RESUMEN
The mutable collagenous tissue (MCT) of echinoderms possesses biological peculiarities that facilitate native collagen extraction and employment for biomedical applications such as regenerative purposes for the treatment of skin wounds. Strategies for skin regeneration have been developed and dermal substitutes have been used to cover the lesion to facilitate cell proliferation, although very little is known about the application of novel matrix obtained from marine collagen. From food waste we isolated eco-friendly collagen, naturally enriched with glycosaminoglycans, to produce an innovative marine-derived biomaterial assembled as a novel bi-layered skin substitute (Marine Collagen Dermal Template or MCDT). The present work carried out a preliminary experimental in vivo comparative analysis between the MCDT and Integra, one of the most widely used dermal templates for wound management, in a rat model of full-thickness skin wounds. Clinical, histological, and molecular evaluations showed that the MCDT might be a valuable tool in promoting and supporting skin wound healing: it is biocompatible, as no adverse reactions were observed, along with stimulating angiogenesis and the deposition of mature collagen. Therefore, the two dermal templates used in this study displayed similar biocompatibility and outcome with focus on full-thickness skin wounds, although a peculiar cellular behavior involving the angiogenesis process was observed for the MCDT.
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Eliminación de Residuos , Piel Artificial , Animales , Ratas , Alimentos , Cicatrización de Heridas , Piel , Colágeno/farmacología , EquinodermosRESUMEN
In peripheral nerve injuries (PNI) with substance loss, where tensionless end-to-end suture is not achievable, the positioning of a graft is required. Available options include autografts (e.g., sural nerve, medial and lateral antebrachial cutaneous nerves, superficial branch of the radial nerve), allografts (Avance®; human origin), and hollow nerve conduits. There are eleven commercial hollow conduits approved for clinical, and they consist of devices made of a non-biodegradable synthetic polymer (polyvinyl alcohol), biodegradable synthetic polymers (poly(DL-lactide-ε-caprolactone); polyglycolic acid), and biodegradable natural polymers (collagen type I with/without glycosaminoglycan; chitosan; porcine small intestinal submucosa); different resorption times are available for resorbable guides, ranging from three months to four years. Unfortunately, anatomical/functional nerve regeneration requirements are not satisfied by any of the possible alternatives; to date, focusing on wall and/or inner lumen organization/functionalization seems to be the most promising strategy for next-generation device fabrication. Porous or grooved walls as well as multichannel lumens and luminal fillers are the most intriguing options, eventually also including the addition of cells (Schwann cells, bone marrow-derived, and adipose tissue derived stem cells) to support nerve regeneration. This review aims to describe common alternatives for severe PNI recovery with a highlight of future conduits.
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Traumatismos de los Nervios Periféricos , Nervio Ciático , Animales , Humanos , Porcinos , Nervio Ciático/lesiones , Poliésteres , Nervios Periféricos/fisiología , Prótesis e Implantes , Ácido Poliglicólico , Regeneración Nerviosa/fisiologíaRESUMEN
Oxidized polyvinyl alcohol (OxPVA) is a new polymer for the fabrication of nerve conduits (NCs). Looking for OxPVA device optimization and coupling it with a natural sheath may boost bioactivity. Thus, OxPVA/chitosan sponges (ChS) as hybrid scaffolds were investigated to predict in the vivo behaviour of two-layered NCs. To encourage interaction with cells, ChS were functionalized with the self-assembling-peptide (SAP) EAK, without/with the laminin-derived sequences -IKVAV/-YIGSR. Thus, ChS and the hybrid scaffolds were characterized for mechanical properties, ultrastructure (Scanning Electron Microscopy, SEM), bioactivity, and biocompatibility. Regarding mechanical analysis, the peptide-free ChS showed the highest values of compressive modulus and maximum stress. However, among +EAK groups, ChS+EAK showed a significantly higher maximum stress than that found for ChS+EAK-IKVAV and ChS+EAK-YIGSR. Considering ultrastructure, microporous interconnections were tighter in both the OxPVA/ChS and +EAK groups than in the others; all the scaffolds induced SH-SY5Y cells' adhesion/proliferation, with significant differences from day 7 and a higher total cell number for OxPVA/ChS+EAK scaffolds, in accordance with SEM. The scaffolds elicited only a slight inflammation after 14 days of subcutaneous implantation in Balb/c mice, proving biocompatibility. ChS porosity, EAK 3D features and neuro-friendly attitude (shared with IKVAV/YIGSR motifs) may confer to OxPVA certain bioactivity, laying the basis for future appealing NCs.
