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Research
Allan Dietz
Mayo Clinic College of Medicine
Corresponding Author
ORCiD: https://orcid.org/0000-0003-3410-9621
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive pulmonary disease characterized by aberrant tissue remodeling, formation of scar tissue within the lungs and continuous loss of lung function. The areas of fibrosis seen in lungs of IPF patients share many features with normal aging lung with cellular senescence being one. The contribution of the immune system to the etiology of IPF remains poorly understood. Evidence obtained from animal models and human studies suggests that innate and adaptive immune processes can orchestrate existing fibrotic responses. Currently, there is only modestly effective pharmacotherapy for IPF. Mesenchymal stem cells (MSCs)-based therapies have emerged as a potential option treatment of IPF. This study explores the possibility of using autologous MSCs as an IPF therapy and present an attempt to determine if the disease occurring in the lungs can be reflected on peripheral blood immune status.
Methods
Comprehensive characterization of autologous adipose derived MSCs (aMSCs) from five IPF patients and five age and gender matched Healthy Controls (HC) was done using flow cytometry, Droplet Digital PCR (ddPCR), Multiplex Luminex xMAP technology and confocal microscopy. For assessing the self renewal capacity and osteogenic differentiation IncuCyte Live Cell Imaging technology was used.
Multi-parameter quantitative flow cytometry of un-manipulated whole blood of another group of 15 IPF patients and 87 (30 age and gender matched) HC was used to analyze 110 peripheral phenotypes.
Results
There are no differences between autologous aMSCs from IPF patients and HC in their stem cell properties, self renewal capacity, plasticity for osteogenic differentiation, secretome content, cell cycle inhibitor marker levels and mitochondrial health.
IPF patients had altered peripheral blood immunophenotype including reduced B cells subsets, increased T cell subsets, and increased granulocytes among others demonstrating clear and significant differences.
Conclusions
Our results indicate that there is no difference in aMSCs properties from IPF patients and HC, suggesting that autologous aMSCs may be an acceptable option for IPF therapy.
Characterization of peripheral immune phenotype may be a valuable indicator for successful therapy, and for potentially staging the disease.
Keywords
Autologous adipose mesenchymal stem cells, Characterization, Idiopathic Pulmonary Fibrosis, Treatment, Immunologic profiling
Figure 1
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This is a list of supplementary files associated with this preprint. Click to download.
- Additionalfile1.pdf
Additional File 1. List of antibodies used for immunophenotyping. * Gustafson, M.P., et al., A method for identification and analysis of non-overlapping myeloid immunophenotypes in humans. PloS One, 2015. 10(3); reference [29] ** Gustafson, M.P., et al., A systems biology approach to investigating the influence of exercise and fitness on the composition of leukocytes in peripheral blood. Journal for immunotherapy of cancer, 2017. 5(1): p.30; reference [30]. *** The antibodies are added into the samples without dilution, as per manufacturer instructions.
- Additionalfile2.pdf
Additional File 2. Growth rate kinetic, population doubling time and R2 for the linear regressions. k represents cell growth rate per hour; td represents cell doubling time per hour, and R2 represents the coefficient of determination for the linear regression.
- Additionalfile3A.pdf
Secretome Profile of IPFaMSCs and HCaMSCs: Unpaired Student t-test. A. Secreted Growth Factors.
- AdditionalFile3B.pdf
B. Secreted Ant Inflammatory Cytokines.
- Additionalfile3Ca.pdf
C. Secreted Pro Inflammatory Cytokines, group 1 (a) and Secreted Pro Inflammatory Cytokines, group 2
- Additionalfile3Cb.pdf
(b). Data are expressed as mean and dots represent individual cell line values. Differences were considered significant when *p<0.05.
- AdditionalFile4.pdf
Additional File 4. Comparison of immunophenotypes between healthy controls (HC, age 50+) and IPF patients. In Blue: decreased mean values in IPF patients; in Red: increased mean values in IPF patients. *p<0.05, ** p<0.01, *** p<0.001, *** p<0.0001, compared with control group. The phenotypes which correlate with the pulmonary functions are denoted with a red asterisk.
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Review #1 received
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Reviewer #2 agreed
On 10 Jul, 2021
Reviews received
Received 08 Jul, 2021
Reviewer #1 agreed
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Submission checks complete
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On 05 Jun, 2021
First submitted
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A new preprint design is coming!
