Publications by authors named "Brian Johnstone"

86 Publications

Substance P and Alpha-Calcitonin Gene-Related Peptide Differentially Affect Human Osteoarthritic and Healthy Chondrocytes.

Front Immunol 2021 27;12:722884. Epub 2021 Aug 27.

Department of Orthopaedic Surgery, Experimental Orthopaedics, Center for Medical Biotechnology, University of Regensburg, Regensburg, Germany.

Osteoarthritis (OA) is a degenerative joint disease that not only causes cartilage loss but also structural damage in all joint tissues. Joints are innervated by alpha-calcitonin gene-related peptide (αCGRP) and substance P (SP)-positive sensory nerve fibers. Alteration of sensory joint innervation could be partly responsible for degenerative changes in joints that contribute to the development of OA. Therefore, our aim was to analyze and compare the molecular effects of SP and αCGRP on the metabolism of articular chondrocytes from OA patients and non-OA cartilage donors. We treated the cells with SP or αCGRP and analysed the influence of these neuropeptides on chondrocyte metabolism and modulation of signaling pathways. In chondrocytes from healthy cartilage, SP had minimal effects compared with its effects on OA chondrocytes, where it induced inflammatory mediators, inhibited chondrogenic markers and promoted apoptosis and senescence. Treatment with αCGRP also increased apoptosis and senescence and reduced chondrogenic marker expression in OA chondrocytes, but stimulated an anabolic and protective response in healthy chondrocytes. The catabolic influence of SP and αCGRP might be due to activation of ERK signaling that could be counteracted by an increased cAMP response. We suggest that a switch between the G-subunits of the corresponding receptors after binding their ligands SP or αCGRP plays a central role in mediating the observed effects of sensory neuropeptides on chondrocytes.
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http://dx.doi.org/10.3389/fimmu.2021.722884DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8430215PMC
December 2021

Surface topography as a tool to detect early changes in a posttraumatic equine model of osteoarthritis.

J Orthop Res 2021 Aug 27. Epub 2021 Aug 27.

Department of Clinical Sciences, Orthopaedic Research Center & C. Wayne McIlwraith Translational Medicine Institute, Colorado State University, Fort Collins, Colorado, USA.

The equine model of posttraumatic osteoarthritis (OA) mimics certain aspects of the naturally occurring disease, both in horses and humans. The objective of this study was to assess articular cartilage degeneration in a posttraumatic OA model using the established macroscopic and microscopic scoring systems and compare them with a novel surface topography analysis. OA was induced in the carpal joint of 15 (n = 15) mixed breed horses. Surface changes on the articular cartilage were characterized using osteochondral blocks from the third carpal bone (C3) and radial carpal bone using surface topography, standard histological grading, and gross evaluation of the joints. Significant differences were observed between OA and non-OA joints for gross evaluation scores. Microscopic scores of hematoxylin and eosin and Safranin O and Fast Green-stained sections demonstrated no differences between OA and non-OA joints. However, articular cartilage from the induced OA joint had significantly greater surface topography measurements compared with the sham treatment group, consistent with the changes seen on gross evaluation of joints. No significant correlations were noted between surface roughness measurements, histological assessment, and gross evaluation scores. The results suggest that surface topography analysis may provide a reliable objective approach to assess early changes in the cartilage surface in OA.
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http://dx.doi.org/10.1002/jor.25175DOI Listing
August 2021

Curcumin-primed human BMSC-derived extracellular vesicles reverse IL-1β-induced catabolic responses of OA chondrocytes by upregulating miR-126-3p.

Stem Cell Res Ther 2021 04 29;12(1):252. Epub 2021 Apr 29.

Department of Orthopaedic Surgery, Experimental Orthopaedics, Centre for Medical Biotechnology (ZMB/Biopark 1), University of Regensburg, Regensburg, Germany.

Background: Curcumin has anti-inflammatory effects and qualifies as a potential candidate for the treatment of osteoarthritis (OA). However, curcumin has limited bioavailability. Extracellular vesicles (EVs) are released by multiple cell types and act as molecule carrier during intercellular communication. We assume that EVs can maintain bioavailability and stability of curcumin after encapsulation. Here, we evaluated modulatory effects of curcumin-primed human (h)BMSC-derived EVs (Cur-EVs) on IL-1β stimulated human osteoarthritic chondrocytes (OA-CH).

Methods: CellTiter-Blue Viability- (CTB), Caspase 3/7-, and live/dead assays were used to determine range of cytotoxic curcumin concentrations for hBMSC and OA-CH. Cur-EVs and control EVs were harvested from cell culture supernatants of hBMSC by ultracentrifugation. Western blotting (WB), transmission electron microscopy, and nanoparticle tracking analysis were performed to characterize the EVs. The intracellular incorporation of EVs derived from PHK26 labeled and curcumin-primed or control hBMSC was tested by adding the labeled EVs to OA-CH cultures. OA-CH were pre-stimulated with IL-1β, followed by Cur-EV and control EV treatment for 24 h and subsequent analysis of viability, apoptosis, and migration (scratch assay). Relative expression of selected anabolic and catabolic genes was assessed with qRT-PCR. Furthermore, WB was performed to evaluate phosphorylation of Erk1/2, PI3K/Akt, and p38MAPK in OA-CH. The effect of hsa-miR-126-3p expression on IL-1β-induced OA-CH was determined using CTB-, Caspase 3/7-, live/dead assays, and WB.

Results: Cur-EVs promoted viability and reduced apoptosis of IL-1β-stimulated OA-CH and attenuated IL-1β-induced inhibition of migration. Furthermore, Cur-EVs increased gene expression of BCL2, ACAN, SOX9, and COL2A1 and decreased gene expression of IL1B, IL6, MMP13, and COL10A1 in IL-1β-stimulated OA-CH. In addition, phosphorylation of Erk1/2, PI3K/Akt, and p38 MAPK, induced by IL-1β, is prevented by Cur-EVs. Cur-EVs increased IL-1β-reduced expression of hsa-miR-126-3p and hsa-miR-126-3p mimic reversed the effects of IL-1β.

Conclusion: Cur-EVs alleviated IL-1β-induced catabolic effects on OA-CH by promoting viability and migration, reducing apoptosis and phosphorylation of Erk1/2, PI3K/Akt, and p38 MAPK thereby modulating pro-inflammatory signaling pathways. Treatment of OA-CH with Cur-EVs is followed by upregulation of expression of hsa-miR-126-3p which is involved in modulation of anabolic response of OA-CH. EVs may be considered as promising drug delivery vehicles of curcumin helping to alleviate OA.
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http://dx.doi.org/10.1186/s13287-021-02317-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8082633PMC
April 2021

A Large-Scale Bank of Organ Donor Bone Marrow and Matched Mesenchymal Stem Cells for Promoting Immunomodulation and Transplant Tolerance.

Front Immunol 2021 26;12:622604. Epub 2021 Feb 26.

