Uso de membrana de colágeno y placa de protección mecánica como instrumento de conservación alveolar / Use of collagen membrane and mechanical protective plate as an instrument for alveolar preservation

Luego de una extracción dental ocurren inevitablemente procesos de reabsorción y remodelación ósea, donde la dimensión y morfología de la cresta alveolar se ve modificada, representando un problema para la rehabilitación de la zona. Estudios clínicos han documentado un promedio de 4,0 a 4,5 mm de reabsorción ósea horizontal luego de una exodoncia, como así también cambios dimensionales significativos en los alrededores del hueso alveolar. El propósito fue evaluar y comparar clínica y tomográficamente los procesos de reparación y conservación del reborde alveolar post-extracción de paredes intactas, con y sin la utilización de esponjas de colágeno intraalveolar como relleno y placa termoformada como barrera física, durante el proceso de cicatrización. Se seleccionaron pacientes con indicación de exodoncia de elementos dentarios normalmente implantados y clínicamente aceptables, aplicando los criterios de exclusión, se realiza toma de impresión del terreno para la confección de una placa de protección rígida 0,8 termoformada para ser colocada posterior a la extracción durante la masticación por un período de 30 días. Se indica tomografía cone-beam post operatoria inmediata y a los tres meses para analizar, medir y comparar alto y ancho de crestas alveolares residuales. En la evaluación clínica y tomográfica de los casos estudiados, el grupo control donde se usó únicamente placa de protección alveolar arrojó mejores resultados que el grupo donde se colocaron esponja de colágeno en el interior del alvéolo. Palabras clave: Regeneración ósea, esponja de colágeno, cicatrización ósea, alvéolo postextracción, placa de protección alveolar (AU)

After a tooth extraction, bone resorption and remodeling processes inevitably occurs, where size and morphology of the alveolar crest is modified, representing a problem for the rehabilitation of the area. Clinical studies have documented an average of 4.0 to 4.5 mm of horizontal bone resorption after an extraction, us well us substantial dimensional changes around the alveolar bone. The purpose was to evaluate and compare clinical and tomographically both repair and preservation of post extraction alveolar ridge of intact walls processes, with and without the use of intraalveolar collagen sponges as filler and a thermoformed protective plaque, us physica? barrier, during healing process. Patients with normally implanted and clinically acceptable tooth with extraction indication were selected, applying the exclusion criteria, impression of the field is taken to build a 0.8 rigid thermoformed protective plaque in order to be placed after extraction and used during chewing for a period of 30 days. Immediate and three months post-operative cone beam tomography are indicated to analyze, measure and compare height and width of residual alveolar crests. In the clinical and tomographic evaluation of the cases treated, control group where only alveolar protective plaque was used, showed better results than the group with intraalveolar collagen sponge (AU)

Stimulatory Effects of KPR-A148 on Osteoblast Differentiation and Bone Regeneration

BACKGROUND: Xanthine derivatives have been used to treat a variety of medical conditions including respiratory disease and neural degeneration. However, few studies have reported their effects on bone regeneration. Therefore, we investigated the effects of KPR-A148, a synthetic xanthine derivative on osteoblast differentiation in vitro and bone regeneration in vivo. METHODS: The cytotoxicity of KPR-A148 was evaluated using MC3T3-E1 cells by the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltertrazolium bromide assay. The effects of KPR-A148 on osteoblast differentiation were examined by alkaline phosphatase staining, Alizarin red S staining, and real-time PCR of osteoblast differentiation marker genes. To investigate the effects of KPR-A148 on in vivo bone regeneration, a KPR-A148-containing collagen sponge was implanted into a mouse calvarial defect and KPR-A148 was injected twice, weekly. Bone regeneration was evaluated quantitatively by micro-CT and qualitatively by hematoxylin and eosin, as well as Masson's Trichrome staining. RESULTS: KPR-A148 did not show toxicity in the MC3T3-E1 cells and promoted osteoblast differentiation in a concentration-dependent manner. 10 µM of KPR-A148 showed the most significant effect on alkaline phospatase staining and matrix mineralization. KPR-A148 increased the expression of osteoblast marker genes in both the early and late stages of differentiation. In addition, KPR-A148 significantly induced new bone formation in the calvarial defect model. CONCLUSION: These results demonstrate that KPR-A148 strongly induces osteoblast differentiation and new bone formation. Therefore, it could be used as a potential therapeutic agent for regenerating bone following its destruction by disease or trauma.

