Publications by authors named "Rajalakshmanan Eswaramoorthy"

15 Publications

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Molecular docking analysis and evaluation of the antibacterial and antioxidant activities of the constituents of Ocimum cufodontii.

Sci Rep 2021 May 12;11(1):10101. Epub 2021 May 12.

Department of Applied Chemistry, School of Applied Natural Science, Adama Science and Technology University, P.O.Box 1888, Adama, Ethiopia.

Ocimum cufodontii ((Lanza) A.J.Paton) has been traditionally used in Ethiopia against bacteria. The extracts of the leaves and roots of O. cufodontii after silica gel column chromatography furnished compounds 1-5, compounds 3 and 4 are new natural products. The oil from the hydro-distillation of the leaves, after analyzed with GC-MS, has led to the identification of β-caryophyllene as a principal component, suggesting the essential oil as medicine and spices to enhance the taste of food. The constituents of O. cufodontii were assessed for their antibacterial activity against E. coli, K. pneumonia, S. typhymurium and S. aureus. The best activity was displayed against S. aureus by the hexane extract of the roots, compound 4, and the essential oil with an inhibition zone of 17, 15, and 19 mm, respectively. Molecular docking analysis revealed that compound 1 has better docking efficiency and forms hydrophobic interactions with five amino acids (ARG192, PHE196, GLU185, GLU193, and LYS189). This suggests that the compounds may act as potential inhibitors of DNA gyrase. The constituents were also assessed for their antioxidant activities using DPPH, ferric thicyanate and ferric reducing power assay. The hexane extracts of the roots inhibited the DPPH radical and peroxide formation by 90.5 and 83%, respectively, suggesting the potential of the extract as an antioxidant. Furthermore, the hexane extract of the roots of O. cufodontii exhibited the maximum reducing power compared with the EtOAc and methanol extracts. Hence, the activity displayed herein indicated as the plant has great potential as a remedy for diseases caused by bacteria and radicals.
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http://dx.doi.org/10.1038/s41598-021-89557-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8115310PMC
May 2021

Antibacterial and antioxidant activities of extracts and isolated compounds from the roots extract of Cucumis prophetarum and in silico study on DNA gyrase and human peroxiredoxin 5.

BMC Chem 2021 May 6;15(1):32. Epub 2021 May 6.

Department of Applied Chemistry, School of Applied Natural Science, Adama Science and Technology University, P.O.Box 1888, Adama, Ethiopia.

Background: Cucumis prophetarum is traditionally used to treat liver and lung disorders, heart failure, diarrhea, gonorrhea, skin infections, intestinal problems and cancer. In the present work, the isolation of two novel compounds along with their antibacterial and antioxidant activities is reported for the first time.

Methods: Silica gel column chromatography was applied to separate constituents of the roots of C. prophetarum. The structures of isolated compounds were established using H NMR, C NMR, DEPT-135, COSY, HSQC and HMBC. Agar well diffusion, DPPH assay and ferric thiocyante methods were used for antibacterial, radical scavenging and anti-lipid peroxidation activities, respectively. AutoDock Vina open source program was used for molecular docking analysis.

Results: Evaluation of the in vitro antibacterial activity of the constituents against S. aureus, B. subtilis, E. coli and S. thyphimurium revealed that the hexane extract were active against E. coli with IZ of 15.0 ± 1.41 mm, whereas an IZ of 14.6 ± 1.70 mm for MeOH extract was observed against S. aureus. Compound 1 displayed IZ of 13.6 ± 0.94 mm against E. coli and curcurbiatin 2 showed activity against B. subtilis with IZ of 13.3 ± 0.54 mm. The molecular docking analysis showed that cucurbitacins 2 and 3 have binding energy of -6.7 and -6.9 kcal/mol, respectively. The methanol and the hexane extracts of the roots of C. prophetarum inhibited DPPH radical by 70.4 and 63.3% at 100 µg/mL, respectively. On the other hand, the methanol extract inhibited lipid peroxidation by 53.0%.