Ossium Health, Indianapolis, IN, United States.

Induction of immune tolerance for solid organ and vascular composite allografts is the Holy Grail for transplantation medicine. This would obviate the need for life-long immunosuppression which is associated with serious adverse outcomes, such as infections, cancers, and renal failure. Currently the most promising means of tolerance induction is through establishing a mixed chimeric state by transplantation of donor hematopoietic stem cells; however, with the exception of living donor renal transplantation, the mixed chimerism approach has not achieved durable immune tolerance on a large scale in preclinical or clinical trials with other solid organs or vascular composite allotransplants (VCA). Ossium Health has established a bank of cryopreserved bone marrow (BM), termed "hematopoietic progenitor cell (HPC), Marrow," recovered from deceased organ donor vertebral bodies. This new source for hematopoietic cell transplant will be a valuable resource for treating hematological malignancies as well as for inducing transplant tolerance. In addition, we have discovered and developed a large source of mesenchymal stem (stromal) cells (MSC) tightly associated with the vertebral body bone fragment byproduct of the HPC, Marrow recovery process. Thus, these vertebral bone adherent MSC (vBA-MSC) are matched to the banked BM obtained from each donor, as opposed to third-party MSC, which enhances safety and potentially efficacy. Isolation and characterization of vBA-MSC from over 30 donors has demonstrated that the cells are no different than traditional BM-MSC; however, their abundance is >1,000-fold higher than obtainable from living donor BM aspirates. Based on our own unpublished data as well as reports published by others, MSC facilitate chimerism, especially at limiting hematopoietic stem and progenitor cell (HSPC) numbers and increase safety by controlling and/or preventing graft-vs.-host-disease (GvHD). Thus, vBA-MSC have the potential to facilitate mixed chimerism, promote complementary peripheral immunomodulatory functions and increase safety of BM infusions. Both HPC, Marrow and vBA-MSC have potential use in current VCA and solid organ transplant (SOT) tolerance clinical protocols that are amenable to "delayed tolerance." Current trials with HPC, Marrow are planned with subsequent phases to include vBA-MSC for tolerance of both VCA and SOT.
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http://dx.doi.org/10.3389/fimmu.2021.622604DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7959805PMC
September 2021

Norms for Clinical Use of CXM, a Real-Time Marker of Height Velocity.

J Clin Endocrinol Metab 2021 01;106(1):e255-e264

Research Center, Shriners Hospitals for Children, Portland, Oregon.

Context: Height velocity (HV) is difficult to assess because growth is very slow. The current practice of calculating it from measurements taken at several-month intervals is insufficient for managing children with growth disorders. We identified a bone growth by-product (collagen X biomarker, CXM) in blood that in preliminary analysis in healthy children correlated strongly with conventionally determined HV and displayed a pattern resembling published norms for HV vs age.

Objective: The goal was to confirm our initial observations supporting the utility of CXM as an HV biomarker in a larger number of individuals and establish working reference ranges for future studies.

Design, Settings, And Participants: CXM was assessed in archived blood samples from 302 healthy children and 10 healthy adults yielding 961 CXM measurements. A total of 432 measurements were plotted by age, and sex-specific reference ranges were calculated. Serial values from 116 participants were plotted against observed HV. Matched plasma, serum, and dried blood spot readings were compared.

Results: A correlation of blood CXM with conventional HV was confirmed. Scatter plots of CXM vs age showed a similar pattern to current HV norms, and CXM levels demarcated the pubertal growth spurt both in girls and boys. CXM levels differed little in matched serum, plasma, and dried blood spot samples.

Conclusions: Blood CXM offers a potential means to estimate HV in real time. Our results establish sex-specific, working reference ranges for assessing skeletal growth, especially over time. CXM stability in stored samples makes it well suited for retrospective studies.
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http://dx.doi.org/10.1210/clinem/dgaa721DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765635PMC
January 2021

Identification and characterization of a large source of primary mesenchymal stem cells tightly adhered to bone surfaces of human vertebral body marrow cavities.

Cytotherapy 2020 11 30;22(11):617-628. Epub 2020 Aug 30.

Ossium Health, Inc, Indianapolis, Indiana, USA; Department of Biomedical Sciences, College of Osteopathic Medicine, Marian University, Indianapolis, Indiana, USA; Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, Indiana, USA. Electronic address:

Background: Therapeutic allogeneic mesenchymal stromal cells (MSCs) are currently in clinical trials to evaluate their effectiveness in treating many different disease indications. Eventual commercialization for broad distribution will require further improvements in manufacturing processes to economically manufacture MSCs at scales sufficient to satisfy projected demands. A key contributor to the present high cost of goods sold for MSC manufacturing is the need to create master cell banks from multiple donors, which leads to variability in large-scale manufacturing runs. Therefore, the availability of large single donor depots of primary MSCs would greatly benefit the cell therapy market by reducing costs associated with manufacturing.

Methods: We have discovered that an abundant population of cells possessing all the hallmarks of MSCs is tightly associated with the vertebral body (VB) bone matrix and only liberated by proteolytic digestion. Here we demonstrate that these vertebral bone-adherent (vBA) MSCs possess all the International Society of Cell and Gene Therapy-defined characteristics (e.g., plastic adherence, surface marker expression and trilineage differentiation) of MSCs, and we have therefore termed them vBA-MSCs to distinguish this population from loosely associated MSCs recovered through aspiration or rinsing of the bone marrow compartment.

Results: Pilot banking and expansion were performed with vBA-MSCs obtained from 3 deceased donors, and it was demonstrated that bank sizes averaging 2.9 × 10 ± 1.35 × 10 vBA-MSCs at passage 1 were obtainable from only 5 g of digested VB bone fragments. Each bank of cells demonstrated robust proliferation through a total of 9 passages, without significant reduction in population doubling times. The theoretical total cell yield from the entire amount of bone fragments (approximately 300 g) from each donor with limited expansion through 4 passages is 100 trillion (1 × 10) vBA-MSCs, equating to over 10 doses at 10 × 10 cells/kg for an average 70-kg recipient.

Discussion: Thus, we have established a novel and plentiful source of MSCs that will benefit the cell therapy market by overcoming manufacturing and regulatory inefficiencies due to donor-to-donor variability.
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http://dx.doi.org/10.1016/j.jcyt.2020.07.003DOI Listing
November 2020

Physioxia Expanded Bone Marrow Derived Mesenchymal Stem Cells Have Improved Cartilage Repair in an Early Osteoarthritic Focal Defect Model.

Biology (Basel) 2020 Aug 17;9(8). Epub 2020 Aug 17.

Laboratory of Experimental Trauma Surgery, Department of Trauma Surgery, University Hospital Regensburg, Franz Josef Strauss Allee 11, 93053 Regensburg, Germany.