An experimental study of simvastatin-collagen composite sponge as a direct pulp capping material in rat molars / 华西口腔医学杂志

OBJECTIVE@#To assess the feasibility and efficacy of simvastatin-collagen composite sponge as a novel, direct pulp capping material.@*METHODS@#A total of 120 Sprague-Dawley rats were randomly divided into three groups: the simvastatin-collagen composite sponge group (SIM group), the collagen sponge group (CS group), and the Ca(OH)2 group (CH group). An endodontic entry cavity was prepared on the occlusion of the first molar on the left maxillary of each rat. The contralateral teeth were utilized as the normal control group. The rats were experimented after 1, 3, 7, 14, and 28 days. X-ray observations were conducted and the specimens underwent hematoxylin-eosin (HE) and Masson's Thichrome staining. Dentin bridge formations and pulpal biology reactions were evaluated histopathologically.@*RESULTS@#X-ray results: high-density images could be observed on the pulp exposure sites in the CH group on the 28th day. In the SIM group, high-density images could be observed after 14 and 28 days, whereas in the CS group, high-density images were not observable in the exposed area. HE and Masson's Thichrome staining results: different degrees of inflammation under the cavity were detected in the three groups at different time points. The inflammatory reaction of the CS group was the most serious. The degree of the inflammatory reaction varied significantly between the SIM and the CS groups on the 14th and 28th days (P<0.01). The inflammatory reaction in the SIM group was lighter than in the CH group. There was a statistical difference between the SIM and the CH groups on the 14th day (P<0.05). During the observation period, the SIM group induced the best and fastest formation of reparative dentin. As for dentin bridge formation, a significantly higher complete bridge rate was observed in the SIM group than in the CH and in the CS groups on the 14th day (P<0.05) and for the SIM and the CH groups compared with the CS group on the 28th day (P<0.05).@*CONCLUSIONS@#The simvastatin-collagen composite sponge exhibited satisfactory biocompatibility with the pulp tissue and promoted the formation of reparative dentin. The application of simvastatin-collagen composite sponge as a pulp-capping material has satisfactory potential.

Skin stretching device combined with collagen sponge for wound repair / 中华创伤骨科杂志

Objective To evaluate the feasibility and clinical efficacy of our self-designed simple skin stretching device combined with collagen sponge for management of severe soft tissue wounds.Methods From September 2015 to October 2017,a consecutive series of 43 patients whose soft tissue wounds could not be closed primarily were enrolled for a therapy using a simple skin stretching device made of round osseous pins and wire combined with collagen sponge.They were 27 males and 16 females,with a mean age of 31.5 years (from 5 to 56 years).There were 18 fresh wounds and 25 old ones.Their skin defects ranged from 5.5 cm × 3.0 cm to 18.0 cm × 7.5 cm.After debridement and vacuum sealing drainage,2 round osseous pins with a diameter of 2.0 mm or 2.5 mm were driven through the dermis about 1 to 2 cm from both edges of the wound,in parallel with the longitudinal axis of the wound.After the parts of 2 pins exposed outside the skin were bent,they were fixed respectively with a fine wire with 2 twisted strands.The wounds were continuously stitched with eversion suture.The wires and sutures were gradually tightened to contract the wounds until the skin color changed and capillary filling reaction started.Then medical collagen sponge was used to cover the wounds.Next,the wires and sutures were tightened continuously until the wound edges were pulled together.Details of this therapy and its complications were recorded.Follow-up visits were paid until wound healing.Results Of the 43 cases,the wounds were directly closed immediately after primary stretching procedure in 8,closed after skin stretching for 4 to 12 days (average,7.5 days) in 30,and significantly reduced in 5 which were cured following skin graft.Eventually,40 cases were followed up for an average of 6.8 months (from 3 to 18 months) and 3 were lost.Aesthetic reoperation was performed in 3 patients who were inflicted with postoperative scar formation after skin graft.Linear healing of the wound edges was achieved in 37 patients without complications like skin necrosis,pathological hyperplasia scar,skin sensation deletion or wound infection,leading to fine appearance and functional recovery.Conclusion Our self-designed simple skin stretching device combined with collagen sponge provides a cost-effective and practical technique for clinical treatment of soft tissue defects,with an advantage of reducing or even avoiding secondary repair with skin graft or skin flap.

