RESUMO
The discovery of several unexpected complex biological roles of hyaluronic acid (HA) has promoted new research impetus for biologists and, the clinical interest in several fields of medicine, such as ophthalmology, articular pathologies, cutaneous repair, skin remodeling, vascular prosthesis, adipose tissue engineering, nerve reconstruction and cancer therapy. In addition, the great potential of HA in medicine has stimulated the interest of pharmaceutical companies which, by means of new technologies can produce HA and several new derivatives in order to increase both the residence time in a variety of human tissues and the anti-inflammatory properties. Minor chemical modifications of the molecule, such as the esterification with benzyl alcohol (Hyaff-11® biomaterials), have made possible the production of water-insoluble polymers that have been manufactured in various forms: membranes, gauzes, nonwoven meshes, gels, tubes. All these biomaterials are used as wound-covering, anti-adhesive devices and as scaffolds for tissue engineering, such as epidermis, dermis, micro-vascularized skin, cartilage and bone. In this review, the essential biological functions of HA and the applications of its derivatives for pharmaceutical and tissue regeneration purposes are reviewed.
Assuntos
Ácido Hialurônico/metabolismo , Tecido Adiposo/metabolismo , Animais , Humanos , Ácido Hialurônico/biossíntese , Neoplasias/patologia , Neoplasias/terapia , Receptores de Superfície Celular/metabolismo , Pele/metabolismo , Engenharia TecidualRESUMO
BACKGROUND: Vernal keratoconjunctivitis (VKC) is a chronic ocular allergic inflammation characterized by corneal complications and the formation of giant papillae. Sma- and Mad-related proteins (Smad) modulate extracellular matrix gene expression during wound healing, inflammation and tissue remodelling. OBJECTIVE: To investigate the relationship between allergic inflammation and TGF-ß/Smad signalling pathway, expression in VKC patients and in primary cultured conjunctival fibroblasts exposed to mediators found previously over-expressed in VKC. METHODS: Smad-2, -3, -7, phospho-(p)Smads, TGF-ß1 and -ß2 were evaluated in the conjunctiva of normal subjects (CT) and VKC patients by immunohistochemistry. The expression of Smads, pro-collagen I (PIP), TGF-ß1, -ß2, mitogen-activated protein kinase (p38/MAPK), c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK1/2) were also determined in conjunctival fibroblast cultures exposed to histamine, IL-4, -13, TGF-ß1, IFN-γ and TNF-α using immunostaining or RT-PCR. RESULTS: Immunostaining for Smad-2, -3, pSmad-2, -3, TGF-ß1, -ß2 and PIP was significantly increased in VKC stroma compared with CT. In conjunctival fibroblast cultures, Smad-3 and PIP were stimulated by histamine, IL-4, -13 and TGF-ß1 exposure, while PIP was reduced by IFN-γ, and TNF-α mRNA expression of Smad-3 was increased by histamine, while Smad-7 was reduced by IL-4. In addition, histamine, IL-4 and TNF-α increased JNK and ERK1/2 expression. CONCLUSION AND CLINICAL RELEVANCE: The TGF-ß/Smad signalling pathway is over-expressed in VKC tissues and modulated in conjunctival fibroblasts by histamine, IL-4, TGF-ß1 and TNF-α. These mechanisms may be involved in fibrillar collagen production, giant papillae formation and tissue remodelling typical of VKC and might provide new therapeutic targets for its treatment.
Assuntos
Túnica Conjuntiva/imunologia , Conjuntivite Alérgica/imunologia , Modelos Imunológicos , Transdução de Sinais/imunologia , Proteínas Smad/imunologia , Fator de Crescimento Transformador beta/imunologia , Adolescente , Criança , Conjuntivite Alérgica/diagnóstico , Conjuntivite Alérgica/genética , Feminino , Humanos , Masculino , RNA Mensageiro/genética , RNA Mensageiro/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Smad/genética , Fator de Crescimento Transformador beta/genéticaRESUMO
Atypical antipsychotics (APDs) are currently used in clinical practice for a variety of mental disorders such as schizophrenia, bipolar disorder and severe behavioral disturbances. A well-known disadvantage of using these compounds is a propensity for weight gain, resulting frequently in obesity. The mechanisms underlying pharmacologically induced weight gain are still controversial. The objective of this study was to evaluate in vitro the effects of different APDs on adipogenic events in cultured human pre-adipocytes and in rat muscle-derived stem cells (MDSCs), aiming to identify a common intracellular event contributable to these drugs. Culture behavior was evaluated in terms of cell proliferation, lipid accumulation, gene expression and morphological features. Results indicate that APDs influence adipogenic events through changes in the differentiation and proliferation of pre-adipocytes and MDSCs that are brought on by protein kinase C-ß (PKC-ß) activation. These data identify a signaling route that could be a potential target of pharmacological approaches for preventing the weight gain associated with APD treatment.
