Rodent Lung Research Articles

Development of a nanometre scale X-ray speckle-based CT technique through the 3-D histological assessment of an acute respiratory distress syndrome model

The study of biological soft tissue structures at the micron scale details the function of healthy and pathological tissues, which is vital in the diagnosis and treatment of diseases. Speckle based X-ray phase contrast tomographic scans at a nanometer scale have the potential to thoroughly analyse such tissues in a quantitative and qualitative manner. Diamond light source, the UKs national synchrotron facility developed and refined a 1-D X-ray speckle-based imaging technique, referred to as Fly scan mode. This novel image acquisition technique was used to perform a rapid structural composition scan of rodent lung histology samples. The rodent samples were taken from healthy and Staphylococcus aureus induced acute respiratory distress syndrome models. The analysis and cross comparison of the fly scan method, absorption-based tomography and conventional histopathology H&E staining microscopy are discussed in this paper. This analysis and cross comparison outline the ways the speckle-based technique can be of benefit. These advantages include improved soft tissue contrast, 3-D volumetric rendering, segmentation of specific gross tissue structures, quantitative analysis of gross tissue volume. A further advantage is the analysis of cellular distribution throughout the volumetric rendering of the tissue sample. The study also details the current limitations of this technique and points to ways in which future work on this imaging modality may progress.

Optimizing miRNA transfection for screening in precision cut lung slices.

Precision cut lung slices(PCLS) are complex 3D lung tissue models, which preserve the native microenvironment, including cell diversity and cell-matrix interactions. They are an innovative ex vivo platform that allows studying disease as well as the effects of therapeutic agents or regulatory molecules(e.g. miRNA). The aim of our study was to develop a protocol to transfect PCLS with miRNA using lipid nanoparticles (LNPs) to enable higher throughput screening of miRNA, obviating the need for custom stabilization and internalization approaches. 4mm diameter PCLS were generated using agarose-filled rodent lungs and a vibratome. TYE665 labelled scrambled miRNA was used to evaluate transfection efficacy of six different commercially available LNPs. Transfection efficacy was visualised using live high content fluorescence microscopy, followed by higher resolution confocal fluorescence microscopy in fixed PCLS. Metabolic activity and cellular damage were assessed using WST-1 and lactate dehydrogenase(LDH) release. Using a live staining kit containing a cell membrane impermeant nuclear dye, RedDot2TM, we established that cellular membranes in PCLS are permeable in the initial 24 hours of slicing but diminished thereafter. Therefore, all transfection experiments occurred at least 24 hours after slicing. All six commercially available LNPs enabled transfection without inducing significant cytotoxicity or impaired metabolic function. However, RNAiMAX and INTERFERin led to increases in transfection efficacy as compared to other LNPs, with detection possible as low as 25nM. Therefore, LNP-based transfection of miRNA is possible and can be visualized in live or fixed PCLS, enabling future higher throughput studies using diverse miRNAs.

Lung quantitative ultrasound to stage and monitor interstitial lung diseases

Chronic interstitial lung diseases (ILDs) require frequent point-of-care monitoring. X-ray-based methods lack resolution and are ionizing. Chest computerized tomographic (CT) scans are expensive and provide more radiation. Conventional ultrasound can detect severe lung damage via vertical artifacts (B-lines). However, this information is not quantitative, and the appearance of B-lines is operator- and system-dependent. Here we demonstrate novel ultrasound-based biomarkers to assess severity of ILDs. Lung alveoli scatter ultrasound waves, leading to a complex acoustic signature, which is affected by changes in alveolar density due to ILDs. We exploit ultrasound scattering in the lung and combine quantitative ultrasound (QUS) parameters, to develop ultrasound-based biomarkers that significantly correlate (p = 1e−4 for edema and p = 3e−7 for fibrosis) to the severity of pulmonary fibrosis and edema in rodent lungs. These innovative QUS biomarkers will be very significant for monitoring severity of chronic ILDs and response to treatment, especially in this new era of miniaturized and highly portable ultrasound devices.

