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Project Title:  Induction of Early Stages of Osteoarthritis After Exposure to Microgravity (Postdoctoral Fellowship) Reduce
Fiscal Year: FY 2015 
Division: Human Research 
Research Discipline/Element:
HRP HHC:Human Health Countermeasures
Start Date: 11/01/2011  
End Date: 02/28/2015  
Task Last Updated: 07/07/2015 
Download report in PDF pdf
Principal Investigator/Affiliation:   Mellor, Liliana F. Ph.D. / North Carolina State University 
Address:  Department of Biomedical Engineering 
Cell Mechanics Laboratory 
Raleigh , NC 27695-7115 
Email: lfmellor@ncsu.edu 
Phone: 208-426-2238  
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: North Carolina State University 
Joint Agency:  
Comments: NOTE: formerly at Boise State University until fall 2013 (Ed., Jan 2014)  
Co-Investigator(s)
Affiliation: 
Loboa, Elizabeth  MENTOR/ North Carolina State University 
Project Information: Grant/Contract No. NCC 9-58-PF02601 
Responsible Center: NSBRI 
Grant Monitor:  
Center Contact:   
Solicitation / Funding Source: 2011 NSBRI-RFA-11-01 Postdoctoral Fellowships 
Grant/Contract No.: NCC 9-58-PF02601 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) HHC:Human Health Countermeasures
Human Research Program Risks: (1) Osteo:Risk Of Early Onset Osteoporosis Due To Spaceflight (No longer used, July 2020)
Human Research Program Gaps: (1) Osteo04:We do not know the contribution of each risk factor on bone loss and recovery of bone strength, and which factors are the best targets for countermeasure application (IRP Rev E)
Flight Assignment/Project Notes: NOTE: End date change to 2/28/2015 per NSBRI (Ed., 12/2/14)

NOTE: End date is now 11/30/2014 with PostDoc change in institution, per NSBRI (Ed., 1/15/14)

NOTE: End date changed to 11/30/2013 per NSBRI (Ed., 10/24/13)

Task Description: POSTDOCTORAL FELLOWSHIP

Little is known about the effects of spaceflight on articular cartilage health. Unlike bone and muscle, cartilage lacks the ability to regenerate. Once a catabolic cascade is triggered, it usually results in osteoarthritis. Some studies have shown degradation of the articular cartilage in response to unloading and prolonged bedrest. However, the underlying molecular mechanisms of articular cartilage degradation in response to unloading are still elusive. The overall goal of my research is to understand and elucidate key molecular mechanisms involved in response of cartilage to changes in gravitational forces. The majority of my focus is at the cell level, with analyses of potential approaches to translate these changes to tissue and joint level. To begin to address this problem, I investigated the effects of simulated microgravity on chondrocytes using the rotating wall vessel (RWV) bioreactor. Also, in collaboration with Dr. Jeff Willey, I studied the combined effects of radiation and simulated microgravity on articular cartilage health. I found that chondrocytes exposed to simulated microgravity undergo morphological rearrangement of the actin cytoskeleton. This change was not observed in irradiated cells; however, when cells were exposed to both radiation and simulated microgravity, they developed long stress-like fibers, resembling a more fibroblast morphology as compared to the cortical control chondrocytes. Gene expression analyses confirmed that cells exposed to both radiation and simulated microgravity express more collagen I and less collagen II and aggrecan, which is characteristic for de-differentiated chondrocytes. We also determined that irradiated chondrocytes upregulate R-spondin1. R-spondin1 is a positive regular of Wnt signaling that has been shown to protect against radiation-induced damage to the oral mucosa and intestine. R-spondin1 has also been shown to protect against inflammatory bone damage in a mouse model of arthritis. Our results are interesting given R-spondin1 is upregulated in response to radiation, but not radiation and microgravity. If chondrocytes in normal gravity respond to radiation by producing "radioprotectant" genes, simulated microgravity is inhibiting that effect, making cartilage cells more susceptible to radiation-induced damage in microgravity conditions. This data shows that microgravity may inhibit the radio-protective mechanism of chondrocytes, suggesting a combined synergistic, degradatory effect of simulated microgravity and radiation, which mimics the environment of spaceflight. The mechanisms responsible for these morphological and signal transduction changes in response to simulated microgravity and radiation are still under investigation. Another unexpected and compelling finding was that chondrocytes respond to simulated microgravity by up-regulating sclerostin, an inhibitor of Wnt signaling known to induce bone density loss in space. However, according to a recent study, sclerostin may prevent cartilage from further degradation by decreasing production of matrix degrading genes, suggesting a chondro-protective role for sclerostin in this tissue. Therapeutic treatments using an antibody against sclerostin have shown promising results to prevent bone density loss in unloading conditions. However, the effects of blocking sclerostin on neighboring tissue, such as articular cartilage have not been addressed and may have adverse effects. My goal for next year is to compare results of cells exposed to microgravity to cells incubated in hydrostatic pressure, an environment known to induce expression of anabolic genes in articular cartilage. In addition, I will investigate the role of primary cilia in microgravity, an important organelle involved in mechanotransduction that has also been associated with the Wnt signaling pathway.

