Factors influencing Frey syndrome after parotidectomy with acellular dermal matrix

BACKGROUND Frey syndrome,also known as ototemporal nerve syndrome or gustatory sweating syndrome,is one of the most common complications of parotid gland surgery.This condition is characterized by abnormal sensations in the facial skin accompanied by episodes of flushing and sweating triggered by cognitive processes,visual stimuli,or eating.AIM To investigate the preventive effect of acellular dermal matrix(ADM)on Frey syndrome after parotid tumor resection and analyzed the effects of Frey syndrome across various surgical methods and other factors involved in parotid tumor resection.METHODS Retrospective data from 82 patients were analyzed to assess the correlation between sex,age,resection sample size,operation time,operation mode,ADM usage,and occurrence of postoperative Frey syndrome.RESULTS Among the 82 patients,the incidence of Frey syndrome was 56.1%.There were no significant differences in sex,age,or operation time between the two groups(P>0.05).However,there was a significant difference between ADM implantation and occurrence of Frey syndrome(P<0.05).ADM application could reduce the variation in the incidence of Frey syndrome across different operation modes.CONCLUSION ADM can effectively prevent Frey syndrome and delay its onset.

Small-diameter acellular porcine corneal stroma for peripheral corneal ulceration treatment

AIM:To evaluate the clinical efficacy of small-diameter acellular porcine corneal stroma(SAPS)for the treatment of peripheral corneal ulceration(PCU).METHODS:This retrospective clinical study included 18 patients(18 eyes)with PCU between April 2018 and December 2020.All patients had PCU and underwent lamellar keratoplasty with SAPS.Observation indicators included preoperative and postoperative best-corrected visual acuity(BCVA)and transparency of SAPS.The infection control rate in the surgical eye-lesion area was also calculated.RESULTS:Eighteen patients underwent lamellar keratoplasty with SAPS to treat PCU.None of the patients experienced rejection after 6mo(18/18)and 12mo(16/16)of follow-up.The BCVA(0.47±0.30)at the 6mo followup after operation was significantly improved compared with the baseline(0.99±0.80),and the difference was statistically significant(Z=-3.415,P<0.05).The BCVA at the 12mo follow-up after operation was not statistically significant compared to the 6mo(Z=0,P=1).With time,the SAPS graft gradually became transparent.At the 6mo(18/18)and 12mo(16/16)follow-up,none of the patients had recurrent corneal infection.CONCLUSION:SAPS is clinically effective in the treatment of PCU,improving the patient’s BCVA and reducing the incidence of rejection after keratoplasty.

The First Reported Case of Co-Infection with Neisseria Meningitidis and Bordetella Pertussis in the Cerebrospinal Fluid Specimen of a Normal Four-Year-Old Child

Pertussis is an acute and highly contagious respiratory disease caused by Bordetella pertussis(B.pertussis),which occurs in people of all ages and is most dangerous for young children,especially infants.The introduction of whole-cell pertussis vaccines(wPVs)in the 1950s dramatically reduced the incidence of pertussis worldwide[1].However,over the past two decades,many studies have reported the resurgence of pertussis in different countries[2].Epidemiological surveillance in Hubei Province over the last 3 years revealed a clear increasing trend in the incidence of pertussis during the coronavirus disease 2019(COVID-19)epidemic.

Antibody Levels and Infection Status of Pertussis in the Population under Pertussis Resurgence in Guangxi in 2018:A Cross-Sectional Survey

