Let-7 restrains an oncogenic epigenetic circuit in AT2 cells to prevent ectopic formation of fibrogenic transitional cell intermediates and pulmonary fibrosis.

Analysis of lung alveolar type 2 (AT2) progenitor stem cells has highlighted fundamental mechanisms that direct their differentiation into alveolar type 1 cells (AT1s) in lung repair and disease. However, microRNA (miRNA) mediated post-transcriptional mechanisms which govern this nexus remain understudied. We show here that the let-7 miRNA family serves a homeostatic role in governance of AT2 quiescence, specifically by preventing the uncontrolled accumulation of AT2 transitional cells and by promoting AT1 differentiation to safeguard the lung from spontaneous alveolar destruction and fibrosis. Using mice and organoid models with genetic ablation of let-7a1/let-7f1/let-7d cluster ( let-7afd ) in AT2 cells, we demonstrate prevents AT1 differentiation and results in aberrant accumulation of AT2 transitional cells in progressive pulmonary fibrosis. Integration of enhanced AGO2 UV-crosslinking and immunoprecipitation sequencing (AGO2-eCLIP) with RNA-sequencing from AT2 cells uncovered the induction of direct targets of let-7 in an oncogene feed-forward regulatory network including BACH1/EZH2 which drives an aberrant fibrotic cascade. Additional analyses by CUT&RUN-sequencing revealed loss of let-7afd hampers AT1 differentiation by eliciting aberrant histone EZH2 methylation which prevents the exit of AT2 transitional cells into terminal AT1s. This study identifies let-7 as a key gatekeeper of post-transcriptional and epigenetic chromatin signals to prevent AT2-driven pulmonary fibrosis.

The Role of Mitochondrial Dysfunction in Idiopathic Pulmonary Fibrosis: New Perspectives for a Challenging Disease.

Mitochondrial biology has always been a relevant field in chronic diseases such as fibrosis or cancer in different organs of the human body, not to mention the strong association between mitochondrial dysfunction and aging. With the development of new technologies and the emergence of new methodologies in the last few years, the role of mitochondria in pulmonary chronic diseases such as idiopathic pulmonary fibrosis (IPF) has taken an important position in the field. With this review, we will highlight the latest advances in mitochondrial research on pulmonary fibrosis, focusing on the role of the mitochondria in the aging lung, new proposals for mechanisms that support mitochondrial dysfunction as an important cause for IPF, mitochondrial dysfunction in different cell populations of the lung, and new proposals for treatment of the disease.

Intrinsic Dynamics of the ClpXP Proteolytic Machine Using Elastic Network Models.

ClpXP complex is an ATP-dependent mitochondrial matrix protease that binds, unfolds, translocates, and subsequently degrades specific protein substrates. Its mechanisms of operation are still being debated, and several have been proposed, including the sequential translocation of two residues (SC/2R), six residues (SC/6R), and even long-pass probabilistic models. Therefore, it has been suggested to employ biophysical-computational approaches that can determine the kinetics and thermodynamics of the translocation. In this sense, and based on the apparent inconsistency between structural and functional studies, we propose to apply biophysical approaches based on elastic network models (ENM) to study the intrinsic dynamics of the theoretically most probable hydrolysis mechanism. The proposed models ENM suggest that the ClpP region is decisive for the stabilization of the ClpXP complex, contributing to the flexibility of the residues adjacent to the pore, favoring the increase in pore size and, therefore, with the energy of interaction of its residues with a larger portion of the substrate. It is predicted that the complex may undergo a stable configurational change once assembled and that the deformability of the system once assembled is oriented, to increase the rigidity of the domains of each region (ClpP and ClpX) and to gain flexibility of the pore. Our predictions could suggest under the conditions of this study the mechanism of the interaction of the system, of which the substrate passes through the unfolding of the pore in parallel with a folding of the bottleneck. The variations in the distance calculated by molecular dynamics could allow the passage of a substrate with a size equivalent to ∼3 residues. The theoretical behavior of the pore and the stability and energy of binding to the substrate based on ENM models suggest that in this system, there are thermodynamic, structural, and configurational conditions that allow a possible translocation mechanism that is not strictly sequential.

