients with Legionella pneumonia than in those with other bacterial pneumonia, pulmonary tuberculosis, and controls. The HGF levels were compared with white blood cell counts, C-reactive protein, Alanine amino- transferase, and lactate dehydrogenase (LDH). The HGF levels were correlated to serum LDH levels. Moreover, serum HGF levels were significantly higher in non-survivors than in survivors.ConclusionsHGF levels increased in severer pneumonia caused by Legionella, suggesting that HGF might play a significant role in the Legionella pneumonia.

In vivo and in vitro effects of salbutamol on alveolar epithelial repair in acute lung injury

In vivo and in vitro effects of salbutamol on alveolar epithelial repair in acute lung injury

In vivo and in vitro effects of salbutamol on alveolar epithelial repair in acute lung injury

Background: Acute lung injury is an important cause of respiratory failure in the critically ill patient. It is caused by damage to the alveolar barrier with subsequent alveolar flooding leading...

Mechanisms of beta-receptor stimulation-induced improvement of acute lung injury and pulmonary edema

Acute lung injury (ALI) and the acute respiratory distress syndrome are complex syndromes because both inflammatory and coagulation cascades cause lung injury. Transport of salt and water, repair and remodeling of the lung, apoptosis, and necrosis are additional important mechanisms of injury. Alveolar edema is cleared by active transport of salt and water from the alveoli into the lung interstitium by complex cellular mechanisms. Beta-2 agonists act on the cellular mechanisms of pulmonary edema clearance as well as other pathways relevant to repair in ALI. Numerous studies suggest that the beneficial effects of beta-2 agonists in ALI include at least enhanced fluid clearance from the alveolar space, anti-inflammatory actions, and bronchodilation. The purposes of the present review are to consider the effects of beta agonists on three mechanisms of improvement of lung injury: edema clearance, anti-inflammatory effects, and bronchodilation. This update reviews specifically the evidence on the effects of beta-2 agonists in human ALI and in models of ALI. The available evidence suggests that beta-2 agonists may be efficacious therapy in ALI. Further randomized controlled trials of beta agonists in pulmonary edema and in acute lung injury are necessary.

Mechanisms of alveolar epithelial repair in acute lung injury--a translational approach.

In patients with acute lung injury (ALI) or acute respiratory distress syndrome (ARDS), extensive damage to the alveolar epithelial and endothelial barrier is observed, resulting in the influx of protein-rich oedema fluid into the air spaces. Efficient alveolar epithelial repair is crucial to ALI/ARDS patients' recovery. Future therapeutic strategies may therefore include acceleration of the epithelial repair process in the injured lung. However, a better understanding of the cellular and molecular mechanisms that promote alveolar epithelial repair is needed if novel therapeutic strategies are to be developed. Pulmonary oedema fluid from patients with ALI/ARDS and from patients with hydrostatic oedema as control was obtained, and the effect on alveolar epithelial repair in vitro using our alveolar epithelial wound repair bioassay was studied. In contrast to the initial hypothesis, pulmonary oedema fluid from ALI/ARDS patients increased alveolar epithelial repair in vitro by an interleukin-1beta (IL-1beta)-dependent mechanism, demonstrating a novel, possibly beneficial role for IL-1beta in patients with ALI/ARDS. Further studies using primary alveolar epithelial cells from rats revealed that IL-1beta induced alveolar epithelial repair by an epidermal growth factor (EGF)/transforming growth factor-alpha (TGF-alpha)-dependent pathway. Besides EGF and TGF-alpha, keratinocyte growth factor (KGF) and hepatocyte growth factor (HGF)--both present in pulmonary oedema fluid obtained from patients with ALI/ARDS--stimulate alveolar epithelial repair in vitro. Further experimental and clinical studies will show whether acceleration of alveolar epithelial repair by modulating cytokines and growth factors in the injured lung represents a promising new therapeutic strategy in patients with ALI/ARDS.

Decreased Intracellular Iron Availability Suppresses Epithelial Cell Surface Plasmin Generation

Iron and iron metabolism are critical in a variety of physiologic and pathophysiologic processes, including lung injury and repair. The plasmin/plasminogen activator (PA) system is involved in the extensive remodeling process that follows acute lung injury, and alveolar epithelial cells play a key role in this repair process. Herein we report that decreased intracellular iron availability markedly suppresses cell-surface plasmin generation by A549 human carcinoma-derived pulmonary epithelial cells. This effect is mediated by concomitant downregulation of urokinase-type PA and upregulation of PA inhibitor-type 1 expression. Northern analyses, runoff transcription assays, and messenger RNA half-life experiments using actinomycin demonstrate that transcriptional and post-transcriptional mechanisms are operative. Given these potent in vitro effects on the plasmin/PA system, we speculate that adequate intracellular iron stores are important for successful repair of acute lung injury.

Hyperoxia increases keratinocyte growth factor mRNA expression in neonatal rabbit lung.