Focal early osteoarthritis (OA) or degenerative lesions account for 60% of treated cartilage defects each year. The current cell-based regenerative treatments have an increased failure rate for treating degenerative lesions compared to traumatic defects. Mesenchymal stem cells (MSCs) are an alternative cell source for treating early OA defects, due to their greater chondrogenic potential, compared to early OA chondrocytes. Low oxygen tension or physioxia has been shown to enhance MSC chondrogenic matrix content and could improve functional outcomes of regenerative therapies. The present investigation sought to develop a focal early OA animal model to evaluate cartilage regeneration and hypothesized that physioxic MSCs improve in vivo cartilage repair in both, post-trauma and focal early OA defects. Using a rabbit model, a focal defect was created, that developed signs of focal early OA after six weeks. MSCs cultured under physioxia had significantly enhanced in vitro MSC chondrogenic GAG content under hyperoxia with or without the presence of interleukin-1β (IL-1β). In both post-traumatic and focal early OA defect models, physioxic MSC treatment demonstrated a significant improvement in cartilage repair score, compared to hyperoxic MSCs and respective control defects. Future investigations will seek to understand whether these results are replicated in large animal models and the underlying mechanisms involved in in vivo cartilage regeneration.
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http://dx.doi.org/10.3390/biology9080230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7463623PMC
August 2020

Ischemia considerations for the development of an organ and tissue donor derived bone marrow bank.

J Transl Med 2020 08 5;18(1):300. Epub 2020 Aug 5.

Ossium Health, Inc., 5742 W. 74th St, Indianapolis, IN, 46278, USA.

Background: Deceased organ donors represent an untapped source of therapeutic bone marrow (BM) that can be recovered in 3-5 times the volume of that obtained from living donors, tested for quality, cryopreserved, and banked indefinitely for future on-demand use. A challenge for a future BM banking system will be to manage the prolonged ischemia times that are inevitable when bones procured at geographically-dispersed locations are shipped to distant facilities for processing. Our objectives were to: (a) quantify, under realistic field conditions, the relationship between ischemia time and the quality of hematopoietic stem and progenitor cells (HSPCs) derived from deceased-donor BM; (b) identify ischemia-time boundaries beyond which HSPC quality is adversely affected; (c) investigate whole-body cooling as a strategy for preserving cell quality; and (d) investigate processing experience as a variable affecting quality.

Methods: Seventy-five bones from 62 donors were analyzed for CD34+ viability following their exposure to various periods of warm-ischemia time (WIT), cold-ischemia time (CIT), and body-cooling time (BCT). Regression models were developed to quantify the independent associations of WIT, CIT, and BCT, with the viability and function of recovered HSPCs.

Results: Results demonstrate that under "real-world" scenarios: (a) combinations of warm- and cold-ischemia times favorable to the recovery of high-quality HSPCs are achievable (e.g., CD34+ cell viabilities in the range of 80-90% were commonly observed); (b) body cooling prior to bone recovery is detrimental to cell viability (e.g., CD34+ viability < 73% with, vs. > 89% without body cooling); (c) vertebral bodies (VBs) are a superior source of HSPCs compared to ilia (IL) (e.g., %CD34+ viability > 80% when VBs were the source, vs. < 74% when IL were the source); and (d) processing experience is a critical variable affecting quality.

Conclusions: Our models can be used by an emerging BM banking system to formulate ischemia-time tolerance limits and data-driven HSPC quality-acceptance standards.
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http://dx.doi.org/10.1186/s12967-020-02470-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7405448PMC
August 2020

Predicting and Promoting Human Bone Marrow MSC Chondrogenesis by Way of TGFβ Receptor Profiles: Toward Personalized Medicine.

Front Bioeng Biotechnol 2020 26;8:618. Epub 2020 Jun 26.

Regenerative Orthopaedics, AO Research Institute Davos, Davos, Switzerland.

The use of human mesenchymal stromal cells (hMSCs) for cartilage regeneration has been hampered by the inherent donor variation of primary monolayer expanded cells. Although CD markers are typically used to characterize cell populations, there is no correlation between CD marker profile and functional outcomes. Therefore, we aimed to discover novel predictive MSC chondrogenesis markers. The chondrogenic potential of primary human bone marrow MSCs (hBMSCs) over multiple passages was assessed by standard pellet culture. We confirmed that the ratio of TGFβ-RI/TGFβ-RII at the time of cell recovery from the tissue culture plastic reliably predicted chondrogenic potential. Furthermore, it is possible to prospectively characterize any human BMSC cell population as responders or non-responders with respect to chondrogenic differentiation potential. Transient increase of the ratio with siRNA knockdown of TGFβ-RII reproducibly recovered the chondrogenic differentiation ability of non-responsive MSCs. Together this offers an opportunity to produce a more functionally characterized cell population for use in autologous cartilage repair therapies.
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http://dx.doi.org/10.3389/fbioe.2020.00618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7333220PMC
June 2020

Comparing cellular bone matrices for posterolateral spinal fusion in a rat model.

JOR Spine 2020 Jun 15;3(2):e1084. Epub 2020 Mar 15.

Department of Orthopaedics and Rehabilitation Oregon Health & Science University Portland Oregon USA.

Introduction: Cellular bone matrices (CBM) are allograft products that provide three components essential to new bone formation: an osteoconductive scaffold, extracellular growth factors for cell proliferation and differentiation, and viable cells with osteogenic potential. This is an emerging technology being applied to augment spinal fusion procedures as an alternative to autografts.

Methods: We aim to compare the ability of six commercially-available human CBMs (Trinity ELITE®, ViviGen®, Cellentra®, Osteocel® Pro, Bio4® and Map3®) to form a stable spinal fusion using an athymic rat model of posterolateral fusion. Iliac crest bone from syngeneic rats was used as a control to approximate the human gold standard. The allografts were implanted at L4-5 according to vendor specifications in male athymic rats, with 15 rats in each group. MicroCT scans were performed at 48 hours and 6 weeks post-implantation. The rats were euthanized 6 weeks after surgery and the lumbar spines were harvested for X-ray, manual palpation and histology analysis by blinded reviewers.

Results: By manual palpation, five of 15 rats of the syngeneic bone group were fused at 6 weeks. While Trinity ELITE had eight of 15 and Cellentra 11 of 15 rats with stable fusion, only 2 of 15 of ViviGen-implanted spines were fused and zero of 15 of the Osteocel Pro, Bio4 and Map3 produced stable fusion. MicroCT analysis indicated that total bone volume increased from day 0 to week 6 for all groups except syngeneic bone group. Trinity ELITE (65%) and Cellentra (73%) had significantly greater bone volume increases over all other implants, which was consistent with the histological analysis.

Conclusion: Trinity ELITE and Cellentra were significantly better than other implants at forming new bone and achieving spinal fusion in this rat model at week 6. These results suggest that there may be large differences in the ability of different CBMs to elicit a successful fusion in the posterolateral spine.
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http://dx.doi.org/10.1002/jsp2.1084DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7323463PMC
June 2020

A quantitative serum biomarker of circulating collagen X effectively correlates with endochondral fracture healing.