Assuntos
Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Antipsicóticos/efeitos adversos , Proteína Quinase C/metabolismo , Células-Tronco/efeitos dos fármacos , Aumento de Peso/efeitos dos fármacos , Adipócitos/citologia , Adipócitos/enzimologia , Adipogenia/genética , Animais , Animais Recém-Nascidos , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Ativação Enzimática , Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/genética , Proteína Quinase C beta , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Ratos , Células-Tronco/citologia , Células-Tronco/enzimologia , Aumento de Peso/genéticaRESUMO
The present study describes the production of hyaluronan based porous microparticles by a semi-continuous gas anti-solvent (GAS) precipitation process to be used as a growth factor delivery system for in vivo treatment of ulcers. Operative process conditions, such as pressure, nozzle diameter and HYAFF11 solution concentrations, were adjusted to optimize particle production in terms of morphology and size. Scanning electron microscopy (SEM) and light scattering demonstrated that porous nano-structured particles with a size of 300 and 900 nm had a high specific surface suitable for absorption of growth factors from the aqueous environment within the polymeric matrix. Water acted as a plasticizer, enhancing growth factor absorption. Water contents within the HYAFF11 matrix were analyzed by differential scanning calorimetry (DSC). The absorption process was developed using fluorescence dyes and growth factors. Immunohistochemical analysis confirmed the high efficiency of absorption of growth factor and a mathematical model was generated to quantify and qualify the in vitro kinetics of growth factor release within the polymeric matrix. In vivo experiments were performed with the aim to optimize timed and focal release of PDGF to promote optimal tissue repair and regeneration of full-thickness wounds.
Assuntos
Materiais Biocompatíveis/química , Substâncias de Crescimento/administração & dosagem , Ácido Hialurônico/química , Nanopartículas/química , Nanopartículas/uso terapêutico , Úlcera Cutânea/terapia , Absorção , Animais , Masculino , Teste de Materiais , Microscopia Eletrônica de Varredura , Nanopartículas/ultraestrutura , Nanotecnologia/instrumentação , Fator de Crescimento Derivado de Plaquetas/administração & dosagem , Ratos , Ratos Wistar , Úlcera Cutânea/tratamento farmacológico , Úlcera Cutânea/patologia , Alicerces Teciduais , Fator de Crescimento Transformador beta/administração & dosagem , Água/química , Cicatrização/efeitos dos fármacosRESUMO
AIM: Thrombosis secondary to renal closed abdominal trauma is a rare event, most of the time it is clinically silent. We report here our experience. MATERIALS AND METHODS. This is the case of a boy came to our observation after a road trauma with motorbike fall-out. The boy arrived in ED for head injury. The patient, stable for haemodynamics, had lacerated and contused injuries at pelvis and right buttock level. He underwent chest x-rays, brain CT and neurosurgery examination: all resulted negative. There was no macrohematuria, nor lumbar pain. Objectively abdomen was treatable. The patient was referred to temporary observation for 12 hours when he was asked to undergo abdomen ultrasound, which showed no documented lesions except for fluid collection at the pelvic level. To rule out all doubts, the patient had an abdominal CT scan, which showed a silent left kidney with suspected thrombosis at left renal level. The patient was sent to our attention after 15 hours: we decided to perform immediately selective arteriography with thrombus lysis. The arteriography documented a massive thrombosis. The thrombus lysis was impossible to be performed. To maintain the perfect functionality of the contralateral kidney we decided not to proceed further, but to perform only left nephrectomy. During surgery mesocolon laceration occurred, so the patient underwent also colic resection. DISCUSSION. Thrombosis secondary to a closed renal abdominal trauma is an uncommon event, with little clinical expression. It is the consequence of an injury. Deceleration produces arterial dissection, which alters the blood flow to the kidney, which is then twisted and complicated with renal thrombosis. Quite common is the association with diaphragmatic rupture or urethral detachment. The alterations of renal parenchyma in the early hours are detectable only through CT scan, which represents the method of election, and which can highlight a functionally silent kidney. CONCLUSIONS. Renal thrombosis requires that diagnosis is done within the first 12 hours; a rapid revascularization should be promptly attempted.
