Alveolus analysis: a web browser-based tool to analyze lung intravital microscopy.

BACKGROUND: Acute lung injury and the acute respiratory distress syndrome are characterized by pulmonary inflammation, reduced endothelial barrier integrity and filling of the alveolar space with protein rich edema fluid and infiltrating leukocytes. Animal models are critical to uncovering the pathologic mechanisms of this devastating syndrome. Intravital imaging of the intact lung via two-photon intravital microscopy has proven a valuable method to investigate lung injury in small rodent models through characterization of inflammatory cells and vascular changes in real time. However, respiratory motion complicates the analysis of these time series images and requires selective data extraction to stabilize the image. Consequently, analysis of individual alveoli may not provide a complete picture of the integrated mechanical, vascular and inflammatory processes occurring simultaneously in the intact lung. To address these challenges, we developed a web browser-based visualization application named Alveolus Analysis to process, analyze and graphically display intravital lung microscopy data. RESULTS: The designed tool takes raw temporal image data as input, performs image preprocessing and feature extraction offline, and visualizes the extracted information in a web browser-based interface. The interface allows users to explore multiple experiments in three panels corresponding to different levels of detail: summary statistics of alveolar/neutrophil behavior, characterization of alveolar dynamics including lung edema and inflammatory cells at specific time points, and cross-experiment analysis. We performed a case study on the utility of the visualization with two members or our research team and they found the tool useful because of its ability to preprocess data consistently and visualize information in a digestible and informative format. CONCLUSIONS: Application of our software tool, Alveolus Analysis, to intravital lung microscopy data has the potential to enhance the information gained from these experiments and provide new insights into the pathologic mechanisms of inflammatory lung injury.

Group V Phospholipase A2 Mediates Endothelial Dysfunction and Acute Lung Injury Caused by Methicillin-Resistant Staphylococcus Aureus.

Lung endothelial dysfunction is a key feature of acute lung injury (ALI) and clinical acute respiratory distress syndrome (ARDS). Previous studies have identified the lipid-generating enzyme, group V phospholipase A2 (gVPLA2), as a mediator of lung endothelial barrier disruption and inflammation. The current study aimed to determine the role of gVPLA2 in mediating lung endothelial responses to methicillin-resistant Staphylococcus aureus (MRSA, USA300 strain), a major cause of ALI/ARDS. In vitro studies assessed the effects of gVPLA2 inhibition on lung endothelial cell (EC) permeability after exposure to heat-killed (HK) MRSA. In vivo studies assessed the effects of intratracheal live or HK-MRSA on multiple indices of ALI in wild-type (WT) and gVPLA2-deficient (KO) mice. In vitro, HK-MRSA increased gVPLA2 expression and permeability in human lung EC. Inhibition of gVPLA2 with either the PLA2 inhibitor, LY311727, or with a specific monoclonal antibody, attenuated the barrier disruption caused by HK-MRSA. LY311727 also reduced HK-MRSA-induced permeability in mouse lung EC isolated from WT but not gVPLA2-KO mice. In vivo, live MRSA caused significantly less ALI in gVPLA2 KO mice compared to WT, findings confirmed by intravital microscopy assessment in HK-MRSA-treated mice. After targeted delivery of gVPLA2 plasmid to lung endothelium using ACE antibody-conjugated liposomes, MRSA-induced ALI was significantly increased in gVPLA2-KO mice, indicating that lung endothelial expression of gVPLA2 is critical in vivo. In summary, these results demonstrate an important role for gVPLA2 in mediating MRSA-induced lung EC permeability and ALI. Thus, gVPLA2 may represent a novel therapeutic target in ALI/ARDS caused by bacterial infection.

Secretory Phospholipase A2 Enzymes in Acute Lung Injury.

