Impaired Wound Healing of Alveolar Lung Epithelial Cells in a Breathing Lung-On-A-Chip

The lung alveolar region experiences remodeling during several acute and chronic lung diseases, as for instance idiopathic pulmonary fibrosis (IPF), a fatal disease, whose onset is correlated with repetitive microinjuries to the lung alveolar epithelium and abnormal alveolar wound repair. Although a high degree of mechanical stress (>20% linear strain) is thought to potentially induce IPF, the effect of lower, physiological levels of strain (5–12% linear strain) on IPF pathophysiology remains unknown. In this study, we examined the influence of mechanical strain on alveolar epithelial wound healing. For this purpose, we adopted the “organ-on-a-chip” approach, which provides the possibility of reproducing unique aspects of the in vivo cellular microenvironment, in particular its dynamic nature. Our results provide the first demonstration that a wound healing assay can be performed on a breathing lung-on-a-chip equipped with an ultra-thin elastic membrane. We cultured lung alveolar epithelial cells to confluence, the cells were starved for 24 h, and then wounded by scratching with a standard micropipette tip. Wound healing was assessed after 24 h under different concentrations of recombinant human hepatic growth factor (rhHGF) and the application of cyclic mechanical stretch. Physiological cyclic mechanical stretch (10% linear strain, 0.2 Hz) significantly impaired the alveolar epithelial wound healing process relative to culture in static conditions. This impairment could be partially ameliorated by administration of rhHGF. This proof-of-concept study provides a way to study of more complex interactions, such as a co-culture with fibroblasts, endothelial cells, or immune cells, as well as the study of wound healing at an air–liquid interface

The potential of microfluidic lung epithelial wounding: towards in vivo-like alveolar microinjuries

Idiopathic pulmonary fibrosis (IPF) remains a major clinical challenge to date. Repeated alveolar epithelial microinjuries are considered as the starting point and the key event in both the development and the progression of IPF. Various pro-fibrotic agents have been identified and shown to cause alveolar damage. In IPF, however, no leading cause of alveolar epithelial microinjuries can be identified and the exact etiology remains elusive. New results from epidemiologic studies suggest a causal relation between IPF and frequent episodes of gastric refluxes resulting in gastric microaspirations into the lung. The effect of gastric contents on the alveolar epithelium has not been investigated in detail. Here, we present a microfluidic lung epithelial wounding system that allows for the selective exposure of alveolar epithelial cells to gastric contents. The system is revealed to be robust and highly reproducible. The thereby created epithelial microwounds are of tiny dimensions and best possibly reproduce alveolar damage in the lung. We further demonstrate that exposure to gastric contents, namely hydrochloric acid (HCl) and pepsin, directly damages the alveolar epithelium. Together, this novel in vitro wounding system allows for the creation of in vivo-like alveolar microinjuries with the potential to study lung injury and alveolar wound repair in vitro

Evaluation of an Animal Warning System Effectiveness

The problem of vehicle/animal crashes is being addressed in this research. There have been a few new technologies that claim to accurately detect the large animals that cross our roadways. Each one has its own strengths and shortcomings. A close attention must be given to the selected site and the technology deployed based on its weather, vegetation, topography, and local animal types and sizes. In this project, we have reviewed a number of animal detection systems and selected one system with the most potential to serve the characteristics of the selected site and the local Deer. We did a preliminary test of the reliability of this system in a testbed in Lewistown, Montana. The results were encouraging. We also carefully selected a site that we felt could benefit the most from this safety improvement based on its physical and climatic characteristics as well as its high number of vehicle/animal crashes. We also designed and developed a data monitoring and recording system that records and archives the response of the driver to our designed animal warning signs. This system incorporates radars, video cameras, communication links, and computer hardware and software. In the next phase of this project, we will analyze the effectiveness of our entire system by analyzing the driver’s response to the animal warning signs. We will also continue our evaluation of reliability of the selected animal detection system both in study site and in Lewistown’s testbed

Microimpedance tomography system to monitor cell activity and membrane movements in a breathing lung-on-chip

