人脐带间充质干细胞与蚕丝素多孔支架的体外复合培养

目的观察蚕丝素多孔支架对人脐带间充质干细胞（human umbilical cord mesenchymal stem cells,hUCMSCs）吸附作用及支架对hUCMSCs形态、功能及活性的影响,为脂肪组织工程支架选择提供实验依据。方法将hUCMSCs制成细胞悬液接种在蚕丝素多孔支架,荧光倒置相差显微镜、扫描电镜和四甲基偶氮唑蓝（MTT）法观察hUCMSCs的吸附和生长情况。结果培养1～2d后可见hUCMSCs与蚕丝素多孔支架充分附着。培养5～7d细胞生长增殖十分活跃,10d左右时,蚕丝素多孔支架孔中hUCMSCs成片状融合。荧光倒置相差显微镜和扫描电镜见细胞与支架黏附良好并有大量基质分泌,且活性指标与正常培养的hUCMSCs比较差异无统计学意义（P〉0．05）。结论蚕丝素多孔支架对hUCMSCs具有良好的吸附作用,并能维持其正常形态、功能及活性,蚕丝素多孔支架是hUCMSCs三维立体培养时的良好天然支架。     

Composite mandibular allografts in canines

Objective:To evaluate the feasibility of transplanting composite mandibular allografts to repair large mandibular defects.Methods:Three composite mandibular transplantation models were established.The first model consisted of hemimandible with the attached teeth,muscle and skin,and oral mucosa.The second model was transplanted in the same way with the first one excluding oral mucosa and some teeth,and third one excluding the oral mucosa and all dental crowns.Fourteen transplanting operations were performed in canines.Cyclosporine A and methylprednisone were given for immunosuppression.Results:The composite mandibular organs had an effective and closed return circuit.Transplantation of vascularized allograft or mandibular compound organs was feasible.Two longest time survivors of 67d and 76d were in the third model group.Cyclosporine A was successful in suppressing rejection of transplanted composite allograft and prolonging survival time of transplantation models.Conclusions:The composite mandibular allografts were available with large block of living composite tissue,and helpful in restoration of appearance and function for severe mandibular defects.     

Evaluation of an antibacterial peptide-loaded amniotic membrane/silk fibroin electrospun nanofiber in wound healing

Antimicrobial peptides (AMPs) are among the compounds that have significant potential to deal with infectious skin wounds. Using wound dressings or skin scaffolds containing AMPs can be an effective way to overcome infections caused by antibiotic-resistant strains. In this study, we developed an amniotic membrane-based skin scaffold using silk fibroin to improve mechanical properties and CM11 peptide as an antimicrobial peptide. The peptide was coated on the scaffold using the soaking method. The fabricated scaffold was characterised by SEM and FTIR, and their mechanical strength, biodegradation, peptide release, and cell cytotoxicity analyses were performed. Then, their antimicrobial activity was measured against antibiotic-resistant strains of Pseudomonas aeruginosa and Staphylococcus aureus. The in vivo biocompatibility of this scaffold was evaluated by subcutaneously implanting it under the skin of the mouse and counting lymphocytes and macrophages in the implanted area. Finally, the regenerative ability of the scaffold was analyzed in the mouse full-thickness wound model by measuring the wound diameter, H&E staining, and examining the expression rate of genes involved in the wound healing process. The developed scaffolds exerted an inhibiting effect on the bacteria growth, indicating their proper antimicrobial property. In vivo biocompatibility results showed no significant count of macrophages and lymphocytes between the test and control groups. The wound closure rate was significantly higher in the wound covered with fibroin electrospun-amniotic membrane loaded with 32 μg/mL CM11, where the relative expression rates of collagen I, collagen III, TGF-β1 and TGF-β3 were higher compared with the other groups.     