Conclusion: The present study identified five compounds from the root extracts of C. prophetarum, of which two are novel cucurbitacins (1, 2). The in vitro antibacterial activity of the hexane and methanol extracts was better than the activity displayed by the isolated compounds. This is probably due to the synergistic effects of the constituents present in the root extract. The in silico molecular docking study results showed that, compounds 2 and 3 have minimum binding energy and have good affinity toward the active pocket, thus, they may be considered as good inhibitor of DNA gyrase B. Furthermore, the "drug-likeness" and ADMET prediction of compounds 2-5 nearly showed compliance with the Lipinski rule, with good absorption, distribution, metabolism, and excretion generally. The radical scavenging and anti-lipid peroxidation activities of the extracts were better than the isolated compounds. This is attributed to the presence of phenolics and flavonoids as minor constituents in the extracts of these species. Therefore, the in vitro antibacterial activity and molecular docking analysis suggest the potential use of the isolated compounds as medicine which corroborates the traditional use of the roots of C. prophetarum.
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http://dx.doi.org/10.1186/s13065-021-00758-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8103605PMC
May 2021

Synthesis, Molecular Docking Analysis, and Evaluation of Antibacterial and Antioxidant Properties of Stilbenes and Pinacol of Quinolines.

Adv Pharmacol Pharm Sci 2021 2;2021:6635270. Epub 2021 Mar 2.

Department of Applied Chemistry, Adama Science and Technology University, Adama, Ethiopia.

Emergence of antimicrobial resistance to standard commercial drugs has become a critical public health concern worldwide. Hence, novel antimicrobials with improved biological activities are urgently needed. In this regard, a series of quinoline-stilbene derivatives were synthesized from substituted quinoline and benzyltriphenylphosphonium chloride using Wittig reaction. Furthermore, a novel pinacol of quinoline was synthesized by pinacolinazation of 2-methoxyquinoline-3-carbaldehyde which was achieved by aluminum powder-potassium hydroxide reagent combination at ambient temperature in methanol. The structures of the synthesized compounds were established based on their spectral data. The antibacterial activities of the synthesized compounds were evaluated in vitro by the paper disc diffusion method against two Gram-positive bacteria ( and ) and two Gram-negative bacteria ( and ). The best activity was displayed by compound against E. coli with an inhibition zone of 16.0 ± 0.82 mm and 14.67 ± 0.94 mm at 500 and 250 g/mL, respectively. This is close to ciprofloxacin which is used as a positive control. The results of in silico molecular docking evaluation of the compounds against DNA gyraseB were in good agreement with the in vitro antibacterial analysis. Compounds 19 (-6.9 kcal/mol) and (-7.1 kcal/mol) showed the maximum binding affinity close to ciprofloxacin (-7.3 kcal/mol) used as positive control. Therefore, the antibacterial activity displayed by these compounds is encouraging for further investigation to improve the activities of quinoline-stilbenes by incorporating various bioisosteric groups in one or more positions of the phenyl nuclei for their potential pharmacological use. Findings of the DPPH radical scavenging assay indicated that some of the quinolone stilbenes and pinacol possess moderate antioxidant properties compared to ascorbic acid used as a natural antioxidant.
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http://dx.doi.org/10.1155/2021/6635270DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7946464PMC
March 2021

In silico Molecular Docking, DFT Analysis and ADMET Studies of Carbazole Alkaloid and Coumarins from Roots of : A Potent Inhibitor for Quorum Sensing.

Adv Appl Bioinform Chem 2021 5;14:13-24. Epub 2021 Feb 5.

Department of Applied Chemistry, School of Applied Natural Science, Adama Science and Technology University, Adama, 1888, Ethiopia.