Prolonged Particulate Hexavalent Chromium Exposure Induces DNA Double-Strand Breaks and Inhibits Homologous Recombination Repair in Primary Rodent Lung Cells.

Hexavalent chromium [Cr(VI)] is a known lung carcinogen and a driving mechanism in human lung cells for Cr(VI)-induced lung cancer is chromosome instability, caused by prolonged Cr(VI) exposure inducing DNA double-strand breaks, while simultaneously inhibiting the repair of these breaks. In North Atlantic right whales, Cr(VI) induces breaks but does not inhibit repair. It is unclear if this repair inhibition is specific to human lung cells or occurs in other species, as it has only been considered in humans and North Atlantic right whales. We evaluated these outcomes in rodent cells, as rodents are an experimental model for metal-induced lung carcinogenesis. We used a guinea pig lung fibroblast cell line, JH4 Clone 1, and rat lung fibroblasts. Cells were exposed to two different particulate Cr(VI) compounds, ranging from 0 to 0.5 ug/cm2, for 24 or 120h and assessed for cytotoxicity, DNA double-strand breaks, and DNA double-strand break repair. Both particulate Cr(VI) compounds induced a concentration-dependent increase in cytotoxicity and DNA double-strand breaks after acute and prolonged exposures. Notably, while the repair of Cr(VI)-induced DNA double-strand breaks increased after acute exposure, the repair of these breaks was inhibited after prolonged exposure. These results are consistent with outcomes in human lung cells indicating rodent cells respond like human cells, while whale cells have a markedly different response.

Distribution and suitability of pulmonary surfactants as a vehicle for topically applied antibodies in healthy and SARS-CoV-2 infected rodent lungs

The use of natural pulmonary surfactants (PS) as a drug delivery vehicle for biologics is a more recent therapeutic modality. Herein, we tested different contents of PS regarding their physicochemical properties under stress conditions. The PS content of 12.25 mg/ml (Formulation B) showed desired properties such as an isotonic osmolality ∼300 mOsm/kg and an acceptable viscosity of 8.61 cSt, being lower than in commercially available PS solutions. Formulation B passed the specifications of surface lowering capacities of >80 % total lung capacity and physiologically desired formulation properties were independent of the antibody used in the composition. The identified formulation showed the capability of significantly increasing the oxygen saturation in ex vivo isolated perfused rat lungs, compared to a control and up to 30 min post lavage. In the in vivo setting, we showed that intratracheal administration of a human mAB with and without pulmonary surfactant led to higher amounts of delivered antibody within the alveolar tissue compared to intravenous administration. The antibody with the PS formulation remained longer in the alveolar tissues than the antibody without the PS formulation. Further, SARS-CoV-2 infected Golden Syrian hamsters showed that the intranasally applied antibody reached the site of infection in the alveoli and could be detected in the alveolar region 24 h after the last administration. With this work, we demonstrated that pulmonary surfactants can be used as a pulmonary drug delivery mechanism for antibodies and may subsequently improve the antibody efficacy by increasing the residence time at the desired site of action in the alveolar tissue.

Endothelial phosphodiesterase 4B inactivation ameliorates endothelial-to-mesenchymal transition and pulmonary hypertension

Pulmonary hypertension (PH) is a fatal disorder characterized by pulmonary vascular remodeling and obstruction. The phosphodiesterase 4 (PDE4) family hydrolyzes cyclic AMP (cAMP) and is comprised of four subtypes (PDE4A–D). Previous studies have shown the beneficial effects of pan-PDE4 inhibitors in rodent PH; however, this class of drugs is associated with side effects owing to the broad inhibition of all four PDE4 isozymes. Here, we demonstrate that PDE4B is the predominant PDE isozyme in lungs and that it was upregulated in rodent and human PH lung tissues. We also confirmed that PDE4B is mainly expressed in the lung endothelial cells (ECs). Evaluation of PH in Pde4b wild type and knockout mice confirmed that Pde4b is important for the vascular remodeling associated with PH. In vivo EC lineage tracing demonstrated that Pde4b induces PH development by driving endothelial-to-mesenchymal transition (EndMT), and mechanistic studies showed that Pde4b regulates EndMT by antagonizing the cAMP-dependent PKA–CREB–BMPRII axis. Finally, treating PH rats with a PDE4B-specific inhibitor validated that PDE4B inhibition has a significant pharmacological effect in the alleviation of PH. Collectively, our findings indicate a critical role for PDE4B in EndMT and PH, prompting further studies of PDE4B-specific inhibitors as a therapeutic strategy for PH.