丝素蛋白/羟基磷灰石组织工程化骨修复兔桡骨节段性骨缺损

目的 探讨丝素蛋白/羟基磷灰石(SF/HA)组织工程化骨的成骨作用,以期为临床治疗骨缺损提供新的人工骨材料.方法 将SF/HA与成骨诱导的兔骨髓基质干细胞(BMSCs)复合,构建组织工程化骨.取54只兔于左侧桡骨中上段制备15 mm节段性骨缺损.实验分3组(A、B组各24只,C组6只):A组:植入SF/HA组织工程化骨,B组:单纯植入SF/HA;C组:骨缺损区不植入任何材料.于术后4、8、12及16周摄X线片,并于16周行螺旋CT扫描重建,观察骨缺损修复及骨塑形情况,参照Lane-Sandhu X线评分标准对各组骨缺损的骨修复程度评分.骨痂标本行 HE染色组织学观察,按照Lane-Sandhu组织学评分法比较12周和16周时各组的骨修复情况. 结果 术后16周,X线片示A组髓腔通畅,新骨塑形好,骨皮质连续;B组缺损区有缩小,两断端不连接;C组缺损区无明显骨痂生长.16周时螺旋CT扫描重建显示:A组骨塑形明显,骨缺损完全修复;B组有部分皮质骨形成,缺损区不能完全修复;C组骨缺损基本无修复.每组术后4、8、12、16周不同时间点的放射学评分差异均有统计学意义(P＜0.05).术后12、16周时3组间Lane-Sandhu组织学评分差异均有统计学意义(P＜0.05). 结论 SF/HA组织工程化骨具有良好的节段性骨缺损修复能力,但SF/HA本身缺乏骨诱导作用,单独修复节段性骨缺损作用有限.     

人脐带间充质干细胞与蚕丝蛋白支架构建组织工程脂肪的研究

目的：将体外以人脐带间充质干细胞（hUCMSCs）与蚕丝蛋白支架初步构建的组织工程脂肪移植到大鼠体内,观察其演变过程。方法：hUCMSCs与蚕丝蛋白支架复合培养10天后,进行成脂诱导;6周后将其移植到Wi st ar大鼠后肢肌肉内,同时,以同体积支架材料作为对照;分别于移植后4周和8周取材,行油红O染色、HE染色以及扫描电镜观察。结果：hUCMSCs与蚕丝蛋白支架复合培养及成脂诱导6周后,见大量成脂样细胞生成,并与支架牢固粘附。移植4周,移植物体积略小,质稍硬,表面有透明薄膜形成,膜中分布新生血管网;油红O染色见支架内新生脂肪组织及细胞呈橙红色;HE染色显示支架网眼内有新生脂肪组织,并可见少量炎性细胞浸润;扫描电镜见支架网眼内有球形、表面光滑的脂肪细胞。移植8周,移植物体积进一步缩小,质变软,表面薄膜内血管网丰富;油红O染色见支架中着橙红色组织较前明显增多,部分呈片状融合;HE染色显示新生脂肪明显增多,仍有少量炎性细胞浸润;扫描电镜显示脂肪细胞较前增生明显。对照组同样可见炎性细胞浸润,未见新生脂肪组织生成,支架材料8周时较4周时降解更加明显。结论：随着时间推移,蚕丝蛋白支架网眼内脂肪细胞逐渐增多,支架材料在体内呈现逐步降解趋势,说明体内环境有利于组织工程化脂肪的进一步形成。同时,也提示支架材料在组织相容性方面尚存不足。     

Improving the properties of fish skin collagen/silk fibroin dressing by chemical treatment for corneal wound healing

Natural biomaterials are crucial in ocular tissue engineering because they allow cells to proliferate, differentiate, and stratify while maintaining the typical epithelial phenotype. In this study, membranes as dressings were formed from silk fibroin and collagen (Co) extracted from fish skin and then modified with carbodiimide chemical cross linker in different concentrations. The samples were evaluated by different analyses such as structural, physical (optical, swelling, denaturation temperature, degradation), mechanical, and biological (viability, cell adhesion, immunocytochemistry) assays. The results showed that all membranes have excellent transparency, especially with higher silk fibroin content. Increasing the cross linker concentration and the ratio of silk fibroin to Co increased the denaturation temperature and mechanical strength and, conversely, reduced the degradation rate and cell adhesion. The samples did not show a significant difference in toxicity with increasing cross linker and silk fibroin ratio. In general, samples with a low silk fibroin ratio combined with cross linker can provide desirable properties as a membrane for corneal wound healing.     