Research Impact/Earth Benefits: Astronauts are exposed to prolonged periods of microgravity during spaceflight. Lack of gravity is known to negatively affect the musculoskeletal system, mainly bone and skeletal muscle, which are constantly exposed to mechanical loading on Earth. However, another important component of the musculoskeletal system, the synovial joint, has not been fully investigated in space-like conditions. Unlike bone and muscle, the articular cartilage of the synovial joint has limited regenerative capacity, and has a relatively slow turn over compared to the fast remodeling of bone. Cartilage degradation leads to a severe disease known as osteoarthritis (OA). OA is the leading cause of disability in the US, and one of the few chronic diseases of aging without a cure. Little is known about the effects of microgravity on the synovial joint and changes in cartilage homeostasis. Bone remodeling occurs much faster. This is the first study to use an in vitro and ex vivo approach to understand the effects of reduced gravity on cartilage health and to elucidate the signaling pathways responsive to changes in gravitational forces. Understanding changes in cartilage homeostasis and molecular pathways in reduced gravity conditions, will help us elucidate the mechanisms involved in disuse OA here on Earth, and develop novel therapeutic targets.

Task Progress & Bibliography Information FY2015 
Task Progress: This is the first study to show cartilage degradation at both the cell and tissue level in response to simulated microgravity via changes in gene expression, protein expression, and GAG (glycosaminoglycan) content. Although it appears that cartilage degradation in response to microgravity may take longer than bone density loss and skeletal muscle atrophy, cartilage tissue does not regenerate. Loss or degeneration of articular cartilage results in painful and debilitating joint disease that can limit mobility of the affected joint. We also targeted a potential signaling pathway that responds to mechanical unloading in chondrocytes. Active canonical Wnt signaling is essential in bone to maintain a balance between osteoclast and osteoblast activity. Inhibition of Wnt signaling has been correlated with bone loss. We detected inhibition of Wnt signaling in response to simulated microgravity in chondrocytes by up-regulation of sclerostin and other Wnt inhibitors. However, in cartilage, Wnt inhibition has been reported to be a chondro-protective mechanism to prevent further cartilage degradation, and canonical Wnt signaling has been associated with cartilage degradation and OA. We showed in our chondrocyte cell line that up-regulation of catabolic genes in response to simulated microgravity was similar to that of IL-1ß treatment. However, when recombinant sclerostin was added to IL-1ß treated cells, it decreased MMP (matrix metalloproteinase) expression as previously suggested, providing supportive evidence of the chondroprotective role of sclerostin in cartilage tissue. Therefore, it appears that sclerostin has opposite roles in these two adjacent tissues, and therapeutic targets using a sclerostin antibody to reduce bone density loss in space and osteoporosis need to be further evaluated given the potential of sclerostin inhibition resulting in the progression of OA. Funding for this project has helped provide new evidence for another tissue potentially affected by lack of gravity. Osteoarthritis and joint degradation may take longer than osteopenia or sarcopenia to develop, but unlike bone and muscle, cartilage degradation is irreversible and therefore, should be taken into consideration as a possible risk during prolonged spaceflight.