Introduction: In modern drug design, in silico methods are largely used to understand drug-receptor interactions and quantum chemical properties. In the present study, a computational de novo design approach was used to confirm mode of binding for antibacterial activity, elucidating quantum chemical properties and ADMET-drug-likeness of carbazole alkaloid () and three coumarins (-) isolated from roots of .

Methods: Docking studies were performed with DNA-Gyrase (6F86) and LasR binding domain (2UV0) employing a flexible ligand docking approach using AutoDock Vina. SwissADME prediction and toxicological predictions were performed by ADMET. The optimized structures and molecular electrostatic potential surface of the isolated compounds were predicted by DFT analysis using B3LYP/6-31G basis levels.

Results And Discussion: The docking results revealed that compound showed better docking scores against both DNA gyrase B and LasR binding domain compared with ciprofloxacin with potential as an inhibitor of bacterial DNA gyrase and quorum sensing LasR binding domain. The SwissADME prediction results showed that all the isolated compounds () satisfy Lipinski's rule of five with zero violations. Toxicological prediction results suggested that all compounds and ciprofloxacin are non-hepatotoxic, non-carcinogenic, non-irritant, immunogenic, and non-cytotoxic. The DFT analysis results revealed that compound has large electronegativity (χeV), global softness (σ eV), global electrophilicity (ωeV), and mutagenicity value closer to ciprofloxacin (with LD value of 480 mg/kg) suggesting better bioactivity and chemical reactivity with considerable intra-molecular charge transfer between electron-donor to electron-acceptor groups.

Conclusion: Overall, compound may serve as a lead molecule that could be developed into a potent DNA gyrase B inhibitor and efficient inhibitor for quorum sensing auto-inducer LasR binding domain of .
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http://dx.doi.org/10.2147/AABC.S290912DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7875078PMC
February 2021

Enhancement of chondrogenesis of adipose-derived stem cells in HA-PNIPAAm-CL hydrogel for cartilage regeneration in rabbits.

Sci Rep 2018 Jul 12;8(1):10526. Epub 2018 Jul 12.

Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.

Injectable thermoresponsive hydrogels have the advantages of effective cell delivery and minimal invasion for tissue engineering applications. In this study, we investigated the chondroinductive potential of newly developed hyaluronic acid (HA)-modified thermoresponsive poly(N-isopropylacrylamide) (HA-PNIPAAm-CL) hydrogels on enhancing rabbit ADSC (rADSC) chondrogenesis in vitro and in the synovial cavity of rabbit. The HA-mixed PNIPAAm (HA-PNIPAAm-CP) and HA-cross-linked PNIPAAm (HA-PNIPAAm-CL) were fabricated using physical interaction and chemical cross-linking methods, respectively. The in vitro results showed that, compared to unmodified PNIPAAm, both HA-modified hydrogels significantly increased cell viability, chondrogenic marker gene (aggrecan and type II collagen) expression and sulfide glycosaminoglycan (sGAG) formation in embedded rADSCs. However, HA-PNIPAAm-CL showed the highest rADSC viability and chondrogenesis. The chondrogenic effects of HA-modified hydrogels on rADSCs were confirmed in vivo by the intraarticular injection of hydrogel-embedded rADSC constructs into rabbit synovial cavities for 3 weeks and tracing with CM-DiI labeling. Neocartilage formation in the hydrogels was determined by histomorphological staining of GAG and type II collagen. In vivo injected rADSC/HA-PNIPAAm-CL constructs showed more hyaline cartilage formation than that of rADSC/HA-PNIPAAm-CP and rADSC/PNIPAAm constructs in the synovial cavity of rabbit. These results suggest that the HA-PNIPAAm-CL provides a suitable microenvironment to enhance ADSC chondrogenesis for articular cartilage tissue engineering applications.
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http://dx.doi.org/10.1038/s41598-018-28893-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6043528PMC
July 2018

Low-oxygen conditions promote synergistic increases in chondrogenesis during co-culture of human osteoarthritic stem cells and chondrocytes.