Objective Pertussis cases have increased markedly since 2018 in Guangxi.The aim of this study was to evaluate antibody levels and the infection status of pertussis in the resident population.Method A total of 10,215 serum samples from residents were collected from August-November 2018 and tested for anti-pertussis IgG and toxin IgG using the enzyme-linked immunosorbent assay(ELISA).Results Of the collected samples,1,833(17.94%)tested positive for anti-pertussis IgG,with the median concentration of 16.06 IU/mL.Antibody level<10 IU/mL accounted for more than 60%in children under 4 years of age,but declined with age,whereas the percentages of the other three levels(10-40,40-50,and≥50 IU/mL)increased almost with age(P<0.001).Moreover,7,924 samples were selected for anti-pertussis toxin IgG,of which 653(8.24%)tested positive(≥40 IU/mL)with the median concentration of 5.89 IU/mL,and 204 participants(2.56%)had recent pertussis infection(≥100 IU/mL).Among the different age groups,the highest rates of positivity and recent infection were observed at 11-20 years of age,the lowest positivity rate at 5 years of age,and the lowest recent infection rate at 4 years of age(P<0.001,P=0.005,respectively).Conclusion The survey results showed that all age groups in Guangxi lacked immunity against pertussis,which was one of the main factors contributing to the resurgence of pertussis in 2018.In addition,the prevalence of pertussis is relatively high in Guangxi,and its incidence is seriously underestimated,especially in adolescents and adults.

Acellular dermal matrices in reconstructive surgery;history,current implications and future perspectives for surgeons

Large-scale defects of body in the reconstructive surgical practice,and the help-lessness of their repair with autologous tissues,have been an important factor in the development of artificial biological products for the temporary,definitive,or staged repair of these defects.A major advance in the field of plastic and other reconstructive surgery in this regard has been the introduction and successful use of acellular dermal matrices(ADMs).In recent years,not only the type of tissue from which ADMs are produced,product range,diversity and areas of use have increased,but their use in reconstructive fields,especially in post oncologic breast surgery,has become highly regarded and this has favored ADMs to be a potential cornerstone in specific and well-defined surgical fields in future.It is essential that reconstructive surgeons become familiar with some of the ADM’s as well as the advantages and limitations to their use.This review not only provides basic science and clinical evidence of the current use of ADMs in wide range of surgical fields but also targets to keep them as an important backdrop in the arma-mentarium of reconstructive surgeons.Brief considerations of possible future directions for ADMs are also conducted in the end.

脱细胞软骨细胞外基质匀浆联合氧化苦参碱治疗大鼠骨关节炎

背景:脱细胞软骨细胞外基质治疗骨关节炎具有一定的效果,但其单独应用时的治疗效果有限。氧化苦参碱可缓解白细胞介素1β诱导的软骨细胞凋亡和细胞外基质降解。目的:观察不同剂量氧化苦参碱与脱细胞软骨细胞外基质联合膝关节腔注射对大鼠骨关节炎的治疗作用。方法:获取SD大鼠股骨软骨,采用物理、化学与酶相结合的方法制备脱细胞软骨细胞外基质。取50只SD大鼠,利用随机数字表法分为假手术组、骨关节炎组、单纯材料组、低剂量药物组、高剂量药物组,每组10只,后4组采用改良Hulth法建立大鼠骨关节炎模型。造模成功后,单纯材料组大鼠膝关节腔注射脱细胞软骨细胞外基质匀浆,低剂量药物组、高剂量药物组膝关节腔分别注射含50,100μg氧化苦参碱的脱细胞软骨细胞外基质匀浆。注射4周后取材进行相关检测。结果与结论:①与假手术组比较,骨关节炎组关节腔积液中白细胞介素1β、肿瘤坏死因子ɑ、丙二醛水平升高(P<0.05),超氧化物歧化酶水平降低(P<0.05);与骨关节炎组比较,低剂量药物组、高剂量药物组关节腔积液中白细胞介素1β、肿瘤坏死因子ɑ、丙二醛水平降低(P<0.05),超氧化物歧化酶水平升高(P<0.05),其中高剂量药物组变化更显著。②Tunel染色显示,与假手术组相比,骨关节炎组凋亡软骨细胞增多;与骨关节炎组相比,单纯材料组、低剂量药物组、高剂量药物组凋亡软骨细胞减少,其中高剂量药物组减少更显著。③苏木精-伊红与免疫组化染色显示,骨关节炎组大鼠膝关节软骨严重退变,软骨组织中Ⅱ型胶原表达降低(P<0.05)、基质金属蛋白酶9表达升高(P<0.05);与骨关节炎组相比,单纯材料组、低剂量药物组、高剂量药物组大鼠膝关节软骨退变明显改善,软骨组织中Ⅱ型胶原表达升高(P<0.05)、基质金属蛋白酶9表达降低(P<0.05),其中高剂量药物组改善最显著。④Western blot检测显示,与假手术组相比,骨关节炎组软骨组织中Nrf2、HO-1、Bcl-2蛋白表达降低(P<0.05),Cleaved-caspase-3、Cleaved-caspase-9、Bax蛋白表达升高(P<0.05);与骨关节炎组相比,低剂量药物组、高剂量药物组Nrf2、HO-1、Bcl-2蛋白表达升高(P<0.05),Cleaved-caspase-3、Cleaved-caspase-9、Bax蛋白表达降低(P<0.05),其中高剂量药物组改善更显著。⑤结果表明,膝关节腔内联合注射脱细胞软骨细胞外基质与氧化苦参碱可通过促进细胞外基质的合成、抑制炎症与氧化应激反应、抑制软骨细胞凋亡发挥骨关节炎治疗作用,该治疗作用可能与激活Nrf2/HO-1通路有关。

