pinecone/core-2020-05-10-deduplication huggingface dataset:
Novel method for the isolation of adipose stem cells (ASCs)

Adipose stem cells (ASCs) represent a cell population with great potential for tissue engineering
applications. Several articles have been published showing the proliferation and differentiation
potential, the markers and the wide range of potential applications of these cells. In the majority
of these studies the ASCs are isolated using a basic enzymatic procedure, which results in a quite
heterogeneous cell population that compromises their proliferation and differentiation. This paper
reports the development and optimization of a new isolation/purification method that allows
populations of ASCs to be obtained, which significantly reduces (and eventually eliminates) the
‘contamination’ of other cell types. This method is based on the use of immunomagnetic beads
coated with specific antibodies. The first part of the study described here analysed the expression of
marker genes for stem cells and the colony-forming unit (CFU) capacity of the cells isolated, while
the second part is dedicated to the osteogenic differentiation potential of the isolated cells. The
results showed that, using the isolation method based on immunomagnetic beads, it was possible to
obtain ASCs and also underline the existence of several subpopulations of stem cells in the adipose
tissue
Novel injectable gel (system) as a vehicle for human articular chondrocytes in cartilage tissue regeneration

We developed a novel injectable carrageenan/fibrin/hyaluronic acid-based hydrogel with in situ
gelling properties to be seeded with chondrogenic cells and used for cartilage tissue engineering
applications. We first analysed the distribution within the hydrogel construct and the phenotype of
human articular chondrocytes (HACs) cultured for 3 weeks in vitro. We observed a statistically
significant increase in the cell number during the first 2 weeks and maintenance of cell
viability throughout the cell culture, together with the deposition/formation of a cartilage-specific
extracellular matrix (ECM). Taking advantage of a new in vivo model that allows the integration
between newly formed and preexisting cartilage in immunodeficient mice to be investigated, we
showed that injectable hydrogel seeded with human articular chondrocytes was able to regenerate
and repair an experimentally made lesion in bovine articular cartilage, thus demonstrating the
potential of this novel cell delivery system for cartilage tissue engineering.The authors are grateful to Recco orthopaedic staff members for the collaboration and patients for bioptic material donation as well as to Mrs Daniela Marubbi for histological assistance. This work was supported by funds from the Italian MUR (FIRB-Tissuenet project), the European Union-funded STREP project, HIPPOCRATES (Grant No. NMP3-CT-2003-505758) and the European NoE EXPERTISSUES project (Grant No. NMP3-CT-2004-500283)
Engineering tendon and ligament tissues : present developments towards successful clinical products

Musculoskeletal diseases are one of the leading causes of disability worldwide. Among them, tendon
and ligament injuries represent an important aspect to consider in both athletes and active working
people. Tendon and ligament damage is an important cause of joint instability, and progresses into
early onset of osteoarthritis, pain, disability and eventually the need for joint replacement surgery.
The social and economical burden associated with these medical conditions presents a compelling
argument for greater understanding and expanding research on this issue. The particular physiology
of tendons and ligaments (avascular, hypocellular and overall structural mechanical features) makes
it difficult for currently available treatments to reach a complete and long-term functional repair of
the damaged tissue, especially when complete tear occurs. Despite the effort, the treatmentmodalities
for tendon and ligament are suboptimal, which have led to the development of alternative therapies,
such as the delivery of growth factors, development of engineered scaffolds or the application of stem
cells, which have been approached in this review
Proteoglycan production is required in initial stages of new cartilage matrix formation but inhibits integrative cartilage repair