J Orthop Res 2021 01 20;39(1):53-62. Epub 2020 Jul 20.

Department of Orthopaedic Surgery, Orthopaedic Trauma Institute, Zuckerberg San Francisco General Hospital (ZSFG), University of California, San Francisco (UCSF), San Francisco, California.

Currently, there are no standardized methods for quantitatively measuring fracture repair. Physicians rely on subjective physical examinations and qualitative evaluation of radiographs to detect mineralized tissue. Since most fractures heal indirectly through a cartilage intermediate, these tools are limited in their diagnostic utility of early repair. Prior to converting to the bone, cartilage undergoes hypertrophic maturation, characterized by the deposition of a provisional collagen X matrix. The objective of this study was to characterize the kinetics of a novel collagen X biomarker relative to other biological measurements of fracture healing using a murine model of endochondral fracture repair in which a closed, mid-shaft tibia fracture was created using the classic drop-weight technique. Serum was collected 5 to 42 days post-fracture in male and female mice and compared to uninjured controls (n = 8-12). Collagen X in the serum was quantified using a recently validated ELISA-based bioassay ("Cxm") and compared to genetic and histological markers of fracture healing and inflammation. We found the Cxm biomarker reliably increased from baseline to a statistically unique peak 14 days post-fracture that then resolved to pre-fracture levels by 3 weeks following injury. The shape and timing of the Cxm curve followed the genetic and histological expression of collagen X but did not show a strong correlation with local inflammatory states. Assessment of fracture healing progress is crucial to making correct and timely clinical decisions for patients. This Cxm bioassay represents a minimally invasive, inexpensive technique that could provide reliable information on the biology of the fracture to significantly improve clinical care.
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http://dx.doi.org/10.1002/jor.24776DOI Listing
January 2021

A Comparative Evaluation of Commercially Available Cell-Based Allografts in a Rat Spinal Fusion Model.

Int J Spine Surg 2020 Apr 30;14(2):213-221. Epub 2020 Apr 30.

Department of Orthopaedics and Rehabilitation, Oregon Health & Science University, Portland, Oregon.

Background: To evaluate the comparative abilities of commercially available, viable, cellular bone allografts to promote posterolateral spinal fusion.

Methods: Human allografts containing live cells were implanted in the athymic rat model of posterolateral spine fusion. Three commercially available allogeneic cellular bone matrices (Trinity Evolution, Trinity ELITE and Osteocel Plus) were compared with syngeneic iliac crest bone as the control. All spines underwent radiographs, manual palpation, and micro-computed tomography (CT) analysis after excision at 6 weeks. Histological sections of randomly selected spines were subjected to semiquantitative histopathological scoring for bone formation.

Results: By manual palpation, posterolateral fusion was detected in 40% (6/15) of spines implanted with syngeneic bone, whereas spines implanted with Trinity Evolution and Trinity ELITE allografts yielded 71% (10/14) and 77% (10/13) fusion, respectively. Only 7% (1/14) of spines implanted with Osteocel Plus allografts were judged fused by manual palpation (statistically significantly less than ELITE, < .0007, and Evolution, < .0013). The mineralized cancellous bone component of the allografts confounded radiographic analysis, but Trinity Evolution (0.452 ± 0.064) and Trinity ELITE (0.536 ± 0.109) allografts produced statistically significantly higher bone fusion mass volumes measured by quantitative micro-CT than did syngeneic bone (0.292 ± 0.109, < .0001 for ELITE and < .003 for Evolution) and Osteocel Plus (0.258 ± 0.103, < .0001). Semiquantitative histopathological scores supported these findings because the total bone and bone marrow scores reflected significantly better new bone and marrow formation in the Trinity groups than in the Osteocel Plus group.

Conclusions: The Trinity Evolution and Trinity ELITE cellular bone allografts were more effective at creating posterolateral fusion than either the Osteocel Plus allografts or syngeneic bone in this animal model.

Clinical Relevance: The superior fusion rate of Trinity cellular bone allografts may lead to better clinical outcome of spinal fusion surgeries.
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http://dx.doi.org/10.14444/7026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7188100PMC
April 2020

Mesenchymal Stem Cell Based Regenerative Treatment of the Knee: From Basic Science to Clinics.

Stem Cells Int 2019 9;2019:7608718. Epub 2019 Sep 9.

Center of Experimental Orthopaedics, Saarland University Medical Center and Saarland University, Homburg, Saar, Germany.

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http://dx.doi.org/10.1155/2019/7608718DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6754960PMC
September 2019

Multi-Disciplinary Approaches for Cell-Based Cartilage Regeneration.

J Orthop Res 2020 03 18;38(3):463-472. Epub 2019 Sep 18.

Integrative Research Institute for Regenerative Biomedical Engineering, Dongguk University, Goyang, Republic of Korea.

Articular cartilage does not regenerate in adults. A lot of time and resources have been dedicated to cartilage regeneration research. The current understanding suggests that multi-disciplinary approach including biologic, genetic, and mechanical stimulations may be needed for cell-based cartilage regeneration. This review summarizes contents of a workshop sponsored by International Combined Orthopaedic Societies during the 2019 annual meeting of the Orthopaedic Research Society held in Austin, Texas. Three approaches for cell-based cartilage regeneration were introduced, including cellular basis of chondrogenesis, gene-enhanced cartilage regeneration, and physical modulation to divert stem cells to chondrogenic cell fate. While the complicated nature of cartilage regeneration has not allowed us to achieve successful regeneration of hyaline articular cartilage so far, the utilization of multi-disciplinary approaches in various fields of biomedical engineering will provide means to achieve this goal faster. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:463-472, 2020.
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http://dx.doi.org/10.1002/jor.24458DOI Listing
March 2020

Physioxia Has a Beneficial Effect on Cartilage Matrix Production in Interleukin-1 Beta-Inhibited Mesenchymal Stem Cell Chondrogenesis.

Cells 2019 08 20;8(8). Epub 2019 Aug 20.

Laboratory of Experimental Trauma Surgery, Department of Trauma Surgery, University Hospital Regensburg, Franz Josef Strauss Allee 11, 93053 Regensburg, Germany.

