RESUMEN
The collagen fibres of rabbit and human ureter were exposed by digestion with trypsin and hyaluronidase. The fibre structure was examined using an SEM and examples of the inner and outer fibre structures are shown together with the effects of different types of mechanical strain. An interesting difference between the arrangements of the inner fibres of human and rabbit was seen where the human ureter had a cross-ply structure while in the rabbit it was helical.
Asunto(s)
Colágeno/ultraestructura , Uréter/ultraestructura , Animales , Colágeno/química , Colágeno/fisiología , Humanos , Hialuronoglucosaminidasa , Microscopía Electrónica de Rastreo , Conejos , Especificidad de la Especie , Estrés Mecánico , Tripsina , Uréter/química , Uréter/fisiologíaRESUMEN
Collagen was dehydrothermally treated (heat cured) by heating dry under vacuum at 60, 80, 100 and 120 degrees C. The change in stability was determined by subjecting to measurement of gross crosslinking, content of lysino-alanine and naturally occurring collagen crosslinks, shrinkage temperature (TM), susceptibility to digestion by lysosomal thiol proteases, and susceptibility to pepsin and trypsin. Morphological changes were examined by electron microscopy. The in vivo biodegradation of dehydrothermally treated collagen sponges was investigated using a rat lumbar muscle implantation model for up to 28 days. For all heat-cured collagens, the data strongly indicated that both crosslinking and denaturation/degradation was present in increasing quantities with increasing temperature of treatment, its level was too low (maximum 179 pmol mg-1) to account for the decreased solubility and increased molecular weight gross changes observed. Increasing resistance of treated collagen to both lysosomal cathepsins and pepsin correlated well with increased crosslinking and increasing temperature of the heat-curing process. However, increased denaturation/degradation of the collagen at higher temperatures was revealed by electrophoretic analysis, trypsin hydrolysis data and by electron microscopy. Differential scanning calorimetry (d.s.c.) correlated well with these results showing an increased level of denaturation in heated samples. The in vivo study showed little difference between control and heat-cured samples except for the material treated at 120 degrees C which was biodegraded in vivo at a significantly faster rate. The data shows, therefore, that crosslinking induced by the dehydrothermal treatment of collagen decreases its rate of proteolysis at acid pH in vitro. However, the simultaneous denaturation/degradation of the protein during the heat-cure process appears to be a more important factor in determining the fate of the material implanted into rat muscle.
Asunto(s)
Colágeno/análogos & derivados , Animales , Biodegradación Ambiental , Colágeno/metabolismo , Colágeno/ultraestructura , Endopeptidasas/metabolismo , Reacción a Cuerpo Extraño , Liofilización , Calor , Lisosomas/enzimología , Microscopía Electrónica , Músculos/metabolismo , Desnaturalización Proteica , Ratas , Solubilidad , Tapones Quirúrgicos de GazaRESUMEN
No ideal dural grafting material is currently available. Many materials have been evaluated in this role, and for many neurosurgeons cadaveric human lyophilized dura has been popular. Recently this material has been putatively associated with Creutzfeldt-Jakob disease. In this study we compared three degradable materials, collagen vicryl (Bovine collagen coated vicryl mesh), Zenoderm (Porcine dermis) and Lyodura (Lyophilized human cadaveric dura) as dural substitutes. In an experimental model using the New Zealand White Rabbit the materials were implanted into dural defects of dimensions 1.7 cm by 1 cm. In the control group the dura was not repaired. In total 47 animals were used and sacrificed at time intervals of 2, 4, 8, 12, and 24 weeks. On gross examination the collagen vicryl produced few adhesions to the cerebral cortex and was replaced by a neomembrane which showed good union with the host dura. In the control group no new layer was formed and there were severe cortical adhesions. Zenoderm and Lyodura remained undegraded and produced more adherence to the cerebral cortex than the collagen vicryl implant. The histological examination showed collagen vicryl to support ingrowth of fibroblasts and the production of a new collagen layer which by 3 months resembled the original host dura. The inflammatory response to the implant did not persist after 3 months. The other substitutes were revitalized by host cells but remained undegraded at 6 months with ingrowth of woven bone and persistence of inflammatory and foreign body response. The results show collagen vicryl to be a suitable dural substitute with potential advantages over other currently used degradable materials, and should be evaluated more fully, both in laboratory studies and clinically.