The optimal stimulus to repair or regenerate cartilage is not known. We therefore modulated collagen deposition, collagen crosslinking and GAG deposition simultaneously during cartilage matrix production and integrative repair, creating more insight into their role in cartilage repair processes. Insulin-like growth factor 1 (IGF-1; increases proteoglycan and collagen synthesis), beta-aminopropionitrile (BAPN; a reversible inhibitor of collagen crosslinking) and para-nitrophenyl-beta-D-xyloside (PNPX; interferes with proteoglycan production) were used. Bovine articular chondrocytes were cultured in alginate beads for 3 weeks with or without IGF-1, BAPN or PNPX alone and in all possible combinations, followed by 3 weeks in control medium. DNA content, GAG and collagen deposition and collagen crosslinks were determined. Cartilage constructs were cultured under the same conditions and histologically analysed for integration of two opposing cartilage matrices. In alginate cultures, inhibition of collagen crosslinking with BAPN, in combination with promotion of matrix synthesis using IGF1, was most beneficial for matrix deposition. Addition of PNPX was always detrimental for matrix deposition. For integration of opposing cartilage constructs, the combination of BAPN, IGF1 and temporary prevention of proteoglycan formation with PNPX was most beneficial. When a new matrix is produced, proteoglycans are important to retain collagen in the matrix. When two already formed cartilage matrices have to integrate, a temporary absence of proteoglycans and temporary inhibition of collagen crosslinking might be more beneficial in combination with stimulation of collagen production, e.g. by IGF1. Therefore, the choice of soluble factors to promote cartilage regeneration depends on the type of therapy that will be used
Evaluation of a novel collagen-gelatin scaffold for achieving the sustained release of basic fibroblast growth factor in a diabetic mouse model.

The objective of this study was to evaluate the ability of a scaffold, collagen-gelatin sponge (CGS), to release basic fibroblast growth factor (bFGF) in a sustained manner, using a pressure-induced decubitus ulcer model involving genetically diabetic mice. We confirmed that CGSs impregnated with a bFGF concentration of up to 50 µg/cm(2) were able to sustain the release of bFGF throughout their biodegradation. We prepared decubitus ulcers on diabetic mice. After debriding the ulcers, we implanted CGSs (diameter 8 mm) impregnated with normal saline solution (NSS) or bFGF solution (7, 14, 28 or 50 µg/cm(2)). At 1 and 2 weeks after implantation, the mice were sacrificed and tissue specimens were obtained. The wound area, neoepithelium length and numbers and total area of newly formed capillaries were evaluated. The CGSs impregnated with NSS became infected and degraded, whereas the CGSs impregnated with 7 or 14 µg/cm(2) bFGF displayed accelerated dermis-like tissue formation and the CGSs impregnated with 14 µg/cm(2) bFGF produced significant improvements in the remaining wound area, neoepithelium length and numbers and total area of newly formed capillaries compared with the NSS group. No significant difference was observed between the NSS and 50 µg/cm(2) bFGF groups. CGSs impregnated with 7-14 µg/cm(2) bFGF accelerated wound healing, and an excess amount of bFGF did not increase the wound-healing efficacy of the CGSs. Our CGS is a scaffold that can release positively charged growth factors such as bFGF in a sustained manner and shows promise as a scaffold for skin regeneration
Platelet lysates scaffolds prepared by supercritical fluid technology as autologous templates for cartilage regeneration

The tissue engineering strategy proposed in this work regards the
development of a novel autologous scaffold based on platelet lysates
(PLs) with the ultimate goal of promoting the regeneration of an orthopaedic
osteochondral interface. PLs are a high concentration of platelets
in a small volume of plasma that, when activated, release several
growth factors (GFs). Most of current PLs-based hydrogels present several
limitations, specifically the lack of stability, the constant shrinking
in culture and the need of activation with animal-derived thrombin.
This study represents a major breakthrough as it demonstrates that a
stable scaffold can be prepared only from PLs, thus acting simultaneously
as a template for cell colonization and as multiple GF release
system. The PL scaffolds, crosslinked with genipin were prepared by
supercritical fluid assisted phase inversion at 100 bar and 40 °C. The
morphological properties of the scaffolds were assessed and in vitro GF
release profile was studied by micro BCA and ELISA assays.Scaffolds
were seeded with human adipose-derived stem cells (hASCs) and cultured
in vitro up to 28 days. Cell viability and proliferation were
assessed as well as histology and immunohistochemistry. Results showed the deposition of cartilage extracellular matrix and the expression
of chondrogenic gene makers, demonstrating the feasibility of the
constructs to simultaneously provide architectural support and biological
cues to promote chondrogenic differentiation
Biological evaluation of intervertebral disc cells in different formulations of gellan gum-based hydrogels