J Tissue Eng Regen Med 2018 04 10;12(4):1074-1084. Epub 2018 Jan 10.

Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland.

There has been increased interest in co-cultures of stem cells and chondrocytes for cartilage tissue engineering as there are the limitations associated with using either cell type alone. Drawbacks associated with the use of chondrocytes include the limited numbers of cells available for isolation from damaged or diseased joints, their dedifferentiation during in vitro expansion, and a diminished capacity to synthesise cartilage-specific extracellular matrix components with age and disease. This has motivated the use of adult stem cells with either freshly isolated or culture-expanded chondrocytes for cartilage repair applications; however, the ideal combination of cells and environmental conditions for promoting robust chondrogenesis remains unclear. In this study, we compared the effect of combining a small number of freshly isolated or culture-expanded human chondrocytes with infrapatellar fat pad-derived stem cells (FPSCs) from osteoarthritic donors on chondrogenesis in altered oxygen (5% or 20%) and growth factor supplementation (TGF-β3 only or TGF-β3 and BMP-7) conditions. Both co-cultures, but particularly those including freshly isolated chondrocytes, were found to promote cell proliferation and enhanced matrix accumulation compared to the use of FPSCs alone, resulting in the development of a tissue that was compositionally more similar to that of the native articular cartilage. Local oxygen levels were found to impact chondrogenesis in co-cultures, with more robust increases in proteoglycan and collagen deposition observed at 5% O . Additionally, collagen type I synthesis was suppressed in co-cultures maintained at low-oxygen conditions. This study demonstrates that a co-culture of freshly isolated human chondrocytes and FPSCs promotes robust chondrogenesis and thus is a promising cell combination for cartilage tissue engineering.
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http://dx.doi.org/10.1002/term.2608DOI Listing
April 2018

Engineering Tissues That Mimic the Zonal Nature of Articular Cartilage Using Decellularized Cartilage Explants Seeded with Adult Stem Cells.

ACS Biomater Sci Eng 2017 Sep 31;3(9):1933-1943. Epub 2016 Mar 31.

Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, 152-160 Pearse Street, Dublin 2, Ireland.

Articular cartilage (AC) possesses uniquely complex mechanical properties; for example its stiffness increases with depth through the tissue and it softens when compressed. These properties are integral to the function of AC and can be attributed to the tissue's collagen network and how it interacts with negatively charged proteoglycans. In this study, scaffolds containing arrays of channels were produced from decellularized AC explants derived from skeletally immature and mature pigs. These scaffolds were then repopulated with human infrapatellar fat pad derived stem cells (FPSCs). After 4 weeks in culture, FPSCs filled channels within the decellularized explants with a matrix rich in proteoglycans and collagen. Cellular and neo-matrix alignment within these scaffolds appeared to be influenced by the underlying collagen architecture of the decellularized cartilage. Repopulating scaffolds derived from decellularized skeletally mature cartilage with FPSCs led to the development of engineered cartilage with depth-dependent mechanical properties mimicking aspects of native tissue. Furthermore, these constructs displayed the characteristic strain softening behavior of AC. These findings highlight the importance of the collagen network to engineering mechanically functional cartilage grafts.
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http://dx.doi.org/10.1021/acsbiomaterials.6b00020DOI Listing
September 2017

PLGA-linked alendronate enhances bone repair in diaphysis defect model.

J Tissue Eng Regen Med 2017 09 3;11(9):2603-2612. Epub 2016 Jun 3.

Orthopedic Research Centre, Kaohsiung Medical University, Kaohsiung, Taiwan.