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Quitosano , Neuroblastoma , Ratones , Animales , Humanos , Alcohol Polivinílico/química , Ingeniería de Tejidos , Quitosano/química , Laminina , Porosidad , Polímeros/química , Andamios del Tejido/química , Materiales BiocompatiblesRESUMEN
In recent decades, the use of adult multipotent stem cells has paved the way for the identification of new therapeutic approaches for the treatment of monogenic diseases such as Haemophilia A. Being already studied for regenerative purposes, adipose-derived mesenchymal stem cells (Ad-MSCs) are still poorly considered for Haemophilia A cell therapy and their capacity to produce coagulation factor VIII (FVIII) after proper stimulation and without resorting to gene transfection. In this work, Ad-MSCs were in vitro conditioned towards the endothelial lineage, considered to be responsible for coagulation factor production. The cells were cultured in an inductive medium enriched with endothelial growth factors for up to 21 days. In addition to significantly responding to the chemotactic endothelial stimuli, the cell populations started to form capillary-like structures and up-regulated the expression of specific endothelial markers (CD34, PDGFRα, VEGFR2, VE-cadherin, CD31, and vWF). A dot blot protein study detected the presence of FVIII in culture media collected from both unstimulated and stimulated Ad-MSCs. Remarkably, the activated partial thromboplastin time test demonstrated that the clot formation was accelerated, and FVIII activity was enhanced when FVIII deficient plasma was mixed with culture media from the untreated/stimulated Ad-MSCs. Overall, the collected evidence supported a possible Ad-MSC contribution to HA correction via specific stimulation by the endothelial microenvironment and without any need for gene transfection.
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Hemofilia A , Células Madre Mesenquimatosas , Adulto , Pruebas de Coagulación Sanguínea , Diferenciación Celular , Células Cultivadas , Medios de Cultivo/metabolismo , Hemofilia A/genética , Hemofilia A/metabolismo , Hemofilia A/terapia , Humanos , Células Madre Mesenquimatosas/metabolismo , Tiempo de Tromboplastina ParcialRESUMEN
PURPOSE: To evaluate the accuracy of the trampoline and hook tests, used in the arthroscopic assessment of triangular fibrocartilage complex (TFCC) tears compared with arthroscopic direct visualization of the radiocarpal joint (RCJ) and of the distal radial ulnar joint (DRUJ). METHODS: In total, 135 patients (97 male, 38 female, mean age 43.5 years) were divided into 2 groups: (1) 80 patients with chronic ulnar-sided wrist pain and positive fovea sign and (2) 55 patients with other complaints. TFCC was assessed by RCJ and DRUJ arthroscopy and by the trampoline and hook tests to detect rupture of distal and proximal components of the TFCC. Accuracy, specificity, sensitivity, and likelihood ratio of the 2 diagnostic methods were measured and compared, using RCJ and DRUJ arthroscopy as reference. RESULTS: The trampoline and the hook tests showed an overall accuracy of 70.37% and 86.67%, respectively. The accuracy of the trampoline test was similar for distal (69%), proximal (66%), and complete (73%) TFCC tears. The hook test was more accurate when evaluating proximal (97%) and complete (98%) tears, rather than distal lesions (75%). Sensitivity for the trampoline and hook tests was 75.00% and 0.00% (P < .001) for distal tears and 78.85% and 100.00% (P < .001) and 58.33% and 100.00% (P < .001) for complete or isolated proximal tears, respectively. Specificity for the trampoline and hook tests was 67.27% and 96.36% (P < .001) respectively. CONCLUSIONS: The trampoline and hook tests can assure accurate diagnosis of peripheral TFCC tear. The hook test shows greater specificity and sensitivity to recognize foveal TFCC tears. Values of positive likelihood ratio suggest a greater probability to detect foveal laceration of peripheral TFCC for the hook test than for the trampoline test. These findings suggest that DRUJ arthroscopy is not necessary to confirm foveal incompetence of the TFCC, if the hook test is positive. LEVEL OF EVIDENCE: Level II, retrospective diagnostic trial.