小肠黏膜下层脱细胞基质复合外泌体构建组织工程尿道

背景:小肠黏膜下层脱细胞基质已被临床与基础研究证实可用于尿道的修复重建,但其单独应用时存在宿主细胞生长缓慢、支架血管化不足而成活困难、重建尿道狭窄梗阻等问题,仅适用于较短的尿道狭窄。目的:探讨应用小肠黏膜下层脱细胞基质复合外泌体构建组织工程尿道的可行性。方法:从新西兰大白兔骨髓间充质干细胞中分离提取外泌体;制备猪小肠黏膜下层脱细胞基质,将外泌体负载于小肠黏膜下层脱细胞基质上。将小肠黏膜下层脱细胞基质-外泌体(PKH26染料标记)复合物与脐静脉内皮细胞共培养12 h,观察细胞摄取外泌体情况。选取生长状态良好的脐静脉内皮细胞,分3组培养:空白组常规培养,对照组加入小肠黏膜下层脱细胞基质,实验组加入小肠黏膜下层脱细胞基质-外泌体复合物,通过划痕实验、成管实验、血管生成因子分泌检测评估血管生成情况。取30只新西兰大白兔,建立长段(3 cm)尿道缺损模型,采用随机数字表法分3组干预(n=10):单独材料组植入小肠黏膜下层脱细胞基质,对照组植入小肠黏膜下层脱细胞基质-骨髓间充质干细胞复合物,实验组植入小肠黏膜下层脱细胞基质-外泌体复合物,植入后12周进行尿道造影、尿动力学检查及重建尿道切片病理观察。结果与结论:(1)荧光显微镜下可见小肠黏膜下层脱细胞基质中的外泌体可被脐静脉内皮细胞摄取。(2)与空白组、对照组相比,实验组材料可促进脐静脉内皮细胞的迁移、成血管能力以及成血管因子血管内皮生长因子、肝细胞生长因子、白细胞介素8的分泌(P <0.05)。(3)尿道造影结果显示,单独材料组10只兔均出现尿道狭窄,对照组10只兔中2只出现尿道狭窄,实验组10只兔均未出现尿道狭窄。尿动力检查结果显示,单独材料组兔材料植入后12周的最大尿道压高于术前(P <0.05),对照组、实验组兔植入后12周的最大尿道压均低于空白组(P <0.05)。苏木精-伊红、Masson与免疫组化染色显示,单独材料组可见明显的再生上皮层、少量的皮下平滑肌与血管,以纤维组织增生为主,伴有明显炎性细胞浸润;对照组可见较完整的再生上皮与少量胶原,可见大量的皮下血管与平滑肌,伴有炎性细胞浸润;实验组可见完整的再生上皮层与大量的皮下血管、平滑肌,未见明显炎性细胞浸润,实验组AE1/AE3、ɑ-平滑肌肌动蛋白、CD31阳性表达均高于单独材料组、对照组(P <0.05)。(4)结果表明,小肠黏膜下层脱细胞基质-外泌体组织工程尿道可通过促进血管生成来修复尿道缺损。