Biomacromolecules as carriers in drug delivery and tissue engineering

Natural biomacromolecules have attracted increased attention as carriers in biomedicine in recent years because of their inherent biochemical and biophysical properties including renewability, nontoxicity, biocompatibility, biodegradability, long blood circulation time and targeting ability. Recent advances in our understanding of the biological functions of natural-origin biomacromolecules and the progress in the study of biological drug carriers indicate that such carriers may have advantages over synthetic material-based carriers in terms of half-life, stability, safety and ease of manufacture. In this review, we give a brief introduction to the biochemical properties of the widely used biomacromolecule-based carriers such as albumin, lipoproteins and polysaccharides. Then examples from the clinic and in recent laboratory development are summarized. Finally the current challenges and future prospects of present biological carriers are discussed.

Effects of salvia miltiorrhiza polyphenols for injection and collagen sponge in diabetic foot / 中国生化药物杂志

Objective To investigate the effect of polyphenol of salvia miltiorrhiza polyphenols combined with collagen sponge in the treatment of diabetic foot.Methods A total of 115 patients with type 2 diabetic foot were randomly divided into treatment group (55 cases) and control group (60 cases).Salvia miltiorrhiza polyphenols and collagen sponge treatment group were treated with conventional treatment plus type 2 diabetes treatment of salvia miltiorrhiza polyphenol combined with collagen sponge;4 weeks as a treatment course, observation of 8 weeks, ulcer and other adverse reactions were recorded and compared.Results After 4 weeks of treatment, 17 cases (30.91%) with 30% or more reduction in foot ulcer area in treatment group,33 cases (60.00%) after 6 weeks treatment and the number of cases with 30% or more reduction in foot ulcer area,49 cases (89.09%) after 8 weeks of treatment, and the number of cases and the proportion of cases with 30% or more decrease of ulcer area in the control group after 4, 6 and 8 weeks were 12(20.00%), 22 (36.67%) and 39 (65.00%).There was no statistical difference between the two groups for the healing time of foot ulcer at three time points.After 8 weeks of treatment, the cure rate was 38.18% in salvia miltiorrhiza polyphenol combined with collagen sponge group. The effective rate was 50.91% and 45.00% for the two groups, the difference was not statistically significant; while the total effective rate was also significantly higher(89.09% vs.65.00%,P<0.01).Conclusion Salvia miltiorrhiza polyphenol combined with collagen sponge has a good effect on diabetic foot treatment, especially it can improve the cure rate, which is superior to the routine therapy.

Effect of collagen sponge and fibrin glue on bone repair

ABSTRACT The ability of hemostatic agents to promote bone repair has been investigated using in vitro and in vivo models but, up to now, the results are inconclusive. Objective In this context, the aim of this study was to compare the potential of bone repair of collagen sponge with fibrin glue in a rat calvarial defect model. Material and Methods Defects of 5 mm in diameter were created in rat calvariae and treated with either collagen sponge or fibrin glue; untreated defects were used as control. At 4 and 8 weeks, histological analysis and micro-CT-based histomorphometry were carried out and data were compared by two-way ANOVA followed by Student-Newman-Keuls test when appropriated (p≤0.05). Results Three-dimensional reconstructions showed increased bone formation in defects treated with either collagen sponge or fibrin glue compared with untreated defects, which was confirmed by the histological analysis. Morphometric parameters indicated the progression of bone formation from 4 to 8 weeks. Additionally, fibrin glue displayed slightly higher bone formation rate when compared with collagen sponge. Conclusion Our results have shown the benefits of using collagen sponge and fibrin glue to promote new bone formation in rat calvarial bone defects, the latter being discreetly more advantageous.