RESUMO
Hyaluronan-based scaffold were used for in vitro commitment of human and rat bone marrow mesenchymal stem cells (MSC). Cells were cultured either in monolayer and in 3D conditions up to 35 days. In order to monitor the differentiating processes molecular biology and morphological studies were performed at different time points. All the reported data supported the evidence that both human and rat MSC grown onto hyaluronan-derived three-dimensional scaffold were able to acquire a unique phenotype of chondrocytes and osteocytes depending on the presence of specific differentiation inducing factors added into the culture medium without significative differences in term of time expression of extracellular matrix proteins.
Assuntos
Células-Tronco Adultas/citologia , Condrócitos/citologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Adultas/metabolismo , Células-Tronco Adultas/ultraestrutura , Animais , Células da Medula Óssea , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Condrócitos/metabolismo , Condrogênese/fisiologia , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Colágeno Tipo II/genética , Colágeno Tipo II/metabolismo , Citometria de Fluxo , Humanos , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/ultraestrutura , Osteocalcina/genética , Osteocalcina/metabolismo , Osteogênese , Osteonectina/genética , Osteonectina/metabolismo , Osteopontina/genética , Osteopontina/metabolismo , Ratos , Ratos Endogâmicos F344 , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
BACKGROUND: Allergic conditions in different organs share many similarities in their inflammatory response. Vernal keratoconjunctivitis (VKC), asthma and nasal polyps exhibit several similar, but site-specific mucosal structural changes. The aim of the study was to investigate whether matrix metalloproteases contribute to different tissue remodelling aspects in different organs. METHODS: Mucosal biopsies were obtained from conjunctiva of healthy donors, tarsal conjunctiva of vernal patients, bronchi of non-asthmatic subjects, bronchi of mild stable asthmatic patients, nasal mucosa of non-allergic donors and nasal polyps of allergic patients. Distribution of metalloprotease-1, -3, -9, -13, tissue inhibitor of metalloproteases-1, collagens I and III and the presence of eosinophils and CD4+ cells were evaluated by immunohistochemistry. RESULTS: Collagens were highly diffuse in the giant papillae of VKC and in nasal polyps, and yet less increased in the subepithelium of asthmatic patients. Immunostaining for metalloprotease-1, -3, -9 and -13 was significantly higher in VKC compared with normal conjunctiva. Metalloprotease-9 staining was higher in the stroma of polyps vs. normal nasal mucosa, and only metalloprotease-13 was significantly more expressed in asthmatic vs. non-asthmatic subjects. Metalloprotease-9 immunostaining was more intense in vernal compared with other tissues. In all pathological tissues, metalloprotease-9-positive staining was in association with eosinophils and CD4+ cells. CONCLUSIONS: Expression of metalloproteases may play an important role in inducing the structural changes seen in VKC, nasal polyps and asthma. Tissue remodelling and gelatinase immunoexpression was more dramatic in giant papillae of vernal patients compared with other tissue sites of chronic allergic inflammation.