Asunto(s)
Duramadre/cirugía , Poliglactina 910 , Prótesis e Implantes , Mallas Quirúrgicas , Animales , Corteza Cerebral/patología , Colágeno , Duramadre/patología , Reacción a Cuerpo Extraño/patología , Masculino , Conejos , Adherencias TisularesRESUMEN
A series of chemically modified collagens were subjected to proteolysis by lysozomal cathepsins, pepsin and trypsin. Modifications of the collagens included acetylation, succinylation, methylation and borohydride reduction. Changes in the integrity of the materials were also monitored by differential scanning calorimetry (DSC). All modified collagens were implanted intramuscularly to assess their relative biodegradation rates in vivo. Methylation of the collagen showed extensive denaturation as confirmed by DSC, pepsin solublization to small fragments and by increased susceptibility to trypsin. However, methylation and succinylation made little difference to hydrolysis by cathepsins. Acetylation and borohydride reduction gave increased resistance to cathepsins as well as to pepsin, this latter also being found with the succinylated substrate. In-vivo implantation data showed both succinylation and methylation increased the rate of biodegradation but that the other modifications did not affect the rate of breakdown when compared with control unmodified collagen. The results of this study showed that chemical modification of collagen can alter in vivo degradation rates and could aid in designing collagen-based prostheses.
Asunto(s)
Colágeno/metabolismo , Acetilación , Animales , Borohidruros/farmacología , Rastreo Diferencial de Calorimetría , Catepsinas/metabolismo , Colágeno/análogos & derivados , Concentración de Iones de Hidrógeno , Metilación , Músculos/metabolismo , Oxidación-Reducción , Pepsina A/metabolismo , Desnaturalización Proteica , Ratas , Succinatos/metabolismo , Ácido Succínico , Tripsina/metabolismoRESUMEN
Following preliminary in vitro and in vivo experiments a new collagen Vicryl mesh has been devised. The membrane has been extensively tested in the laboratory and has been found to resist the passage of urine. It holds sutures well and has been shown on an experimental animal to be biodegradable. The membrane has now been applied in human subjects. It has been particularly efficacious in renal surgery and as a means of sealing a bladder after it has been opened. With respect to the ureter the sealing effect is excellent, to the extent of having produced a urinoma. The membrane has been found to be easily applied in vesicovaginal fistulae but difficulties have been experienced in the presence of infection. It is also an ideal membrane for repair of a urethrovaginal fistula--an area where natural tissues are less easily available. This is the first recorded use of this new biodegradable membrane in the human subject.
Asunto(s)
Membranas Artificiales , Mallas Quirúrgicas , Enfermedades Urológicas/cirugía , Biodegradación Ambiental , Femenino , Humanos , Enfermedades Renales/cirugía , Nefrectomía , Adhesivos Tisulares , Enfermedades de la Vejiga Urinaria/cirugía , Fístula Vesicovaginal/cirugíaRESUMEN
This experimental study assessed the use of a collagen-coated vicryl mesh tube to reconstruct the esophagus in growing piglets. Initial experiments, in which a segment of thoracic esophagus was excised and replaced by this prosthetic tube, resulted in all the animals succumbing to mediastinitis within the first 3-4 days. This was shown, at post-mortem, to be due to leakage of the prosthesis, secondary to acid reflux, resulting in dissolution of the prosthesis. The collagen-coated vicryl mesh was thereafter treated with glutaraldehyde and in-vitro studies showed that the glutaraldehyde-treated material exhibited a higher resistance to 0.01 M HCl at pH 2.0, when compared with untreated material. In addition, the glutaraldehyde-treated material also showed an increased resistance to digestion by bacterial collagenase. Further experiments to reconstruct the cervical esophagus in growing piglets were performed using the revised glutaraldehyde cross-linked collagen coated vicryl mesh tube. All the animals survived the procedure and the prosthesis were leakproof immediately post-operatively. The animals, however, developed severe stenosis at a mean of 11 days post-operatively. Attempts at neo-epithelialization of the graft were seen histologically. There was considerable granulation tissue and scar tissue formation on the mediastinal aspect of the graft. It is suggested that collagen coated vicryl mesh tube may find applications in the treatment of esophageal atresia only if the problem of stenosis of the prosthesis can be solved.