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Fibrocartílago Triangular , Traumatismos de la Muñeca , Adulto , Artroscopía , Femenino , Humanos , Masculino , Estudios Retrospectivos , Muñeca , Traumatismos de la Muñeca/diagnóstico , Articulación de la MuñecaRESUMEN
The carotid body (CB) plays a contributory role in the pathogenesis of various respiratory, cardiovascular, renal, and metabolic diseases through reflex changes in ventilation and sympathetic output. On the basis of available data about peripheral arterial chemoreception and severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2), a potential involvement in the coronavirus disease 2019 (COVID-19) may be hypothesized through different mechanisms. The CB could be a site of SARS-CoV-2 invasion, due to local expression of its receptor [angiotensin-converting enzyme (ACE) 2] and an alternative route of nervous system invasion, through retrograde transport along the carotid sinus nerve. The CB function could be affected by COVID-19-induced inflammatory/immune reactions and/or ACE1/ACE2 imbalance, both at local or systemic level. Increased peripheral arterial chemosensitivity and reflex sympatho-activation may contribute to the increased morbidity and mortality in COVID-19 patients with respiratory, cardiovascular, renal, or metabolic comorbidities.
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Cuerpo Carotídeo/metabolismo , Sistema Nervioso Central/virología , Infecciones por Coronavirus/patología , Peptidil-Dipeptidasa A/metabolismo , Neumonía Viral/patología , Enzima Convertidora de Angiotensina 2 , Betacoronavirus , COVID-19 , Seno Carotídeo/inervación , Seno Carotídeo/virología , Humanos , Pandemias , Sistema Renina-Angiotensina/fisiología , SARS-CoV-2RESUMEN
The carotid body may undergo plasticity changes during development/ageing and in response to environmental (hypoxia and hyperoxia), metabolic, and inflammatory stimuli. The different cell types of the carotid body express a wide series of growth factors and corresponding receptors, which play a role in the modulation of carotid body function and plasticity. In particular, type I cells express nerve growth factor, brain-derived neurotrophic factor, neurotrophin 3, glial cell line-derived neurotrophic factor, ciliary neurotrophic factor, insulin-like-growth factor-I and -II, basic fibroblast growth factor, epidermal growth factor, transforming growth factor-α and -ß, interleukin-1ß and -6, tumor necrosis factor-α, vascular endothelial growth factor, and endothelin-1. Many specific growth factor receptors have been identified in type I cells, indicating autocrine/paracrine effects. Type II cells may also produce growth factors and express corresponding receptors. Future research will have to consider growth factors in further experimental models of cardiovascular, metabolic, and inflammatory diseases and in human (normal and pathologic) samples. From a methodological point of view, microarray and/or proteomic approaches would permit contemporary analyses of large groups of growth factors. The eventual identification of physical interactions between receptors of different growth factors and/or neuromodulators could also add insights regarding functional interactions between different trophic mechanisms.