蚕丝丝素蛋白在骨科领域的应用进展

骨缺损、软骨损伤以及肌腱和韧带损伤等由于移植物来源短缺或者移植后存在多种副反应,治疗上非常困难,成为骨科领域亟待攻克的难题。目前的研究主要集中在可降解支架材料方面（包括人工合成材料和天然材料）,这些材料均存在自身缺点：如人工合成材料力学     

新型丝素蛋白多孔支架复合兔髓核细胞体外构建组织工程化髓核的实验研究

[目的]观察以新型丝素蛋白多孔支架复合兔髓核细胞体外构建组织工程化髓核的可行性。[方法]分离培养兔髓核细胞,与丝素蛋白多孔支架在体外复合培养,建立组织工程化髓核模型,通过扫描电镜、HE染色、甲苯胺蓝染色、Ⅱ型胶原免疫组化以及酶联免疫吸附测定观察细胞在支架上1周和3周的生长及增殖情况。[结果]培养1周后,扫描电镜显示细胞呈球状均匀地贴附在支架内部。细胞-支架复合体HE染色可见支架内部有大量髓核细胞,甲苯胺兰染色阳性,Ⅱ型胶原免疫组化染色阳性。培养3周后,扫描电镜显示细胞成层黏附于支架表面,细胞重叠生长,分泌大量细胞外基质,HE染色可见支架内部有大量髓核细胞填满支架孔隙并分泌大量细胞外基质,甲苯胺兰染色阳性,Ⅱ型胶原免疫组化染色阳性。酶联免疫吸附测定:3周组蛋白多糖含量(52.4±4.5)ng/ml明显高于1周组(29.3±3.6)ng/ml,P<0.05,3周组的II型胶原含量(24.3±1.8)ng/ml明显高于1周组(15.16±1.5)ng/ml,P<0.05。[结论]新型丝素蛋白多孔支架复合兔髓核细胞体外培养生长良好,分泌大量类似髓核样细胞外基质,可以用于体外构建组织工程化髓核。     

丝素蛋白应用于椎间盘纤维环组织工程的研究进展

[目的]综述国内外有关丝素蛋白应用于椎间盘纤维环组织工程的研究进展,为下一步的实验研究提供理论依据。[方法]广泛查阅近年来国内外有关丝素蛋白的研究成果,尤其是应用于椎间盘纤维环组织工程的相关文献,归纳总结其发展现状和研究进展。[结果]目前丝素蛋白提纯工艺已臻成熟,并且在多个领域得到应用,比如纤维环、软骨、肌腱、韧带组织工程和药物缓释材料等方面。以丝素蛋白为材料制成的支架,结构稳定,力学强度高,还具有良好的生物相容性和可控的生物降解速率,并且能够通过简单的化学修饰来改进各方面的性能。[结论]丝素蛋白作为椎间盘纤维环组织工程支架的理想材料,表现出了良好的发展前景,值得进一步研究和推广。     

A rib-containing free flap to reconstruct mandibular defects

A composite free flap based on intercostal vessels may be used to reconstruct in one stage defects around the mandible in which skin, soft tissue and bone are missing. A delay procedure is recommended one week prior to transfer.     

Novel fabrication of antibiotic containing multifunctional silk fibroin injectable hydrogel dressing to enhance bactericidal action and wound healing efficiency on burn wound: In vitro and in vivo evaluations

The development of biologically active multifunctional hydrogel wound dressings can assist effectively to wound regeneration and also has influenced multiple functions on wound injury. Herein, we designed a carbon‐based composited injectable silk fibroin hydrogel as multifunctional wound dressing to provide effective anti‐bacterial, cell compatibility and in vivo wound closure actions. Importantly, the fabricated injectable hydrogel exhibit sustained drug delivery properties, anti‐oxidant and self‐healing abilities, which confirm that composition of hydrogel is highly beneficial to tissue adhesions and burn wound regeneration ability. Frequently, designed injectable hydrogel can be injected into deep and irregular burn wound sites and would provide rapid self‐healing and protection from infection environment with thoroughly filled wound area. Meanwhile, incorporated carbon nanofillers improve injectable hydrogel strength and also offer high fluid uptake to hydrogel when applied on the wound sites. In vitro MTT cytotoxicity assay on human fibroblast cell lines establish outstanding cytocompatibility of the injectable hydrogel and also have capability to support cell growth and proliferations. In vivo burn wound animal model results demonstrate that the hydrogel dressings predominantly influenced enhanced wound contraction and also promoted greater collagen deposition, granulation tissue thickness and vascularization. This investigation''s outcome could open a new pathway to fabricate multifunctional biopolymeric hydrogel for quicker burn wound therapy and effectively prevents microenvironment bacterial infections.     