Asunto(s)
Colágeno/uso terapéutico , Esofagoplastia/métodos , Poliglactina 910 , Prótesis e Implantes , Mallas Quirúrgicas , Anastomosis Quirúrgica , Animales , Atresia Esofágica/cirugía , Complicaciones Posoperatorias , PorcinosRESUMEN
This study demonstrates how the mechanical strength of a series of collagen/composite gels can be measured using a penetrometer. It was found that the presence of fibrin in collagen gels resulted in increased gel strength. Similarly hyaluronic acid was found to increase the strength of collagen gels. Addition of heparin weakened collagen gels as did chondroitin-6-sulphate. Neutrophil migration into collagen gels was found to be inversely proportional to gel strength. Fibrin and hyaluronic acid containing gels inhibited neutrophil migration while the presence of heparin and chondroitin sulphate increased neutrophil migration. BHK gel contraction experiments demonstrated how the presence of fibrin prevents gel contraction. Despite increasing gel strength the presence of hyaluronic acid appeared to have no effect on BHK contraction of collagen gels. Similarly the presence of heparin or chondroitin sulphate had no effect on gel contraction by BHK cells.
Asunto(s)
Fibroblastos/citología , Neutrófilos/citología , Animales , Línea Celular , Movimiento Celular , Células Cultivadas , Sulfatos de Condroitina , Colágeno , Cricetinae , Fibrina , Geles , Ácido Hialurónico , Riñón , Mesocricetus , Estrés MecánicoRESUMEN
Native collagen, acetylated collagen, collagen/10% chondroitin sulphate, collagen/2.5% hyaluronic acid and collagen/20% hyaluronic acid were implanted both as film and as sponge into rat lumbar muscle for 7 and 14 d. After 7 d implantation, all materials elicited an acute inflammatory cell response characterized by numerous polymorphs and histocytes. The cell population after 14 d was principally mononuclear, i.e. leucocytes, neutrophils, macrophages, lymphocytes and fibroblasts. Both films and sponges followed a similar pattern. Native collagen elicited a subacute inflammatory response after 7 d. However, 14 d after implantation, a marked infiltration by neutrophils was apparent with subsequent degradation of existing collagen material. Acetylated collagen film evoked a much greater inflammatory cell response than native collagen. Both collagen/hyaluronic acid composites elicited a similar response. The collagen/10% chondroitin sulphate composite elicited the least inflammatory cell response at 7 d, whereas infiltration by host fibroblasts after 14 d implantation was clearly seen.
Asunto(s)
Materiales Biocompatibles , Colágeno , Glicosaminoglicanos , Polímeros , Tapones Quirúrgicos de Gaza , Acetilación , Animales , Materiales Biocompatibles/efectos adversos , Colágeno/metabolismo , Colágeno/toxicidad , Glicosaminoglicanos/metabolismo , Glicosaminoglicanos/toxicidad , Ácido Hialurónico/análisis , Inflamación/etiología , Ensayo de Materiales , Colagenasa Microbiana/metabolismo , Músculos/cirugía , Polímeros/metabolismo , Polímeros/toxicidad , Ratas , Tapones Quirúrgicos de Gaza/efectos adversosRESUMEN
The attachment and growth of an established cell line derived from mouse fibroblasts on collagen, chemically modified collagen, and collagen composite surfaces were compared. Tissue culture polystyrene dishes provided a suitable control. The substrates included native bovine dermal collagen, succinylated, acetylated and methylated collagen, and a series of composite materials formed from collagen and the glycosaminoglycans hyaluronic acid, chondroitin 4-sulphate and chondroitin 6-sulphate and the glycoprotein fibronectin. Attachment and growth of cells on each of these substrates were assessed by visual inspection under optical microscopy, by detachment of the cells using trypsinization and subsequent counting in a Coulter counter; and by 3H-thymidine incorporation studies. A very good correlation between the results was obtained by the three methods employed which showed that collagen, in comparison to polystyrene, is a relatively poor substrate for cellular attachment, growth and proliferation, but it may be improved by chemical modification and by incorporation of either fibronectin, chondroitin sulphate (5 and 10%), or low levels (less than 5%) of hyaluronic acid into the collagen matrix. Concentrations in excess of 5% hyaluronic acid into the collagen matrix, however, appeared to inhibit cellular attachment and growth and such materials provided a poorer substrate than native collagen.