Syndecan-2 regulates PAD2 to exert antifibrotic effects on RA-ILD fibroblasts.

Rheumatoid arthritis (RA)-associated interstitial lung disease (RA-ILD) is the most common pulmonary complication of RA, increasing morbidity and mortality. Anti-citrullinated protein antibodies have been associated with the development and progression of both RA and fibrotic lung disease; however, the role of protein citrullination in RA-ILD remains unclear. Here, we demonstrate that the expression of peptidylarginine deiminase 2 (PAD2), an enzyme that catalyzes protein citrullination, is increased in lung homogenates from subjects with RA-ILD and their lung fibroblasts. Chemical inhibition or genetic knockdown of PAD2 in RA-ILD fibroblasts attenuated their activation, marked by decreased myofibroblast differentiation, gel contraction, and extracellular matrix gene expression. Treatment of RA-ILD fibroblasts with the proteoglycan syndecan-2 (SDC2) yielded similar antifibrotic effects through regulation of PAD2 expression, phosphoinositide 3-kinase/Akt signaling, and Sp1 activation in a CD148-dependent manner. Furthermore, SDC2-transgenic mice exposed to bleomycin-induced lung injury in an inflammatory arthritis model expressed lower levels of PAD2 and were protected from the development of pulmonary fibrosis. Together, our results support a SDC2-sensitive profibrotic role for PAD2 in RA-ILD fibroblasts and identify PAD2 as a promising therapeutic target of RA-ILD.

Glutamine restores mitochondrial respiration in bleomycin-injured epithelial cells.

Whether from known or unknown causes, loss of epithelial repair plays a central role in the pathogenesis of pulmonary fibrosis. Recently, diminished mitochondrial function has been implicated as a factor contributing to the loss of epithelial repair but the mechanisms mediating these changes have not been defined. Here, we investigated the factors contributing to mitochondrial respiratory dysfunction after bleomycin, a widely accepted agent for modeling pulmonary fibrosis in mice and in vitro systems. In agreement with previous reports, we found that mitochondrial respiration was decreased in lung epithelial cells exposed to bleomycin, but also observed that responses differed depending on the type of metabolic fuel available to cells. For example, we found that mitochondrial respiration was dramatically reduced when glucose served as the primary fuel. Moreover, this associated with a marked decrease in glucose uptake, expression of glucose uptake transport 1 and capacity to augment glycolysis to either glucose or oligomycin. Conversely, mitochondrial respiration was largely preserved if glutamine was present in culture medium. The addition of glutamine also led to increased intracellular metabolite levels, including multiple TCA cycle intermediates and the glycolytic intermediate lactate, as well as reduced DNA damage and cell death to bleomycin. Taken together, these findings indicate that glutamine, rather than glucose, supports mitochondrial respiration and metabolite production in injured lung epithelial cells, and suggest that this shift away from glucose utilization serves to protect the lung epithelium from injury.

A model of the aged lung epithelium in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is an age-related disorder that carries a universally poor prognosis and is thought to arise from repetitive micro injuries to the alveolar epithelium. To date, a major factor limiting our understanding of IPF is a deficiency of disease models, particularly in vitro models that can recapitulate the full complement of molecular attributes in the human condition. In this study, we aimed to develop a model that more closely resembles the aberrant IPF lung epithelium. By exposing mouse alveolar epithelial cells to repeated, low doses of bleomycin, instead of usual one-time exposures, we uncovered changes strikingly similar to those in the IPF lung epithelium. This included the acquisition of multiple phenotypic and functional characteristics of senescent cells and the adoption of previously described changes in mitochondrial homeostasis, including alterations in redox balance, energy production and activity of the mitochondrial unfolded protein response. We also uncovered dramatic changes in cellular metabolism and detected a profound loss of proteostasis, as characterized by the accumulation of cytoplasmic protein aggregates, dysregulated expression of chaperone proteins and decreased activity of the ubiquitin proteasome system. In summary, we describe an in vitro model that closely resembles the aberrant lung epithelium in IPF. We propose that this simple yet powerful tool could help uncover new biological mechanisms and assist in developing new pharmacological tools to treat the disease.