First published online: 2012Gellan gum (GG)-based hydrogels are advantageous in tissue engineering not only due to their ability
to retain large quantities of water and provide a similar environment to that of natural extracellular
matrix (ECM), but also because they can gelify in situ in seconds. Their mechanical properties can be
fine-tuned to mimic natural tissues such as the nucleus pulposus (NP). This study produced different
formulations of GG hydrogels by mixing varying amounts of methacrylated (GG-MA) and high-acyl
gellan gums (HA-GG) for applications as acellular and cellular NP substitutes. The hydrogels were
physicochemically characterized by dynamic mechanical analysis. Degradation and swelling abilities
were assessed by soaking in a phosphate buffered saline solution for up to 170 h. Results showed that
as HA-GG content increased, the modulus of the hydrogels decreased. Moreover, increases in HA-GG
content induced greater weight loss in the GG-MA/HA-GG formulation compared to GG-MA hydrogel.
Potential cytotoxicity of the hydrogel was assessed by culturing rabbit NP cells up to 7 days. An MTS
assay was performed by seeding rabbit NP cells onto the surface of 3D hydrogel disc formulations.
Viability of rabbit NP cells encapsulated within the different hydrogel formulations was also
evaluated by Calcein-AM and ATP assays. Results showed that tunable GG-MA/HA-GG hydrogels were
non-cytotoxic and supported viability of rabbit NP cells.The authors thank the Portuguese Foundation for Science and Technology (FCT) for financial support through POCTI and FEDER programmes. This work was also conducted with the support of the European Union funded Collaborative Project Disc Regeneration (NMP3-LA-2008-213904), MBC(A04003) and the World Class University project (R31-20029)
Characterization of human skeletal stem and bone cell populations using dielectrophoresis

© 2012 John Wiley & Sons, Ltd. Dielectrophoresis (DEP) is a non-invasive cell analysis method that uses differences in electrical properties between particles and surrounding medium to determine a unique set of cellular properties that can be used as a basis for cell separation. Cell-based therapies using skeletal stem cells are currently one of the most promising areas for treating a variety of skeletal and muscular disorders. However, identifying and sorting these cells remains a challenge in the absence of unique skeletal stem cell markers. DEP provides an ideal method for identifying subsets of cells without the need for markers by using their dielectric properties. This study used a 3D dielectrophoretic well chip device to determine the dielectric characteristics of two osteosarcoma cell lines (MG-63 and SAOS-2) and an immunoselected enriched skeletal stem cell fraction (STRO-1 positive cell) of human bone marrow. Skeletal cells were exposed to a series of different frequencies to induce dielectrophoretic cell movement, and a model was developed to generate the membrane and cytoplasmic properties of the cell populations. Differences were observed in the dielectric properties of MG-63, SAOS-2 and STRO-1 enriched skeletal populations, which could potentially be used to sort cells in mixed populations. This study provide evidence of the ability to characterize different human skeletal stem and mature cell populations, and acts as a proof-of-concept that dielectrophoresis can be exploited to detect, isolate and separate skeletal cell populations from heterogeneous bone marrow cell populations
Cartilage regeneration using zonal chondrocyte subpopulations : a promising approach or an overcomplicated strategy?

Cartilage defects heal imperfectly and osteoarthritic changes develop frequently as a result. Although the existence of specific behaviours of chondrocytes derived from various depth-related zones in vitro has been known for over 20 years, only a relatively small body of in vitro studies has been performed with zonal chondrocytes and current clinical treatment strategies do not reflect these native depth-dependent (zonal) differences. This is surprising since mimicking the zonal organization of articular cartilage in neo-tissue by the use of zonal chondrocyte subpopulations could enhance the functionality of the graft. Although some research groups including our own have made considerable progress in tailoring culture conditions using specific growth factors and biomechanical loading protocols, we conclude that an optimal regime has not yet been determined. Other unmet challenges include the lack of specific zonal cell sorting protocols and limited amounts of cells harvested per zone. As a result, the engineering of functional tissue has not yet been realized and no long-term in vivo studies using zonal chondrocytes have been described. This paper critically reviews the research performed to date and outlines our view of the potential future significance of zonal chondrocyte populations in regenerative approaches for the treatment of cartilage defects. Secondly, we briefly discuss the capabilities of additive manufacturing technologies that can not only create patient-specific grafts directly from medical imaging data sets but could also more accurately reproduce the complex 3D zonal extracellular matrix architecture using techniques such as hydrogel-based cell printing
Chondrogenic potential of injectable κ-carrageenan hydrogel with encapsulated adipose stem cells for cartilage tissue-engineering applications

