Non-coding RNAs Analysis of Eosinophil Subtypes in Asthma

September 7, 2020 updated by: Kestutis Malakauskas, Lithuanian University of Health Sciences

Chronic airway inflammation rich in eosinophils is an important feature seen in asthma. Airway and blood eosinophilia is associated with increased rates of asthma exacerbations and more intense treatment.

Recently, the existence of two distinct eosinophils subtypes was revealed-lung-resident eosinophils (rEOS), which maturate independently to interleukin (IL) 5, with the primary function to maintain tissue homeostasis, and inflammatory eosinophils (iEOS), which mature in IL-5-dependent manner and are mainly involved in immune responses. Eosinophils' effect on the airway remodeling in asthma depends not only on the activity but also by their viable number in the lungs. Blood iEOS infiltrate the airways mainly after the environmental stimulus like allergen and leave the airways with bronchial secretions. However, rEOS reside lung tissue for their entire lifetime regulating local immunity. Blood rEOS and iEOS ratio alters in asthma, compared with healthy controls. It is known that the predominant eosinophils subtype in allergic asthma are iEOS, while rEOS are basic subtype in severe eosinophilic asthma patients, moreover, they are different in adhesive properties and survivability as well. Distinct biological properties allows to speculate about their different functions in asthma, however, there are still little information. Data about differently expressed microRNA (miRNA) profiles in eosinophils in asthma suggests, that eosinophils subtypes can be distinct in non-coding RNA (ncRNA) - microRNA (miRNA), piwi-interacting RNA (piRNA) and long non-coding RNA (IncRNA) profiles that could describe their role in asthma pathogenesis and act as biomarkers to discern asthma phenotypes.

Study Overview

Status

Recruiting

Conditions

Intervention / Treatment

Detailed Description

Asthma is not cured, and only well-balanced treatment can control the course and severity of the disease. Most clinical symptoms rise from aberrant chronic airway inflammation mostly eosinophilic. Eosinophils are terminally differentiated granulocytes that actively contribute to innate and adaptive inflammatory cascades through the production and release of diverse chemokines, cytokines, lipid mediators and other growth factors. IL-5 plays a fundamental role in eosinophils maturation in the bone marrow, their recruitment, and activation at sites of inflammation.

Historically eosinophils were described as a critical player in host defense, including parasites, viruses, fungi, or bacteria, giving them a destructive inflammatory cell label. However, it became clear that steady-state eosinophils can contribute to the immunoregulation and tissue homeostasis as well. Studies revealed that there are distinct eosinophils subtypes - immunoregulatory lung-resident eosinophils (rEOS) and inflammatory eosinophils (iEOS), involved in immune responses. Distinct eosinophils subtypes with different functions determines the separate treatment. There are still only a few studies describing distinct eosinophils subtypes in the lungs or blood. It is the beginning of a new promising research area for better individualized eosinophilic asthma treatment, moreover, other eosinophilic diseases as well.

Peripheral blood eosinophils studies are sufficiently relevant to the tissue eosinophils studies, as blood eosinophils are released into the bloodstream in a fully maturated form. Moreover, peripheral blood study could give additional information with possibilities to prevent eosinophils effects in the early stage, before migration to the airways. Furthermore, the existence of tissue-resident eosinophils in peripheral blood is confirmed and primary research for eosinophil subtypes surface markers was made according to the data of human blood eosinophils.

Data about differently differently expressed microRNA (miRNA) profiles in eosinophils in asthma suggests, that eosinophils subtypes can be distinct in non-coding RNA (ncRNA) - microRNA (miRNA), piwi-interacting RNA (piRNA) and long non-coding RNA (IncRNA) profiles that could describe their role in asthma pathogenesis and act as biomarkers to discern asthma phenotypes.

Researchers have plan to expand research by analyzing non-coding RNA (ncRNA) - miRNA, piRNA and lncRNA profiles of rEOS and iEOS as well as selected ncRNA signatures in blood plasma estimating their diagnostic value. Moreover, additional investigation of ncRNA in eosinophil-derived exosomes will provide important data about possible effect of eosinophils subtypes on airway remodeling via secreted ncRNA. ncRNAs are key regulators for gene transcription. However, there is evidence about their dysregulation in eosinophils during asthma. It will give important information about molecular signaling pathways that regulate the activity of distinct eosinophil subtypes during health and asthma, and provide the essential information about possible new therapeutic targets for their control. Additionally researchers will investigate the biological differences between rEOS and iEOS, including surface integrins and eosinophilopoietins receptors expression, adhesive properties, survivability, synthesized reactive oxygen species and apoptosis, as well as their effect on pulmonary structural cells physiological activity as proliferation, apoptosis, migration, contractility and proteins production, and will relate it with molecular signaling pathways, regulated by distinct expressed ncRNAs. ncRNAs can be stored in eosinophils exosomes and expressed to the surrounding environment. Information about ncRNAs in eosinophils-derived exosomes will demonstrate their function by affecting the other cells, especially after migration to airways. Moreover, ncRNAs are stable and resistant to blood RNases and differentially expressed in several pathologies. Researchers suppose that altered blood levels of ncRNAs could act as a possible new diagnostic biomarker in asthma.