Establishment of a rabbit model of lumbar laminectomy and bone grains replantation / 中国比较医学杂志

Objective To establish a rabbit model of lumbar laminectomy and bone grains replantation and provide experimental evidence for the clinical application.Methods Eighteen healthy male New Zealand rabbits were selected and randomly divided into two groups:the control group (n=6) and experimental group (n=12).The rabbits of control group were given general anesthesia, and taking the L5 spinous process as the center to perfom left L5 laminectomy, using a micro lancet forceps to slowly bite the lamina and ligamentum flavum for fenestration and exposed to an approximately 0.8 cm x 0.3 cm sized bone window and then sutured the skin.The rabbits of experimental group were exposed to an approximately 0.8 x 0.3 cm sized bone window as well, and bone fragments were cut into small grains.Then the small bone grains were embedded in medical collagen sponge, to form an arch shape, and replanted them to the site of epidural fenestration.CT scan and histological changes were observed at 4, 8 and 12 weeks after operation.Results At 8 weeks after operation, CT examination showed that in the experimental group, a thin bone plate was formed by the bone grains.At 12 weeks after operation, the bone plate became thicker and was connected with the vertebral bone, and with continuous bone trabeculae.The spinal canal and volume were not obviously changed, and no spinal cord compression was observed. The rabbits of control group showed segment lamina defects, a small scar protruding into the spinal canal, and the vertebral canal was not completely reconstructed. Conclusions The bone grains replantation can effectively promote bone reconstruction in the laminectomized rabbits, and the formed bone plate can prevent epidural scar from intruding into the spinal canal, and can reduce the postlaminectomy adhesion.

Complication rates in patients using absorbable collagen sponges in third molar extraction sockets: a retrospective study

OBJECTIVES: The purpose of this study is to retrospectively evaluate the postoperative complication rates for absorbable type-I collagen sponge (Ateloplug; Bioland) use in third molar extraction. MATERIALS AND METHODS: From January to August 2013, 2,697 total patients undergoing third molar extraction and type-I collagen sponge application in the Department of Oral and Maxillofacial Surgery at Yonsei University Dental Hospital (1,163 patients) and Dong-A University Hospital (1,534 patients) were evaluated in a retrospective study using their operation and medical records. RESULTS: A total of 3,869 third molars in 2,697 patients were extracted and the extraction sockets packed with type-I collagen sponges to prevent postoperative complications. As a result, the overall complication rate was 4.52%, with 3.00% experiencing surgical site infection (SSI), 1.14% showing alveolar osteitis, and 0.39% experiencing hematoma. Of the total number of complications, SSI accounted for more than a half at 66.29%. CONCLUSION: Compared to previous studies, this study showed a relatively low incidence of complications. The use of type-I collagen sponges is recommended for the prevention of complications after third molar extraction.

Evaluation of bFGF collagen composite sponge promoting vascular ingrowth in orbital implantation by 99Tcm-MDP scan / 中华实验眼科杂志

Background Intraorbital implantation of coralline porous hydroxyapatite (CHA) is a favorable cosmetic method after enucleation.However,the low degree of vascularizatiou in implant results in implant infection and exposure.Studies showed that a collagen composite sponge treated by basic fibroblast growth factor (bFGF/ collagen composite sponge) can promote angiogenesis.However,whether bFGF/collagen composite sponge improves the vascularization of CHA implants is unclear.Objective This study was to investigate the accelerating effect of bFGF collagen composite sponge on vascularization of orbital implant made of CHA using 99Tcm-methylene diphosphate (MDP) scan.Methods Forty-five New Zealand rabbits were randomly assigned to 3 groups.Evisceration of eyeball was performed on the left eyes of rabbits,and naked CHA,collagen composite sponge wrapped CHA and bFGF/collagen composite sponge wrapped CHA were implanted into the orbit respectively in 3 groups.99Tcm-MDP of 3 mCi was injected in the rabbits via ear vein in 2,4,6,8 and 12 weeks,and the vascular enhancement intensity on implants was observed 3 hours after injection.The ratio of average radioactive count from the area of interest with the same size between the left eyes and the right eyes was calculated.The implants were extracted for histopathological examination in the 12 weeks.Results As the lapse of postoperative time,the inflammation response gradually disappeared and no exposure of implants was seen during the 12-week duration.A similar vascular development strength was found in the area of interest among the 3 groups 2 weeks after surgery.However,the vascular development was significantly enhanced in the left eyes compared the right eyes from 4 to 6 weeks,with the highest intensity in the 8th week in the naked CHA group and collagen composite sponge wrapped CHA group.In the bFGF/ collagen composite sponge wrapped CHA group,the strongest image was in the 6th week after operation.The ratios of average radioactive count between the left eyes and the right eyes were significantly higher in the bFGF/collagen somposite sponge wrapped CHA group compared with the naked CHA group and collagen composite sponge wrapped CHA group (all at P＜0.05),and ratios of average radioactive count of the collagen composite sponge wrapped CHA group was significantly higher than that of the naked CHA group (all at P＜0.05).New blood vessels ingrowed toward the center of the implants through the coralline porous under the optical microscope.Conclusions Both bFGF (20 μg)/collagen composite sponge and collagen composite sponge can accelerate the ingrowth of vessel in the CHA,but the promoting effect of bFGF collagen composite sponge is prominent.