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This preprint is under consideration at Stem Cell Research & Therapy. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research
Allan Dietz
Mayo Clinic College of Medicine
Corresponding Author
ORCiD: https://orcid.org/0000-0003-3410-9621
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Review
Version 1
Posted 09 Jun, 2021
No community comments so far
Editorial decision: Major Revision
On 12 Oct, 2021
Review #2 received
Received 10 Oct, 2021
Reviewer #3 agreed
On 26 Sep, 2021
Review #1 received
Received 19 Jul, 2021
Reviewer #2 agreed
On 10 Jul, 2021
Reviews received
Received 08 Jul, 2021
Reviewer #1 agreed
On 07 Jul, 2021
Reviewers invited
Invitations sent on 13 Jun, 2021
Editor assigned
On 05 Jun, 2021
Submission checks complete
On 05 Jun, 2021
Editor invited
On 05 Jun, 2021
First submitted
On 27 May, 2021
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Stem Cell & Developmental Cell Biology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive pulmonary disease characterized by aberrant tissue remodeling, formation of scar tissue within the lungs and continuous loss of lung function. The areas of fibrosis seen in lungs of IPF patients share many features with normal aging lung with cellular senescence being one. The contribution of the immune system to the etiology of IPF remains poorly understood. Evidence obtained from animal models and human studies suggests that innate and adaptive immune processes can orchestrate existing fibrotic responses. Currently, there is only modestly effective pharmacotherapy for IPF. Mesenchymal stem cells (MSCs)-based therapies have emerged as a potential option treatment of IPF. This study explores the possibility of using autologous MSCs as an IPF therapy and present an attempt to determine if the disease occurring in the lungs can be reflected on peripheral blood immune status.
Methods
Comprehensive characterization of autologous adipose derived MSCs (aMSCs) from five IPF patients and five age and gender matched Healthy Controls (HC) was done using flow cytometry, Droplet Digital PCR (ddPCR), Multiplex Luminex xMAP technology and confocal microscopy. For assessing the self renewal capacity and osteogenic differentiation IncuCyte Live Cell Imaging technology was used.
Multi-parameter quantitative flow cytometry of un-manipulated whole blood of another group of 15 IPF patients and 87 (30 age and gender matched) HC was used to analyze 110 peripheral phenotypes.
Results
There are no differences between autologous aMSCs from IPF patients and HC in their stem cell properties, self renewal capacity, plasticity for osteogenic differentiation, secretome content, cell cycle inhibitor marker levels and mitochondrial health.
IPF patients had altered peripheral blood immunophenotype including reduced B cells subsets, increased T cell subsets, and increased granulocytes among others demonstrating clear and significant differences.
Conclusions
Our results indicate that there is no difference in aMSCs properties from IPF patients and HC, suggesting that autologous aMSCs may be an acceptable option for IPF therapy.
Characterization of peripheral immune phenotype may be a valuable indicator for successful therapy, and for potentially staging the disease.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
This is a list of supplementary files associated with this preprint. Click to download.
- Additionalfile1.pdf
Additional File 1. List of antibodies used for immunophenotyping. * Gustafson, M.P., et al., A method for identification and analysis of non-overlapping myeloid immunophenotypes in humans. PloS One, 2015. 10(3); reference [29] ** Gustafson, M.P., et al., A systems biology approach to investigating the influence of exercise and fitness on the composition of leukocytes in peripheral blood. Journal for immunotherapy of cancer, 2017. 5(1): p.30; reference [30]. *** The antibodies are added into the samples without dilution, as per manufacturer instructions.
- Additionalfile2.pdf
Additional File 2. Growth rate kinetic, population doubling time and R2 for the linear regressions. k represents cell growth rate per hour; td represents cell doubling time per hour, and R2 represents the coefficient of determination for the linear regression.
- Additionalfile3A.pdf
Secretome Profile of IPFaMSCs and HCaMSCs: Unpaired Student t-test. A. Secreted Growth Factors.
- AdditionalFile3B.pdf
B. Secreted Ant Inflammatory Cytokines.
- Additionalfile3Ca.pdf
C. Secreted Pro Inflammatory Cytokines, group 1 (a) and Secreted Pro Inflammatory Cytokines, group 2
- Additionalfile3Cb.pdf
(b). Data are expressed as mean and dots represent individual cell line values. Differences were considered significant when *p<0.05.
- AdditionalFile4.pdf
Additional File 4. Comparison of immunophenotypes between healthy controls (HC, age 50+) and IPF patients. In Blue: decreased mean values in IPF patients; in Red: increased mean values in IPF patients. *p<0.05, ** p<0.01, *** p<0.001, *** p<0.0001, compared with control group. The phenotypes which correlate with the pulmonary functions are denoted with a red asterisk.
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