Bibliography Type: Description: (Last Updated: 11/12/2020)  Show Cumulative Bibliography Listing
 
Articles in Peer-reviewed Journals Mellor L, Mohiti-Asli M, Williams J, Kannan A, Dent MR, Guilak F, Loboa E. "Extracellular calcium modulates chondrogenic and osteogenic differentiation of human derived adipose stem cells: A novel approach for osteochondral tissue engineering using a single stem cell source." Tissue Eng Part A. 2015 Jun 2. [Epub ahead of print] PubMed PMID: 26035347 ; http://dx.doi.org/10.1089/ten.TEA.2014.0572 , Jun-2015
Articles in Peer-reviewed Journals Mellor LF, Baker TL, Brown RJ, Catlin LW, Oxford JT. "Optimal 3D culture of primary articular chondrocytes for use in the rotating wall vessel bioreactor." Aviation, Space, and Environmental Medicine. 2014 Aug;85(8):798-804. PubMed PMID: 25199120; PubMed Central PMCID: PMC4207436 ; http://dx.doi.org/10.3357/ASEM.3905.2014 , Aug-2014
Articles in Peer-reviewed Journals Mellor LF, Steward AJ, Nordberg RC, Taylor MA, Loboa EG. "Comparison of simulated microgravity and hydrostatic pressure for chondrogenesis of hASC." Aerosp Med Hum Perform. 2017 Apr;88(4):377-84. https://doi.org/10.3357/AMHP.4743.2017 ; PubMed PMID: 28518000 , Apr-2017
Articles in Peer-reviewed Journals Mellor LF, Nordberg RC, Huebner P, Mohiti-Asli M, Taylor MA, Efird W, Oxford JT, Spang JT, Shirwaiker RA, Loboa EG. "Investigation of multiphasic 3D-bioplotted scaffolds for site-specific chondrogenic and osteogenic differentiation of human adipose-derived stem cells for osteochondral tissue engineering applications." J Biomed Mater Res B Appl Biomater. 2020 Jul;108(5):2017-30. Epub 2019 Dec 27. https://doi.org/10.1002/jbm.b.34542 ; PMID: 31880408; PMCID: PMC7217039 , Jul-2020
Articles in Peer-reviewed Journals Nordberg RC, Mellor LF, Krause AR, Donahue HJ, Loboa EG. "LRP receptors in chondrocytes are modulated by simulated microgravity and cyclic hydrostatic pressure." PLoS One. 2019 Oct 4;14(10):e0223245. https://doi.org/10.1371/journal.pone.0223245 ; PMID: 31584963; PMCID: PMC6777824 , Oct-2019
Awards Mellor LF. "Career advancement award, National Space Biomedical Research Institute, December 2014." Dec-2014
Awards Mellor LF. "Fellow Travel Award, Biomedical Engineering Society (BMES)-CMBE annual meeting, January 2015." Jan-2015
Awards Mellor LF. "Poster competition award, 3rd place Postdoctoral Research Symposium at NC State University, May 2014." May-2014
Awards Mellor LF. "Professional Development Award, NCSU (NC State University) Postdoctoral Association, September 2014." Sep-2014
Project Title:  Induction of Early Stages of Osteoarthritis After Exposure to Microgravity (Postdoctoral Fellowship) Reduce
Fiscal Year: FY 2014 
Division: Human Research 
Research Discipline/Element:
HRP HHC:Human Health Countermeasures
Start Date: 11/01/2011  
End Date: 02/28/2015  
Task Last Updated: 12/24/2013 
Download report in PDF pdf
Principal Investigator/Affiliation:   Mellor, Liliana F. Ph.D. / North Carolina State University 
Address:  Department of Biomedical Engineering 
Cell Mechanics Laboratory 
Raleigh , NC 27695-7115 
Email: lfmellor@ncsu.edu 
Phone: 208-426-2238  
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: North Carolina State University 
Joint Agency:  
Comments: NOTE: formerly at Boise State University until fall 2013 (Ed., Jan 2014)  
Co-Investigator(s)
Affiliation: 
Loboa, Elizabeth  MENTOR/ North Carolina State University 
Project Information: Grant/Contract No. NCC 9-58-PF02601 
Responsible Center: NSBRI 
Grant Monitor:  
Center Contact:   
Solicitation / Funding Source: 2011 NSBRI-RFA-11-01 Postdoctoral Fellowships 
Grant/Contract No.: NCC 9-58-PF02601 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) HHC:Human Health Countermeasures
Human Research Program Risks: (1) Osteo:Risk Of Early Onset Osteoporosis Due To Spaceflight (No longer used, July 2020)
Human Research Program Gaps: (1) Osteo04:We do not know the contribution of each risk factor on bone loss and recovery of bone strength, and which factors are the best targets for countermeasure application (IRP Rev E)
Flight Assignment/Project Notes: NOTE: End date change to 2/28/2015 per NSBRI (Ed., 12/2/14)