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Factor Neurotrófico Derivado del Encéfalo/genética , Cuerpo Carotídeo/metabolismo , Hiperoxia/genética , Hipoxia/genética , Factor de Crecimiento Nervioso/genética , Receptores de Factores de Crecimiento/genética , Animales , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Cuerpo Carotídeo/citología , Factor Neurotrófico Ciliar/genética , Factor Neurotrófico Ciliar/metabolismo , Factor de Crecimiento Epidérmico/genética , Factor de Crecimiento Epidérmico/metabolismo , Factor 2 de Crecimiento de Fibroblastos/genética , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Regulación de la Expresión Génica , Factor Neurotrófico Derivado de la Línea Celular Glial/genética , Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Humanos , Hiperoxia/metabolismo , Hiperoxia/patología , Hipoxia/metabolismo , Hipoxia/patología , Factor I del Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/metabolismo , Factor de Crecimiento Nervioso/metabolismo , Neurotrofina 3/genética , Neurotrofina 3/metabolismo , Receptores de Factores de Crecimiento/metabolismo , Factor de Crecimiento Transformador alfa/genética , Factor de Crecimiento Transformador alfa/metabolismo , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismo , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Partial oxidation of polyvinyl alcohol (PVA) with potassium permanganate turned out to be an efficient method to fabricate smart scaffolds for tissue engineering, endowed with biodegradation and protein delivery capacity. This work considered for the first time the use of halogens (bromine, chlorine and iodine) as less aggressive agents than potassium permanganate to perform controlled PVA oxidation, in order to prevent degradation of polymer molecular size upon chemical modification. Oxidized PVA solutions were chemically characterized (i.e., dinitrophenylhydrazine assay, viscosity measurements, molecular size distribution) before preparing physically cross-linked hydrogels. Scaffolds were assessed for their mechanical properties and cell/tissue biocompatibiliy through cytotoxic extract test on IMR-90 fibroblasts and subcutaneous implantation into BALB/c mice. According to chemical investigations, bromine and iodine allowed for minor alteration of polymer molecular weight. Uniaxial tensile tests demonstrated that oxidized scaffolds had decreased mechanical resistance to deformation, suggesting tunable hydrogel stiffness. Finally, oxidized hydrogels exhibited high biocompatibility both in vitro and in vivo, resulting neither to be cytotoxic nor to elicit severe immunitary host reaction in comparison with atoxic PVA. In conclusion, PVA hydrogels oxidized by halogens were successfully fabricated in the effort of adapting polymer characteristics to specific tissue engineering applications.
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Halógenos/química , Oxidación-Reducción , Alcohol Polivinílico/química , Ingeniería de Tejidos , Animales , Materiales Biocompatibles , Biopsia , Espectroscopía de Resonancia Magnética con Carbono-13 , Inmunohistoquímica , Ensayo de Materiales , Fenómenos Mecánicos , Ratones , Estructura Molecular , Andamios del Tejido , ViscosidadRESUMEN
Nowadays, research in Tissue Engineering and Regenerative Medicine is focusing on the identification of instructive scaffolds to address the requirements of both clinicians and patients to achieve prompt and adequate healing in case of injury. Among biomaterials, hemocomponents, and in particular Platelet-rich Fibrin matrices, have aroused widespread interest, acting as delivery platforms for growth factors, cytokines and immune/stem-like cells for immunomodulation; their autologous origin and ready availability are also noteworthy aspects, as safety- and cost-related factors and practical aspects make it possible to shorten surgical interventions. In fact, several authors have focused on the use of Platelet-rich Fibrin in cartilage and tendon tissue engineering, reporting an increasing number of in vitro, pre-clinical and clinical studies. This narrative review attempts to compare the relevant advances in the field, with particular reference being made to the regenerative role of platelet-derived growth factors, as well as the main pre-clinical and clinical research on Platelet-rich Fibrin in chondrogenesis and tenogenesis, thereby providing a basis for critical revision of the topic.