The pulmonary effects of nickel-containing nanoparticles: Cytotoxicity, genotoxicity, carcinogenicity, and their underlying mechanisms.

With the exponential growth of the nanotechnology field, the global nanotechnology market is on an upward track with fast-growing jobs. Nickel (Ni)-containing nanoparticles (NPs), an important class of transition metal nanoparticles, have been extensively used in industrial and biomedical fields due to their unique nanostructural, physical, and chemical properties. Millions of people have been/are going to be exposed to Ni-containing NPs in occupational and non-occupational settings. Therefore, there are increasing concerns over the hazardous effects of Ni-containing NPs on health and the environment. The respiratory tract is a major portal of entry for Ni-containing NPs; thus, the adverse effects of Ni-containing NPs on the respiratory system, especially the lungs, have been a focus of scientific study. This review summarized previous studies, published before December 1, 2023, on cytotoxic, genotoxic, and carcinogenic effects of Ni-containing NPs on humans, lung cells in vitro, and rodent lungs in vivo, and the potential underlying mechanisms were also included. In addition, whether these adverse effects were induced by NPs themselves or Ni ions released from the NPs was also discussed. The extra-pulmonary effects of Ni-containing NPs were briefly mentioned. This review will provide us with a comprehensive view of the pulmonary effects of Ni-containing NPs and their underlying mechanisms, which will shed light on our future studies, including the urgency and necessity to produce engineering Ni-containing NPs with controlled and reduced toxicity, and also provide the scientific basis for developing nanoparticle exposure limits and policies.

Molecular study of the status of Angiostrongylus cantonensis in rats in Haiti

Angiostrongylus cantonensis, commonly known as the rat lungworm, causes Eosinophilic meningitis in humans. Our study aimed to investigate the prevalence and distribution of this parasite in rats in Haiti. Rats were trapped at 8 sites, 7 in Artibonite (rural region) and one in an urban area of Port-au-Prince. After euthanasia, hearts and lungs were sampled and preserved in 70% ethanol. Subsequently, the organs were dissected to detect adult worms. Parasite DNA was amplified using PCR targeting either the nematode ITS2 gene for rodent lung tissue or cox1 for isolated worms. Subsequent sequencing allowed parasite identification. A total of 70 rats were captured, i.e. 23 Rattus norvegicus and 47 Rattus rattus. Adult nematodes morphologically compatible with A. cantonensis were isolated from 5/70 rats (7%) and identification was confirmed by sequencing. Molecular analysis of lung tissue revealed a parasite prevalence of 31.4% (22/70), and its presence at 4 of the 8 sites investigated, including Port-au-Prince. The molecular approach on lung tissue targeting the ITS2 gene enabled us to detect a prevalence 4 times higher than the visual search for adult worms alone. Only one COX1 haplotype was identified, belonging to genotype II-G, widely distributed in Brazil, the French Antilles (Guadeloupe), French Polynesia, Hawaii, and Japan. These results confirm that A. cantonensis is an endemic parasite in Haiti not only in the capital Port-au-Prince, but also in several rural areas. Direct molecular screening for Angiostrongylus DNA in rat lung tissue showed higher sensitivity than visual detection of worms during dissection and could be useful for further prevalence studies.

Kynurenine pathway metabolism evolves with development of preclinical and scleroderma-associated pulmonary arterial hypertension.