We report about a new microimpedance tomography (MITO) system integrated in a lung-on-chip that recapitulates the thin alveolar barrier including the cyclic mechanical strain of the breathing movements. The system enables to detect changes that take place in the lung alveolar barrier located at 1 mm from the detection system. This leaves space for the three-dimensional deflection of the lung alveolar barrier. The aim of the MITO is to monitor both the electrochemical and the mechanical changes occurring in the lung alveolar barrier using impedimetric coplanar electrodes. Distant and real-time monitoring of changes in the resistivity of a human lung epithelial cell monolayer challenged with Triton X-100 could easily be detected. An exponential drop of 7% in impedance magnitude was recorded following the permeabilization of the monolayer. While the membrane is deflected to mimic the respiratory movements, the impedance readout can be correlated to the mechanical strain in the alveolar barrier. Small variations of the mechanical strain due to the density of the cell population can be detected. A mechanical strain difference of 0.4% was monitored between epithelial cells just seeded on the alveolar membrane and the resulting confluent layer 24 h later. The system is produced using a flexible printed circuit board (PCB) bonded to the lung-on-chip device made of polydimethylsiloxane (PDMS). It brings impedance-based cell and organ function monitoring onto organs-on-chips on a single, cost-efficient and integrable layer that does not interfere with the biomimetic capabilities of the chip

Lung on Chip: In vitro HGF effects on injured alveolar A549 epithelial cells in microfluidic system

Microfluidic systems have become competitive tools in the invitro modelling of diseases and promising alternatives to animal studies. They allow obtaining more invivo like conditions for cellular assays. Research in idiopathic pulmonary fibrosis could benefit from this novel methodological approach to understand the pathophysiology of the disease & develop efficient therapies. The use of hepatocyte growth factor (HGF) for alveolar reepithelisation is a promising approach. In this study, we show a new microfluidic system to analyse the effects of HGF on injured alveolar epithelial cells. Microfluidic systems in polydimethylsiloxane were fabricated by soft lithography. The alveolar A549 epithelial cells (10,000 cells) were seeded and studied in these microfluidic systems with media perfusion (1μl/30min). Injury tests were made on the cells by the perfusion with media containing H2O2 or bleomycin. The degree of injury was then assessed by a metabolic and an apoptotic assays. Wound assays were also performed with a central laminar flow of trypsin. Monitoring of wound closure with HGF vs control media was assessed. The alveolar A549 epithelial cells grew and proliferated in the microfluidic system. In the wound closure assay, the degree of wound closure after 5 hours was (53.3±1.3%) with HGF compared to (9.8±2.4%) without HGF (P <0.001). We present a novel microfluidic model that allows culture, injury and wounding of A549 epithelial cells and represents the first step towards the development of an invitro reconstitution of the alveolar-capillary interface. We were also able to confirm that HGF increased alveolar epithelial repair in this system

Microfluidic wound-healing assay to assess the regenerative effect of HGF on wounded alveolar epithelium

We present a microfluidic epithelial wound-healing assay that allows characterization of the effect of hepatocyte growth factor (HGF) on the regeneration of alveolar epithelium using a flow-focusing technique to create a regular wound in the epithelial monolayer. The phenotype of the epithelial cell was characterized using immunostaining for tight junction (TJ) proteins and transmission electron micrographs (TEMs) of cells cultured in the microfluidic system, a technique that is reported here for the first time. We demonstrate that alveolar epithelial cells cultured in a microfluidic environment preserve their phenotype before and after wounding. In addition, we report a wound-healing benefit induced by addition of HGF to the cell culture medium (19.2 vs. 13.5 μm h(-1) healing rate)

Intravenous iron alone resolves anemia in patients with functional iron deficiency and lymphoid malignancies undergoing chemotherapy

This randomized trial evaluated ferric carboxymaltose without erythropoiesis-stimulating agents (ESA) for correction of anemia in cancer patients with functional iron deficiency. Patients on treatment for indolent lymphoid malignancies, who had anemia [hemoglobin (Hb) 8.5-10.5 g/dL] and functional iron deficiency [transferrin saturation (TSAT) &lt;= 20 %, ferritin &gt;30 ng/mL (women) or &gt;40 ng/mL (men)], were randomized to ferric carboxymaltose (1,000 mg iron) or control. Primary end point was the mean change in Hb from baseline to weeks 4, 6 and 8 without transfusions or ESA. Difficulties with patient recruitment led to premature termination of the study. Seventeen patients (8 ferric carboxymaltose and 9 control) were included in the analysis. In the ferric carboxymaltose arm, mean Hb increase was significantly higher versus control at week 8 (p = 0.021). All ferric carboxymaltose- treated patients achieved an Hb increase &gt;1 g/dL (control 6/9; p = 0.087), and mean TSAT was &gt;20 % from week 2 onwards. No treatment-related adverse events were reported. In conclusion, ferric carboxymaltose without ESA effectively increased Hb and iron status in this small patient population