Assuntos
Asma/imunologia , Linfócitos T CD4-Positivos/imunologia , Conjuntivite Alérgica/imunologia , Eosinófilos/imunologia , Metaloproteinases da Matriz/imunologia , Pólipos Nasais/imunologia , Adolescente , Adulto , Asma/enzimologia , Asma/patologia , Linfócitos T CD4-Positivos/enzimologia , Linfócitos T CD4-Positivos/patologia , Criança , Colágeno Tipo I/imunologia , Colágeno Tipo I/metabolismo , Colágeno Tipo III/imunologia , Colágeno Tipo III/metabolismo , Conjuntivite Alérgica/enzimologia , Conjuntivite Alérgica/patologia , Eosinófilos/enzimologia , Eosinófilos/patologia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Metaloproteinases da Matriz/metabolismo , Pessoa de Meia-Idade , Mucosa/enzimologia , Mucosa/imunologia , Mucosa/patologia , Pólipos Nasais/enzimologia , Pólipos Nasais/patologia , Especificidade de Órgãos/imunologia , Inibidor Tecidual de Metaloproteinase-1/imunologia , Inibidor Tecidual de Metaloproteinase-1/metabolismoRESUMO
In this study we coated a new biocompatible, nanostructured titanium alloy, Ti13Nb13Zr, with a thin layer of hydroxyapatite nanocrystals and we investigated the response of human bone-marrow-derived mesenchymal cells. The coating was realized using a slightly supersaturated CaP solution, which provokes a fast deposition of nanocrystalline hydroxyapatite. A thin layer of deposition is appreciable on the etched Ti13Nb13Zr substrates after just 1.5 h soaking in the CaP solution, and it reaches a thickness of 1-2 mum after 3 h soaking. The coating seems thinner than that deposited on Ti6Al4V, which was examined for comparison, likely because of the different roughness profiles of the two etched alloys, and it is constituted of elongated HA nanocrystals, with a mean length of about 100 nm. Mesenchymal stem cells were seeded onto coated and uncoated Ti alloys and cultured for up to 35 days. Cell morphology, proliferation and differentiation were evaluated. The cells display good adhesion and proliferation on the uncoated substrates, whereas the presence of hydroxyapatite coating slightly reduces cell proliferation and induces differentiation of MSCs towards a phenotypic osteoblastic lineage, in agreement with the increase of the expression of osteopontin, osteonectin and collagen type I, evaluated by means of rt-PCR. Type I collagen expression is higher in Ti13Nb13Zr MSC culture compared to Ti6Al4V, standing for a more efficient extracellular matrix deposition.
Assuntos
Materiais Revestidos Biocompatíveis/química , Durapatita/química , Células-Tronco Mesenquimais/citologia , Nanopartículas/química , Titânio/química , Ligas/química , Sequência de Bases , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Adesão Celular , Técnicas de Cultura de Células/métodos , Diferenciação Celular , Proliferação de Células , Colágeno Tipo I/genética , Primers do DNA/genética , Expressão Gênica , Humanos , Teste de Materiais , Células-Tronco Mesenquimais/metabolismo , Microscopia Eletrônica de Varredura , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteonectina/genética , Osteopontina/genéticaRESUMO
OBJECTIVE: To evaluate a new Hyaluronan-based graft. MATERIAL AND METHODS: Hyaluronan-based grafts (HYAFF 11trade mark tube, diameter 2 mm, length 1.5 cm) were implanted in an end-to-end fashion in the abdominal aorta of 15 rats. Histology, immunohistochemistry and electron microscopy were used to evaluate the results at 7, 21, and 90 days. RESULTS: At day 7, new tissue was observed in the graft coming from both the proximal and distal ends of the aorta. The luminal surface of the regenerating tissue was covered by endothelial cells (CD34(+), VEGFR-2(+), vWF(+)). At day 21, regenerating tissue joined at the centre of the tube. The neo-vessel was formed by smooth muscle cells (Myosin Light Chain Kinase) as well as elastic, and collagen fibres. At day 90 a stable artery segment was formed and the biomaterial was almost completely degraded. Infiltration of neutrophils and lymphocytes was not observed. All animals survived the observation period and there were no signs of stenoses or aneurysms. CONCLUSION: The hyaluronan-based graft allowed complete regeneration of a newly formed vascular tube in which all the cellular and extracellular components are present and organized in a well defined architecture similar to native artery.
Assuntos
Implantes Absorvíveis , Prótese Vascular , Ácido Hialurônico , Engenharia Tecidual , Animais , Aorta Abdominal/cirurgia , Artérias/cirurgia , Materiais Biocompatíveis , Implante de Prótese Vascular , Células Cultivadas , Endotélio Vascular/citologia , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Tissue engineering is a promising approach to developing hepatic tissue suitable for the functional replacement of a failing liver. The aim of the present study was to investigate whether an extracellular cell matrix obtained from fibroblasts-cultured within scaffolds of hyaluronic acid (HYAFF) could influence the proliferation rate and survival of rat hepatocytes both during long-term culture and after in vivo transplantation. Cultures were evaluated by histological and morphological analysis, a proliferation assay and metabolic activity (albumin secretion). Hepatocytes cultured in extracellular matrix-enriched scaffolds exhibited a round cellular morphology and re-established cell-cell contacts, growing into aggregates of several cells along and/or among fibers in the fabric. Hepatocytes were able to secrete albumin up to 14 days in culture. In vivo results demonstrated the biocompatibility of HYAFF-11 implanted in nude mice, in which hepatocytes maintained small well-organised aggregates until the 35th day. In conclusion, the presence of a fibroblast-secreted extracellular matrix improved the biological properties of the hyaluronan scaffold, favoring the survival and morphological integrity of hepatocytes in vitro and in vivo.