Osteoarthritis (OA) is a degenerative condition that involves the production of inflammatory cytokines (e.g., interleukin-1β (IL-1β), tumour necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6)) that stimulate degradative enzymes, matrix metalloproteinases (MMPs) and aggrecanases (ADAMTS) resulting in articular cartilage breakdown. The presence of interleukin-1β (IL-1β) is one reason for poor clinical outcomes in current cell-based tissue engineering strategies for treating focal early osteoarthritic defects. Mesenchymal stem cells (MSCs) are a potential cell source for articular cartilage regeneration, although IL-1β has been shown to inhibit in vitro chondrogenesis. In vivo, articular chondrocytes reside under a low oxygen environment between 2-5% oxygen (physioxia) and have been shown to enhance in vitro MSC chondrogenic matrix content with reduced hypertrophic marker expression under these conditions. The present investigation sought to understand the effect of physioxia on IL-1β inhibited MSC chondrogenesis. MSCs expanded under physioxic (2% oxygen) and hyperoxic (20%) conditions, then chondrogenically differentiated as pellets in the presence of TGF-β1 and either 0.1 or 0.5 ng/mL IL-1β. Results showed that there were donor variations in response to physioxic culture based on intrinsic GAG content under hyperoxia. In physioxia responsive donors, MSC chondrogenesis significantly increased GAG and collagen II content, whilst hypertrophic markers were reduced compared with hyperoxia. In the presence of IL-1β, these donors showed a significant increase in cartilage matrix gene expression and GAG content relative to hyperoxic conditions. In contrast, a set of MSC donors were unresponsive to physioxia and showed no significant increase in matrix production independent of IL-1β presence. Thus, physioxia has a beneficial effect on MSC cartilage matrix production in responsive donors with or without IL-1β application. The mechanisms controlling the MSC chondrogenic response in both physioxia responsive and unresponsive donors are to be elucidated in future investigations.
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http://dx.doi.org/10.3390/cells8080936DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6721827PMC
August 2019

The Importance of Physioxia in Mesenchymal Stem Cell Chondrogenesis and the Mechanisms Controlling Its Response.

Int J Mol Sci 2019 Jan 23;20(3). Epub 2019 Jan 23.

Laboratory of Experimental Trauma Surgery, Department of Trauma Surgery, University Hospital Regensburg, Franz Josef Strauss Allee 11, 93053 Regensburg, Germany.

Articular cartilage covers the surface of synovial joints and enables joint movement. However, it is susceptible to progressive degeneration with age that can be accelerated by either previous joint injury or meniscectomy. This degenerative disease is known as osteoarthritis (OA) and it greatly affects the adult population. Cell-based tissue engineering provides a possible solution for treating OA at its earliest stages, particularly focal cartilage lesions. A candidate cell type for treating these focal defects are Mesenchymal Stem Cells (MSCs). However, present methods for differentiating these cells towards the chondrogenic lineage lead to hypertrophic chondrocytes and bone formation in vivo. Environmental stimuli that can stabilise the articular chondrocyte phenotype without compromising tissue formation have been extensively investigated. One factor that has generated intensive investigation in MSC chondrogenesis is low oxygen tension or physioxia (2⁻5% oxygen). In vivo articular cartilage resides at oxygen tensions between 1⁻4%, and in vitro results suggest that these conditions are beneficial for MSC expansion and chondrogenesis, particularly in suppressing the cartilage hypertrophy. This review will summarise the current literature regarding the effects of physioxia on MSC chondrogenesis with an emphasis on the pathways that control tissue formation and cartilage hypertrophy.
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http://dx.doi.org/10.3390/ijms20030484DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6387316PMC
January 2019

Serum C-peptide level correlates with the course of muscle tissue healing in the rabbit model of critical limb ischemia.

Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub 2019 Jun 7;163(2):132-140. Epub 2018 Sep 7.

Radiodiagnostic Institute, University Hospital Ostrava, Czech Republic Corresponding author: Vaclav Prochazka, e-mail.

Aim: The therapeutic potential of adipose-derived stem cell conditioned medium (ASC-CM) was studied in the rabbit model of critical limb ischemia (CLI).

Methods: Rabbits received treatment with ASC-CM or placebo. Gastrocnemius muscle tissue was collected 35 days after ischemia induction. Ischemic changes were evaluated in hematoxylin-eosin stained tissues for early (necrotic lesions/granulation tissue) and late (fibrous scars) phases of tissue repair. The expression of proangiogenic miR-126 was also evaluated using in situ hybridization. The levels of cytokines, insulin, and C-peptide were measured in blood.

Results: Early repair phases were observed more often in placebo-treated samples (45.5%) than in ASC-CM-treated ones (22.2%). However, the difference was not statistically significant. We demonstrated a statistically significant positive correlation between the early healing phases in tissue samples and C-peptide levels in peripheral blood. The expression of proangiogenic miR-126 was also shown in a number of structures in all phases of ischemic tissue healing.

Conclusion: Based on our results, we believe that treatment with ASC-CM has the potential to accelerate the healing process in ischemic tissues in the rabbit model of CLI. The whole healing process was accompanied by miR-126 tissue expression. C-peptide could be used to monitor the course of the tissue healing process.
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http://dx.doi.org/10.5507/bp.2018.048DOI Listing
June 2019

Regenerative medicine will make orthopaedic implants obsolete in our time Orthopaedic Research Society first annual meeting debate, San Diego, March 21st, 2017.

J Orthop Res 2018 10 24;36(10):2579-2585. Epub 2018 May 24.

Department of Oncology and Metabolism, University of Sheffield, Sheffield, United Kingdom.

The mission of the Orthopaedic Research Society is to promote and advance musculoskeletal research worldwide. With this in mind, the Annual Meeting Program Committee sought to establish a debate as a key component of the meeting. Our purpose was to provoke discussion on topics that are core to our mission and to engage all constituencies within the society by examining questions of broad relevance. To this end, the topic "Regenerative medicine will make orthopaedic implants obsolete in our time" was selected as the title of the inaugural debate. The arguments for and against the motion are presented in this perspectives article. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2579-2585, 2018.
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http://dx.doi.org/10.1002/jor.24033DOI Listing
October 2018

The Role of miR-126 in Critical Limb Ischemia Treatment Using Adipose-Derived Stem Cell Therapeutic Factor Concentrate and Extracellular Matrix Microparticles.

Med Sci Monit 2018 Jan 26;24:511-522. Epub 2018 Jan 26.

NeuroFX, LLC, Indianapolis, IN, USA.

BACKGROUND Paracrine factors secreted by adipose-derived stem cells can be captured, fractionated, and concentrated to produce therapeutic factor concentrate (TFC). The present study examined whether TFC effects could be enhanced by combining TFC with a biological matrix to provide sustained release of factors in the target region. MATERIAL AND METHODS Unilateral hind limb ischemia was induced in rabbits. Ischemic limbs were injected with either placebo control, TFC, micronized small intestinal submucosa tissue (SIS), or TFC absorbed to SIS. Blood flow in both limbs was assessed with laser Doppler perfusion imaging. Tissues harvested at Day 48 were assessed immunohistochemically for vessel density; in situ hybridization and quantitative real-time PCR were employed to determine miR-126 expression. RESULTS LDP ratios were significantly elevated, compared to placebo control, on day 28 in all treatment groups (p=0.0816, p=0.0543, p=0.0639, for groups 2-4, respectively) and on day 36 in the TFC group (p=0.0866). This effect correlated with capillary density in the SIS and TFC+SIS groups (p=0.0093 and p=0.0054, respectively, compared to placebo). A correlation was observed between miR-126 levels and LDP levels at 48 days in SIS and TFC+SIS groups. CONCLUSIONS A single bolus administration of TFC and SIS had early, transient effects on reperfusion and promotion of ischemia repair. The effects were not additive. We also discovered that TFC modulated miR-126 levels that were expressed in cell types other than endothelial cells. These data suggested that TFC, alone or in combination with SIS, may be a potent therapy for patients with CLI that are at risk of amputation.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5795917PMC
http://dx.doi.org/10.12659/msm.905442DOI Listing
January 2018

Dynamic Mechanical Compression of Chondrocytes for Tissue Engineering: A Critical Review.