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Cartílago/fisiología , Fibrina Rica en Plaquetas/química , Medicina Regenerativa , Tendones/fisiología , Andamios del Tejido/química , Investigación Biomédica Traslacional , HumanosRESUMEN
Tissue engineering and regenerative medicine involve many different artificial and biologic materials, frequently integrated in composite scaffolds, which can be repopulated with various cell types. One of the most promising scaffolds is decellularized allogeneic extracellular matrix (ECM) then recellularized by autologous or stem cells, in order to develop fully personalized clinical approaches. Decellularization protocols have to efficiently remove immunogenic cellular materials, maintaining the nonimmunogenic ECM, which is endowed with specific inductive/differentiating actions due to its architecture and bioactive factors. In the present paper, we review the available literature about the development of grafts from decellularized human tissues/organs. Human tissues may be obtained not only from surgery but also from cadavers, suggesting possible development of Human Tissue BioBanks from body donation programs. Many human tissues/organs have been decellularized for tissue engineering purposes, such as cartilage, bone, skeletal muscle, tendons, adipose tissue, heart, vessels, lung, dental pulp, intestine, liver, pancreas, kidney, gonads, uterus, childbirth products, cornea, and peripheral nerves. In vitro recellularizations have been reported with various cell types and procedures (seeding, injection, and perfusion). Conversely, studies about in vivo behaviour are poorly represented. Actually, the future challenge will be the development of human grafts to be implanted fully restored in all their structural/functional aspects.
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Trasplante de Células Madre/tendencias , Células Madre/citología , Ingeniería de Tejidos/tendencias , Andamios del Tejido , Matriz Extracelular/trasplante , Humanos , Medicina de Precisión , Trasplantes/citología , Trasplantes/trasplanteRESUMEN
Haemophilic arthropathy is the major cause of disability in patients with haemophilia and, despite prophylaxis with coagulation factor concentrates, some patients still develop articular complications. We evaluate the feasibility of a tissue engineering approach to improve current clinical strategies for cartilage regeneration in haemophiliacs by using autologous chondrocytes (haemophilic chondrocytes; HaeCs). Little is known about articular chondrocytes from haemophilic patients and no characterisation has as yet been performed. An investigation into whether blood exposure alters HaeCs should be interesting from the perspective of autologous implants. The typical morphology and expression of specific target genes and surface markers were therefore assessed by optical microscopy, reverse transcription plus the polymerase chain reaction (PCR), real-time PCR and flow-cytometry. We then considered chondrocyte behaviour on a bio-hybrid scaffold (based on polyvinyl alcohol/Wharton's jelly) as an in vitro model of articular cartilage prosthesis. Articular chondrocytes from non-haemophilic donors were used as controls. HaeC morphology and the resulting immunophenotype CD44(+)/CD49c(+)/CD49e(+)/CD151(+)/CD73(+)/CD49f(-)/CD26(-) resembled those of healthy donors. Moreover, HaeCs were active in the transcription of genes involved in the synthesis of the extracellular matrix proteins of the articular cartilage (ACAN, COL1A, COL2A, COL10A, COL9A, COMP, HAS1, SOX9), although the over-expression of COL1A1, COL10A1, COMP and HAS was observed. In parallel, the composite scaffold showed adequate mechanical and biological properties for cartilage tissue engineering, promoting chondrocyte proliferation. Our preliminary evidence contributes to the characterisation of HaeCs, highlighting the opportunity of using them for autologous cartilage implants in patients with haemophilia.
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Condrocitos/citología , Condrogénesis , Hemofilia A/patología , Cartílago Articular/efectos de los fármacos , Cartílago Articular/patología , Proliferación Celular/efectos de los fármacos , Forma de la Célula/efectos de los fármacos , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Condrocitos/ultraestructura , Condrogénesis/efectos de los fármacos , Módulo de Elasticidad/efectos de los fármacos , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Hemofilia A/genética , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Alcohol Polivinílico/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Estrés Mecánico , Andamios del Tejido , Trasplante AutólogoRESUMEN
In regenerative neurobiology, Ciliary Neurotrophic Factor (CNTF) is raising high interest as a multifunctional neurocytokine, playing a key role in the regeneration of injured peripheral nerves. Despite its promising trophic and regulatory activity, its clinical application is limited by the onset of severe side effects, due to the lack of efficient intracellular trafficking after administration. In this study, recombinant CNTF linked to the transactivator transduction domain (TAT) was investigated in vitro and found to be an optimized fusion protein which preserves neurotrophic activity, besides enhancing cellular uptake for therapeutic advantage. Moreover, a compelling protein delivery method was defined, in the future perspective of improving nerve regeneration strategies. Following determination of TAT-CNTF molecular weight and concentration, its specific effect on neural SH-SY5Y and PC12 cultures was assessed. Cell proliferation assay demonstrated that the fusion protein triggers PC12 cell growth within 6h of stimulation. At the same time, the activation of signal transduction pathway and enhancement of cellular trafficking were found to be accomplished in both neural cell lines after specific treatment with TAT-CNTF. Finally, the recombinant growth factor was successfully loaded on oxidized polyvinyl alcohol (PVA) scaffolds, and more efficiently released when polymer oxidation rate increased. Taken together, our results highlight that the TAT domain addiction to the protein sequence preserves CNTF specific neurotrophic activity in vitro, besides improving cellular uptake. Moreover, oxidized PVA could represent an ideal biomaterial for the development of nerve conduits loaded with the fusion protein to be delivered to the site of nerve injury.