Due to the limited self-repair capacity of cartilage, regenerative medicine therapies for the treatment
of cartilage defects must use a signiﬁcant amount of cells, preferably applied using a hydrogel system
that can promise their delivery and functionality at the speciﬁc site. This paper discusses the potential
use of k-carrageenan hydrogels for the delivery of stem cells obt ained from adipose tissue in the
treatment of cartilage tissue defects. The developed hydrogels were produced by an ionotropic
gelation met hod and human adipose stem cells (hASCs) were encapsulated in 1.5% w/v k-carrageenan
solution at a cell density of 5  10
6
cells/ml. The results from the analysis of the cell-encapsulating
hydrogels, cultured for up to 21 days, indicated that k-carrageenan hydrogels support the viability,
proliferation and chondrogenic differentiation of hASCs. Additionally, the mec hanical analysis
demonstrated an increase in stiffness and viscoelastic properties of k-carrageenan gels with their
encapsulated cells with increasing time in culture with chondrogenic medium. These results allowed
the conclusion that k-carrageenan exhibits properties t hat enable the in vitro functionality of
encapsulated hASCs and thus may provide the basis for new successful approaches for the treatment
of cartilage defects.Elena G. Popa would like to acknowledge the Portuguese Foundation for Science and Technology (PhD Grant No. SFRH/BD/64070/2009)
Electrical stimulation of cardiac adipose tissue-derived progenitor cells modulates cell phenotype and genetic machinery

A major challenge of cardiac tissue engineering is directing cells to establish the physiological structure and function of the myocardium being replaced. Our aim was to examine the effect of electrical stimulation on the cardiodifferentiation potential of cardiac adipose tissue-derived progenitor cells (cardiac ATDPCs). Three different electrical stimulation protocols were tested; the selected protocol consisted of 2ms monophasic square-wave pulses of 50mV/cm at 1Hz over 14days. Cardiac and subcutaneous ATDPCs were grown on biocompatible patterned surfaces. Cardiomyogenic differentiation was examined by real-time PCR and immunocytofluorescence. In cardiac ATDPCs, MEF2A and GATA-4 were significantly upregulated at day 14 after stimulation, while subcutaneous ATDPCs only exhibited increased Cx43 expression. In response to electrical stimulation, cardiac ATDPCs elongated, and both cardiac and subcutaneous ATDPCs became aligned following the linear surface pattern of the construct. Cardiac ATDPC length increased by 11.3%, while subcutaneous ATDPC length diminished by 11.2% (p=0.013 and p=0.030 vs unstimulated controls, respectively). Compared to controls, electrostimulated cells became aligned better to the patterned surfaces when the pattern was perpendicular to the electric field (89.71±28.47o for cardiac ATDPCs and 92.15±15.21o for subcutaneous ATDPCs). Electrical stimulation of cardiac ATDPCs caused changes in cell phenotype and genetic machinery, making them more suitable for cardiac regeneration approaches. Thus, it seems advisable to use electrical cell training before delivery as a cell suspension or within engineered tissue.Peer ReviewedPreprin
Progenitor and stem cells for bone and cartilage regeneration

Research in regenerative medicine is developing at a significantly quick pace. Cell-based bone and
cartilage replacement is an evolving therapy aiming at the treatment of patients who suffer from
limb amputation, damaged tissues and various bone and cartilage-related disorders. Stem cells are
undifferentiated cells with the capability to regenerate into one or more committed cell lineages.
Stem cells isolated from multiple sources have been finding widespread use to advance the field of
tissue repair. The present review gives a comprehensive overview of the developments in stem cells
originating from different tissues and suggests future prospects for functional bone and cartilage
tissue regeneration.The European Network of Excellence EXPERTISSUES (Project No. NMP3-CT-2004-500283), under which this work was carried out, is acknowledged
Silk fibroin scaffolds enhance cell commitment of adult rat cardiac progenitor cells.