Front Bioeng Biotechnol 2017 11;5:76. Epub 2017 Dec 11.

Department of Orthopaedics and Rehabilitation, Oregon Health & Science University, Portland, OR, United States.

Articular cartilage functions to transmit and translate loads. In a classical structure-function relationship, the tissue resides in a dynamic mechanical environment that drives the formation of a highly organized tissue architecture suited to its biomechanical role. The dynamic mechanical environment includes multiaxial compressive and shear strains as well as hydrostatic and osmotic pressures. As the mechanical environment is known to modulate cell fate and influence tissue development toward a defined architecture , dynamic mechanical loading has been hypothesized to induce the structure-function relationship during attempts at regeneration of articular cartilage. Researchers have designed increasingly sophisticated bioreactors with dynamic mechanical regimes, but the response of chondrocytes to dynamic compression and shear loading remains poorly characterized due to wide variation in study design, system variables, and outcome measurements. We assessed the literature pertaining to the use of dynamic compressive bioreactors for generation of cartilaginous tissue from primary and expanded chondrocytes. We used specific search terms to identify relevant publications from the PubMed database and manually sorted the data. It was very challenging to find consensus between studies because of species, age, cell source, and culture differences, coupled with the many loading regimes and the types of analyses used. Early studies that evaluated the response of primary bovine chondrocytes within hydrogels, and that employed dynamic single-axis compression with physiologic loading parameters, reported consistently favorable responses at the tissue level, with upregulation of biochemical synthesis and biomechanical properties. However, they rarely assessed the cellular response with gene expression or mechanotransduction pathway analyses. Later studies that employed increasingly sophisticated biomaterial-based systems, cells derived from different species, and complex loading regimes, did not necessarily corroborate prior positive results. These studies report positive results with respect to very specific conditions for cellular responses to dynamic load but fail to consistently achieve significant positive changes in relevant tissue engineering parameters, particularly collagen content and stiffness. There is a need for standardized methods and analyses of dynamic mechanical loading systems to guide the field of tissue engineering toward building cartilaginous implants that meet the goal of regenerating articular cartilage.
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http://dx.doi.org/10.3389/fbioe.2017.00076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5732133PMC
December 2017

Physioxia Promotes the Articular Chondrocyte-Like Phenotype in Human Chondroprogenitor-Derived Self-Organized Tissue.

Tissue Eng Part A 2018 02 7;24(3-4):264-274. Epub 2017 Jul 7.

1 Department of Orthopaedics & Rehabilitation, Oregon Health & Science University , Portland, Oregon.

Introduction: Biomaterial-based tissue engineering has not successfully reproduced the structural architecture or functional mechanical properties of native articular cartilage. In scaffold-free tissue engineering systems, cells secrete and organize the entire extracellular matrix over time in response to environmental signals such as oxygen level. In this study, we investigated the effect of oxygen on the formation of neocartilage from human-derived chondrogenic cells.

Materials And Methods: Articular chondrocytes (ACs) and articular cartilage progenitor cells (ACPs) derived from healthy human adults were guided toward cell condensation by centrifugation onto plate inserts that were uncoated or coated with either agarose or fibronectin. Neocartilage discs were cultured at hyperoxic (20%) or physioxic (5%) oxygen levels, and biochemical, biomechanical, and molecular analyses were used to compare the cartilage produced by ACs versus ACPs.

Results: Fibronectin-coated inserts proved optimal for growing cartilaginous discs from both cell types. In comparison with culture in hyperoxia, AC neocartilage cultured at physioxia exhibited a significant increase in chondrogenic gene expression, proteoglycan production, and mechanical properties with a concomitant decrease in collagen content. At both oxygen levels, ACP-derived neocartilage produced tissue with significantly enhanced mechanical properties and collagen content relative to AC-derived neocartilage. Both ACs and ACPs produced substantial collagen II and reduced levels of collagens I and X in physioxia relative to hyperoxia. Neocartilage from ACPs exhibited anisotropic organization characteristic of native cartilage with respect to collagen VI of the pericellular matrix when compared with AC-derived neocartilage; however, only ACs produced abundant surface-localized lubricin.

Discussion And Conclusions: Guiding human-derived cells toward condensation and subsequent culture in physioxia promoted the articular cartilage tissue phenotype for ACs and ACPs. Unlike ACs, ACPs are clonable and highly expandable while retaining chondrogenicity. The ability to generate large tissues utilizing a scaffold-free approach from a single autologous progenitor cell may represent a promising source of neocartilage destined for cartilage repair.
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http://dx.doi.org/10.1089/ten.TEA.2016.0510DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5792248PMC
February 2018

Responses to altered oxygen tension are distinct between human stem cells of high and low chondrogenic capacity.

Stem Cell Res Ther 2016 10 20;7(1):154. Epub 2016 Oct 20.

Oregon Health & Science University, 3181 SW Sam Jackson Park Road, OP31, Portland, OR, 97239, USA.

Background: Lowering oxygen from atmospheric level (hyperoxia) to the physiological level (physioxia) of articular cartilage promotes mesenchymal stem cell (MSC) chondrogenesis. However, the literature is equivocal regarding the benefits of physioxic culture on preventing hypertrophy of MSC-derived chondrocytes. Articular cartilage progenitors (ACPs) undergo chondrogenic differentiation with reduced hypertrophy marker expression in hyperoxia but have not been studied in physioxia. This study sought to delineate the effects of physioxic culture on both cell types undergoing chondrogenesis.

Methods: MSCs were isolated from human bone marrow aspirates and ACP clones were isolated from healthy human cartilage. Cells were differentiated in pellet culture in physioxia (2 % oxygen) or hyperoxia (20 % oxygen) over 14 days. Chondrogenesis was characterized by biochemical assays and gene and protein expression analysis.