CD148 Deficiency in Fibroblasts Promotes the Development of Pulmonary Fibrosis.

Rationale: CD148/PTRJ (receptor-like protein tyrosine phosphatase η) exerts antifibrotic effects in experimental pulmonary fibrosis via interactions with its ligand syndecan-2; however, the role of CD148 in human pulmonary fibrosis remains incompletely characterized.Objectives: We investigated the role of CD148 in the profibrotic phenotype of fibroblasts in idiopathic pulmonary fibrosis (IPF).Methods: Conditional CD148 fibroblast-specific knockout mice were generated and exposed to bleomycin and then assessed for pulmonary fibrosis. Lung fibroblasts (mouse lung and human IPF lung), and precision-cut lung slices from human patients with IPF were isolated and subjected to experimental treatments. A CD148-activating 18-aa mimetic peptide (SDC2-pep) derived from syndecan-2 was evaluated for its therapeutic potential.Measurements and Main Results: CD148 expression was downregulated in IPF lungs and fibroblasts. In human IPF lung fibroblasts, silencing of CD148 increased extracellular matrix production and resistance to apoptosis, whereas overexpression of CD148 reversed the profibrotic phenotype. CD148 fibroblast-specific knockout mice displayed increased pulmonary fibrosis after bleomycin challenge compared with control mice. CD148-deficient fibroblasts exhibited hyperactivated PI3K/Akt/mTOR signaling, reduced autophagy, and increased p62 accumulation, which induced NF-κB activation and profibrotic gene expression. SDC2-pep reduced pulmonary fibrosis in vivo and inhibited IPF-derived fibroblast activation. In precision-cut lung slices from patients with IPF and control patients, SDC2-pep attenuated profibrotic gene expression in IPF and normal lungs stimulated with profibrotic stimuli.Conclusions: Lung fibroblast CD148 activation reduces p62 accumulation, which exerts antifibrotic effects by inhibiting NF-κB-mediated profibrotic gene expression. Targeting the CD148 phosphatase with activating ligands such as SDC2-pep may represent a potential therapeutic strategy in IPF.

Hermansky-Pudlak syndrome-2 alters mitochondrial homeostasis in the alveolar epithelium of the lung.

BACKGROUND: Mitochondrial dysfunction has emerged as an important player in the pathogenesis of idiopathic pulmonary fibrosis (IPF), a common cause of idiopathic interstitial lung disease in adults. Hermansky-Pudlak syndrome (HPS) is a rare autosomal recessive disorder that causes a similar type of pulmonary fibrosis in younger adults, although the role of mitochondrial dysfunction in this condition is not understood. METHODS: We performed a detailed characterization of mitochondrial structure and function in lung tissues and alveolar epithelial cells deficient in the adaptor protein complex 3 beta 1 (Ap3b1) subunit, the gene responsible for causing subtype 2 of HPS (HPS-2). RESULTS: We observed widespread changes in mitochondrial homeostasis in HPS-2 cells, including the acquisition of abnormally shaped mitochondria, with reduced number of cristae, and markedly reduced activity of the electron transport chain and the tricarboxylic acid cycle. We also found that mitochondrial redox imbalance and activity of the mitochondrial unfolded protein response were dysregulated in HPS-2 cells and this associated with various other changes that appeared to be compensatory to mitochondrial dysfunction. This included an increase in glycolytic activity, an upregulation in the expression of mitochondrial biogenesis factors and enhanced activation of the energy-conserving enzyme AMP-activated protein kinase. CONCLUSION: In summary, our findings indicate that mitochondrial function is dramatically altered in HPS-2 lung tissues, suggesting dysfunction of this organelle might be a driver of HPS lung disease.