Understanding metabolic evolution underlying pulmonary arterial hypertension (PAH) development may clarify pathobiology and reveal disease-specific biomarkers. Patients with systemic sclerosis (SSc) are regularly surveilled for PAH, presenting an opportunity to examine metabolic change as disease develops in an at-risk cohort. We performed mass spectrometry-based metabolomics on longitudinal serum samples collected before and near SSc-PAH diagnosis, compared with time-matched SSc subjects without PAH, in a SSc surveillance cohort. We validated metabolic differences in a second cohort and determined metabolite-phenotype relationships. In parallel, we performed serial metabolomic and hemodynamic assessments as the disease developed in a preclinical model. For differentially expressed metabolites, we investigated corresponding gene expression in human and rodent PAH lungs. Kynurenine and its ratio to tryptophan (kyn/trp) increased over the surveillance period in patients with SSc who developed PAH. Higher kyn/trp measured two years before diagnostic right heart catheterization increased the odds of SSc-PAH diagnosis (OR 1.57, 95% CI 1.05-2.36, P = 0.028). The slope of kyn/trp rise during SSc surveillance predicted PAH development and mortality. In both clinical and experimental PAH, higher kynurenine pathway metabolites correlated with adverse pulmonary vascular and RV measurements. In human and rodent PAH lungs, expression of TDO2, which encodes tryptophan 2,3 dioxygenase (TDO), a protein that catalyzes tryptophan conversion to kynurenine, was significantly upregulated and tightly correlated with pulmonary hypertensive features. Upregulated kynurenine pathway metabolism occurs early in PAH, localizes to the lung, and may be modulated by TDO2. Kynurenine pathway metabolites may be candidate PAH biomarkers and TDO warrants exploration as a potential novel therapeutic target.NEW & NOTEWORTHY Our study shows an early increase in kynurenine pathway metabolism in at-risk subjects with systemic sclerosis who develop pulmonary arterial hypertension (PAH). We show that kynurenine pathway upregulation precedes clinical diagnosis and that this metabolic shift is associated with increased disease severity and shorter survival times. We also show that gene expression of TDO2, an enzyme that generates kynurenine from tryptophan, rises with PAH development.

Discussion to: Gene delivery followed by ex vivo lung perfusion using an adeno-associated viral vector in a rodent lung transplant model

Discussion to: Gene delivery followed by ex vivo lung perfusion using an adeno-associated viral vector in a rodent lung transplant model

Gene delivery followed by ex vivo lung perfusion using an adeno-associated viral vector in a rodent lung transplant model

ObjectiveEx vivo lung perfusion has emerged as a platform for organ preservation, evaluation, and restoration. Gene delivery using a clinically relevant adeno-associated vector during ex vivo lung perfusion may be useful in optimizing donor allografts while the graft is maintained physiologically active. We evaluated the feasibility of adeno-associated vector–mediated gene delivery during ex vivo lung perfusion in a rat transplant model. Additionally, we assessed off-target effects and explored different routes of delivery. MethodsRat heart-lung blocks were procured and underwent 1-hour ex vivo lung perfusion. Before ex vivo lung perfusion, 4e11 viral genome luciferase encoding adeno-associated vector 9 was administered via the left bronchus (Br group, n = 4), via the left pulmonary artery (PA group, n = 3), or directly into the circuit (Circuit group, n = 3). Donor lungs in the Control group (n = 3) underwent ex vivo lung perfusion without adeno-associated vector 9. Only the left lung was transplanted. Animals underwent bioluminescence imaging weekly before being killed at 2 weeks. Tissues were collected for luciferase activity measurement. ResultsAll recipients tolerated the transplant well. At 2 weeks post-transplant, luciferase activity in the transplanted lung was significantly higher among animals in the Br group compared with the other 3 groups (Br: 1.1 × 106 RLU/g, PA: 8.3 × 104 RLU/g, Circuit: 3.8 × 103 RLU/g, Control: 2.5 × 103 RLU/g, P = .0003). No off-target transgene expression was observed. ConclusionsIn this work, we demonstrate that a clinically relevant adeno-associated vector 9 vector mediates gene transduction during ex vivo lung perfusion in rat lung grafts when administered via the airway and potentially the pulmonary artery. Our preliminary results suggest a higher transduction efficiency when adeno-associated vector 9 was delivered via the airway, and delivery during ex vivo lung perfusion reduces off-target effects after graft implant.