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Factor Neurotrófico Ciliar/uso terapéutico , Productos del Gen tat/química , Regeneración Nerviosa , Nervios Periféricos/fisiología , Animales , Línea Celular Tumoral , Proliferación Celular , Factor Neurotrófico Ciliar/química , Humanos , Ratas , Transducción de SeñalRESUMEN
Oxidative stress and fibrosis are important stress responses that characterize bronchopulmonary dysplasia (BPD), a disease for which only a therapy but not a cure has been developed. In this work, we investigated the effects of mesenchymal stromal cells-derived extracellular vesicles (MSC-EVs) on lung and brain compartment in an animal model of hyperoxia-induced BPD. Rat pups were intratracheally injected with MSC-EVs produced by human umbilical cord-derived MSC, following the Good Manufacturing Practice-grade (GMP-grade). After evaluating biodistribution of labelled MSC-EVs in rat pups left in normoxia and hyperoxia, oxidative stress and fibrosis investigation were performed. Oxidative stress protection by MSC-EVs treatment was proved both in lung and in brain. The lung epithelial compartment ameliorated glycosaminoglycan and surfactant protein expression in MSC-EVs-injected rat pups compared to untreated animals. Pups under hyperoxia exhibited a fibrotic phenotype in lungs shown by increased collagen deposition and also expression of profibrotic genes. Both parameters were reduced by treatment with MSC-EVs. We established an in vitro model of fibrosis and another of oxidative stress, and we proved that MSC-EVs suppressed the induction of αSMA, influencing collagen deposition and protecting from the oxidative stress. In conclusion, intratracheal administration of clinical-grade MSC-EVs protect from oxidative stress, improves pulmonary epithelial function, and counteracts the development of fibrosis. In the future, MSC-EVs could represent a new cure to prevent the development of BPD.
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Displasia Broncopulmonar , Vesículas Extracelulares , Hiperoxia , Células Madre Mesenquimatosas , Recién Nacido , Ratas , Animales , Humanos , Displasia Broncopulmonar/terapia , Distribución Tisular , Vesículas Extracelulares/metabolismo , Fibrosis , Cordón Umbilical/metabolismo , Células Madre Mesenquimatosas/metabolismo , Estrés Oxidativo , Colágeno/metabolismo , Modelos Animales de EnfermedadRESUMEN
Angiotensin-converting enzyme 2 receptor (ACE2R) is a transmembrane protein expressed in various tissues throughout the body that plays a key role in the regulation of blood pressure. Recently, ACE2R has gained significant attention due to its involvement in the pathogenesis of COVID-19, the disease caused by the Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2). While ACE2 receptors serve as entry points for the novel coronavirus, Transmembrane Serine Protease 2 (TMPRSS2), an enzyme located on the cell membrane, is required for SARS-CoV-2 S protein priming. Even though numerous studies have assessed the effects of COVID-19 on the brain, very little information is available concerning the distribution of ACE2R and TMPRSS2 in the human brain, with particular regard to their topographical expression in the brainstem. In this study, we investigated the expression of ACE2R and TMPRSS2 in the brainstem of 18 adult subjects who died due to pneumonia/respiratory insufficiency. Our findings indicate that ACE2R and TMPRSS2 are expressed in neuronal and glial cells of the brainstem, particularly at the level of the vagal nuclei of the medulla and the midbrain tegmentum, thus confirming the expression and anatomical localization of these proteins within specific human brainstem nuclei. Furthermore, our findings help to define anatomically susceptible regions to SARS-CoV-2 infection in the brainstem, advancing knowledge on the neuropathological underpinnings of neurological manifestations in COVID-19.