Asunto(s)
Colágeno/análogos & derivados , Fibroblastos/citología , Animales , Adhesión Celular/fisiología , Recuento de Células , División Celular/fisiología , Línea Celular , TimidinaRESUMEN
Collagen films were prepared by three different methods involving acid homogenization at pH 3.0 of a collagen suspension for 1.5 min (film A) and 15 min (film B) and alkaline homogenization at pH 10 for 15 min (film C), after which the resulting slurries were degassed and dried over sterile air. Subsequent examination by scanning electron microscope (SEM) revealed that all samples showed a distinctly layered structure which was much finer in the acid films. Implantation into the lumbar muscle of rats followed by histology and SEM studies revealed that film B was completely resorbed at 14 d whereas remnants of film A could still be seen at this period. The slowest rate of resorption was observed with film C, traces of which could still be found at 70 d. Invading inflammatory cells moved into the collagen films between the layers from the edges only causing the whole structure to 'ribbon out'. The surfaces of all three films appeared to be impenetrable to cells. Incubation of the three films with bacterial collagenase revealed similar relative rates of degradation to those observed in vivo.
Asunto(s)
Colágeno/metabolismo , Músculos/cirugía , Prótesis e Implantes , Absorción , Animales , Biodegradación Ambiental , Bioprótesis , Hidrólisis , Técnicas In Vitro , Colagenasa Microbiana/metabolismo , Microscopía Electrónica de Rastreo , Músculos/patología , RatasRESUMEN
A novel procedure, the adhesive film test, is proposed as a screening method to predict potential cytotoxicity of biomaterials. This in vitro test utilizes a sterile strip of acrylate-based medical adhesive as an anchorage substrate in cytotoxicity studies. The adhesive film allows direct fixation of test samples to the base of the petri dish, ensuring close contact between sample and cells. The test is based on the principle that toxic components present in the test material will readily leach out into the culture medium and adversely affect the local cell population. The main advantage of the adhesive film test is that a viable cell population can be added directly to the test plate and after an incubation period of 24 h, the cellular response can be recorded as either cytotoxic or cytocompatible. Microscopic examination can be followed by quantifying the results using a micrometer to measure cellular attachment areas, migration distances and zones of inhibition. In addition, the adhesive film used to attach the test samples is shown to support extensive fibroblast growth and attachment to its surface and hence can also function as a negative nontoxic control in cytotoxicity studies.
Asunto(s)
Materiales Biocompatibles/efectos adversos , Ensayo de Materiales/métodos , Adhesivos , Animales , División Celular/efectos de los fármacos , Células Cultivadas , Fibroblastos/fisiología , Ensayo de Materiales/instrumentación , RatonesRESUMEN
The regeneration of smooth muscle appears to take place within the fibrous tissue characteristically found when a biodegradable collagen/Vicryl prosthesis is used to repair full thickness defects in the rabbit urinary bladder. The question of whether the central smooth muscle was the result of myoblastic differentiation in the fibrous tissue or arose from healthy pre-existing detrusor muscle was resolved by serial sectioning and specific staining. Only in situ transmutation, or differentiation, explains the morphology, and the results therefore strongly suggest that this central smooth muscle regenerated from within the repair area.