Comparison of the bone healing capacity of autogenous bone, demineralized freeze dried bone allograft, and collagen sponge in repairing rabbit cranial defects

OBJECTIVES: This study sought to evaluate the efficacy of collagen graft materials, as compared to other graft materials, for use in healing calvarial defects in rabbits. MATERIALS AND METHODS: Ten mm diameter calvarial defects were made in ten rabbits. The rabbits were then divided into 4 groups: control, autogenous bone graft, SureOss graft, and Teruplug graft. Bone regeneration was evaluated using histological and radiographic methods. RESULTS: Based on visual examination, no distinct healing profile was observed. At 4 weeks after treatment, histological analysis showed there was no bone regeneration in the control group; however, at 8 weeks after treatment, new bone formation was observed around the margin of the defective sites. In the autogenous bone graft group, new bone formation was observed at 4 weeks after treatment and mature bone was detected around the grafted bone after 8 weeks. In the SureOss graft group, at 4 weeks after treatment, acute inflammatory and multinuclear cells were noted around the grafted materials; at 8 weeks after treatment, a decrease in graft materials coupled with new bone formation were observed at the defective sites. In the Teruplug graft group, new bone formation was detected surrounding the bone margin and without signs of inflammation. There were statistically significant differences observed between the graft and control group in terms of bone density as evidenced by radiographic analysis using computed tomography (P<0.05), particularly for the autogenous bone graft group (P<0.001). CONCLUSION: These results suggested that autogenous bone, SureOss and Teruplug have the ability to induce bone regeneration as compared to an untreated control group. The osteogenic potential of Teruplug was observed to be lower than that of autogenous bone, but similar to that of SureOss.

The effects of low frequency electromagnetic fields on the proliferation of epidermal stem cells cultured on two different scaffolds / 中华物理医学与康复杂志

Objective To observe the effects of low frequency electromagnetic fields (LFEMFs) on the proliferation of human epidermal stem cells (hESCs) cultured in a three dimensional environment so as to provide an experimental basis for applying LFEMF in skin tissue engineering.Methods hESCs from human prepuces were isolated and purified by the method of rapid adherence to collagen type ⅣV. They were grafted into a type-I collagen sponge or chitosan scaffold in vitro, and then stimulated with different frequencies of LFEMF ( 1 Hz, 10 Hz or 50 Hz) at a magnetic field intensity of 5 mT for 30 min/d. The cells' growth and proliferation were tracked using hematoxylin and eosin (HE) and diamine pheny1 indole (DAPI) staining and observed under the scanning electron microscope at different time points ( on 2nd, 7th, 10th and 14th days of LFEMF intervention). The amounts of cell proliferation at every time point were analyzed and compared.Results LFEMFs of different frequencies showed significantly different efficacy in promoting hESC proliferation. The two scaffolds also showed significantly different effects.By the 10th day, hESCs had grown significantly better on collagen sponge scaffolds than on the chitosan ones. All LFEMF frequencies could promote proliferation of hESCs, but the differences in their effects were statistically significant.Conclusion Collagen sponge may be a preferable scaffold for hESCs cultured in vitro. Rapid proliferation of ESCs in three-dimensional settings can be promoted by LFEMF intervention. LFEMF has relatively great potential in skin tissue engineering.