Acute hyperoxic lung injury remains a major factor in the development of chronic lung disease in neonates. A critical step in the repair of acute lung injury is the proliferation of type II alveolar epithelial cells. Type II cell proliferation is stimulated by keratinocyte growth factor (KGF), an epithelial cell-specific mitogen. We sought to investigate KGF mRNA expression in relation to type II cell proliferation during hyperoxic lung injury. We studied a previously described newborn (NB) rabbit model of acute and chronic hyperoxic injury [C. T. D'Angio, J. N. Finkelstein, M. B. LoMonaco, A. Paxhia, S. A. Wright, R. B. Baggs, R. H. Notter, and R. M. Ryan. Am. J. Physiol. 272 (Lung Cell. Mol. Physiol. 16): L720-L730, 1997]. NB rabbits were placed in 100% O2 for 9 days and then recovered in 60% O2. RT-PCR was used to synthesize and amplify a 267-bp fragment of rabbit KGF cDNA from whole lung RNA. KGF mRNA expression was analyzed by ribonuclease protection assay, and mRNA abundance was quantified by phosphorimaging. Proliferating cell nuclear antigen immunohistochemistry was used on lung sections to identify proliferating cells. The rabbit partial cDNA sequenced was >95% homologous to human cDNA, and all amino acids were conserved. Whole lung KGF mRNA expression was increased 12-fold after 6 days of hyperoxia compared with control lungs, and remained increased throughout the 100% O2 exposure period. Proliferating cell nuclear antigen immunohistochemistry showed an increase in type II cell proliferation after 8-12 days of hyperoxia. NB rabbits exposed to hyperoxic injury exhibit increased whole lung KGF mRNA expression preceding type II cell proliferation. KGF may be an important mitogen in the regulation of alveolar epithelial repair after hyperoxic lung injury.

Repair of chronic hyperoxic lung injury: Changes in lung ultrastructure and matrix

We studied changes in lung ultrastructure, fibronectin, and collagen during repair of chronic hyperoxic lung injury induced by exposure of rats to 85% oxygen for 14 days. Morphologically, the most persistent changes were in the alveolar interstitium. After 28 days of repair, the extracellular matrix volume was still twofold normal. Total interstitial cell numbers also remained high and intersititial myofibroblast number actually doubled between Days 7 and 14. These changes contrast markedly with repair of acute lung injury induced by 100% oxygen ( Thet et al. (1986) , Exp. Lung Res. 11, 209–228) in which matrix volume and interstitial myofibroblast number increased initially but then returned to normal. Biochemically, tissue-associated fibronectin was high initially and peaked at 3 days before slowly declining. Tissue collagen content began to increase after the peak in fibronectin content and was over 150% of controls at 28 days; this correlated with an increase in visible collagen fibers. We conclude that changes in lung morphology and matrix after chronic hyperoxic lung injury are more persistent than after acute hyperoxic lung injury and result in a greater degree of chronic interstitial fibrosis.

Changes in pulmonary calpain activity following treatment of mice with butylated hydroxytoluene

The antioxidant, butylated hydroxytoluene (BHT), causes lung toxicity in mice followed by regenerative repair, and can also modulate the development of carcinogen-induced lung adenomas. We are investigating changes in pulmonary biochemistry following BHT treatment in order to understand the mechanisms of BHT-induced pulmonary regenerative repair. BHT administration lowered cytosolic Ca 2+-activated neutral protease (calpain) activity, increased the activity of the endogenous calpain inhibitor, calpastatin, increased the extent of photoincorporation of 8- N 3-[ 32P]cAMP into a M r 37,000 proteolytic product derived from cAMP-dependent protein kinase regulatory (R) subunits, and increased membrane-associated protease activity. All of these changes were dependent on the BHT dosage; the altered proteolytic activities occurred at a dose lower than that which caused observable lung toxicity as assessed by the lung weight/body weight ratio. Decreased cytosolic calpain activity was detectable within 1 day after BHT administration, was lowest at 4–7 days, and had not returned to control levels by Day 21, a time when normal lung morphology had been regained. The decrease in calpain activity cannot fully be accounted for by increased calpastatin activity; upon separation of these proteins by DEAE chromatography, the amount of calpain activity from BHT-treated mice remained lower than the corresponding peak from control mice. Increased photolabeling of the M r 37,000 protein began at 1 day and continued to increase up to 4 days after BHT. All of the cytosolic changes preceded the increased particulate proteolytic activity by 1–2 days. R-subunits which have dissociated from their catalytic subunits are more susceptible to degradation by calpain, but BHT treatment did not enhance subunit dissociation as determined by the elution profile of 8- N 3[ 32P]cAMP-labeled R-subunits following DEAE chromatography. A large percentage of the particulate protease activity was inhibited by calpastatin, leupeptin, and E-64, all of which are known to inhibit calpain activity; this suggested that calpain accounted for most of this activity. Changes in the activities of proteases which catalyze limited proteolysis reactions may play an important role in the repair of acute lung injury.

Sequential changes in lung morphology during the repair of acute oxygen-induced lung injury in adult rats.

We studied changes in lung ultrastructure and collagen content during the repair of acute lung injury in adult rats exposed to 100% O2 for 60 h and recovering in ambient air. In the interstitium, during the first 3 days of repair, the number of neutrophils decreased 16-fold, and monocytes and lymphocytes increased to 7-fold and 4-fold the respective control values. Myofibroblasts increased about 5-fold and the volume of the interstitial matrix remained high. By 7 days, the differential count of inflammatory cells was normal although the number of total interstitial cells and myofibroblasts decreased more slowly. In the capillary endothelium, after 3 days of repair, the cells were hypertrophied and had organelle-rich cytoplasm, and total cell number had increased back to control values; endothelial cell number increased an additional 63% between 3 and 7 days of repair. In the epithelium, type 2 cells increased 150% during the first 3 days of repair before decreasing; type 1 cell number did not change. After 28 days of repair, the lungs appeared qualitatively almost normal; however, interstitial cell number and collagen content were still increased. We conclude that the repair of oxygen-induced lung injury involves a complex pattern of morphologic changes that has important similarities to those occurring during repair on other tissues such as the skin.