Assuntos
Matriz Extracelular/metabolismo , Fibroblastos/metabolismo , Hepatócitos/citologia , Hepatócitos/transplante , Ácido Hialurônico/química , Fígado Artificial , Engenharia Tecidual/métodos , Animais , Materiais Biocompatíveis/química , Técnicas de Cultura de Células/métodos , Proliferação de Células , Sobrevivência Celular/fisiologia , Células Cultivadas , Hepatócitos/fisiologia , Humanos , Teste de Materiais , Camundongos , Camundongos SCID , Ratos , Ratos WistarRESUMO
With the aim to detect what kind of cells, in addition to erythroid progenitors, could be involved in the pathogenesis of B19 infection in some connective tissue diseases, primary cultures of human fibroblasts (HF) and endothelial cells (HUVEC) were exposed to a B19 positive serum (350 genome copies/cell). The presence of NS1 and VP1 mRNA, in both HF and HUVEC cultures 1, 2 and 6 days after the exposure, indicated infection by B19 virus. However, no significant increase of B19 DNA level in the infected HF and HUVEC cultures was detectable through the entire incubation period of 6 days. It is possible that HF and HUVEC are not permissive for B19 virus replication or, alternatively, that few cells only get infected by B19 virus. HF and HUVEC stimulation with different growth factors or cytokines could be required for a B19 productive infection to occur.
Assuntos
Células Endoteliais/virologia , Fibroblastos/virologia , Parvovirus B19 Humano/patogenicidade , Células Cultivadas , DNA Viral/análise , Humanos , Infecções por Parvoviridae/microbiologia , Parvovirus B19 Humano/genética , Parvovirus B19 Humano/isolamento & purificação , RNA Mensageiro/metabolismo , RNA Viral/metabolismo , Pele/citologia , Veias Umbilicais/citologia , Proteínas Virais/genética , Proteínas Virais/metabolismoRESUMO
PURPOSE: To quantify the presence of the proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) in allergic conjunctivitis. MATERIALS AND METHODS: Tears and peripheral blood samples were collected from patients with seasonal allergic conjunctivitis (SAC, n=6), vernal keratoconjunctivitis (VKC, n=12), and normal subjects (CT, n=12). From an additional six nonactive allergic patients, tears were collected before and after specific conjunctival allergen challenge (CAC). Upper tarsal conjunctival biopsies were obtained from five CT and five VKC patients. TNF-alpha in tears was measured by enzyme-linked immunoassay and identified in tissues by immunohistochemistry. RESULTS: Tear TNF-alpha levels in VKC patients were significantly increased compared to CT (p=0.03), and were significantly correlated with the severity of the disease. No differences were found between SAC and CT tear samples. TNF-alpha serum levels were higher in VKC than CT, however, this difference was not statistically significant. After CAC, tear TNF-alpha levels were found increased in only one of six patients. In VKC tissues, TNF-alpha positive cells were significantly increased compared to CT (p=0.03). CONCLUSIONS: TNF-alpha may have a significant role in severe forms of allergic conjunctivitis.
Assuntos
Conjuntivite Alérgica/sangue , Lágrimas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adolescente , Adulto , Alérgenos/efeitos adversos , Biomarcadores/análise , Criança , Conjuntivite Alérgica/etiologia , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Estações do AnoRESUMO
OBJECTIVE: To identify the CD44-receptor-mediated effects of 5-7 x 10(5)MW hyaluronan (HA, Hyalgan) on cell viability in normal and damaged human chondrocyte primary cultures isolated from articular cartilage. DESIGN: Primary cultures of human chondrocytes were established from normal articular biopsies and expanded to the second culture passage. The dose-response effects of HA on the viability of normal cultures were identified. Chondrocytes were then treated with either hypoxanthine (2 mM) and xanthine oxidase (20-60 mU), or with activated polymorphonuclear leukocytes (PMNs) to induce injury. Damaged and control cells were then treated with 5-7 x 10(5)HA in the previously identified optimal dose of 0.05 mg/ml. Viability was assessed at specific time periods for the chemically and PMN-damaged cells. To identify if HA effects were mediated by the CD44 receptor, chondrocytes were incubated with anti-CD44 antibody at saturating concentrations (5 microg/ml for 100,000 cells) to produce a maximum inhibition of HA binding. Cells were evaluated using the MTT viability assay, histology, electron microscopy and immunohistochemistry. RESULTS: Direct addition of HA (optimal dose, 0.5 mg/ml) significantly increased cell survival in normal chondrocyte primary cultures (P<0.05). Similarly, addition of this same dose of HA to cultures of free radical-damaged chondrocytes, restored the viability to baseline conditions. Cell viability rates dropped significantly (P<0.05) when CD44 receptor binding was inhibited, indicating that cell growth was mediated by the CD44 receptor. CONCLUSIONS: HA (0.5 mg/ml of 5-7 x 10(5)) significantly increased the viability of normal human chondrocytes in primary culture and restored cell viability to near normal levels after oxidative cell injury.