Cellular metabolomics of pulmonary fibrosis, from amino acids to lipids.

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive lung disease of unknown etiology with limited treatment options. It is characterized by repetitive injury to alveolar epithelial cells and aberrant activation of numerous signaling pathways. Recent evidence suggests that metabolic reprogramming, metabolic dysregulation, and mitochondria dysfunction are distinctive features of the IPF lungs. Through numerous mechanisms, metabolomic abnormalities in alveolar epithelial cells, myofibroblast, macrophages, and fibroblasts contribute to the abnormal collagen synthesis and dysregulated airway remodeling described in lung fibrosis. This review summarizes the metabolomic changes in amino acids, lipids, glucose, and heme seen in IPF lungs. Simultaneously, we provide new insights into potential therapeutic strategies by targeting a variety of metabolites.

A Tale of Two Proteolytic Machines: Matrix Metalloproteinases and the Ubiquitin-Proteasome System in Pulmonary Fibrosis.

Pulmonary fibrosis is a chronic and progressive lung disease characterized by the activation of fibroblasts and the irreversible deposition of connective tissue matrices that leads to altered pulmonary architecture and physiology. Multiple factors have been implicated in the pathogenesis of lung fibrosis, including genetic and environmental factors that cause abnormal activation of alveolar epithelial cells, leading to the development of complex profibrotic cascade activation and extracellular matrix (ECM) deposition. One class of proteinases that is thought to be important in the regulation of the ECM are the matrix metalloproteinases (MMPs). MMPs can be up- and down- regulated in idiopathic pulmonary fibrosis (IPF) lungs and their role depends upon their location and function. Furthermore, alterations in the ubiquitin-proteosome system (UPS), a major intracellular protein degradation complex, have been described in aging and IPF lungs. UPS alterations could potentially lead to the abnormal accumulation and deposition of ECM. A better understanding of the specific roles MMPs and UPS play in the pathophysiology of pulmonary fibrosis could potentially drive to the development of novel biomarkers that can be as diagnostic and therapeutic targets. In this review, we describe how MMPs and UPS alter ECM composition in IPF lungs and mouse models of pulmonary fibrosis, thereby influencing the alveolar epithelial and mesenchymal cell behavior. Finally, we discuss recent findings that associate MMPs and UPS interplay with the development of pulmonary fibrosis.

Mitochondrial Quality Control in Age-Related Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is age-related interstitial lung disease of unknown etiology. About 100,000 people in the U.S have IPF, with a 3-year median life expectancy post-diagnosis. The development of an effective treatment for pulmonary fibrosis will require an improved understanding of its molecular pathogenesis and the "normal" and "pathological' hallmarks of the aging lung. An important characteristic of the aging organism is its lowered capacity to adapt quickly to, and counteract, disturbances. While it is likely that DNA damage, chronic endoplasmic reticulum (ER) stress, and accumulation of heat shock proteins are capable of initiating tissue repair, recent studies point to a pathogenic role for mitochondrial dysfunction in the development of pulmonary fibrosis. These studies suggest that damage to the mitochondria induces fibrotic remodeling through a variety of mechanisms including the activation of apoptotic and inflammatory pathways. Mitochondrial quality control (MQC) has been demonstrated to play an important role in the maintenance of mitochondrial homeostasis. Different factors can induce MQC, including mitochondrial DNA damage, proteostasis dysfunction, and mitochondrial protein translational inhibition. MQC constitutes a complex signaling response that affects mitochondrial biogenesis, mitophagy, fusion/fission and the mitochondrial unfolded protein response (UPRmt) that, together, can produce new mitochondria, degrade the components of the oxidative complex or clearance the entire organelle. In pulmonary fibrosis, defects in mitophagy and mitochondrial biogenesis have been implicated in both cellular apoptosis and senescence during tissue repair. MQC has also been found to have a role in the regulation of other protein activity, inflammatory mediators, latent growth factors, and anti-fibrotic growth factors. In this review, we delineated the role of MQC in the pathogenesis of age-related pulmonary fibrosis.