Study Type

Interventional

Enrollment (Anticipated)

80

Phase

  • Not Applicable

Contacts and Locations

This section provides the contact details for those conducting the study, and information on where this study is being conducted.

Study Contact

Study Contact Backup

Study Locations

  • Lithuania
      • Kaunas, Lithuania, LT-50009
        • Recruiting
        • Lithuanian University of Health Sciences, Pulmonology Department
        • Contact:

Participation Criteria

Researchers look for people who fit a certain description, called eligibility criteria. Some examples of these criteria are a person's general health condition or prior treatments.

Eligibility Criteria

Ages Eligible for Study

14 years to 66 years (Adult, Older Adult)

Accepts Healthy Volunteers

No

Genders Eligible for Study

All

Description

Inclusion Criteria:

  • Men and women between the ages of 18-70 years;
  • Allergic asthma and sensitization to house dust mites (D. pteronyssinus) allergen, approved with: 1) medical history and symptoms more than one year; 2) skin prick test positive for D. pteronyssinus (positive wheals are those exceeding 3 mm in diameter greater than the negative control); 3) positive bronchial challenge with methacholine or documented reversible bronchial obstruction;
  • Severe eosinophilic asthma;
  • Premenopausal women if pregnancy test is negative;
  • Healthy subjects without allergic and other chronic respiratory diseases (control group);
  • Participants who gave his/her informed written consent.

Exclusion Criteria:

  • Asthma exacerbation 1 month prior to study;
  • Clinically significant permanent allergy symptoms (ex. cat or dog dander induced allergy);
  • Contraindications to perform an allergy skin test and/or bronchial provocation test: 1) active airway infection 1 month prior the study; 2) used medicaments: inhaled glucocorticoids intake 1 month prior the study, antihistamines intake 7 days prior the study; 3) short acting β2 agonists 12 hours prior the study; 4) long acting β2 agonists 2 days prior the study; 5) leukotriene receptor antagonists prior 14 days;
  • Contraindications for epinephrine;
  • Other significant mental and / or internal diseases and conditions, which could be as exclusion criteria due to the opinion of the researcher;
  • Alcohol or narcotic abuse;
  • Pregnancy;
  • Breast-feeding.

Study Plan

This section provides details of the study plan, including how the study is designed and what the study is measuring.

How is the study designed?

Design Details

  • Primary Purpose: Basic Science
  • Allocation: Randomized
  • Interventional Model: Parallel Assignment
  • Masking: None (Open Label)

Arms and Interventions

Participant Group / Arm
Intervention / Treatment
Experimental: Allergic asthma patients
Allergic asthma patients and sensitization to house dust mites (D. pteronyssinus) allergen.
Biological: Dermatophagoides pteronyssinus allergen
Dermatophagoides pteronyssinus allergen is required to perform allergen bronchial challenge test.
Procedure: Blood sampling
An amount of a person's blood taken from their body for use in medical.
Procedure: Bronchial challenge with allergen
Bronchial challenge is performed with D. pteronyssinus allergen. Measurements of differences in eosinophils activity after allergen challenge.
Experimental: Severe eosinophilic asthma patients
Procedure: Blood sampling
An amount of a person's blood taken from their body for use in medical.
Active Comparator: Healthy subjects as a control group
Healthy subjects without allergic and other chronic respiratory diseases (control group).
Procedure: Blood sampling
An amount of a person's blood taken from their body for use in medical.

What is the study measuring?

Primary Outcome Measures

Outcome Measure
Measure Description
Time Frame
Fold changes of ncRNA expression between eosinophils subtypes
Time Frame: From 6 to 12 months
Validated ncRNA expression of rEOS and iEOS in severe and non-severe eosinophilic asthma patients and healthy subjects.
From 6 to 12 months
ncRNA levels in rEOS- and iEOS-derived exosomes
Time Frame: From 12 to 18 months
Qualitative and quantitative selected ncRNA levels in rEOS- and iEOS-derived exosomes of all investigated groups.
From 12 to 18 months