Results: MSC preparations and ACP clones of high intrinsic chondrogenicity (termed high-GAG) produced abundant matrix in hyperoxia and physioxia. Poorly chondrogenic cells (low-GAG) demonstrated a significant fold-change matrix increase in physioxia. Both high-GAG and low-GAG groups of MSCs and ACPs significantly upregulated chondrogenic genes; however, only high-GAG groups had a concomitant decrease in hypertrophy-related genes. High-GAG MSCs upregulated many common hypoxia-responsive genes in physioxia while low-GAG cells downregulated most of these genes. In physioxia, high-GAG MSCs and ACPs produced comparable type II collagen but less type I collagen than those in hyperoxia. Type X collagen was detectable in some ACP pellets in hyperoxia but reduced or absent in physioxia. In contrast, type X collagen was detectable in all MSC preparations in hyperoxia and physioxia.

Conclusions: MSC preparations and ACP clones had a wide range of chondrogenicity between donors. Physioxia significantly enhanced the chondrogenic potential of both ACPs and MSCs compared with hyperoxia, but the magnitude of response was inversely related to intrinsic chondrogenic potential. Discrepancies in the literature regarding MSC hypertrophy in physioxia can be explained by the use of low numbers of preparations of variable chondrogenicity. Physioxic differentiation of MSC preparations of high chondrogenicity significantly decreased hypertrophy-related genes but still produced type X collagen protein. Highly chondrogenic ACP clones had significantly lower hypertrophic gene levels, and there was little to no type X collagen protein in physioxia, emphasizing the potential advantage of these cells.
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http://dx.doi.org/10.1186/s13287-016-0419-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5073443PMC
October 2016

Bone marrow cell characteristics associated with patient profile and cardiac performance outcomes in the LateTIME-Cardiovascular Cell Therapy Research Network (CCTRN) trial.

Am Heart J 2016 09 6;179:142-50. Epub 2016 Jul 6.

Texas Heart Institute, CHI St. Luke's Health Baylor College of Medicine Medical Center, Houston, TX.

Background: Although several preclinical studies have shown that bone marrow cell (BMC) transplantation promotes cardiac recovery after myocardial infarction, clinical trials with unfractionated bone marrow have shown variable improvements in cardiac function.

Methods: To determine whether in a population of post-myocardial infarction patients, functional recovery after BM transplant is associated with specific BMC subpopulation, we examined the association between BMCs with left ventricular (LV) function in the LateTIME-CCTRN trial.

Results: In this population, we found that older individuals had higher numbers of BM CD133(+) and CD3(+) cells. Bone marrow from individuals with high body mass index had lower CD45(dim)/CD11b(dim) levels, whereas those with hypertension and higher C-reactive protein levels had higher numbers of CD133(+) cells. Smoking was associated with higher levels of CD133(+)/CD34(+)/VEGFR2(+) cells and lower levels of CD3(+) cells. Adjusted multivariate analysis indicated that CD11b(dim) cells were negatively associated with changes in LV ejection fraction and wall motion in both the infarct and border zones. Change in LV ejection fraction was positively associated with CD133(+), CD34(+), and CD45(+)/CXCR4(dim) cells as well as faster BMC growth rates in endothelial colony forming assays.

Conclusions: In the LateTIME population, BM composition varied with patient characteristics and treatment. Irrespective of cell therapy, recovery of LV function was greater in patients with greater BM abundance of CD133(+) and CD34(+) cells and worse in those with higher levels of CD11b(dim) cells. Bone marrow phenotype might predict clinical response before BMC therapy and administration of selected BM constituents could potentially improve outcomes of other future clinical trials.
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http://dx.doi.org/10.1016/j.ahj.2016.06.018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5014395PMC
September 2016

Use of MicroRNA biomarkers to distinguish enchondroma from low-grade chondrosarcoma.

Connect Tissue Res 2017 Mar 7;58(2):155-161. Epub 2016 Jun 7.

a Bone and Joint Center , Henry Ford Hospital , Detroit , MI , USA.

Establishing a definitive diagnosis between benign enchondroma versus low-grade chondrosarcoma presents a potential challenge to both clinicians and pathologists. microRNAs (small non-coding RNAs) have proven to be effective biomarkers for the identification of tumors and tumor progression. We present analysis, both array and quantitative PCR, that shows consistently and substantially increased expression of two microRNAs, miRs-181a and -138, in low-grade chondrosarcomas compared with enchondromas. The data suggest these microRNAs would provide an analytical distinction between the chondrosarcoma and benign neoplasms that can be performed in formalin-fixed paraffin-embedded specimens. Together with recent publications, these data indicate that miRs-181a and -138 also play a role in tumor development and homeostasis and may provide new targets for the development of much needed therapeutic intervention.
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http://dx.doi.org/10.1080/03008207.2016.1197212DOI Listing
March 2017

GDNF secreted from adipose-derived stem cells stimulates VEGF-independent angiogenesis.

Oncotarget 2016 Jun;7(24):36829-36841

Department of Neurology, Indiana University School of Medicine, Indianapolis, IN 46202, USA.

Adipose tissue stroma contains a population of mesenchymal stem cells (MSC) promote new blood vessel formation and stabilization. These adipose-derived stem cells (ASC) promote de novo formation of vascular structures in vitro. We investigated the angiogenic factors secreted by ASC and discovered that glial-derived neurotrophic factor (GDNF) is a key mediator for endothelial cell network formation. It was found that both GDNF alone or present in ASC-conditioned medium (ASC-CM) stimulated capillary network formation by using human umbilical vein endothelial cells (HUVECs) and such an effect was totally independent of vascular endothelial growth factor (VEGF) activity. Additionally, we showed stimulation of capillary network formation by GDNF, but not VEGF, could be blocked by the Ret (rearranged during transfection) receptor antagonist RPI-1, a GDNF signaling inhibitor. Furthermore, GDNF were found to be overexpressed in cancer cells that were resistant to the anti-angiogenic treatment using the VEGF antibody. Cancer cells in the liver hepatocellular carcinoma (HCC), a non-nervous related cancer, highly overexpressed GDNF as compared to normal liver cells. Our data strongly suggest that, in addition to VEGF, GDNF secreted by ASC and HCC cells, may be another important factor promoting pathological neovascularization. Thus, GDNF may be a potential therapeutic target for HCC and obesity treatments.
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http://dx.doi.org/10.18632/oncotarget.9208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5095042PMC
June 2016

Identification of Bone Marrow Cell Subpopulations Associated With Improved Functional Outcomes in Patients With Chronic Left Ventricular Dysfunction: An Embedded Cohort Evaluation of the FOCUS-CCTRN Trial.

Cell Transplant 2016 19;25(9):1675-1687. Epub 2015 Nov 19.

Texas Heart Institute, CHI St. Luke's Health, Houston, TX, USA.