NOTE: End date is now 11/30/2014 with PostDoc change in institution, per NSBRI (Ed., 1/15/14)

NOTE: End date changed to 11/30/2013 per NSBRI (Ed., 10/24/13)

Task Description: POSTDOCTORAL FELLOWSHIP

Little is known about the effects of spaceflight on articular cartilage health, and the synovial joint in general. Unlike bone and muscle, cartilage lacks the ability to regenerate and once a catabolic cascade is triggered, it usually results in osteoarthritis. Some studies have shown degradation of the articular cartilage in response to unloading (1) and prolonged bedrest (2). However, the underlying molecular mechanisms of articular cartilage degradation in response to unloading are still elusive. My study aims to investigate the effects of simulated microgravity on chondrocytes using the RWV bioreactor. Further, in collaboration with Dr. Jeff Willey, we are studying the combined effects of radiation and simulated microgravity in articular cartilage health. My goal is to understand the molecular mechanisms involved in response to changes in gravitational forces at the cell level, and translate these changes to tissue and whole joint level. One interesting finding is that chondrocytes exposed to simulated microgravity undergo morphological rearrangement of the actin cytoskeleton. This change was not observed in irradiated cells; however, when cells were exposed to both radiation and simulated microgravity, they developed long stress-like fibers, resembling a more fibroblast morphology compared to the cortical control chondrocytes. Gene expression analyses confirmed that cells exposed to both radiation and simulated microgravity express more collagen I and less collagen II and aggrecan, which is characteristic for de-differentiated chondrocytes. The mechanotransduction mechanisms responsible for these morphological changes in response to simulated microgravity are still under investigation. Another unexpected and compelling finding is that chondrocytes respond to simulated microgravity by up-regulating sclerostin, an inhibitor of Wnt signaling known to induce bone density loss in space. However, according to a recent study, sclerostin may prevent cartilage from further degradation by decreasing production of matrix degrading genes, suggesting a chondro-protective mechanism in this tissue (3). Therapeutic treatments using an antibody against sclerostin have shown promising results to prevent bone density loss in unloading conditions; however, the effects of blocking sclerostin on neighboring tissue, such as articular cartilage, have not been addressed and may have adverse effects. In addition, irradiated chondrocytes upregulate R-spondin1, a positive regulator of Wnt signaling. R-spondin1 has been shown to protect against radiation-induced damage to the oral mucosa and intestine. Moreover, R-spondin1 has been shown to protect against inflammatory bone damage in a mouse model of arthritis (4). Our results are interesting given R-spondin1 is upregulated in response to radiation but not radiation and microgravity. If chondrocytes in normal gravity respond to radiation by producing "radioprotectant" genes, simulated microgravity is inhibiting that effect, making cartilage cells more susceptible to radiation-induced damage in microgravity conditions. This data shows that microgravity may inhibit the radio-protective mechanism of chondrocytes, suggesting a combined synergistic, degrading effect of simulated microgravity and radiation, which mimics the environment of spaceflight.

Our goal for next year is to compare our results to cells incubated in hydrostatic pressure, an environment known to induce expression of anabolic genes in articular cartilage. In addition, we will study the role of primary cilia in microgravity, an important organelle involved in mechanotransduction that has also been associated with Wnt signaling pathway.

