Dataset Information

Rupintrivir reduces RV-induced T_H-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.

ABSTRACT:

Background

Antiviral drugs such as rupintrivir may have an immune-modulatory effect in experimentally induced allergic asthma with subsequent RV infection. We infected lung slices of house-dust mite (HDM)-sensitized asthmatic mice ex vivo with human rhinovirus (RV) and investigated the effect of the antiviral drug rupintrivir on RV-induced cytokine response in lung tissue of HDM-sensitized mice ex vivo.

Methods

Mice were sensitized with HDM. Precision-cut lung slices (PCLS) were prepared from HDM-sensitized or non-sensitized mice. Lung slices were infected ex vivo with RV or RV together with rupintrivir. Modulation of immune responses was evaluated by cytokine secretion 48 h post infection.

Results

In vivo HDM sensitization resulted in a T_H-2/T_H-17-dominated cytokine response that persisted in PCLS ex vivo. RV infection of PCLS from non-sensitized mice resulted in the induction of an antiviral and pro-inflammatory immune response, as indicated by the secretion of IFN-α, IFN-β, IFN-γ, TNF-α, MCP-1, IP-10, IL-10, and IL-17A. In contrast, PCLS from HDM-sensitized mice showed an attenuated antiviral response, but exaggerated IL-4, IL-6, and IL-10 secretion upon infection. Rupintrivir inhibited exaggerated pro-inflammatory cytokine IL-6 and T_H-2 cytokine IL-4 in HDM-sensitized mice.

Conclusions

In summary, this study demonstrates that treatment with rupintrivir influences virus-induced IL-4 and IL-6 cytokine release under experimental conditions ex vivo.

SUBMITTER: Danov O

PROVIDER: S-EPMC6805592 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Rupintrivir reduces RV-induced T<sub>H</sub>-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.

Danov Olga O Lasswitz Lisa L Obernolte Helena H Hesse Christina C Braun Armin A Wronski Sabine S Sewald Katherina K

Respiratory research 20191022 1

<h4>Background</h4>Antiviral drugs such as rupintrivir may have an immune-modulatory effect in experimentally induced allergic asthma with subsequent RV infection. We infected lung slices of house-dust mite (HDM)-sensitized asthmatic mice ex vivo with human rhinovirus (RV) and investigated the effect of the antiviral drug rupintrivir on RV-induced cytokine response in lung tissue of HDM-sensitized mice ex vivo.<h4>Methods</h4>Mice were sensitized with HDM. Precision-cut lung slices (PCLS) were p ...[more]

PMID: 31640701

Similar Datasets

Project description:Precision-cut lung slices (PCLS) are ex vivo models with preserved lung cell populations and maintained tissue architecture. PCLS are, therefore, a powerful tool in respiratory research to study molecular mechanisms that closely reflect whole tissue biology. High-quality RNA and protein extraction from PCLS is, however, challenging as agarose significantly interferes with the yield and purity of extracted material. The present study aimed to optimize QIAzol-based isolation protocol for high-yield and quality RNA, miRNA, and protein extraction from PCLS. PCLS were prepared from 10 to 15-week-old Wistar rats and cultured for 7 days in Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12 (DMEM/F-12) supplemented with 0.1% FBS, penicillin, and streptomycin. LDH release to PCLS culture media was measured to determine cellular cytotoxicity. To select the optimal miRNA/RNA isolation protocol, we tested two different times (10 min, 2 h) and temperatures (room temperature, 4 °C, and -20 °C) of precipitation with isopropanol. Finally, we also assessed isolation with GHCL (guanidinium hydrochloride) extraction buffer. To select the optimal protein isolation protocol, we tested protein precipitation for 10 min at room temperatures (21 ± 1 °C) with 1.5 volumes of isopropanol and 3 volumes of acetone per 1 volume of phenol-ethanol supernatant. Additionally, we also tested protein precipitation for 3 h at -20 °C with 3, 5, and 7 acetone volumes per 1 volume of phenol-ethanol supernatant. We also validated protein precipitation with back extraction buffer instead of 100% ethanol. To measure the general efficiency of the optimized QIAZ-4 protocol, we used native rat lungs. PCLS for the ex vivo model of allergic inflammation were treated with IL-13 at a concentration of 80 ng/ml. Standard QIAzol isolation protocol provided RNA, miRNA, and protein with low yield and poor quality. We found that 2-h isopropanol precipitation at 4 °C with a high concentration of salts significantly increased the yield and quality of extracted RNA and miRNA and provided acceptable qPCR efficiency (between 90 and 110%). Surprisingly, 2-h isopropanol precipitation at -20 °C significantly increased qPCR efficiency above the acceptable range (average efficiency: 120.4%). As for protein extraction, we found that 3-h acetone precipitation at -20 °C provided the highest yield with linear protein detection on Westen Blot. Optimized QIAZ-4 provided significantly higher miRNA and RNA yield compared to standard QIAzol protocols. We also found a significantly increased expression of Eotaxin-1 in PCLS treated with IL-13 as compared to the untreated controls. In our study, we described a simple QIAzol-based method for the simultaneous isolation of RNA, miRNA, and protein from PCLS.

Dataset Information

Rupintrivir reduces RV-induced T_H-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.

Background

Methods

Results

Conclusions

Publications

Rupintrivir reduces RV-induced T<sub>H</sub>-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Dataset Information

Rupintrivir reduces RV-induced TH-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.

Background

Methods

Results

Conclusions

Publications

Rupintrivir reduces RV-induced T<sub>H</sub>-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Rupintrivir reduces RV-induced T_H-2 cytokine IL-4 in precision-cut lung slices (PCLS) of HDM-sensitized mice ex vivo.