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COVID-19 , Adulto , Humanos , SARS-CoV-2 , Enzima Convertidora de Angiotensina 2 , Tronco Encefálico , Serina Endopeptidasas/genéticaRESUMEN
Severe tracheal injuries that cannot be managed by mobilization and end-to-end anastomosis represent an unmet clinical need and an urgent challenge to face in surgical practice; within this scenario, decellularized scaffolds (eventually bioengineered) are currently a tempting option among tissue engineered substitutes. The success of a decellularized trachea is expression of a balanced approach in cells removal while preserving the extracellular matrix (ECM) architecture/mechanical properties. Revising the literature, many Authors report about different methods for acellular tracheal ECMs development; however, only few of them verified the devices effectiveness by an orthotopic implant in animal models of disease. To support translational medicine in this field, here we provide a systematic review on studies recurring to decellularized/bioengineered tracheas implantation. After describing the specific methodological aspects, orthotopic implant results are verified. Furtherly, the only three clinical cases of compassionate use of tissue engineered tracheas are reported with a focus on outcomes.
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In severe peripheral nerve injuries, nerve conduits (NCs) are good alternatives to autografts/allografts; however, the results the available devices guarantee for are still not fully satisfactory. Herein, differently bioactivated NCs based on the new polymer oxidized polyvinyl alcohol (OxPVA) are compared in a rat model of sciatic nerve neurotmesis (gap: 5 mm; end point: 6 weeks). Thirty Sprague Dawley rats are randomized to 6 groups: Reverse Autograft (RA); Reaxon®; OxPVA; OxPVA + EAK (self-assembling peptide, mechanical incorporation); OxPVA + EAK-YIGSR (mechanical incorporation); OxPVA + Nerve Growth Factor (NGF) (adsorption). Preliminarily, all OxPVA-based devices are comparable with Reaxon® in Sciatic Functional Index score and gait analysis; moreover, all conduits sustain nerve regeneration (S100, ß-tubulin) without showing substantial inflammation (CD3, F4/80) evidences. Following morphometric analyses, OxPVA confirms its potential in PNI repair (comparable with Reaxon®) whereas OxPVA + EAK-YIGSR stands out for its myelinated axons total number and density, revealing promising in injury recovery and for future application in clinical practice.
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Tracheal reconstruction represents a challenge when primary anastomosis is not feasible. Within this scenario, the study aim was to develop a new pig-derived decellularized trachea (DecellT) to be compared with the cryopreserved counterpart (CryoT) for a close predictive analysis. Tracheal segments underwent decellularization by a physical + enzymatic + chemical method (12 cycles); in parallel, cryopreserved samples were also prepared. Once decellularized (histology/DNA quantification), the two groups were characterized for Alpha-Gal epitopes/structural proteins (immunohistochemistry/histology/biochemical assays/second harmonic generation microscopy)/ultrastructure (Scanning Electron Microscopy (SEM))/mechanical behaviour. Cytotoxicity absence was assessed in vitro (extract-test assay/direct seeding, HM1SV40 cell line) while biocompatibility was verified in BALB/c mice, followed by histological/immunohistochemical analyses and SEM (14 days). Decellularization effectively removed Alpha-Gal epitopes; cartilage histoarchitecture was retained in both groups, showing chondrocytes only in the CryoT. Cryopreservation maintained few respiratory epithelium sparse cilia, not detectable in DecellT. Focusing on ECM, preserved structural/ultrastructural organization and collagen content were observed in the cartilage of both; conversely, the GAGs were significantly reduced in DecellT, as confirmed by mechanical study results. No cytotoxicity was highlighted by CryoT/DecellT in vitro, as they were also corroborated by a biocompatibility assay. Despite some limitations (cells presence/GAGs reduction), CryoT/DecellT are both appealing options, which warrant further investigation in comparative in vivo studies.