Secondary Outcome Measures

Outcome Measure
Measure Description
Time Frame
Fold changes of ncRNA profiles of distinct eosinophil subtypes
Time Frame: From 6 to 12 months
Non-validated whole ncRNA profiles of distinct eosinophil subtypes in severe and non-severe eosinophilic asthma patients and healthy subjects.
From 6 to 12 months
The fold changes of rEOS and iEOS surface integrins expression
Time Frame: From 6 to 12 months
The gene expression of selected outer-membrane integrins in eosinophil subtypes.
From 6 to 12 months
The fold changes of rEOS and iEOS eosinophilopoietins receptors expression
Time Frame: From 6 to 12 months
The gene expression of interleukin (IL)-5, IL-3, and granulocyte-macrophage colony-stimulating factor in eosinophil subtypes.
From 6 to 12 months
The efficiency of iEOS and rEOS adhesion
Time Frame: From 6 to 12 months
The differences in stable adhered iEOS and rEOS quantity in the combined cell culture with airway smooth muscle (ASM) cells or pulmonary fibroblasts compared between the investigated groups.
From 6 to 12 months
iEOS and rEOS survival differences
Time Frame: From 6 to 12 months
Viable iEOS and rEOS number after an appropriate period of time in combined cell culture with ASM cells or pulmonary fibroblasts.
From 6 to 12 months
Quantity of iEOS and rEOS synthesized reactive oxygen species
Time Frame: From 6 to 12 months
Relative differences between iEOS and rEOS synthesized reactive oxygen species quantity after an appropriate period of incubation alone or with ASM cells or pulmonary fibroblasts.
From 6 to 12 months
Apoptotic iEOS and rEOS number
Time Frame: From 6 to 12 months
The number of apoptotic iEOS and rEOS after an appropriate period of time in combined cell culture with ASM cells or pulmonary fibroblasts.
From 6 to 12 months
Concentrations of iEOS and rEOS produced proteins in investigated subjects' body fluids.
Time Frame: From 6 to 12 months
Selected iEOS and rEOS proteins concentrations, measured in investigated subjects body fluids, expressed as the amount of protein in the respective amount of fluid sample.
From 6 to 12 months
iEOS and rEOS effect on airway smooth muscle cells or pulmonary fibroblasts proliferation
Time Frame: From 6 to 12 months
The quantity of ASM cells or pulmonary fibroblasts after several repeats of proliferation in the presence or absence of eosinophil subtypes.
From 6 to 12 months
iEOS and rEOS effect on apoptotic ASM cells and pulmonary fibroblasts number
Time Frame: From 6 to 12 months
The number of apoptotic ASM cells and pulmonary fibroblasts after an appropriate period of time in combined cell culture with iEOS and rEOS.
From 6 to 12 months
iEOS and rEOS effect on migration of ASM cells.
Time Frame: From 6 to 12 months
The migrated ASM cells number after an appropriate period of time in combined cell culture with iEOS and rEOS
From 6 to 12 months
iEOS and rEOS effect on migration of pulmonary fibroblasts
Time Frame: From 6 to 12 months
The migrated pulmonary fibroblasts number after an appropriate period of time in combined cell culture with iEOS and rEOS
From 6 to 12 months
iEOS and rEOS effect on ASM cells contractility
Time Frame: From 6 to 12 months
The relative efficiency of ASM cells' ability to contract collagen gel after an appropriate period of time in combined cell culture with iEOS and rEOS, expressed as reduced poured gel size in percentage, compared with control ASM cells without incubation with eosinophils.
From 6 to 12 months
iEOS and rEOS effect on pulmonary fibroblasts contractility
Time Frame: From 6 to 12 months
The relative efficiency of pulmonary fibroblasts ability to contract collagen gel after an appropriate period of time in combined cell culture with iEOS and rEOS. expressed as reduced poured gel size in percentage, compared with control pulmonary fibroblasts, without incubation with eosinophils.
From 6 to 12 months
iEOS and rEOS effect on fold changes of ASM cells and pulmonary fibroblasts proteins expression;
Time Frame: From 6 to 12 months
Altered selected ASM cells and pulmonary fibroblasts proteins expression, after incubation with iEOS and rEOS, expressed as fold changes in comparison with control cells, without incubation with eosinophils
From 6 to 12 months
iEOS and rEOS effect on fold changes of ASM cells and pulmonary fibroblasts proteins gene expression;
Time Frame: From 6 to 12 months
Altered selected ASM cells and pulmonary fibroblasts proteins gene expression, after incubation with iEOS and rEOS, expressed as fold changes in comparison with control cells, without incubation with eosinophils
From 6 to 12 months

Collaborators and Investigators

This is where you will find people and organizations involved with this study.

Sponsor

Lithuanian University of Health Sciences

Publications and helpful links

The person responsible for entering information about the study voluntarily provides these publications. These may be about anything related to the study.

General Publications

Study record dates

These dates track the progress of study record and summary results submissions to ClinicalTrials.gov. Study records and reported results are reviewed by the National Library of Medicine (NLM) to make sure they meet specific quality control standards before being posted on the public website.

Study Major Dates

Study Start (Anticipated)

October 1, 2020

Primary Completion (Anticipated)

October 1, 2023

Study Completion (Anticipated)

October 1, 2023

Study Registration Dates

First Submitted

August 25, 2020

First Submitted That Met QC Criteria

September 7, 2020

First Posted (Actual)

September 9, 2020

Study Record Updates

Last Update Posted (Actual)

September 9, 2020

Last Update Submitted That Met QC Criteria

September 7, 2020

Last Verified

August 1, 2020

More Information

Terms related to this study

Keywords

Additional Relevant MeSH Terms

Other Study ID Numbers

  • LAB-1/2020

Plan for Individual participant data (IPD)

Plan to Share Individual Participant Data (IPD)?

No

Drug and device information, study documents

Studies a U.S. FDA-regulated drug product

No

Studies a U.S. FDA-regulated device product

No

This information was retrieved directly from the website clinicaltrials.gov without any changes. If you have any requests to change, remove or update your study details, please contact register@clinicaltrials.gov. As soon as a change is implemented on clinicaltrials.gov, this will be updated automatically on our website as well.

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