Alcohol-induced lipid dysregulation impairs glycolytic responses to LPS in alveolar macrophages.

Several conditions are marked by increased susceptibility to, and enhanced severity of, bacterial infections. Alcohol use disorder, one of these conditions, is known to predispose to bacterial pneumonia by suppressing the lung's innate immune system, and more specifically by disrupting critical alveolar macrophage (AM) functions. Recently, we established that chronic ethanol consumption also perturbs surfactant lipid homeostasis in the lung and that elevated concentrations of free fatty acids contribute to blocking essential AM functions, such as agonist-induced cytokine expression. In this study, we extend these observations by showing that elevated free fatty acid levels impair metabolic responses to lipopolysaccharide (LPS) in AMs. In particular, we show that the glycolytic reprogramming characteristic of LPS-stimulated AMs is blunted by the saturated fatty acid palmitate, whereas oleate, an unsaturated fatty acid, or ethanol alone, had no effect on this adaptive metabolic response. Additionally, we found that elevated concentrations of palmitate induced mitochondrial oxidative stress and that glycolytic reprogramming and cytokine production to LPS could be partially restored in AMs by either pharmacologically blocking palmitate entry into mitochondria or administering a mitochondrial-specific antioxidant. Taken together, these findings suggest that alcohol and elevated levels of saturated fatty acids conspire to impair pulmonary innate immunity by altering metabolic responses in AMs. Additionally, our findings suggest that targeting the mechanisms involved in fatty acid metabolism can restore pulmonary immunity and possibly limit bacterial pneumonia in individuals with alcohol use disorder.

Matrix metalloproteinase activity in the lung is increased in Hermansky-Pudlak syndrome.

BACKGROUND: Hermansky-Pudlak syndrome (HPS) is a rare autosomal recessive disorder characterized by oculocutaneous albinism and platelet dysfunction and can sometimes lead to a highly aggressive form of pulmonary fibrosis that mimics the fatal lung condition called idiopathic pulmonary fibrosis (IPF). Although the activities of various matrix metalloproteinases (MMPs) are known to be dysregulated in IPF, it remains to be determined whether similar changes in these enzymes can be detected in HPS. RESULTS: Here, we show that transcript and protein levels as well as enzymatic activities of MMP-2 and -9 are markedly increased in the lungs of mice carrying the HPS Ap3b1 gene mutation. Moreover, immunohistochemical staining localized this increase in MMP expression to the distal pulmonary epithelium, and shRNA knockdown of the Ap3b1 gene in cultured lung epithelial cells resulted in a similar upregulation in MMP-2 and -9 expression. Mechanistically, we found that upregulation in MMP expression associated with increased activity of the serine/threonine kinase Akt, and pharmacological inhibition of this enzyme resulted in a dramatic suppression of MMP expression in Ap3b1 deficient lung epithelial cells. Similarly, levels and activity of different MMPs were also found to be increased in the lungs of mice carrying the Bloc3 HPS gene mutation and in the bronchoalveolar lavage fluid of subjects with HPS. However, an association between MMP activity and disease severity was not detected in these individuals. CONCLUSIONS: In summary, our findings indicate that MMP activity is dysregulated in the HPS lung, suggesting a role for these proteases as biological markers or pathogenic players in HPS lung disease.

The involvement of GM-CSF deficiencies in parallel pathways of pulmonary alveolar proteinosis and the alcoholic lung.

