Determination of reliable lung function parameters in intubated mice.

BACKGROUND: Animal models and, in particular, mice models, are important tools to investigate the pathogenesis of respiratory diseases and to test potential new therapeutic drugs. Lung function measurement is a key step in such investigation. In mice, it is usually performed using forced oscillation technique (FOT), negative pressure-driven forced expiratory (NPFE) and pressure-volume (PV) curve maneuvers. However, these techniques require a tracheostomy, which therefore only allows end-point measurements. Orotracheal intubation has been reported to be feasible and to give reproducible lung function measurements, but the agreement between intubation and tracheostomy generated-data remains to be tested. METHODS: Using the Flexivent system, we measured lung function parameters (in particular, forced vital capacity (FVC), forced expiratory volume in the first 0.1 s (FEV0.1), compliance (Crs) of the respiratory system, compliance (C) measured using PV loop and an estimate of inspiratory capacity (A)) in healthy intubated BALB/cJ mice and C57BL/6 J mice and compared the results with similar measurements performed in the same mice subsequently tracheostomized after intubation, by means of paired comparison method, correlation and Bland-Altman analysis. The feasibility of repetitive lung function measurements by intubation was also tested. RESULTS: We identified parameters that are accurately evaluated in intubated animals (i.e., FVC, FEV0.1, Crs, C and A in BALB/cJ and FVC, FEV0.1, and A in C57BL/6 J). Repetitive lung function measurements were obtained in C57BL/6 J mice. CONCLUSION: This subset of lung function parameters in orotracheally intubated mice is reliable, thereby allowing relevant longitudinal studies.

Air-liquid interface exposure to aerosols of poorly soluble nanomaterials induces different biological activation levels compared to exposure to suspensions.

BACKGROUND: Recently, much progress has been made to develop more physiologic in vitro models of the respiratory system and improve in vitro simulation of particle exposure through inhalation. Nevertheless, the field of nanotoxicology still suffers from a lack of relevant in vitro models and exposure methods to predict accurately the effects observed in vivo, especially after respiratory exposure. In this context, the aim of our study was to evaluate if exposing pulmonary cells at the air-liquid interface to aerosols of inhalable and poorly soluble nanomaterials generates different toxicity patterns and/or biological activation levels compared to classic submerged exposures to suspensions. Three nano-TiO2 and one nano-CeO2 were used. An exposure system was set up using VitroCell® devices to expose pulmonary cells at the air-liquid interface to aerosols. A549 alveolar cells in monocultures or in co-cultures with THP-1 macrophages were exposed to aerosols in inserts or to suspensions in inserts and in plates. Submerged exposures in inserts were performed, using similar culture conditions and exposure kinetics to the air-liquid interface, to provide accurate comparisons between the methods. Exposure in plates using classical culture and exposure conditions was performed to provide comparable results with classical submerged exposure studies. The biological activity of the cells (inflammation, cell viability, oxidative stress) was assessed at 24 h and comparisons of the nanomaterial toxicities between exposure methods were performed. RESULTS: Deposited doses of nanomaterials achieved using our aerosol exposure system were sufficient to observe adverse effects. Co-cultures were more sensitive than monocultures and biological responses were usually observed at lower doses at the air-liquid interface than in submerged conditions. Nevertheless, the general ranking of the nanomaterials according to their toxicity was similar across the different exposure methods used. CONCLUSIONS: We showed that exposure of cells at the air-liquid interface represents a valid and sensitive method to assess the toxicity of several poorly soluble nanomaterials. We underlined the importance of the cellular model used and offer the possibility to deal with low deposition doses by using more sensitive and physiologic cellular models. This brings perspectives towards the use of relevant in vitro methods of exposure to assess nanomaterial toxicity.

Randomized study of antiseptic application technique in healthy volunteers before vascular access insertion (TApAS trial).

BACKGROUND: In France about 32% of hospitalized patients have a vascular access placement. However, a common complication associated with these is catheter-related bloodstream infection (CRBI) due to the introduction of microorganisms from the skin during catheter insertion. There is no consensus on the best way to clean the skin prior to catheter insertion, which could be a key element of CRBI prevention. The two techniques most commonly used to apply antiseptic to the skin are the concentric circle and back-and-forth techniques, but these have not been compared in clinical trials. Hence, this study conducted this comparison. METHODS: This single-center, non-comparative, randomized, matched pilot study investigated the levels of cutaneous microorganisms before and after antiseptic application using both techniques in a population of healthy French volunteers. The two application methods were used on each participant's arms at the elbow fold, with randomization for the application side (right or left). Quantification of cutaneous microorganisms was performed in a blinded manner with regard to the technique used. FINDINGS: From April 8 to July 17, 2019, 132 healthy volunteers participated in the study. For the whole study population, the mean initial colonization level was 2.68 log10 colony forming units (CFU)/mL (SD 0.82) before the back-and-forth technique, and 2.66 log10 CFU/mL (SD 0.85) before the concentric circle technique. The mean differences in number of microorganisms between the initial sample and the final sample were 2.45 log10 CFU/mL (95% CI: 2.29 to 2.61) for the back-and-forth technique and 2.43 log10 CFU/mL (95% CI: 2.27 to 2.59) for the concentric circle technique. The mean difference in reduction in microorganisms between the back-and-forth technique and the concentric circle technique was 0.02 log10 CFU/mL (95% CI: -0.11 to 0.15). INTERPRETATION: There was no clinically difference in reduction of microorganisms between the concentric circle and back-and-forth techniques at the bend of the healthy volunteer's elbow, after the 30 s of drying of the antiseptic. These findings have a significant impact on time required to achieve antiseptic application before catheter insertion because there is yet no argument to justify application for 30 s, because a single concentric circle pass was much faster with similar results. Future studies should investigate the impact of skin application technique on the prevention of infectious risk associated with catheter insertion on admission to health care facilities (conventional, outpatient, or emergency) and throughout the period of stay in a health care facility.