1. Niu, H.J. et al. (2012) Acta Mechanica Sinica 28, 1488-1493.

2. Liphardt, A.M. et al. (2009) Osteoarthritis Cartilage 17, 1598-1603.

3. Chan, B.Y., et al. (2011) Osteoarthritis Cartilage 19, 874-885.

4. Zhao, J. et al. (2009) Trends Biotechnol 27, 131-136.

Research Impact/Earth Benefits: Spaceflight has a drastic effect on the musculoskeletal system. Several studies have developed different exercise and nutrition countermeasures to prevent or minimize bone density loss and skeletal muscle atrophy after space missions. However, the effects of microgravity and radiation on articular cartilage health of the synovial joints are still elusive, and pathological conditions such as arthritis can result in severely restricted mobility. Currently, there is no cure for arthritis due to the lack of understanding of the molecular mechanisms that trigger cartilage degradation. Our study is the first to investigate the effects of radiation and simulated microgravity at the molecular level using several chondrocyte cell lines and a Rotating Wall Vessel bioreactor to simulate reduced microgravity. We found that chondrocytes respond to reduced gravity by changing their cytoskeletal morphology, and are still investigating the mechanism and mechanoreceptors responsible for re-arranging the cell morphology in response to changes in gravitational forces. Another interesting finding is the change in gene expression of molecules associated with the Wnt signaling pathway in response to simulated microgravity. The role of Wnt signaling in cartilage is well understood in embryology and development, and it has recently emerged as a critical regulator of bone and cartilage homeostasis. However, the role of Wnt signaling in arthritis is not understood, and recent studies found that sclerostin, an inhibitor of Wnt signaling, was up-regulated in mineralized cartilage and end-stage osteoarthritic samples. Sclerostin has been implicated in bone density loss in microgravity, and is now a promising therapeutic target to protect bones during space missions as well as in patients suffering from osteoporosis here on Earth. Our study is the first to detect sclerostin up-regulation in chondrocytes exposed to simulated microgravity. The mechanisms and effects of sclerostin up-regulation and changes to the Wnt signaling pathway on cartilage homeostasis are still under investigation. If Wnt signaling is in fact associated with the onset of osteoarthritis as suggested recently by a few studies, the findings from our current study may help with the understanding of the underlying mechanism involved in osteoarthritis, and can help provide a new therapeutic target for the millions of people suffering from arthritis here on Earth.

Task Progress & Bibliography Information FY2014 
Task Progress: Aim 1: Investigate changes in gene expression after exposure to simulated microgravity. Our data shows several changes in gene expression in response to simulated microgravity and radiation. Cells that are irradiated and exposed to simulated microgravity are de-differentiating as shown by morphological data and over-expression of collagen type I and aggrecan down-regulation. Additionally, we identified Wnt signaling as a target pathway in chondrocytes exposed to simulated microgravity. A wnt pathway PCR array evaluated 84 different genes related with Wnt pathway, and cells exposed to microgravity, expressed up-regulation of several genes that either inhibit or down-regulate Wnt signaling. In addition, qPCR analysis and ELISA confirmed up-regulation of sclerostin, a Wnt inhibitor, produced by chondrocytes in response to simulated microgravity.

Aim 2: Examine changes in cell-matrix interactions in response to simulated microgravity. Preliminary data looking at CD44 expression in chondrocytes, a receptor important in cartilage homeostasis that mediates binding to hyaluronan, did not show any changes after 48 hr exposure to simulated microgravity. A time-course study will evaluate different time points, and other important cell surface receptors, such as integrins, will be evaluated.

Aim 3: Examine the effects of simulated microgravity on the cytoskeletal morphology of chondrocytes. Chondrocytes in simulated microgravity respond by re-arranging their cytoskeletal morphology. This change is more pronounced in cells exposed to radiation and simulated microgravity. The mechanotransduction mechanisms involved with these morphological changes are still under investigation.