Chronic alcohol consumption renders the lung more susceptible to infections by disrupting essential alveolar macrophage functions. Emerging evidence suggests that these functional deficits are due, in part, to a suppression of GM-CSF signaling, which is believed to compromise monocyte growth and maturation in the lung. However, in addition to controlling monocyte behaviors, GM-CSF also regulates surfactant homeostasis. For example, mice with targeted deletion of the gene for GM-CSF accumulate large amounts of surfactant phospholipids in their lungs. Moreover, decreased GM-CSF signaling in humans has been linked to the development of pulmonary alveolar proteinosis (PAP), a rare disorder in which surfactant lipids and proteins accumulate in alveolar macrophages and the lung exhibits enhanced susceptibility to infection. Consistent with parallel mechanisms in the PAP and alcoholic lung, we have recently reported that levels of intrapulmonary lipids, specifically triglycerides and free fatty acids, are increased in BAL fluid, whole lung digests and alveolar macrophages of chronically alcohol exposed rats. Additionally, we showed that uptake of saturated fatty acids alone could induce phenotypic and functional changes in alveolar macrophages that mimicked those in the alcohol-exposed rat and human lung. Herein, we discuss the role of GM-CSF in surfactant homeostasis and highlight the evidence that links decreased GM-CSF signaling to alveolar macrophage dysfunction in both the PAP and alcohol-exposed lung. Moreover, we discuss how lipid accumulation itself might contribute to altering alveolar macrophage function and propose how targeting these mechanisms could be employed for reducing the susceptibility to pulmonary infections in alcoholics.

Activation of the mTORC1/PGC-1 axis promotes mitochondrial biogenesis and induces cellular senescence in the lung epithelium.

Cellular senescence is a biological process by which cells lose their capacity to proliferate yet remain metabolically active. Although originally considered a protective mechanism to limit the formation of cancer, it is now appreciated that cellular senescence also contributes to the development of disease, including common respiratory ailments such as chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis. While many factors have been linked to the development of cellular senescence, mitochondrial dysfunction has emerged as an important causative factor. In this study, we uncovered that the mitochondrial biogenesis pathway driven by the mammalian target of rapamycin/peroxisome proliferator-activated receptor-γ complex 1α/ß (mTOR/PGC-1α/ß) axis is markedly upregulated in senescent lung epithelial cells. Using two different models, we show that activation of this pathway is associated with other features characteristic of enhanced mitochondrial biogenesis, including elevated number of mitochondrion per cell, increased oxidative phosphorylation, and augmented mitochondrial reactive oxygen species (ROS) production. Furthermore, we found that pharmacological inhibition of the mTORC1 complex with rapamycin not only restored mitochondrial homeostasis but also reduced cellular senescence to bleomycin in lung epithelial cells. Likewise, mitochondrial-specific antioxidant therapy also effectively inhibited mTORC1 activation in these cells while concomitantly reducing mitochondrial biogenesis and cellular senescence. In summary, this study provides a mechanistic link between mitochondrial biogenesis and cellular senescence in lung epithelium and suggests that strategies aimed at blocking the mTORC1/PGC-1α/ß axis or reducing ROS-induced molecular damage could be effective in the treatment of senescence-associated lung diseases.

Metabolic Reprogramming as a Driver of Fibroblast Activation in PulmonaryFibrosis.

Pulmonary fibrosis refers to a heterogeneous group of disorders that scar the lung, most often irreversibly. To date, there are limited effective treatments for these conditions, despite decades of research in this area of investigation. In pulmonary fibrosis, the principle cell responsible for producing the vast majority of scar tissue is the fibroblast, making these cells ideally suited for drug targeting. For decades, the major experimental approach to blocking the activity of lung fibroblasts has been either to inhibit the interaction of fibroblast growth factors with their receptors or interfere with downstream effector molecules regulating extracellular matrix production. However, emerging evidence now indicates that lung fibroblasts also undergo dramatic metabolic reprogramming in the setting of growth factor stimulation. These discoveries, along with preclinical investigations showing marked reductions in lung fibrosis after targeting specific metabolic pathways, has led to a total rethinking of drug development in the pulmonary fibrosis field. Here, we review the major metabolic pathways and highlight some of the key metabolic events that occur in the transition of fibroblasts from quiescent to activated states. Moreover, we discuss the emerging evidence linking changes in fibroblast metabolism to pulmonary fibrosis and propose how targeting specific metabolic pathways could be employed in the treatment of fibrotic lung diseases.