The secretory phospholipase A2 (sPLA2) group of secreted enzymes hydrolyze phospholipids and lead to the production of multiple biologically active lipid mediators. sPLA2s and their products (e.g., eicosanoids) play a significant role in the pathophysiology of various inflammatory diseases, including life-threatening lung disorders such as acute lung injury (ALI) and the Acute Respiratory Distress Syndrome (ARDS). The ALI/ARDS spectrum of severe inflammatory conditions is caused by direct (such as bacterial or viral pneumonia) or indirect insults (sepsis) that are associated with high morbidity and mortality. Several sPLA2 isoforms are upregulated in patients with ARDS as well as in multiple ALI preclinical models, and individual sPLA2s exert unique roles in regulating ALI pathophysiology. This brief review will summarize the contributions of specific sPLA2 isoforms as markers and mediators in ALI, supporting a potential therapeutic role for targeting them in ARDS.

Cortical Actin Dynamics in Endothelial Permeability.

The pulmonary endothelial cell forms a critical semi-permeable barrier between the vascular and interstitial space. As part of the blood-gas barrier in the lung, the endothelium plays a key role in normal physiologic function and pathologic disease. Changes in endothelial cell shape, defined by its plasma membrane, determine barrier integrity. A number of key cytoskeletal regulatory and effector proteins including non-muscle myosin light chain kinase, cortactin, and Arp 2/3 mediate actin rearrangements to form cortical and membrane associated structures in response to barrier enhancing stimuli. These actin formations support and interact with junctional complexes and exert forces to protrude the lipid membrane to and close gaps between individual cells. The current knowledge of these cytoskeletal processes and regulatory proteins are the subject of this review. In addition, we explore novel advancements in cellular imaging that are poised to shed light on the complex nature of pulmonary endothelial permeability.

DPP4 inhibition by sitagliptin attenuates LPS-induced lung injury in mice.

Acute respiratory distress syndrome (ARDS) is a severe clinical condition marked by acute respiratory failure and dysregulated inflammation. Pulmonary vascular endothelial cells (PVECs) function as an important pro-inflammatory source in ARDS, suggesting that modulation of inflammatory events at the endothelial level may have a therapeutic benefit. Dipeptidyl peptidase-4 (DPP4) inhibitors, widely used for the treatment of diabetes mellitus, have been reported to have possible anti-inflammatory effects. However, the potential anti-inflammatory effects of DPP4 inhibition on PVEC function and ARDS pathophysiology are unknown. Therefore, we evaluated the effects of sitagliptin, a DPP4 inhibitor in wide clinical use, on LPS-induced lung injury in mice and in human lung ECs in vitro. In vivo, sitagliptin reduced serum DPP4 activity, bronchoalveolar lavage protein concentration, cell number, and proinflammatory cytokine levels after LPS and alleviated histological findings of lung injury. LPS decreased the expression levels of CD26/DPP4 on pulmonary epithelial cells and PVECs isolated from mouse lungs, and the effect was partially reversed by sitagliptin. In vitro, human lung microvascular ECs (HLMVECs) expressed higher levels of CD26/DPP4 than human pulmonary arterial ECs. LPS induced the release of TNFα, IL-6, and IL-8 by HLMVECs that were inhibited by sitagliptin. LPS promoted the proliferation of HLMVECs, and sitagliptin suppressed this response. However, sitagliptin failed to reverse LPS-induced permeability in cultured ECs or lung epithelial cells in vitro. In summary, sitagliptin attenuates LPS-induced lung injury in mice and exerts anti-inflammatory effects on HLMVECs. These novel observations indicate DPP4 inhibitors may have potential as therapeutic drugs for ARDS.

A Review of Quality of Life Measures used in Surgical Outcomes for Stage I Lung Cancers.

This review summarizes the literature on QoL in early stage lung cancer patients who underwent surgery. PubMed and PsycINFO were searched. Twelve articles from 10 distinct studies were identified for a total of 992 patients. Five QoL measures were used. One study reported only on pre-surgical QoL, six only on post-surgical QoL and three studies reported on both pre- and post-surgical QoL. Timing for the administration of post-surgical QoL surveys varied. The literature on QoL in Stage I non-small-cell lung cancer patients is very sparse. Additional research is needed to explore the impact of different surgical approaches on QoL.