Bibliography Type: Description: (Last Updated: 11/12/2020)  Show Cumulative Bibliography Listing
 
 None in FY 2014
Project Title:  Induction of Early Stages of Osteoarthritis After Exposure to Microgravity (Postdoctoral Fellowship) Reduce
Fiscal Year: FY 2013 
Division: Human Research 
Research Discipline/Element:
HRP HHC:Human Health Countermeasures
Start Date: 11/01/2011  
End Date: 11/30/2013  
Task Last Updated: 11/16/2012 
Download report in PDF pdf
Principal Investigator/Affiliation:   Mellor, Liliana F. Ph.D. / North Carolina State University 
Address:  Department of Biomedical Engineering 
Cell Mechanics Laboratory 
Raleigh , NC 27695-7115 
Email: lfmellor@ncsu.edu 
Phone: 208-426-2238  
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: North Carolina State University 
Joint Agency:  
Comments: NOTE: formerly at Boise State University until fall 2013 (Ed., Jan 2014)  
Co-Investigator(s)
Affiliation: 
Oxford, Julia  MENTOR/ Boise State University  
Project Information: Grant/Contract No. NCC 9-58-PF02601 
Responsible Center: NSBRI 
Grant Monitor:  
Center Contact:   
Solicitation / Funding Source: 2011 NSBRI-RFA-11-01 Postdoctoral Fellowships 
Grant/Contract No.: NCC 9-58-PF02601 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) HHC:Human Health Countermeasures
Human Research Program Risks: (1) Osteo:Risk Of Early Onset Osteoporosis Due To Spaceflight (No longer used, July 2020)
Human Research Program Gaps: (1) Osteo04:We do not know the contribution of each risk factor on bone loss and recovery of bone strength, and which factors are the best targets for countermeasure application (IRP Rev E)
Flight Assignment/Project Notes: NOTE: End date changed to 11/30/2013 per NSBRI (Ed., 10/24/13)

Task Description: POSTDOCTORAL FELLOWSHIP

Our goal is to investigate the effects of simulated microgravity on cartilage homeostasis, by exposing two chondrocyte cell lines to a modeled microgravity environment using a rotating wall vessel bioreactor. Cells exposed to microgravity will be compared to cells incubated under normal gravitational conditions as a control, as well as an arthritic-like cell model induced by treating chondrocytes with pro-inflammatory cytokines such as IL-1B and oncostatin M (OSM). Disruptions in cell-matrix interactions, changes in cytoskeletal morphology and gene up-regulation will be evaluated to determine changes in chondrocyte metabolism. We hypothesize that similar to bone, cartilage homeostasis can be compromised during exposure to microgravity, resulting in osteoarthritic-like conditions in astronauts after space missions. This study will give us a better insight into whether exposure to microgravity can increase the risk of developing early stages of osteoarthritis in astronauts.

Due to the limited capacity of regeneration in articular cartilage, early detection and treatment are key components to prevent the advanced stages of cartilage degradation, which is the leading cause of disability in the U.S., limiting the activities of nearly 21 million adults. Cells incubated under simulated microgravity undergo morphological changes in the actin cytoskeleton. These changes were detected after only two days in simulated microgravity and retained for seven days. Cells under normal gravitation forces maintain the characteristic cortical morphology of healthy chondrocytes, while cells under simulated microgravity developed elongated fibers and adopted a more fibroblast-like morphology. Similar changes have been detected before in osteoarthritic chondrocytes, cells exposed to mechanical loading and cells undergoing de-differentiation. In addition, we detected several changes at the RNA level of genes associated with Wnt signaling pathway. Sclerostin, an inhibitor of Wnt signaling associated with bone density loss in space, was up-regulated in chondrocytes under simulated microgravity. There is not much known about the effects of sclerostin in articular cartilage, but a recent report suggested that sclerostin may protect cartilage from degradation because this up-regulation was also associated with decreased expression of several cartilage degrading genes. However, in addition to sclerostin up-regulation we also detected up-regulation of MMP-9 and down-regulation of aggrecan, both previously associated with arthritic conditions.