Asunto(s)
Músculo Liso/fisiología , Regeneración , Vejiga Urinaria/fisiología , Animales , Diferenciación Celular , Músculo Liso/citología , Músculo Liso/cirugía , Conejos , Vejiga Urinaria/citología , Vejiga Urinaria/cirugíaRESUMEN
The evolution of a collagen/vicryl composite membrane designed as a prosthetic material for use in urinary tract surgery is described. The early experiments in which collagen film alone was used to repair experimental ureterotomies are reviewed together with our first experiments with the collagen/vicryl prosthesis in the repair of partial nephrectomies and of full thickness defects created in the urinary bladder of experimental rabbits. These early results led to the preparation of a composite using a more highly purified collagen and employing a method of sterilisation (gamma irradiation) which would be acceptable for regular use in medical products. The results of a further series of partial nephrectomy and full thickness bladder repairs show that irradiation does not compromise the efficacy of the collagen/vicryl composite in vivo.
Asunto(s)
Colágeno , Membranas Artificiales , Poliglactina 910 , Polímeros , Sistema Urinario/cirugía , Animales , Biodegradación Ambiental , Nefrectomía , Prótesis e Implantes , Conejos , Vejiga Urinaria/patología , Vejiga Urinaria/cirugía , Cicatrización de HeridasRESUMEN
A Collagen/Vicryl (Polyglactin) composite membrane (developed for use in urinary tract surgery) has been incubated in cultures of radioactively labelled urinary tract pathogens vis Escherichia coli, Staphylococcus epidermidis, and Proteus mirabilis for up to 1 h. For comparison, collagen film, Vicryl mesh, and a number of absorbable and non-absorbable sutures were similarly tested. Following incubation, samples were also examined by scanning electron microscopy. Under the experimental conditions employed, only minimal adherence of the micro-organisms to the collagen coated Vicryl mesh, its two individual components, as well as Vicryl and nylon sutures was observed. Significantly greater numbers of bacteria, however, adhered to silk and Chromic Catgut. The results suggest that adherence of microorganisms to the prosthesis even in infected urine is unlikely to develop into a microcolony of bacteria. However, it should be emphasised that great care must be exercised when extrapolating from the in-vitro to the in-vivo situation.
Asunto(s)
Adhesión Bacteriana , Materiales Biocompatibles , Poliglactina 910 , Polímeros , Sistema Urinario/cirugía , Escherichia coli , Humanos , Técnicas In Vitro , Ensayo de Materiales , Proteus mirabilis , Staphylococcus epidermidis , SuturasRESUMEN
Collagen coated vicryl mesh has been incubated with a series of urine collections from healthy and stone-forming patients. For comparison, collagen film, vicryl mesh, and a number of absorbable and non-absorbable sutures were similarly tested. Incubation in rabbit urine were also included in the study. Deposition of urinary salts, estimated qualitatively by electron microscopy, were observed on the collagen vicryl composite in approximately two thirds of the urines tested including rabbit urine. Those urines from patients with a high calcium excretion in particular caused urinary deposits on the material. Similar results were obtained with collagen film although the latter was not tested in rabbit urine. Considerably less deposits of urinary constituents were found with other absorbable materials such as vicryl mesh, vicryl sutures, and chromic catgut, whereas a higher proportion of concretions were found with the non-absorbable sutures. The results indicate that urinary salt deposition may be a problem associated with collagen based composite materials after prolonged exposure to urine. It must, however, be emphasised that great care should be exercised when extrapolating any results obtained in-vitro to the in-vivo situation.