Alendronate (ALN) is known as an anti-resorptive drug for the treatment of osteoporosis. Recently, ALN was found to stimulate osteogenic differentiation in mesenchymal stem cells and enhance new bone formation in calvarial bone defects. Previous in vitro and in vivo studies found that the effective concentration of ALN was approximately 1-10   μm. In the present study, a poly (lactic-co-glycolic acid) (PLGA) cross-linked ALN (PLGA-ALN) with a short-term controlled-release property for local application to enhance bone repair was developed. An in vitro drug-release kinetic test showed that PLGA-ALN microspheres released an effective concentration (50-100 nm) of ALN for 9 days. The effect of PLGA-ALN on bone repair was tested in a rat femoral bone defect model. The biomechanical study results showed that the maximal strength, stiffness and energy absorption were significantly increased in the PLGA-ALN group compared with the PLGA group. The microstructure of the newly formed bone at the defect site was analysed using microcomputed tomography. The PLGA-ALN group significantly improved the trabecular bone volume at the defect site compared with the PLGA group. The fibril collagen and immunolocalized bone morphogenetic protein 2 were evident in the newly formed trabecular bone in the PLGA-ALN group. Local use of newly developed PLGA-ALN-enhanced bone repair was attributable to increasing bone matrix formation, which improved the ultrastructure of the newly formed bone and thus increased the biomechanical properties of the repaired bone. It is suggested that PLGA-ALN may be a potential bone graft substitute to enhance bone repair. Copyright © 2016 John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/term.2160DOI Listing
September 2017

Decellularization of porcine articular cartilage explants and their subsequent repopulation with human chondroprogenitor cells.

J Mech Behav Biomed Mater 2015 Mar 22;55:21-31. Epub 2015 Oct 22.

Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland; Department of Mechanical and Manufacturing Engineering, School of Engineering, Trinity College Dublin, Dublin, Ireland; Department of Anatomy, Royal College of Surgeons in Ireland, Dublin, Ireland; Advanced Materials and Bioengineering Research Centre (AMBER), Royal College of Surgeons in Ireland and Trinity College Dublin, Dublin, Ireland. Electronic address:

Engineering tissues with comparable structure, composition and mechanical functionality to native articular cartilage remains a challenge. One possible solution would be to decellularize xenogeneic articular cartilage in such a way that the structure of the tissue is maintained, and to then repopulate this decellularized matrix with human chondroprogenitor cells that will facilitate the reconstitution, maintenance and eventual turnover of the construct following implantation. The overall objective of this study was to develop a protocol to efficiently decellularize porcine articular cartilage grafts and to identify a methodology to subsequently repopulate such explants with human chondroprogenitor cells. To this end, channels were first introduced into cylindrical articular cartilage explants, which were then decellularized with a combination of various chemical reagents including sodium dodecyl sulfate (SDS) and nucleases. The decellularization protocol resulted in a ~90% reduction in porcine DNA content, with little observed effect on the collagen content and the collagen architecture of the tissue, although a near-complete removal of sulfated glycosaminoglycans (sGAG) and a related reduction in tissue compressive properties was observed. The introduction of channels did not have any detrimental effect on the biochemical or the mechanical properties of the decellularized tissue. Next, decellularized cartilage explants with or without channels were seeded with human infrapatellar fat pad derived stem cells (FPSCs) and cultured chondrogenically under either static or rotational conditions for 10 days. Both channeled and non-channeled explants supported the viability, proliferation and chondrogenic differentiation of FPSCs. The addition of channels facilitated cell migration and subsequent deposition of cartilage-specific matrix into more central regions of these explants. The application of rotational culture appeared to promote a less proliferative cellular phenotype and led to an increase in sGAG synthesis within the explants. Rotational culture also appeared to promote higher cell viability and led to a more even distribution of cells within the channels of decellularized explants. To conclude, this study describes an effective protocol for the decellularization of porcine articular cartilage grafts and a novel methodology for the partial recellularization of such explants with human stem cells. Decellularized soft tissue explants that maintain their native collagen architecture may represent promising scaffolds for musculoskeletal tissue engineering applications.
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http://dx.doi.org/10.1016/j.jmbbm.2015.10.002DOI Listing
March 2015

Advances in tissue engineering through stem cell-based co-culture.

