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Performance of PRP associated with porous chitosan as a composite scaffold for regenerative medicine.

ABSTRACT: This study aimed to evaluate the in vitro performance of activated platelet-rich plasma associated with porous sponges of chitosan as a composite scaffold for proliferation and osteogenic differentiation of human adipose tissue-derived mesenchymal stem cells. The sponges were prepared by controlled freezing (-20, -80, or -196°C) and lyophilization of chitosan solutions (1, 2, or 3% w/v). The platelet-rich plasma was obtained from controlled centrifugation of whole blood and activated with calcium and autologous serum. The composite scaffolds were prepared by embedding the sponges with the activated platelet-rich plasma. The results showed the performance of the scaffolds was superior to that of activated platelet-rich plasma alone, in terms of delaying the release of growth factors and increased proliferation of the stem cells. The best preparation conditions of chitosan composite scaffolds that coordinated the physicochemical and mechanical properties and cell proliferation were 3% (w/v) chitosan and a -20°C freezing temperature, while -196°C favored osteogenic differentiation. Although the composite scaffolds are promising for regenerative medicine, the structures require stabilization to prevent the collapse observed after five days.

SUBMITTER: Shimojo AA

PROVIDER: S-EPMC4363634 | biostudies-other | 2015

REPOSITORIES: biostudies-other

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Project description:Providing accurate and up-to-date practical tools enabling oversight of platelet-rich plasma (PRP) legislation and of the appropriate standards to be implemented for its manufacture and use in Europe is a demanding task. This is due to rapid medico-technological advancements, slowness and disparity in legislation updates and enforcement between member states, and many reported gray-zone practices, notably for autologous PRP use. The levels of risk associated with blood manipulation processes generally dictate the manufacturing requirements for PRP preparations, which have gradually shifted toward good manufacturing practices (GMP) for standardization and overall quality enhancement. This work firstly outlines Western European and Swiss legislation for PRP products/preparations, providing key simplified information and recommendations for medical doctors seeking to implement this biological-based therapy for safe use in hospital settings, clinics, or private offices. This work secondly shows the importance of PRP-based product manufacturing standardization, which subsequently enables sound clinical evaluation of therapeutic interventions. Although the applicable legal bases provide guidelines for GMP manufacturing infrastructure and basic process design, paramount importance is set on the definition of workflows, technical specifications, and key parameters for PRP preparation and delivery. Overall, the development of simple and robust technologies and processes for PRP preparation is critical for guaranteeing the high therapeutic quality of the intervention, in collaboration with qualified GMP manufacturing platforms. Importantly, this work aims to serve as a practical tool for clinicians based in Western Europe who are willing to appropriately (i.e., administratively and technically) implement autologous PRP treatments in musculoskeletal regenerative medicine workflows, to ensure they make informed and optimal regulatory or process-based decisions.

Project description:The number of studies evaluating platelet-rich plasma (PRP) concentration has substantially grown in the last fifteen years. A systematic review on this field has been realized by evaluating in the identified studies the in vitro PRP concentration-also analyzing the platelet amount-and the in vivo PRP effects in tissue regeneration compared to any control. The protocol has been developed in agreement with the Preferred Reporting for Items for Systematic Reviews and Meta-Analyses-Protocols (PRISMA-P) guidelines. Multistep research of the PubMed, MEDLINE, Embase, PreMEDLINE, Ebase, CINAHL, PsycINFO, Clinicaltrials.gov, Scopus database and Cochrane databases has permitted to identify articles on different concentrations of PRP in vitro and related in vivo impact for tissue repair. Of the 965 articles initially identified, 30 articles focusing on PRP concentration have been selected and, consequently, only 15 articles have been analyzed. In total, 40% (n = 6) of the studies were related to the fixed PRP Concentration Group used a fixed PRP concentration and altered the platelet concentration by adding the different volumes of the PRP (lysate) to the culture. This technique led to a substantial decrease in nutrition available at higher concentrations. Sixty percent (n = 9) of the studies were related to the fixed PRP Volume Group that used a fixed PRP-to-media ratio (Vol/Vol) throughout the experiment and altered the concentration within the PRP volume. For both groups, when the volume of medium (nutrition) decreases, a lower rate of cell proliferation is observed. A PRP concentration of 1.0 × 106 plt/?L, appears to be optimal thanks to the constant and plentiful capillary nutrition supply and rapid diffusion of growth factors that happen in vivo and it also respects the blood decree-law. The PRP/media ratio should provide a sufficient nutrition supply to prevent cellular starvation, that is, PRP ? 10% (Vol/Vol) and thus best mimic the conditions in vivo.

Project description:Bone tissue may suffer from bone injury and bone defects due to accidents or diseases. Since the demand for autologous bone and allograft tissue far exceeds the supply, bone scaffolds have taken the lead. The use of bone scaffolds is one of the measures to help heal or regenerate bone tissue. Therefore, a new bone scaffold was proposed in this study, which has a simpler preparation process and stronger performance. This study proposes bone scaffolds with an attempt to use polymers that are synthesized separately with three types of minerals as the filler using the microwave foaming method as follows. A 0.1 wt% of montmorillonite (MMT), zinc oxide (ZnO), or titanium dioxide (TiO2) is added to chitosan (CS)/gelatin mixtures, respectively, after which sodium bicarbonate is added as a foaming agent, thereby forming porous gels. The polymer synthesized from three minerals was used as filler. The following microwave foaming method was adopted: 0.1 wt% MMT, ZnO, or TiO2 was added to the CS/gelatin mixture, and then sodium bicarbonate was added as a foaming agent to form a porous gel. Next, porous gels and polycaprolactone were combined in a self-made mold in order to form bone scaffolds. A stereo microscope is used to observe the morphology of bone scaffolds, after which the pore size analysis, pore connectivity, swell property, porosity, and compressive strength are tested, examining the effects of the mineral type on bone scaffolds. The test results indicate that with MMT being the filler and sodium bicarbonate being the foaming agent, the resulting bone scaffolds yield a porous structure with a pore size between 120 μm and 370 μm. Besides, the incorporation of polycaprolactone also provides samples of 1MCG-P, 2MCG-P, and 5MCG-P with a certain compressive strength of 150-170 MPa. To sum up, the test results substantiate that a combination of the microwave foaming method and MMT generates a porous structure for bone scaffolds (1MCG-P, 2MCG-P, and 5MCG-P), involving a porosity of 38%, an inter-connected porous structure, and the compressive strength that exceeds 150 MPa.

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Performance of PRP associated with porous chitosan as a composite scaffold for regenerative medicine.

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