Assuntos
Condrócitos/efeitos dos fármacos , Receptores de Hialuronatos/metabolismo , Ácido Hialurônico/farmacologia , Cartilagem Articular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Humanos , Hipoxantina/farmacologia , Imuno-Histoquímica/métodos , Microscopia Eletrônica , Neutrófilos/fisiologia , Xantina Oxidase/farmacologiaRESUMO
Experiences coming from many cell-culture studies has brought about the concept that tissue and organ reconstruction should be performed in a three-dimensional environment as it normally occurs in vivo. As far as endothelial cell culture is concerned, it has been shown that angiogenesis can be successfully achieved only when cells are cultured in the presence of collagen-based matrices or basal membrane substrates. The aim of the present investigation is to demonstrate that human umbilical vein endothelial cells (HUVEC) can be grown and differentiated on an artificial dermis obtained by fibroblasts cultured on hyaluronic acid-based scaffolds. For this purpose, we have cultured HUVEC, retrieved by collagenase digestion of perfused human umbilical vein either alone and with fibroblast at 1/1 ratio into HYAFF-11 non-woven mesh. Cultures were maintained for up to 3 weeks. Samples were taken at different time points within this period for the MTT proliferation test and for immunohistochemical analysis. Our results demonstrate that hyaluronan-based biomaterials (HYAFF-11 NW mesh) represent a suitable substrate for HUVEC adhesion, proliferation and reorganization in microcapillary network.
Assuntos
Materiais Biocompatíveis , Endotélio Vascular/fisiologia , Fibronectinas/fisiologia , Pele Artificial , Veias Umbilicais/citologia , Adesão Celular/efeitos dos fármacos , Técnicas de Cultura de Células , Colágeno/análogos & derivados , Endotélio Vascular/citologia , Fibronectinas/biossíntese , Fibronectinas/farmacologia , Humanos , Queratinócitos/citologiaRESUMO
In this study we report a preliminary investigation of the feasibility of non-woven/sponge fabrics of a hyaluronan derived biomaterials (benzyl ester of HA (HYAFF-11 FAB, Abano Terme, Italy) for the in vitro culture of rat hepatocytes and rat beta cells. Cell growth on hyaluronan derived biomaterials were tested in the presence of complete medium and in the presence of ECM (extracellular matrix) secreted by fibroblasts previously cultured into the scaffold. Hepatocytes and beta cells were extracted from rat liver/pancreas and seeded either on the HYAFF-11 scaffold alone, or on HYAFF-11 scaffold containing ECM. Direct assay of cell proliferation was performed with MTT test. For morphological observations samples were stained with hematoxylin and eosin. The results obtained by MTT test showed that hepatocytes cultivated in both the above described conditions were able to proliferate up to 14 days and Langerhans islet up to 21 days. After this time, cells started to undergo apoptosis. The morphological analyses showed cell aggregation in three-dimensional structures promoted by the fibers of the biomaterial. Our results confirmed that HYAFF-11 meshes represent a suitable scaffold for hepatocyte adhesion/Langerhans islet organization and proliferation. In particular, the presence of a fibroblast secreted extracellular matrix improves the biological property of the scaffold.