J Tissue Eng Regen Med 2015 May 3;9(5):488-503. Epub 2014 Feb 3.

Department of Biomedical Engineering and Orthopedic Surgery, University of California at Davis, CA, 95616, USA.

Stem cells are the future in tissue engineering and regeneration. In a co-culture, stem cells not only provide a target cell source with multipotent differentiation capacity, but can also act as assisting cells that promote tissue homeostasis, metabolism, growth and repair. Their incorporation into co-culture systems seems to be important in the creation of complex tissues or organs. In this review, critical aspects of stem cell use in co-culture systems are discussed. Direct and indirect co-culture methodologies used in tissue engineering are described, along with various characteristics of cellular interactions in these systems. Direct cell-cell contact, cell-extracellular matrix interaction and signalling via soluble factors are presented. The advantages of stem cell co-culture strategies and their applications in tissue engineering and regenerative medicine are portrayed through specific examples for several tissues, including orthopaedic soft tissues, bone, heart, vasculature, lung, kidney, liver and nerve. A concise review of the progress and the lessons learned are provided, with a focus on recent developments and their implications. It is hoped that knowledge developed from one tissue can be translated to other tissues. Finally, we address challenges in tissue engineering and regenerative medicine that can potentially be overcome via employing strategies for stem cell co-culture use.
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http://dx.doi.org/10.1002/term.1870DOI Listing
May 2015

Self-organization and the self-assembling process in tissue engineering.

Annu Rev Biomed Eng 2013 20;15:115-36. Epub 2013 May 20.

Department of Biomedical Engineering, University of California, Davis, CA 95616, USA.

In recent years, the tissue engineering paradigm has shifted to include a new and growing subfield of scaffoldless techniques that generate self-organizing and self-assembling tissues. This review aims to cogently describe this relatively new research area, with special focus on applications toward clinical use and research models. Particular emphasis is placed on providing clear definitions of self-organization and the self-assembling process, as delineated from other scaffoldless techniques in tissue engineering and regenerative medicine. Significantly, during formation, self-organizing and self-assembling tissues display biological processes similar to those that occur in vivo. These processes help lead to the recapitulation of native tissue morphological structure and organization. Notably, functional properties of these engineered tissues, some of which are already in clinical trials, also approach native tissue values. This review endeavors to provide a cohesive summary of work in this field and to highlight the potential of self-organization and the self-assembling process for providing cogent solutions to currently intractable problems in tissue engineering.
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http://dx.doi.org/10.1146/annurev-bioeng-071812-152423DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4420200PMC
February 2014

Hyaluronan initiates chondrogenesis mainly via CD44 in human adipose-derived stem cells.

J Appl Physiol (1985) 2013 Jun 28;114(11):1610-8. Epub 2013 Feb 28.

Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.