Reconstruction of mandibular defects in irradiated patients

In this prospective study, mandibular reconstruction using titanium plates was evaluated in 31 patients treated between July 1988 and January 1990. Sixteen patients had prior surgery; 13 had prior radiotherapy. In 11 patients, prior radiation and surgery had failed. Sixteen patients received postoperative radiotherapy either in standard or accelerated fractions. Twelve patients had complications of either intraoral (8), extraoral (5), or combined (1) plate exposure or fistula formation (2). Factors significantly related to complications were poor nutrition, accelerated radiation, and recurrence. Sixty-one percent of all patients healed uneventfully. When patients with complications secondary to recurrence who subsequently died were excluded, the success rate was 73%. Only one patient had an unacceptable result that produced a cosmetic and functional deformity despite secondary repair.     

Effects of laser-modified versus osteopromotively coated titanium membranes on bone healing: a pilot study in rat mandibular defects

Regeneration of alveolar bone with membrane techniques has become an integral part of implant dentistry. The aim of the present study was to determine if laser-modified titanium membranes are of value in the regeneration of so-called critical size defects in the rat model compared with titanium membranes that were coated with growth factors. A total of 24 rats were included in the study. Critical size defects were created bilaterally and covered by titanium membranes coated with (1) polylactide, (2) polylactide and clindamycin, (3) polylactide and growth factors, (4) polylactide, clindamycin and growth factors and (5) uncoated but laser-modified titanium membranes. All 18 control defects were covered by titanium membranes without any substrate. Four weeks after treatment the animals were killed. Laser-modified titanium membranes (group 5) showed new bone formation in many areas. Nevertheless, complete bridging was found only in one specimen. In contrast, in groups 3 and 4, most defects showed almost complete bridging of the defects. In particular, clindamycin had no inhibitory effect on bone healing. Furthermore, after 28 days, there was no significant difference between the individual groups (including controls) with respect to the total amount of lamellar bone. Growth-factor-coated membranes can significantly accelerate the healing process of bony defects in the rat mandibular model. Nevertheless, it is not possible to accelerate bone healing with laser-irradiated membranes or to enhance the quality of bone within the time period examined.     

Tissue-engineered buccal mucosa using silk fibroin matrices for urethral reconstruction in a canine model

Background

To investigate the feasibility of urethral reconstruction using tissue-engineered buccal mucosa (TEBM) with silk fibroin (SF) matrices in a canine model.

Materials and methods

Autologous oral keratinocytes and autologous fibroblasts were isolated, expanded, and seeded onto SF matrices to obtain TEBM. The TEBM was assessed using hematoxylin and eosin staining and scanning electron microscopy. A 5-cm urethral mucosal defect was created in 10 female canines. Urethroplasty was performed using TEBM in five canines in the experimental group and with SF matrices without cells in the five canines in the comparison group. Retrograde urethrography was performed after 6 mo of grafting. The urethral grafts were analyzed grossly and histologically.

Results

The oral keratinocytes and fibroblasts exhibited good biocompatibility with the SF matrices. TEBM could be constructed using SF matrices. The canines implanted with the tissue-engineered mucosa voided without difficulty. The retrograde urethrography revealed no sign of stricture. The histologic staining showed that epithelial cells developed gradually and exhibited stratified epithelial layers at 6 mo. In the comparison group, the canines had difficulty voiding, and the retrograde urethrography showed urethra stricture. The histologic staining showed that one to two layers of epithelial cells developed.

Conclusions

The TEBM using SF matrices could be a potential material for urethra reconstruction.