Lastly, in a collaborative effort with Dr. Jeff Willey from Wake Forest Baptist Medical Center, we are investigating the combined effects of radiation and microgravity in articular cartilage. Radiation alone up-regulated many genes involved with OSM/IL-6 signaling pathway, suggesting that these cells are activating a pro-inflammatory reaction. The morphological cytoskeleton of irradiated cells exposed to simulated microgravity had a more drastic change than that of microgravity alone. Our goal for the next year is to look at changes at the tissue level by isolating articular cartilage explants from bovine hooves and exposing them to simulated microgravity. In addition we will continue our collaborative efforts with Dr. Willey, as well as further investigating the effects of Wnt signaling pathway in chondrocytes and simulated microgravity.

Research Impact/Earth Benefits: It has been demonstrated that the musculoskeletal system is highly affected by radiation and microgravity during spaceflight, and there are exercise and nutrition countermeasures to prevent or minimize bone density loss and skeletal muscle atrophy after space missions. However, the effects of microgravity and radiation on articular cartilage health of the synovial joints have not been investigated, and pathological conditions such as arthritis can result in severely restricted mobility. Currently, there is no cure for arthritis due to the lack of understanding of the molecular mechanisms that trigger cartilage degradation. This is the first study to investigate the effects of radiation and simulated microgravity at the molecular level using several chondrocyte cell lines and a Rotating Wall Vessel bioreactor to simulate reduced microgravity. Our study found that chondrocytes respond to reduced gravity by changing their cytoskeletal morphology, similar to the morphological changes shown in studies using mechanical loading. We are still investigating the mechanism and mechanoreceptors responsible for re-arranging the cell morphology in response to changes in gravitational forces. Another interesting finding is the change in gene expression of molecules associated with the Wnt signaling pathway in response to simulated microgravity. The role of Wnt signaling in cartilage is well understood in embryology and development, and it has recently emerged as a critical regulator of bone and cartilage homeostasis. However, the role of Wnt signaling in arthritis is not understood, and recent studies found that sclerostin, an inhibitor of Wnt signaling, was up-regulated in mineralized cartilage and end-stage osteoarthritic samples. Sclerostin has been implicated in bone density loss in microgravity, and is now a promising therapeutic target to protect bones during space missions as well as in patients suffering from osteoporosis here on Earth.

Our study is the first to detect sclerostin up-regulation in chondrocytes exposed to simulated microgravity. The mechanisms and effects of sclerostin up-regulation and changes to the Wnt signaling pathway on cartilage homeostasis will be further investigated. If Wnt signaling is in fact associated with the onset of osteoarthritis as suggested recently by a few studies, the findings from our current study may help with the understanding of the underlying mechanism involved in osteoarthritis, and can help provide a new therapeutic target for the millions of people suffering from arthritis here on Earth.

Lastly, we detected the up-regulation of several genes involved in OSM/IL-6 signaling pathway in samples that received a 1Gy dose of radiation. Similar expression of these same genes have been detected in chondrocytes treated with OSM and IL-1B inflammatory cytokines, suggesting that radiation may trigger a similar reaction to that of cartilage exposed to inflammation. This is another novel finding that will require further investigation.

Task Progress & Bibliography Information FY2013 
Task Progress: Aims: 1: Investigate changes in gene expression after exposure to simulated microgravity. Hypothesis: Simulated microgravity will up-regulate pro-catabolic and anti-anabolic genes similar to the ones expressed after OSM and IL-1B pro-inflammatory cytokine treatments, characteristic of arthritic conditions. These changes will be assessed by RT-PCR and western blotting techniques.

2: Examine changes in cell-matrix interactions in response to simulated microgravity. Hypothesis: Exposure to simulated microgravity will have an effect on cell-matrix interactions resulting in a signaling cascade that alters the catabolic rate of chondrocytes.

3: Examine the effects of simulated microgravity on the cytoskeletal morphology of chondrocytes. Hypothesis: Chondrocytes respond to mechanical loading by changing their cytoskeletal morphology. Cells will have a similar change in morphology in response to unloading. Cytoskeletal changes will be studied by immunofluorescent staining and confocal microscopy techniques.