In the current study, we sought to identify bone marrow-derived mononuclear cell (BM-MNC) subpopulations associated with a combined improvement in left ventricular ejection fraction (LVEF), left ventricular end-systolic volume (LVESV), and maximal oxygen consumption (VO max) in patients with chronic ischemic cardiomyopathy 6 months after receiving transendocardial injections of autologous BM-MNCs or placebo. For this prospectively planned analysis, we conducted an embedded cohort study comprising 78 patients from the FOCUS-Cardiovascular Cell Therapy Research Network (CCTRN) trial. Baseline BM-MNC immunophenotypes and progenitor cell activity were determined by flow cytometry and colony-forming assays, respectively. Previously stable patients who demonstrated improvement in LVEF, LVESV, and VO max during the 6-month course of the FOCUS-CCTRN study (group 1, n = 17) were compared to those who showed no change or worsened in one to three of these endpoints (group 2, n = 61) and to a subset of patients from group 2 who declined in all three functional endpoints (group 2A, n = 11). Group 1 had higher frequencies of B-cell and CXCR4 BM-MNC subpopulations at study baseline than group 2 or 2A. Furthermore, patients in group 1 had fewer endothelial colony-forming cells and monocytes/macrophages in their bone marrow than those in group 2A. To our knowledge, this is the first study to show that in patients with ischemic cardiomyopathy, certain bone marrow-derived cell subsets are associated with improvement in LVEF, LVESV, and VO max at 6 months. These results suggest that the presence of both progenitor and immune cell populations in the bone marrow may influence the natural history of chronic ischemic cardiomyopathy-even in stable patients. Thus, it may be important to consider the bone marrow composition and associated regenerative capacity of patients when assigning them to treatment groups and evaluating the results of cell therapy trials.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5088500PMC
http://dx.doi.org/10.3727/096368915X689901DOI Listing
November 2017

Adipose-derived Stem Cell Conditioned Media Extends Survival time of a mouse model of Amyotrophic Lateral Sclerosis.

Sci Rep 2015 Nov 20;5:16953. Epub 2015 Nov 20.

Department of Neurology, Indiana University School of Medicine, Indianapolis, IN.

Adipose stromal cells (ASC) secrete various trophic factors that assist in the protection of neurons in a variety of neuronal death models. In this study, we tested the effects of human ASC conditional medium (ASC-CM) in human amyotrophic lateral sclerosis (ALS) transgenic mouse model expressing mutant superoxide dismutase (SOD1(G93A)). Treating symptomatic SOD1(G93A) mice with ASC-CM significantly increased post-onset survival time and lifespan. Moreover, SOD1(G93A) mice given ASC-CM treatment showed high motor neuron counts, less activation of microglia and astrocytes at an early symptomatic stage in the spinal cords under immunohistochemical analysis. SOD1(G93A) mice treated with ASC-CM for 7 days showed reduced levels of phosphorylated p38 (pp38) in the spinal cord, a mitogen-activated protein kinase that is involved in both inflammation and neuronal death. Additionally, the levels of α-II spectrin in spinal cords were also inhibited in SOD1(G93A) mice treated with ASC-CM for 3 days. Interestingly, nerve growth factor (NGF), a neurotrophic factor found in ASC-CM, played a significant role in the protection of neurodegeneration inSOD1(G93A) mouse. These results indicate that ASC-CM has the potential to develop into a novel and effective therapeutic treatment for ALS.
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http://dx.doi.org/10.1038/srep16953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4653659PMC
November 2015

Therapeutic Potential of Adipose-Derived Therapeutic Factor Concentrate for Treating Critical Limb Ischemia.

Cell Transplant 2016 30;25(9):1623-1633. Epub 2015 Oct 30.

Radiodiagnostic Institute, University Hospital Ostrava, Ostrava, Czech Republic.

Transplantation of adipose-derived stem cells (ADSCs) is an emerging therapeutic option for addressing intractable diseases such as critical limb ischemia (CLI). Evidence suggests that therapeutic effects of ADSCs are primarily mediated through paracrine mechanisms rather than transdifferentiation. These secreted factors can be captured in conditioned medium (CM) and concentrated to prepare a therapeutic factor concentrate (TFC) composed of a cocktail of beneficial growth factors and cytokines that individually and in combination demonstrate disease-modifying effects. The ability of a TFC to promote reperfusion in a rabbit model of CLI was evaluated. A total of 27 adult female rabbits underwent surgery to induce ischemia in the left hindlimb. An additional five rabbits served as sham controls. One week after surgery, the ischemic limbs received intramuscular injections of either (1) placebo (control medium), (2) a low dose of TFC, or (3) a high dose of TFC. Limb perfusion was serially assessed with a Doppler probe. Blood samples were analyzed for growth factors and cytokines. Tissue was harvested postmortem on day 35 and assessed for capillary density by immunohistochemistry. At 1 month after treatment, tissue perfusion in ischemic limbs treated with a high dose of TFC was almost double (p < 0.05) that of the placebo group [58.8 ± 23 relative perfusion units (RPU) vs. 30.7 ± 13.6 RPU; mean ± SD]. This effect was correlated with greater capillary density in the affected tissues and with transiently higher serum levels of the angiogenic and prosurvival factors vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF). The conclusions from this study are that a single bolus administration of TFC demonstrated robust effects for promoting tissue reperfusion in a rabbit model of CLI and that a possible mechanism of revascularization was promotion of angiogenesis by TFC. Results of this study demonstrate that TFC represents a potent therapeutic cocktail for patients with CLI, many of whom are at risk for amputation of the affected limb.
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http://dx.doi.org/10.3727/096368915X689767DOI Listing
November 2017

Hypoxia-inducible factor 3-alpha expression is associated with the stable chondrocyte phenotype.

J Orthop Res 2015 Nov 14;33(11):1561-70. Epub 2015 Jul 14.

Department of Orthopaedics and Rehabilitation, Oregon Health & Science University, Portland, Oregon.

The hypoxia-inducible factors HIF-1α and HIF-2α are important regulators of the chondrocyte phenotype but little is known about HIF-3α in cartilage. The objective of this study was to characterize HIF-3α (HIF3A) expression during chondrocyte differentiation in vitro and in native cartilage tissues. HIF3A, COL10A1, and MMP13 were quantified in mesenchymal stem cells (MSCs) and articular chondrocytes from healthy and osteoarthritic (OA) tissue in three-dimensional cultures and in human embryonic epiphyses and adult articular cartilage. HIF3A was found to have an inverse association with hypertrophic markers COL10A1 and MMP13 in chondrogenic cells and tissues. In healthy chondrocytes, HIF3A was induced by dexamethasone and increased during redifferentiation. By comparison, HIF3A expression was extremely low in chondrogenically differentiated MSCs expressing high levels of COL10A1 and MMP13. HIF3A was also lower in redifferentiated OA chondrocytes than in healthy chondrocytes. In human embryonic epiphyseal tissue, HIF3A expression was lowest in the hypertrophic zone. Distinct splice patterns were also found in embryonic cartilage when compared with adult articular cartilage and redifferentiated chondrocytes. These in vitro and in vivo findings suggest that HIF3A levels are indicative of the hypertrophic state of chondrogenic cells and one or more splice variants may be important regulators of the chondrocyte phenotype.
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http://dx.doi.org/10.1002/jor.22930DOI Listing
November 2015
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