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Ingeniería de Tejidos , Andamios del Tejido , Ratones , Porcinos , Animales , Ingeniería de Tejidos/métodos , Matriz Extracelular/metabolismo , Colágeno/metabolismo , Criopreservación/métodosRESUMEN
Polyvinyl alcohol (PVA) hydrogels are extensively used as scaffolds for tissue engineering, although their biodegradation properties have not been optimized yet. To overcome this limitation, partially oxidized PVA has been developed by means of different oxidizing agents, obtaining scaffolds with improved biodegradability. The oxidation reaction also allows tuning the mechanical properties, which are essential for effective use in vivo. In this work, the compressive mechanical behavior of native and partially oxidized PVA hydrogels is investigated, to evaluate the effect of different oxidizing agents, i.e., potassium permanganate, bromine, and iodine. For this purpose, PVA hydrogels are tested by means of indentation tests, also considering the time-dependent mechanical response. Indentation results show that the oxidation reduces the compressive stiffness from about 2.3 N/mm for native PVA to 1.1 ÷ 1.4 N/mm for oxidized PVA. During the consolidation, PVA hydrogels exhibit a force reduction of about 40% and this behavior is unaffected by the oxidizing treatment. A poroviscoelastic constitutive model is developed to describe the time-dependent mechanical response, accounting for the viscoelastic polymer matrix properties and the flow of water molecules within the matrix during long-term compression. This model allows to estimate the long-term Young's modulus of PVA hydrogels in drained conditions (66 kPa for native PVA and 34-42 kPa for oxidized PVA) and can be exploited to evaluate their performances under compressive stress in vivo, as in the case of cartilage tissue engineering.
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Meniscal tears are a frequent orthopedic injury commonly managed by conservative strategies to avoid osteoarthritis development descending from altered biomechanics. Among cutting-edge approaches in tissue engineering, 3D printing technologies are extremely promising guaranteeing for complex biomimetic architectures mimicking native tissues. Considering the anisotropic characteristics of the menisci, and the ability of printing over structural control, it descends the intriguing potential of such vanguard techniques to meet individual joints' requirements within personalized medicine. This literature review provides a state-of-the-art on 3D printing for meniscus reconstruction. Experiences in printing materials/technologies, scaffold types, augmentation strategies, cellular conditioning have been compared/discussed; outcomes of pre-clinical studies allowed for further considerations. To date, translation to clinic of 3D printed meniscal devices is still a challenge: meniscus reconstruction is once again clear expression of how the integration of different expertise (e.g., anatomy, engineering, biomaterials science, cell biology, and medicine) is required to successfully address native tissues complexities.
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Polyvinyl alcohol (PVA) hydrogels are synthetic polymers which can be used as scaffolds for tissue engineering due to their biocompatibility and large water content. To improve their biodegradation properties, partial oxidation of PVA is achieved by means of different oxidizing agents, such as potassium permanganate, bromine and iodine. The effect of this process on hydrogels mechanical performance has not been fully investigated in view of tissue engineering applications. In this work, the time-dependent mechanical behavior of unmodified and partially oxidized PVA hydrogels is evaluated by means of uniaxial tensile and stress relaxation tests, to evaluate the effect of different oxidizing agents on the viscoelastic response. Tensile tests show an isotropic and almost-incompressible behavior, with a stiffness reduction after PVA oxidation. The time-dependent response of oxidized PVA is comparable to the one of unmodified PVA and is modeled as a quasi-linear viscoelastic behavior. Finite Element (FE) models of PVA samples are developed and numerical analyses are used to evaluate the effect of different strain rates on the mechanical response under uniaxial tension. This model can be exploited to predict the time-dependent mechanical behavior of partially oxidized PVA in tissue engineering application under tensile loading.