CNSC-03. EXPLORING STRUCTURAL DISRUPTION OF TRICELLULAR JUNCTIONS TO ENHANCE BLOOD-BRAIN-BARRIER PERMEABILITY

Abstract The blood-brain barrier (BBB) hinders CNS chemotherapy entry for malignant glioma treatment. It is predominantly composed of brain endothelium linked by bicellular (BJ) and tricellular (TJ) tight junctions. Previous studies demonstrated,angubindin-1 inhibited TJ angulin-1 to increase brain and CSF targeted therapy in rodent lung cancer models. We hypothesize angubindin-1 can transiently decrease BBB junctional integrity to increase chemotherapy permeability and prolong rodent glioma model survival. Rat brain endothelial cells were treated with TJ and BJ inhibitors against angulin-1 (angubindin-1 600µg/mL), claudin-3 (C-CPE 200µg/mL) or claudin-5 (C-CPE-MT 200µg/mL). Endothelial integrity was assessed by immunoblotting and cell-cell electrical impedance. Effects of angubindin-1 on efflux transporter P-glycoprotein (PGP) and migration were studied using rat and human derived glioma cells. Rat glioma models were treated with doxil (3 mg/kg), angubindin-1 (10 mg/kg or 30 mg/kg), or combination therapy for assessments on tumor volume, BBB permeability and survival. We observed decreased angulin-1 expression 5 hours after angubindin-1 treatment, with return to baseline by 24 hours (p<0.05). Angubindin-1, CCPE, and CCPE-mt globally reduced endothelial cell-cell integrity, maximally at 4 hours with a return to baseline by 12 hours; with angubindin-1 demonstrating the largest decreased cell-adhesion (angubindin-1 vs control, p< 0.0001). Angubindin-1 also decreased PGP efflux of rhodamine in both endothelial and glioma cells, along with demonstrating a pro-migratory dose-dependent effect on rat glioma cells. Combined angubindin-1 and doxil increased survival in rat glioma models compared with doxil alone (24 days vs. 18 days, p < 0.0001). Day 14 tumor volume was significantly decreased with angubindin-1 and combination treatment respectively (77.5 % vs 81.6 %, p<0.05). Ongoing studies are exploring, angubindin-1’s effect on large CNS drug permeability, TJ localization post BBB disruption and additional combinational treatments in glioma models. Collectively, these findings pose a unique opportunity to disrupt BBB integrity and improve malignant glioma treatment options.

Coccidioides undetected in soils from agricultural land and uncorrelated with time or the greater soil fungal community on undeveloped land.

Coccidioidomycosis is a typically respiratory fungal disease that, in the United States, occurs primarily in Arizona and California. In California, most coccidioidomycosis cases occur in the San Joaquin Valley, a primarily agricultural region where the disease poses a risk for outdoor workers. We collected 710 soil samples and 265 settled dust samples from nine sites in the San Joaquin Valley and examined how Coccidioides detection varied by month, site, and the presence and abundance of other fungal species. We detected Coccidioides in 89 of 238 (37.4%) rodent burrow soil samples at five undeveloped sites and were unable to detect Coccidioides in any of 472 surface and subsurface soil samples at four agricultural sites. In what is the largest sampling effort undertaken on agricultural land, our results provide no evidence that agricultural soils in the San Joaquin Valley harbor Coccidioides. We found no clear association between Coccidioides and the greater soil fungal community, but we identified 19 fungal indicator species that were significantly associated with Coccidioides detection in burrows. We also did not find a seasonal pattern in Coccidioides detection in the rodent burrow soils we sampled. These findings suggest both the presence of a spore bank and that coccidioidomycosis incidence may be more strongly associated with Coccidioides dispersal than Coccidioides growth. Finally, we were able to detect Coccidioides in only five of our 265 near-surface settled dust samples, one from agricultural land, where Coccidioides was undetected in soils, and four from undeveloped land, where Coccidioides was common in the rodent burrow soils we sampled. Our ability to detect Coccidioides in few settled dust samples indicates that improved methods are likely needed moving forward, though raises questions regarding aerial dispersal in Coccidioides, whose key transmission event likely occurs over short distances in rodent burrows from soil to naïve rodent lungs.