Cell-matrix adhesion is one of the important interactions that regulates stem cell survival, self-renewal, and differentiation. Our previous report (Wu SC, Chang JK, Wang CK, Wang GJ, Ho ML. Biomaterials 31: 631-640, 2010) indicated that a microenvironment enriched with hyaluronan (HA) initiated and enhanced chondrogenesis in human adipose-derived stem cells (hADSCs). We further hypothesize that HA-induced chondrogenesis in hADSCs is mainly due to the interaction of HA and CD44 (HA-CD44), a cell surface receptor of HA. The HA-CD44 interaction was tested by examining the mRNA expression of hyaluronidase-1 (Hyal-1) and chondrogenic marker genes (SOX-9, collagen type II, and aggrecan) in hADSCs cultured on HA-coated wells. Cartilaginous matrix formation, sulfated glycosaminoglycan, and collagen productions by hADSCs affected by HA-CD44 interaction were tested in a three-dimensional fibrin hydrogel. About 99.9% of hADSCs possess CD44. The mRNA expressions of Hyal-1 and chondrogenic marker genes were upregulated by HA in hADSCs on HA-coated wells. Blocking HA-CD44 interaction by anti-CD44 antibody completely inhibited Hyal-1 expression and reduced chondrogenic marker gene expression, which indicates that HA-induced chondrogenesis in hADSCs mainly acts through HA-CD44 interaction. A 2-h preincubation and coculture of cells with HA in hydrogel (HA/fibrin hydrogel) not only assisted in hADSC survival, but also enhanced expression of Hyal-1 and chondrogenic marker genes. Higher levels of sulfated glycosaminoglycan and total collagen were also found in HA/fibrin hydrogel group. Immunocytochemistry showed more collagen type II, but less collagen type X, in HA/fibrin than in fibrin hydrogels. Our results indicate that signaling triggered by HA-CD44 interaction significantly contributes to HA-induced chondrogenesis and may be applied to adipose-derived stem cell-based cartilage regeneration.
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http://dx.doi.org/10.1152/japplphysiol.01132.2012DOI Listing
June 2013

Electromagnetic fields enhance chondrogenesis of human adipose-derived stem cells in a chondrogenic microenvironment in vitro.

J Appl Physiol (1985) 2013 Mar 13;114(5):647-55. Epub 2012 Dec 13.

Orthopaedic Research Center, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan.

We tested the hypothesis that electromagnetic field (EMF) stimulation enhances chondrogenesis in human adipose-derived stem cells (ADSCs) in a chondrogenic microenvironment. A two-dimensional hyaluronan (HA)-coated well (2D-HA) and a three-dimensional pellet culture system (3D-pellet) were used as chondrogenic microenvironments. The ADSCs were cultured in 2D-HA or 3D-pellet, and then treated with clinical-use pulse electromagnetic field (PEMF) or the innovative single-pulse electromagnetic field (SPEMF) stimulation. The cytotoxicity, cell viability, and chondrogenic and osteogenic differentiations were analyzed after PEMF or SPEMF treatment. The modules of PEMF and SPEMF stimulations used in this study did not cause cytotoxicity or alter cell viability in ADSCs. Both PEMF and SPEMF enhanced the chondrogenic gene expression (SOX-9, collagen type II, and aggrecan) of ADSCs cultured in 2D-HA and 3D-pellet. The expressions of bone matrix genes (osteocalcin and collagen type I) of ADSCs were not changed after SPEMF treatment in 2D-HA and 3D-pellet; however, they were enhanced by PEMF treatment. Both PEMF and SPEMF increased the cartilaginous matrix (sulfated glycosaminoglycan) deposition of ADSCs. However, PEMF treatment also increased mineralization of ADSCs, but SPEMF treatment did not. Both PEMF and SPEMF enhanced chondrogenic differentiation of ADSCs cultured in a chondrogenic microenvironment. SPEMF treatment enhanced ADSC chondrogenesis, but not osteogenesis, when the cells were cultured in a chondrogenic microenvironment. However, PEMF enhanced both osteogenesis and chondrogenesis under the same conditions. Thus the combination of a chondrogenic microenvironment with SPEMF stimulation can promote chondrogenic differentiation of ADSCs and may be applicable to articular cartilage tissue engineering.
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http://dx.doi.org/10.1152/japplphysiol.01216.2012DOI Listing
March 2013

Sustained release of PTH(1-34) from PLGA microspheres suppresses osteoarthritis progression in rats.

Acta Biomater 2012 Jul 10;8(6):2254-62. Epub 2012 Mar 10.

Orthopedic Research Center, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.