RESUMO
PURPOSE: We evaluate the results of an elective cardiopulmonary bypass conceived to minimize the surgical risk related to its use with temporary circulatory arrest and deep hypothermia in the treatment of patients with renal tumor extending into the right atrium. MATERIALS AND METHODS: From July 1996 to December 2000, 19 patients with renal neoplasm and venous involvement were admitted to our department. Three patients 4, 57 and 58 years old with a right (2) and left (1) renal tumor extending into the right atrium underwent radical nephrectomy and tumor thrombus removal using a normothermic cardiopulmonary bypass. The bypass circuit was connected with a vacuum assisted venous drainage giving a negative pressure of 20 to 40 mm. Hg. Neither circulatory arrest nor hypothermia was used. Tumor thrombus was extracted through a longitudinal "cavotomy" and removed along with the kidney. RESULTS: Total cardiopulmonary bypass time was 14, 19 and 22 minutes, respectively. No intraoperative or postoperative complications due to surgical technique occurred. No significant bleeding was observed at the time of cavotomy and all neoplastic tissue was removed. Pathological examination documented renal cell carcinoma in 2 cases and Wilms tumor in 1. All the patients are alive 30, 42 and 15 months, respectively, after the operation. CONCLUSIONS: Normothermic cardiopulmonary bypass with vacuum assisted venous drainage makes circulatory arrest and hypothermia unnecessary and avoids the potential complications associated with these procedures. With respect to veno-venous shunts this technique guarantees complete surgical control of the thrombus and avoids the need for extensive dissection of the retrohepatic vena cava and Pringle maneuver.
Assuntos
Ponte Cardiopulmonar/métodos , Neoplasias Cardíacas/cirurgia , Neoplasias Renais/cirurgia , Pré-Escolar , Átrios do Coração , Neoplasias Cardíacas/patologia , Humanos , Neoplasias Renais/patologia , Pessoa de Meia-Idade , Invasividade Neoplásica , Células Neoplásicas Circulantes , Veia Cava Inferior/patologia , Veia Cava Inferior/cirurgiaRESUMO
Long-term maintenance of viability and expression of differentiated hepatocyte function is crucial for bioartificial liver support. We developed a new bioreactor design (ALEX), associated with a new extracellular autologous hepatocyte biomatrix (Porcine Autologous Biomatrix - PBM) support. To test this new bioreactor, we compared it to a standard BAL (BioArtificial Liver) cartridge in a ex vivo model using human plasma added to bilirubin, ammonium and lidocaine. A pathology study was performed on both bioreactors. The results suggest that ALEX allows a maximal contact between the perfusing plasma and the liver cells and a proper hepatocyte support by a cell-to-matrix attachment. ALEX is a suitable cell support bioreactor, guaranteeing long-term maintenance of the metabolic activity of hepatocytes when compared to a standard BAL cartridge.
Assuntos
Circulação Extracorpórea , Fígado Artificial , Amônia/sangue , Animais , Bilirrubina/sangue , Reatores Biológicos , Hepatócitos , Humanos , Lidocaína/sangue , Tempo de Protrombina , Suínos , Engenharia TecidualRESUMO
OBJECTIVE: Intraarticular injection of native hyaluronan (HA) or a cross-linked derivative are commonly utilized in the treatment of osteoarthritis. Unlike from native hyaluronan, the crosslinked HA derivative is a gel containing also other chemical entities. This study compares the local tolerability of these different preparations in normal rabbit knees, in order to provide further information on their biological effects. METHODS: Synovial fluids were aspirated after single or repeated weekly injections (up to three) of the therapeutic agents and cell count was determined in a Burker chamber and in an automatic cell counter. The percentage of the different cell types was determined by light microscopy in semithin sections of fixed synovial fluid cytocentrifugate. Fragments of synovial membrane were also morphologically analyzed. RESULTS: In the synovial membrane no signs of inflammation were evident either after a single or repeated injections of native Hyaluronan (Hyalgan or Artz). In addition, the cell recruitment and the percentage of cell types in the synovial fluid was not statistically different from saline treated joints. After 3 weekly injections of the crosslinked HA derivative (Hylan G-F20, Synvisc) about 50% of the treated joints appeared slightly inflamed and in these joints a statistically significantly higher cell content was determined in the synovial fluid compared to placebo and native Hyaluronan treatment. In addition an unexpectedly high percentage of eosinophils was found in the synovial fluid and in the synovial membrane of slightly inflamed joints treated with crosslinked HA. CONCLUSION: The data obtained after repeated intra-articular injections in normal rabbit knee joints confirm the safety profile of native Hyaluronan.