ELA/APELA precursor cleaved by furin displays tumor suppressor function in renal cell carcinoma through mTORC1 activation.

Apelin is a well-established mediator of survival and mitogenic signaling through the apelin receptor (Aplnr) and has been implicated in various cancers; however, little is known regarding Elabela (ELA/APELA) signaling, also mediated by Aplnr, and its role and the role of the conversion of its precursor proELA into mature ELA in cancer are unknown. Here, we identified a function of mTORC1 signaling as an essential mediator of ELA that repressed kidney tumor cell growth, migration, and survival. Moreover, sunitinib and ELA showed a synergistic effect in repressing tumor growth and angiogenesis in mice. The use of site-directed mutagenesis and pharmacological experiments provided evidence that the alteration of the cleavage site of proELA by furin induced improved ELA antitumorigenic activity. Finally, a cohort of tumors and public data sets revealed that ELA was only repressed in the main human kidney cancer subtypes, namely clear cell, papillary, and chromophobe renal cell carcinoma. Aplnr was expressed by various kidney cells, whereas ELA was generally expressed by epithelial cells. Collectively, these results showed the tumor-suppressive role of mTORC1 signaling mediated by ELA and established the potential use of ELA or derivatives in kidney cancer treatment.

Inactivation of Proprotein Convertases in T Cells Inhibits PD-1 Expression and Creates a Favorable Immune Microenvironment in Colorectal Cancer.

Proprotein convertases (PC) activate precursor proteins that play crucial roles in various cancers. In this study, we investigated whether PC enzyme activity is required for expression of the checkpoint protein programmed cell death protein 1 (PD-1) on cytotoxic T lymphocytes (CTL) in colon cancer. Although altered expression of the PC secretory pathway was observed in human colon cancers, only furin showed highly diffuse expression throughout the tumors. Inhibition of PCs in T cells using the general protein-based inhibitor α1-PDX or the pharmacologic inhibitor Decanoyl-Arg-Val-Lys-Arg-chloromethylketone repressed PD-1 and exhausted CTLs via induction of T-cell proliferation and apoptosis inhibition, which improved CTL efficacy against microsatellite instable and microsatellite stable colon cancer cells. In vivo, inhibition of PCs enhanced CTL infiltration in colorectal tumors and increased tumor clearance in syngeneic mice compared with immunodeficient mice. Inhibition of PCs repressed PD-1 expression by blocking proteolytic maturation of the Notch precursor, inhibiting calcium/NFAT and NF-κB signaling, and enhancing ERK activation. These findings define a key role for PCs in regulating PD-1 expression and suggest targeting PCs as an adjunct approach to colorectal tumor immunotherapy. SIGNIFICANCE: Protein convertase enzymatic activity is required for PD-1 expression on T cells, and inhibition of protein convertase improves T-cell targeting of microsatellite instable and stable colorectal cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/79/19/5008/F1.large.jpg.

Study of TiO2 P25 Nanoparticles Genotoxicity on Lung, Blood, and Liver Cells in Lung Overload and Non-Overload Conditions After Repeated Respiratory Exposure in Rats.

Inhaled titanium dioxide (TiO2) nanoparticles (NPs) can have negative health effects, and have been shown to cause respiratory tract cancer in rats. Inflammation has been linked to oxidative stress, and both have been described as possible mechanisms for genotoxicity of NPs, but rarely examined side-by-side in animal studies. In the present study, a wide range of complementary endpoints have been performed to study TiO2 P25 NP-induced genotoxicity in lung overload and non-overload conditions. Additionally, lung burden, inflammation, cytotoxicity and oxidative stress have also been evaluated in order to link genotoxicity with these responses. To assess quick and delayed responses after recovery, endpoints were evaluated at two time points: 2 h and 35 days after three repeated instillations. This study confirmed the previously described lung overload threshold at approximately 200-300 cm2 of lung burden for total particle surface area lung deposition or 4.2 µl/kg for volume-based cumulative lung exposure dose, above which lung clearance is impaired and inflammation is induced. Our results went on to show that these overload doses induced delayed genotoxicity in lung, associated with persistent inflammation only at the highest dose. The lowest tested doses had no toxicity or genotoxicity effects in the lung. In blood, no lymphocyte DNA damage, erythrocytes chromosomal damage or gene mutation could be detected. Our data also demonstrated that only overload doses induced liver DNA lesions irrespective of the recovery time. Tested doses of TiO2 P25 NPs did not induce glutathione changes in lung, blood or liver at both recovery times.