The use of three-dimensional (3D) cultures may induce cardiac progenitor cells to synthesize their\ud
own extracellular matrix (ECM) and sarcomeric proteins to initiate cardiac differentiation. 3D\ud
cultures grown on synthetic scaffolds may favour the implantation and survival of stem cells for cell\ud
therapy when pharmacological therapies are not efficient in curing cardiovascular diseases and when\ud
organ transplantation remains the only treatment able to rescue the patient’s life. Silk fibroin-based\ud
scaffolds may be used to increase cell affinity to biomaterials and may be chemically modified to\ud
improve cell adhesion. In the present study, porous, partially orientated and electrospun nanometric\ud
nets were used. Cardiac progenitor cells isolated from adult rats were seeded by capillarity in the 3D\ud
structures and cultured inside inserts for 21 days. Under this condition, the cells expressed a high\ud
level of sarcomeric and cardiac proteins and synthesized a great quantity of ECM. In particular,\ud
partially orientated scaffolds induced the synthesis of titin, which is a fundamental protein in\ud
sarcomere assembly
Computational model for the analysis of cartilage and cartilage tissue constructs

We propose a new non-linear poroelastic model that is suited to the analysis of soft tissues. In this paper the model is tailored to the analysis of cartilage and the engineering design of cartilage constructs. The proposed continuum formulation of the governing equations enables the strain of the individual material components within the extracellular matrix (ECM) to be followed over time, as the individual material components are synthesized, assembled and incorporated within the ECM or lost through passive transport or degradation. The material component analysis developed here naturally captures the effect of time-dependent changes of ECM composition on the deformation and internal stress states of the ECM. For example, it is shown that increased synthesis of aggrecan by chondrocytes embedded within a decellularized cartilage matrix initially devoid of aggrecan results in osmotic expansion of the newly synthesized proteoglycan matrix and tension within the structural collagen network. Specifically, we predict that the collagen network experiences a tensile strain, with a maximum of ~2% at the fixed base of the cartilage. The analysis of an example problem demonstrates the temporal and spatial evolution of the stresses and strains in each component of a self-equilibrating composite tissue construct, and the role played by the flux of water through the tissue.Australian Research Council (Grant ARC DP0988001)National Health and Medical Research Council (Australia) (Grant APP 1051538)National Institutes of Health (U.S.) (Grant AR60331
Design and characterization of a biodegradable double-layer scaffold aimed at periodontal tissue-engineering applications

First published: 1 September 2013The inefficacy of the currently used therapies in achieving the regeneration ad integrum of the
periodontium stimulates the search for alternative approaches, such as tissue-engineering strategies.
Therefore, the core objective of this study was to develop a biodegradable double-layer scaffold for
periodontal tissue engineering. The design philosophy was based on a double-layered construct
obtained from a blend of starch and poly-ε-caprolactone (30:70 wt%; SPCL). A SPCL fibre mesh
functionalized with silanol groups to promote osteogenesis was combined with a SPCL solvent
casting membrane aiming at acting as a barrier against the migration of gingival epithelium into
the periodontal defect. Each layer of the double-layer scaffolds was characterized in terms of
morphology, surface chemical composition, degradation behaviour and mechanical properties.
Moreover, the behaviour of seeded/cultured canine adipose-derived stem cells (cASCs) was assessed.
In general, the developed double-layered scaffolds demonstrated adequate degradation and
mechanical behaviour for the target application. Furthermore, the biological assays revealed that
both layers of the scaffold allow adhesion and proliferation of the seeded undifferentiated cASCs,
and the incorporation of silanol groups into the fibre-mesh layer enhance the expression of a typical
osteogenic marker. This study allowed an innovative construct to be developed, combining a
three-dimensional (3D) scaffold with osteoconductive properties and with potential to assist
periodontal regeneration, carrying new possible solutions to current clinical needs
.The research leading to these results has received funding from the European Union's Seventh Framework Programme (FP7/2007-2013; under Grant Agreement No. REGPOT-CT2012-316331-POLARIS) and from the Portuguese Foundation for Science and Technology (FCT; Grant No, MIT/ECE/0047/2009). Joao Requicha acknowledges the FCT for his PhD scholarship ( Grant No. SFRH/BD/44143/2008)