Clinical observation of collagen sponge for reducing volume of drainage after surgery for lumbar spinal stenosis / 中国医师进修杂志

ObjectiveTo observe the effectiveness of collagen sponge for reducing volume of drainage after surgery for lumbar spinal stenosis. MethodsOne hundred and eighty-six patients who suffered from lumbar spinal stenosis were divided into two groups by random digits table method. The test group(96 cases) used collagen sponge to cover dura mater before placing drainage tube,the control group (90 cases) was treated without collagen sponge. The volume of drainage at 1,12,24 h after surgery were observed, the blood routine test was carried out at before and 48 h after surgery and the volume and ratio of blood transfusion after surgery was also measured and compared between the two groups. ResultsThe volume of drainage were significantly decreased in the test group compared with the control group at 1,12,24h after surgery [( 106.11 ± 20.02 ) ml vs. ( 127.02 ± 25.09) ml, (236.12 ± 34.06) ml vs. (327.31 ± 51.21 )ml, (355.16 ± 49.03 ) ml vs.( 506.36 ± 85.29 ) ml](P ＜ 0.05 ). The volume and the ratio of blood transfusion in the test group were ( 176.27 ± 21.37) ml and 10.42％(10/96) ,which were greatly lower than those in the control group[(445.94 ±24.56) ml and 32.22％ (29/90)](P ＜0.05). The number of RBC and the concentration of Hb were (2.96 ± 0.45 ) × 1012/L and ( 106.75 ± 7.30) g/L, differently in the test group at 48 h after surgery,which were increased significantly compared with the control group[(2.35 + 0.57) × 1012/Land (90.45 ± 5.10) g/L](P ＜ 0.05 ). ConclusionsCollagen sponge provides rapid ,effective and durable hemostasis and decreases the leak of cerebrospinal fluid after surgery for lumbar spinal stenosis. It can be used as an effective and economic method to reduce the volume of drainage after surgery.

Effect of rhBMP-2 produced by Escherichia coli expression system on bone formation in rat calvarial defects / 대한치주과학회지

PURPOSE: Recombinant human bone morphogenetic protein-2(rhBMP-2) has been evaluated as potential candidates for periodontal and bone regenerative therapy. In spite of good prospects in BMP applications, there is economically unavailable for clinical use in dental area. The purpose of this study was to evaluate the osteogenic effect of rhBMP-2 produced by E.coli expression system. MATERIALS AND METHODS: Eight-mm critical-size calvarial defects were created in 48 male Sprague-Dawley rats. The animals were divided into 6 groups of 8 animals each. Each group received one of the following: Negative control(sham-surgery control), positive control(absorbable collagen sponge(ACS) alone) and experimental(ACS loaded with rhBMP-2). Defects were evaluated by histologic and histometric parameters following 2- and 8-week healing intervals. RESULTS: The experimental group showed significant defect closure at 2 and 8weeks than the sham surgery and positive control groups. Moreover, the experimental group showed significantly greater new bone and augmented area than the other groups at both 2 and 8weeks. CONCLUSION: rhBMP-2 produced by E.coli expression system may be effective for bone regeneration.

Repair of Osteochondral Defect Using Chitosan-collagen Sponge intheRabbit's Knee / 대한정형외과학회잡지

PURPOSE: To determine the suitability of using a chatoyant-collagen sponge as a scaffold for transplanting a chondrocyte into a full-thickness articular cartilage defect. MATERIALS AND METHODS: The in vitro characterization of a chatoyant-collagen sponge infiltrated with the chondrocyte was combined with an in vivo assessment of the early articular cartilage repair in a rabbit's knee by H&E and MTT staining. These porous chatoyant-collagen sponges were implanted into the osteochondral defects made in the left patellofemoral grooves of 12 rabbits. The osteochondral defects were untreated in the right side and used as controls. The experimental animals were sacrificed 1, 3, 6 and 12 weeks after implantation and the repaired tissue was evaluated by a gross and histological evaluation using the Wakitani score. RESULTS: More primary cells cultured from the articular cartilage of the rabbit's knee were found to attach to and survive within a porous chatoyant-collagen sponge than with a chatoyant sponge. In gross and histological examination, the experimental group showed indications of repair, which appeared similar in color and texture to the surrounding articular cartilage. The Wakitani scoring in the experimental group at 6 (Ave. 10.7) and 12 (Ave. 7.3) weeks were superior to those in the control group at 6 (Ave. 8.7) and 12 (Ave. 3.7) weeks (6 wk: p=0.03, 12 wk: p=0.02). CONCLUSION: Scaffolds composed of porous a chatoyant-collagen sponge enhance the growth of cartilaginous repair and make a milieu for the survival of chondrogenic cells both in vitro and in vivo.