Bibliography Type: Description: (Last Updated: 11/12/2020)  Show Cumulative Bibliography Listing
 
 None in FY 2013
Project Title:  Induction of Early Stages of Osteoarthritis After Exposure to Microgravity (Postdoctoral Fellowship) Reduce
Fiscal Year: FY 2012 
Division: Human Research 
Research Discipline/Element:
HRP HHC:Human Health Countermeasures
Start Date: 11/01/2011  
End Date: 10/31/2013  
Task Last Updated: 10/26/2011 
Download report in PDF pdf
Principal Investigator/Affiliation:   Mellor, Liliana F. Ph.D. / North Carolina State University 
Address:  Department of Biomedical Engineering 
Cell Mechanics Laboratory 
Raleigh , NC 27695-7115 
Email: lfmellor@ncsu.edu 
Phone: 208-426-2238  
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: North Carolina State University 
Joint Agency:  
Comments: NOTE: formerly at Boise State University until fall 2013 (Ed., Jan 2014)  
Co-Investigator(s)
Affiliation: 
Oxford, Julia  MENTOR/ Boise State University 
Project Information: Grant/Contract No. NCC 9-58-PF02601 
Responsible Center: NSBRI 
Grant Monitor:  
Center Contact:   
Solicitation / Funding Source: 2011 NSBRI-RFA-11-01 Postdoctoral Fellowships 
Grant/Contract No.: NCC 9-58-PF02601 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:  
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) HHC:Human Health Countermeasures
Human Research Program Risks: (1) Osteo:Risk Of Early Onset Osteoporosis Due To Spaceflight (No longer used, July 2020)
Human Research Program Gaps: (1) Osteo04:We do not know the contribution of each risk factor on bone loss and recovery of bone strength, and which factors are the best targets for countermeasure application (IRP Rev E)
Task Description: POSTDOCTORAL FELLOWSHIP

The health of space crews during and after space missions has become a major concern of NASA. Several physiological changes have been associated with long-term exposure to microgravity, including skeletal muscles atrophy, immune system dysfunction, decreased nutrient intake, cardiovascular anomalies and bone density loss. One field that has yet to be explored is the possible effects of microgravity on the health of articular cartilage health, which is constantly exposed to mechanical forces under normal gravitational conditions on Earth. Disruption of cartilage homeostasis leads to pathological conditions such as arthritis, which according to the Center for Disease Control is the leading cause of disability in the United States.

Osteoarthritis (OA) is a degenerative joint disease that limits mobility of the affected joint due to the degradation of articular cartilage, and in advanced stages, can expose the highly innervated bone tissue, producing excruciating pain in the affected joint. Because of the limited regenerative capacity of articular cartilage, it is very important to detect early changes in the catabolic and anabolic rates that can lead to cartilage degradation.

This project’s goal is to investigate the effects of microgravity on cartilage homeostasis by exposing two chondrocyte cell lines to a modeled microgravity environment using a rotating wall vessel bioreactor. Cells exposed to microgravity will be compared to cells incubated under normal gravitational conditions as a control, as well as an arthritic-like cell model induced by treating chondrocytes with pro-inflammatory cytokines such as IL-1B and oncostatin M (OSM). Disruptions in cell-matrix interactions, changes in cytoskeletal morphology and gene up-regulation will be evaluated to determine changes in chondrocyte metabolism.

Hypothesis. Similar to bone, cartilage homeostasis can be compromised during exposure to microgravity, resulting in osteoarthritic-like conditions in astronauts after space missions.

This study will give a better insight to whether exposure to microgravity can make astronauts more prone to develop early osteoarthritis. In addition, since early stages of OA are hard to diagnose due to the lack of symptoms, the researchers also plan to investigate potential markers in the synovial fluid that can help detect early stages of OA.

Due to the limited capacity of regeneration in articular cartilage, early detection and treatment are key components to prevent the advanced stages of cartilage degradation, which is the leading cause of disability in the U.S., limiting the activities of nearly 21 million adults.

Research Impact/Earth Benefits: 0

Task Progress & Bibliography Information FY2012 
Task Progress: New project for FY2012.

Bibliography Type: Description: (Last Updated: 11/12/2020)  Show Cumulative Bibliography Listing
 
 None in FY 2012