Differential Expression of ABC Transporter Genes in Brain Vessels vs. Peripheral Tissues and Vessels from Human, Mouse and Rat.

ATP-binding cassette (ABC) transporters comprise a superfamily of genes encoding membrane proteins with nucleotide-binding domains (NBD). These transporters, including drug efflux across the blood-brain barrier (BBB), carry a variety of substrates through plasma membranes against substrate gradients, fueled by hydrolyzing ATP. The expression patterns/enrichment of ABC transporter genes in brain microvessels compared to peripheral vessels and tissues are largely uncharacterized. In this study, the expression patterns of ABC transporter genes in brain microvessels, peripheral tissues (lung, liver and spleen) and lung vessels were investigated using RNA-seq and WesTM analyses in three species: human, mouse and rat. The study demonstrated that ABC drug efflux transporter genes (including ABCB1, ABCG2, ABCC4 and ABCC5) were highly expressed in isolated brain microvessels in all three species studied; the expression of ABCB1, ABCG2, ABCC1, ABCC4 and ABCC5 was generally higher in rodent brain microvessels compared to those of humans. In contrast, ABCC2 and ABCC3 expression was low in brain microvessels, but high in rodent liver and lung vessels. Overall, most ABC transporters (with the exception of drug efflux transporters) were enriched in peripheral tissues compared to brain microvessels in humans, while in rodent species, additional ABC transporters were found to be enriched in brain microvessels. This study furthers the understanding of species similarities and differences in the expression patterns of ABC transporter genes; this is important for translational studies in drug development. In particular, CNS drug delivery and toxicity may vary among species depending on their unique profiles of ABC transporter expression in brain microvessels and BBB.

Aldo-Keto Reductase 1C15 Characterization and Protection in Ischemic Brain Injury.

Aldo-keto reductase (AKR) 1C15, a member of the AKR superfamily, was recently identified and cloned, and reported to alleviate oxidative stress in endothelial cells in rodent lungs. However, its expression and role in the brain and ischemic brain diseases have not been investigated. AKR1C15 expression was detected with real-time PCR. Mouse ischemic stroke and ischemic preconditioning (IPC) were established with middle cerebral artery occlusion (MCAO) for 1 h or 12 min, respectively. Recombinant AKR1C15 was administered intraperitoneally, and stroke outcome was evaluated with neurobehavioral tests and infarct volumes. Rat primary brain cell cultures were subjected to oxygen-glucose deprivation (OGD) to mimic ischemic injury. Cell survival or in vitro blood-brain barrier (BBB) permeability was measured, and nitric oxide (NO) release was detected. Immunostaining and Western blotting were used to evaluate oxidative-stress-related protein expression. AKR1C15 administration decreased the infarct volume and neurological deficits 2d post-stroke, and its early (1-h) administration after IPC abolished the protection of IPC against stroke. In rat primary brain cell cultures, AKR1C15 was most abundantly expressed in brain microvascular endothelial cells (BMVECs) and microglia. Its expression decreased upon OGD in most cell types except for BMVECs and microglia. In primary neuronal cultures, AKR1C15 treatment prevented OGD-induced cell death accompanied by decreased levels of 4-hydroxynonenal, 8-hydroxy-2'-deoxyguanosine, and heme oxygenase-1. In BMVEC cultures, AKR1C15 treatment protected against OGD-induced cell death and in vitro BBB leakage. In primary microglial cultures, AKR1C15 reduced the release of NO upon proinflammatory stimulation. Our results provide a characterization of the novel antioxidant AKR1C15 and demonstrate its protective role against ischemic injury, both in vivo and in vitro. AKR1C15 may be a promising agent for ischemic stroke treatment.