Osteogenesis of Recombinant BMP-7 Adenovirus-transduced Human Dermal Fibroblasts in Collagen Sponge / 대한해부학회지

As a preceding study to apply recombinant BMP-7 gene to the human, we investigated bone formation in immunodeficient mice by using tissue engineering and gene transplatation. Human dermal fibroblasts were transduced with AdBMP-7 and cultured with type I collagen solution to form collagen sponge. The collagen sponge containing AdBMP-7 transduced fibroblasts was transplanted into hypodermis of the mice and osteogenesis in the spongy was investigated by histochemical, electronmicroscopic, and radiologic methods at 1, 2, 4, 6, and 8 weeks. At one week after transplantation, there were fluent cells infiltration around the collagen sponge and capsular structure was formed with fibers arranged in concentric circles. New vessel formation was observed in the capsule and subcapsular area of the sponge, but there were nucleus condensation and obscure cell boundary in the cells of the central region. Lacuna containing eosinophilic structures were observed in the capsular structure at two weeks. This structures were enlarged with time and were confirmed to be bone tissue by showing positive reaction for Von Kossa stain. Cartilaginous structure was not observed in light microscopic level, but a few chondroblasts were observed in pericapsular area in electron microscopic observation. After 6 weeks, radiopaque shadows were observed at the region of transplantation. Cortical bone was formed in periphery of the sponge while marrow like structure was observed in central region; some trabecula bone, adipocytes, and well developed vessels. The percentage of bone formation in transplanted sponge at 1, 2, 4, 6, and 8 weeks were 0, 63, 88, 100, and 100% (n = 8), respectively. From these results, bone formation by BMP-7 transduced human dermal fibroblasts using collagen sponge scaffolds in immunodeficient mouse shows another potential way of human gene transplantation using recombinant BMP-7 adenovirus.

Bone Formation by rhBMP-7 Transduced HEK 293 Cells in Nude Mouse / 영남의대학술지

To induce bone formation at ectopic site by tissue engineering and gene therapy, we transplanted collagen sponges containing rhBMP-7 transduced HEK 293 cells in the hypodermis of nude mice. Bone formation was investigated by histological and electron microscopic method at 3, 6, and 9 weeks after transplantation. At 9 weeks after transplantation, eosinophilic bony tissue was observed in the implanted collagen sponge and was confirmed as bone tissue by Von Kossa stain. In the transmission electron microscopic observation, the cells in newly formed bone tissue had eccentrically located nucleus and well developed rough endoplasmic reticulum (rER). Therefore, the cells were evaluated as osteoblasts. Those results suggest that it is possible to form a bone tissue in the ectopic site by transplantation of rhBMP-7 transduced HEK 293 cells. This will be contributed to push more advanced gene therapy for bone formation. However, the HEK 293 cell is unable to apply to the clinical gene therapy. Therefore it is worth to find more compatible cells for clinical application. In addition, collagen sponge is considered as an excellent scaffold and/or carrier for gene therapy and a good biomaterial for tissue engineering.

Bone Formation by rhBMP-7 Transduced HEK 293 Cells in Nude Mouse / 영남의대학술지

To induce bone formation at ectopic site by tissue engineering and gene therapy, we transplanted collagen sponges containing rhBMP-7 transduced HEK 293 cells in the hypodermis of nude mice. Bone formation was investigated by histological and electron microscopic method at 3, 6, and 9 weeks after transplantation. At 9 weeks after transplantation, eosinophilic bony tissue was observed in the implanted collagen sponge and was confirmed as bone tissue by Von Kossa stain. In the transmission electron microscopic observation, the cells in newly formed bone tissue had eccentrically located nucleus and well developed rough endoplasmic reticulum (rER). Therefore, the cells were evaluated as osteoblasts. Those results suggest that it is possible to form a bone tissue in the ectopic site by transplantation of rhBMP-7 transduced HEK 293 cells. This will be contributed to push more advanced gene therapy for bone formation. However, the HEK 293 cell is unable to apply to the clinical gene therapy. Therefore it is worth to find more compatible cells for clinical application. In addition, collagen sponge is considered as an excellent scaffold and/or carrier for gene therapy and a good biomaterial for tissue engineering.