Asunto(s)
Proteínas de Insectos , Poliglactina 910 , Polímeros , Suturas , Sistema Urinario/cirugía , Orina , Animales , Calcio/orina , Catgut , Humanos , Ensayo de Materiales , Microscopía Electrónica de Rastreo , Fosfatos/orina , Proteínas , Conejos , Seda , Ácido Úrico/orinaRESUMEN
Glutaraldehyde stabilizes pericardial tissue in prosthetic heart valves by forming covalent cross-links between collagen molecules. If the cross-linking is nonuniform owing to variable penetration of the glutaraldehyde, areas of the tissue may become sites for enzymatic attack or become potentially antigenic. Cross-linking can be easily assessed by measuring the shrinkage (thermal denaturation) temperature of the tissue. We used differential scanning calorimetry to perform a stratigraphic analysis of the shrinkage temperature of glutaraldehyde-treated pericardium. In the fixation conditions used (0.25 per cent glutaraldehyde for 28, seven and two days and for 2 hours), no variation was found in the shrinkage temperature measurement throughout the thickness of the tissue. This indicates uniform penetration of fixative and cross-linking throughout the tissue.
Asunto(s)
Aldehídos/farmacología , Glutaral/farmacología , Pericardio/fisiología , Animales , Rastreo Diferencial de Calorimetría/métodos , Bovinos , Técnicas In Vitro , Cinética , Pericardio/citología , Pericardio/efectos de los fármacos , TemperaturaRESUMEN
Collagen/vicryl (Polyglactin) composite membrane has been used to repair full-thickness defects in the urinary bladder of rabbits. The material has been shown to be biodegradable, prevent leakage of urine, and is readily replaced by collagenous scar tissue lined with a urothelium. Regeneration of smooth muscle has been observed in the repair area of some animals. The results suggest that such a material may well be of use to urologists wishing to augment contracted bladders or in the repair of bladder fistulae in human subjects, thereby avoiding the use of bowel or other material e.g. omentum.
Asunto(s)
Colágeno , Membranas Artificiales , Poliglactina 910 , Polímeros , Vejiga Urinaria/cirugía , Animales , Músculo Liso/fisiología , Prótesis e Implantes , Conejos , Regeneración , Sistema Urinario/cirugíaRESUMEN
A composite membrane produced from collagen and vicryl mesh has been used to cover the kidney surface following partial nephrectomy in rabbits. The membrane readily held sutures, gave satisfactory haemostasis, and prevented leakage of urine. The experiments showed that the prosthesis biodegraded in less than twenty weeks. The only observed long-term effect of the material was thickening of the renal capsule. The results indicate that this membrane may be a suitable reparative material for use in traumatised kidneys in humans.
Asunto(s)
Colágeno , Membranas Artificiales , Nefrectomía , Poliglactina 910 , Polímeros , Animales , Biodegradación Ambiental , Riñón/cirugía , Prótesis e Implantes , Conejos , Sistema Urinario/cirugíaRESUMEN
A Vicryl (Polyglactin)/Collagen composite membrane has been developed for potential use in urinary tract surgery. The compatability of this membrane, together with its two individual components, collagen film and vicryl mesh, has been tested over a three week period in urine obtained from both healthy and from stone forming patients. The rate of degradation as indicated by changes in the mechanical strength was determined at regular intervals. In addition, absorbable suture materials such as plain catgut, chromic catgut, and vicryl were similarly tested; in the latter case, the breaking stress, and actual loss of weight of the material were compared. The whole series of experiments were then repeated in urine obtained from rabbits, the animal chosen for any future in vivo studies.
Asunto(s)
Colágeno , Poliglactina 910 , Polímeros , Suturas , Orina , Animales , Catgut , Humanos , Concentración de Iones de Hidrógeno , Hidrólisis , Conejos , Resistencia a la Tracción , Sistema Urinario/cirugíaRESUMEN
This investigation assessed a biodegradable collagen membrane which can be sutured around the ureter to prevent urine leakage, thus permitting healing to proceed more rapidly while the membrane itself is resorbed. Following an early in vitro investigation in which collagen was assessed, a more comprehensive survey has now been carried out. Tissue compatibility and biodegradation were assessed by implanting the film into the lumbar muscles of rats; it was then used to cover experimental ureterotomies in New Zealand White rabbits. The data obtained from the rabbits confirmed that a collagen membrane can prevent leakage of urine from the ureter during healing while it itself is biodegraded, indicating that a collagen membrane could be used to repair the injured urinary tract.