Fine-tuning the ubiquitin-proteasome system to treat pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is an extremely aggressive lung disease that develops almost exclusively in older individuals, carries a very poor prognosis, and lacks any truly effective therapies. The current conceptual model is that IPF develops because of an age-related decline in the ability of the lung epithelium to regenerate after injury, largely due to death or senescence of epithelial progenitor cells in the distal airways. This loss of regenerative capacity is thought to initiate a chronic and ineffective wound-healing response, characterized by persistent, low-grade lung inflammation and sustained production of collagen and other extracellular matrix materials. Despite recent advances in our understanding of IPF pathobiology, there remains a pressing need to further delineate underlying mechanisms to develop more effective therapies for this disease. In this review, we build the case that many of the manifestations of IPF result from a failure of cells to effectively manage their proteome. We propose that epithelial progenitor cells, as well as immune cells and fibroblasts, become functionally impaired, at least in part, because of an accumulation or a loss in the expression of various crucial proteins. Further, we propose that central to this defect is the dysregulation of the ubiquitin-proteasome system (UPS), which is the major protein-degradation system in eukaryotic cells. Lastly, borrowing concepts from other fields, we discuss how targeting the UPS system could be employed as a novel treatment for IPF and perhaps for other fibrotic lung diseases as well.

MicroRNA-34a Promotes Endothelial Dysfunction and Mitochondrial-mediated Apoptosis in Murine Models of Acute Lung Injury.

Recent evidence has shown that microRNAs (miRs) are involved in endothelial dysfunction and vascular injury in lung-related diseases. However, the potential role of miR-34a in the regulation of pulmonary endothelial dysfunction, vascular injury, and endothelial cells (ECs) apoptosis in acute lung injury (ALI)/acute lung respiratory distress syndrome is largely unknown. Here, we show that miR-34a-5p was upregulated in whole lungs, isolated ECs from lungs, and ECs stimulated with various insults (LPS and hyperoxia). Overexpression of miR-34a-5p in ECs exacerbated endothelial dysfunction, inflammation, and vascular injury, whereas the suppression of miR-34a-5p expression in ECs and miR-34a-null mutant mice showed protection against LPS- and hyperoxia-induced ALI. Furthermore, we observed that miR-34a-mediated endothelial dysfunction is associated with decreased miR-34a direct-target protein, sirtuin-1, and increased p53 expression in whole lungs and ECs. Mechanistically, we show that miR-34a leads to translocation of p53 and Bax to the mitochondrial compartment with disruption of mitochondrial membrane potential to release cytochrome C into the cytosol, initiating a cascade of mitochondrial-mediated apoptosis in lungs. Collectively, these data show that downregulating miR-34a expression or modulating its target proteins may improve endothelial dysfunction and attenuate ALI.

Surface Behavior of BSA/Water/Carbohydrate Systems from Molecular Polarizability Measurements.

The effect of the presence of glucose and sucrose on the nonintrinsic contribution to partial molar volume ⟨Θ⟩ni of bovine serum albumin (BSA) is determined by means of static and dynamic electronic polarizability measurements. For that aim, a combined strategy based on high-resolution refractometry, high exactitude densitometry, and synchronous fluorescence spectroscopy is applied. Both static and dynamic mean electronic molecular polarizability values are found to be sensitive to the presence of glucose. In the case of sucrose, the polarizability of BSA is not appreciably affected. In fact, our results revealed that the electronic changes observed occurred without a modification of the native conformation of BSA. On the contrary, a nonmonotonous behavior with the concentration is observed in presence of glucose. These results advocate the influence of the electronic polarization on the repulsive and attractive protein-carbohydrate interactions. An analysis using the scaled particle theory indicates that the accumulation of glucose on the protein surface promotes dehydration. Inversely, hydration and preferential exclusion occur in the vicinity of the protein surface for sucrose-enriched systems.