Vascular-Parenchymal Cross-Talk Promotes Lung Fibrosis through BMPR2 Signaling.

Rationale: Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease characterized by progressive lung scarring. IPF-related pulmonary vascular remodeling and pulmonary hypertension (PH) result in a particularly poor prognosis. Objectives: To study the pathogenesis of vascular remodeling in fibrotic lungs and its contribution to progression of fibrosis. Methods: We used an experimental model of lung fibrosis associated with PH by transient overexpression of active TGF-β1 (transforming growth factor-β1). Samples from patients with fibrotic lung diseases were analyzed in depth using immunostaining, gene expression, and gene mutations. Measurements and Main Results: We found a reduction in endothelial cells (ECs) and activation of vascular smooth muscle cells (VSMCs) in fibrotic lungs. Coculturing fibroblasts with VSMCs or ECs from fibrotic lungs induced fibrotic phenotypes in fibroblasts. IPF fibroblasts induced EC death and activation of VSMCs in coculture systems. Decreased concentrations of BMPR2 (bone morphogenic protein receptor 2) and its signaling were observed in ECs and VSMCs from fibrotic lungs in both rats and humans. On fibroblasts treated with media from VSMCs, BMPR2 suppression in VSMCs led to fibrogenic effects. Tacrolimus activated BMPR2 signaling and attenuated fibrosis and PH in rodent lungs. Whole-exome sequencing revealed rare mutations in PH-related genes, including BMPR2, in patients with IPF undergoing transplantation. A unique missense BMPR2 mutation (p.Q721R) was discovered to have dysfunctional effects on BMPR2 signaling. Conclusions: Endothelial dysfunction and vascular remodeling in PH secondary to pulmonary fibrosis enhance fibrogenesis through impaired BMPR2 signaling. Tacrolimus may have value as a treatment of advanced IPF and concomitant PH. Genetic abnormalities may determine the development of PH in advanced IPF.

Random matrix theory to quantify micro-structural changes in rodent lungs due to pulmonary diseases

We exploit the random matrix theory to detect changes in rodent lungs exhibiting pulmonary fibrosis and edema. Coherences in the backscattered signals are stronger when single scattering dominates (fibrosis/edema). On the contrary, healthy lungs exhibit more apparent randomness due to multiple scattering. This leads to differences in the distribution of eigenvalues, which can be retrieved using Singular Value Decomposition of the Inter-elementResponse Matrix (IRM). We use features of the eigenvalue distribution (E(x), the expected value, and, the eigenvalue with the highest probability) to quantify changes in lung parenchyma and investigate whether they can improve the specificity of quantitative ultrasound to lung diseases. IRMs were acquired from 51 rat lungs (10 controls, 18 edematous, 17 fibrotic, 6 fibrotic rats, which were treated with Nintedanib) using a 128-element linear array (Verasonics L11-4v, 7.8 MHz). Severity of fibrosis and edema were quantified by histology and the ratio of wet to dry weight. Both parameters showed significant differences between edematous and fibrotic lungs, and between control and fibrotic lungs, which was significantly correlated to both the severity of fibrosis and edema. E(x) was significantly correlated to the severity of fibrosis. This suggests that these parameters could be part of a toolkit for the quantitative assessment of lung diseases.