The effect of chitosan/ACS on bone regeneration in rat calvarial defects / 대한치주과학회지

The ultimate objective of periodontal treatment is to get rid of an on-going periodontal disease and further regenerate the supporting tissue, which is already destroyed, functionally. Currently, the bone grafting operation using various kinds of bone grafting materials and the operation for induced regeneration of periodontal tissue using the blocking membrane are performed for regeneration of the destroyed periodontal tissue. However, there are respective limitations Galenical preparations, which are used for regeneration of periodontal tissue, has less risk of rejective reaction or toxicity that may be incidental to degradation and their effect is sustainable. Thus, in case they are applicable to a clinic, they can be used economically. Chitosan has such compatibility, biological actions including antibacterial activity, acceleration of wound treatment, etc., and excellent mechanical characteristics, which has recently aroused more interest in it. Also, it has been reported that it promotes osteogenesis directly or indirectly by functioning as a matrix to promote migration and differentiation of a specific precussor cell (for example, osteoblast) and further inhibiting the function of such a cell as fibroblast to prevent osteogenesis. In this study, the pure chitosan solution, which was obtained by purifying chitosan, was used. However, since this chitosan is of a liquiform, it is difficult to sustain it in a defective region. It is, therefore, essential to use a carrier for delivering chitosan to, and sustaining it gradually in the defective region. In the calvarial defect model of the Sprague-Dawley rat, it is relatively easy to maintain a space. Therefore, in this study, the chitosan solution with which ACS was wetted was grafted onto the defective region. For an experimental model, a calvarial defect of rat was selected, and a critical size of the defective region was a circular defect with a diameter of 8 mm. A group in which no treatment was conducted for the calvarial defect was set as a negative control group. Another group in which treatment was conducted with ACS only was set as a positive control group (ACS group). And another group in which treatment was conducted by grafting the pure chitosan solution onto the defective region through ACS which was wetted with the chitosan solution was set as an experimental group (Chitosan/ACS group). Chitosan was applied to the Sprague-Dawley rat's calvarial bone by applying ACS which was wetted with the chitosan solution, and each Sprague-Dawley rat was sacrificed respectively 2 weeks and 8 weeks after the operation for such application. Then, the treatment results were compared and observed histologically and histometrically. Thereby, the following conclusions were obtained. 1. In the experimental group, a pattern was shown that from 2 weeks after the operation, vascular proliferation proceeded and osteogenesis proceeded through osteoblast infiltration, and at 8 week after the operation, ACS was almost absorbed, the amount of osteogenesis was increased and many osteoid tissue layers were observed. 2. At 2 weeks after the operation, each amount of osteogenesis appeared to be 8.70.8 %, 13.62.3 % and 4.80.7 % respectively in the experimental group, the positive control group and the negative control group. Accordingly, it appeared to be higher in the Experimental group and the positive control group than in the negative control group, but there was no significant difference statistically (p<0.01). 3. At 8 weeks after the operation, each amount of osteogenesis appeared to be 62.26.1 %, 17.42.5 % and 8.21.4 % respectively in the experimental group, the positive control group and the negative control group. Accordingly, it appeared to be substantially higher in the experimental group than in the positive control group and the negative control group, and there was a significant difference statistically (p<0.01). As a result of conducting the experiment, when ACS was used as a carrier for chitosan, chitosan showed effective osteogenesis in the perforated defective region of the Sprague-Dawley rat's calvarial bone.

Attachment and Growth of Cultured Fibroblasts on Chitosan Sponge

Closure of large skin wounds with split-thickness skin graft requires extensive harvesting of autologous skin. The limitation of available donor areas has sought various kinds of skin equivalents for coverage of skin defects. Concept of living skin equivalent using fibroblast and keratinocyte is the most promising one. For seeking ideal artificial skin, we conducted a research of new dermal alternative using chitosan. Fibroblast scattered on chitosan and chitosan-collagen sponge polymers were cultured for 3 weeks and chitosan and chitosan-collagen sponge polymers were grafted on the back of 250 gm Sprague-Dawley rat. There was statistically significant difference in fibroblast attachment as well as fibroblast proliferation between chitosan and chitosan-collagen sponge. Four weeks after grafting, the grafted area was examined. In the area of chitosan sponge graft, there was no clinical sign of inflammation. However, mild inflammatory infiltration and a few multinucleated giant cells were observed. In contrast, chitosan-collagen sponge grafted area showed no foreign body reaction clinically and histologically. In conclusion, concomitant use of chitosan and collagen resulted in better fibroblast attachment and proliferation and minimal immunologic reaction than chitosan sponge.