We previously reported that PTH(1-34) inhibits the terminal differentiation of articular chondrocytes and, in turn, suppresses the progression of osteoarthritis (OA). However, this treatment requires an injection of PTH(1-34) once every 3 days over the treatment period. In this study, we studied the effect of sustained administration of PTH(1-34) in a papain-induced OA rat model. We developed an effective controlled-release system for prolonging the treatment duration of an intra-articular injection for OA treatment in rats. The effects of released PTH(1-34) from PLGA(65:35)-encapsulated PTH(1-34) microspheres (PTH/PLGA) on papain-induced OA in rat knees were studied. Microsphere morphology was observed in vitro by scanning electron microscopy, and microsphere size was determined with a particle size analyzer. The PTH(1-34) encapsulation efficiency and release profile, as well as the toxicity of PTH/PLGA, were examined. The bioactivity of released PTH(1-34) was tested by examining cAMP levels in MC3T3E1 cells. In vivo, we evaluated the changes of localized GAG, Col II, and Col X in the articular cartilage of rat knees. Our results demonstrated that the surface of the PLGA microspheres was smooth, and the size of the microspheres was in the range of 51-127 μm. PTH/PLGA microspheres sustainably released PTH(1-34) for 19 days with a concentration range of 0.01-100 nM that covered the expected concentration of 10nM at 37°C. The cAMP levels of MC3T3E1 cells were elevated in the response to released PTH(1-34) from PTH/PLGA microspheres, indicating that the released PTH(1-34) is bioactive. Most importantly, intra-articular treatment with either PTH(1-34) (0.1-100 nM) 3 days/injection or PTH/PLGA microspheres (15 days/injection) for 5 weeks revealed the similar effect on suppressing papain-induced OA changes (decreasing GAG and Col II and increasing Col X) in rat knee cartilage. The effect of PTH/PLGA microspheres on suppressing OA progression was similar to that of a once-every-three-day injection of PTH(1-34), indicating that both the sustained and intermittent action of PTH(1-34) effectively suppress OA progression. The developed PLGA microspheres with sustained release and long-term effect may be potent carriers for PTH(1-34) used to treat early OA.
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http://dx.doi.org/10.1016/j.actbio.2012.03.015DOI Listing
July 2012

DDR1 regulates the stabilization of cell surface E-cadherin and E-cadherin-mediated cell aggregation.

J Cell Physiol 2010 Aug;224(2):387-97

Orthopedic Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan.

The stabilization of cell surface E-cadherin is important for the maintenance of apical junction complexes and epithelial polarity. Previously, we reported that discoidin domain receptor 1 (DDR1) forms a complex with E-cadherin at adhesive contacts; however, the regulatory role of DDR1 in the stabilization of cell surface E-cadherin and E-cadherin-mediated cell behaviors remained undefined. To gain insight into these questions, we utilized two stable clones depleted for DDR1 via the small interfering RNA (siRNA) technique, and we over-expressed DDR1 in MDCK cells. We performed Western blotting, immunofluorescence analysis, flow cytometry, and cell aggregation studies to investigate the effect of DDR1 on cell surface E-cadherin. The results showed that both DDR1/2 and E-cadherin use their extracellular domains to form DDR/E-cadherin complexes. Neither the depletion nor the over-expression of DDR1 changed the expression level of E-cadherin in MDCK cells. Collagen disrupted the formation of E-cadherin complexes and caused E-cadherin to accumulate in the cytoplasm; however, over-expression of DDR1 stabilized E-cadherin on the cell surface and decreased its cytoplasmic accumulation. Furthermore, independently of collagen stimulation, the depletion of DDR1 resulted in a decrease in the level of cell surface E-cadherin, which consequently caused its cytoplasmic accumulation and decreased E-cadherin-mediated cell aggregation. These results indicate that DDR1 can increase the stability of cell surface E-cadherin and promote MDCK cell aggregation, which may be mediated through the formation of DDR1/E-cadherin complexes. Overall, these findings have implications for the physiological roles of DDR1 in association with the maintenance of both the adhesion junction and epithelial polarity.
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http://dx.doi.org/10.1002/jcp.22134DOI Listing
August 2010