Random matrix theory (RMT) to quantify scattering behavior in lung mimicking phantoms

As we previously reported in rodent lungs, RMT parameters (Expected value E(x), and λmax, the eigenvalue with the highest probability) extracted from singular value decomposition (SVD) of inter-element response matrix (IRM) show a significant correlation with fibrosis histology scores. The lack of fibrotic models for larger lungs such as pigs motivated us to investigate porous 3D printed PEGDA phantoms with controllable strut size and alveolar density. We hypothesize that E(x), and λmax can distinguish phantoms with different alveolar size, by evaluating multiple scattering in the backscattered signals. IRMs were acquired using a 128-element linear array L7-4 (Verasonics, at 5.2 MHz central frequency) connected to a Verasonics Vantage ultrasound scanner. Phantoms of 1-inches size with different strut diameters of 0.085, 0.17, and 0.26 mm were used. Attenuation constants for these samples were measured using the Substitution Method at 1 MHz, 2.25MHz, and 5 MHz to evaluate scattering attenuation. E(x), and λmax were evaluated using SVD of the IRM. Attenuation constants, E(x), and λmax all show that the phantoms with larger strut size exhibit more multiple scattering than phantoms with smaller strut size. These preliminary results suggest that such phantoms could be used to mimic pulmonary fibrosis.

Low 15d-PGJ2 status is associated with oxidative stress in chronic obstructive pulmonary disease patients.

Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-dependent nuclear receptor and highly expressed in human and rodent lungs. 15-Deoxy-delta-12,14-prostaglandin J2 (15d-PGJ2), known for cyclopentenone prostaglandin, is the endogenous ligand of PPARγ. However, the associations among PPARγ, 15d-PGJ2 and chronic obstructive pulmonary disease (COPD) were unclear. All 130 fasting blood samples and 40 lung specimens were obtained from COPD patients and control subjects. Serum 15d-PGJ2 was detected by ELISA. The expressions of oxidative stress indicators were measured using western blotting and PPARγ nuclei were evaluated with immunohistochemistry in lungs. The associations among serum 15d-PGJ2, pulmonary PPARγ and oxidative stress indicators, and COPD were estimated. Serum 15d-PGJ2 was reduced in COPD patients compared with healthy volunteers. Linear and logistic regression analysis indicated that serum 15d-PGJ2 was positively associated with pulmonary function in COPD patients. In addition, PPARγ-positive nuclei were reduced and oxidative stress indicators, included HO-1 and NOX-4, were increased in lungs of COPD patients. Further correlative analysis suggested that pulmonary function parameters was positively correlated with serum 15d-PGJ2 and pulmonary PPARγ-positive nuclei, inversely related to oxidative stress indicators in lungs of COPD patients. Pretreatment with 15d-PGJ2 obviously attenuated TNFα-induced oxidative stress in BEAS-2B cells. Serum 15d-PGJ2 and pulmonary PPARγ are reduced, and oxidative stress is elevated in COPD patients. Serum 15d-PGJ2 is inversely associated with oxidative stress in COPD patients.

Using random matrix theory to quantify pulmonary fibrosis: Investigating the effect of time window duration

Random matrix theory (RMT) exploits the distribution of singular values of the inter-element response matrix (IRM). If multiple scattering dominates the propagation, the singular value distribution follows the quarter circle law. However, dominance of single scattering results in Henkel function behavior of the singular value distribution. In our previous work, we have shown that this can be exploited to estimate severity of bleomycin-induced fibrosis (measured by histology) in rodent lungs. We showed that E(x), the expected value of the singular value distribution, as well as the singular value with the highest probability, correlated significantly with histology scores. Here, we investigate the sensitivity of these metrics to the time window duration used for the Singular Value Decomposition of the IRM, which is performed in the frequency domain, using overlapping time windows. A linear transducer with a central frequency of 7.8 MHz and a Verasonics scanner were used to obtain IRMs in 24 rat lungs. Different degrees of pulmonary fibrosis were induced using bleomycin in 18 rats while 6 rats were left as controls. The IRMs were time-windowed with different duration of 2T, 4T, and 6T where T is the transmitted pulse period. E(x) and were evaluated for each time window duration for all rat lungs. For all time window durations significant correlations were observed between and E(x), and histology scores. Wilcoxon ranksum tests show that the distributions obtained for and E(x) are not affected by the time window duration.