组织工程化口腔黏膜等效物的血管化特征

背景:前期研究中三维细胞重建组织工程化口腔黏膜等效物结构类似于正常口腔黏膜,即存在类上皮样结构、类固有层样结构、类血管腔样结构,并已初步实现了等效物的血管化建立,但其血管化特征尚不十分明确。目的:采用血管内皮细胞特异性标志物表达谱关联激光捕获显微切割系统靶向获取血管化口腔黏膜等效物的血管样结构,评价其成血管能力,揭示其血管化特征。方法:分别从人牙龈上皮组织和固有层组织原代培养人牙龈上皮细胞、人牙龈成纤维细胞、人牙龈间充质干细胞,人牙龈间充质干细胞经单克隆扩增培养后诱导分化形成血管内皮样细胞。将人牙龈上皮细胞、人牙龈成纤维细胞、血管内皮样细胞分层负载于脱细胞血管基质-0.25%类人Ⅰ型胶原支架上,构建血管化口腔黏膜等效物。将血管化口腔黏膜等效物(实验组)与脱细胞血管基质-0.25%类人Ⅰ型胶原支架(对照组)分别植入裸鼠背部皮下,14 d后两组切口表面涂布生物胶,实验组生物胶表面接种人牙龈上皮细胞,对照组不接种细胞,继续饲养14 d后取材,利用形态学观察口腔黏膜等效物分层结构;采用较为全面的血管内皮细胞特异性标志物表达谱对口腔黏膜等效物中的新生血管样结构进行免疫组化、免疫荧光标记,进行血管化特征分析;采用激光捕获显微切割系统靶向捕获免疫组化特异性标记的口腔黏膜等效物中新生血管样结构,靶向分析其血管化特征。结果与结论:(1)形态学观察显示口腔黏膜等效物细胞层次清晰,结构类似于正常口腔黏膜,即存在类上皮样结构、类固有层样结构、类血管腔样结构,类血管腔样结构内存在散在红细胞;(2)口腔黏膜等效物组中EdU Apollo示踪种子细胞结果显示:EdU Apollo 488标记的人牙龈上皮细胞呈绿色荧光表达;DAPI标记的人牙龈成纤维细胞呈蓝色荧光表达,体内形成类固有层样结构;Ed U Apollo 567标记的血管内皮样细胞呈红色荧光表达,体内形成类血管样结构;(3)血管内皮细胞特异性标志物表达谱免疫荧光标记血管结构显示,与正常口腔黏膜相比,口腔黏膜等效物中CD31、CD51、CD54、CD105、Tie-2、VWF、血管内皮生长因子受体1、血管内皮生长因子受体2表达升高(P<0.0001),CD34表达无明显变化(P>0.05);(4)与特异性标记的口腔黏膜血管结构相比,激光捕获显微切割系统靶向捕获的口腔黏膜等效物血管样结构中CD51、CD54、CD105、Tie-2、VWF、血管内皮生长因子受体1、血管内皮生长因子受体2表达升高(P<0.0001),CD31、CD34表达无明显变化(P>0.05);(5)结果表明,通过三维细胞分层重建的口腔黏膜等效物能够实现良好的血管化,其血管化特征符合新生血管生成的免疫学功能及特点;血管化助力三维细胞分层重建的口腔黏膜等效物再生。

Randomized controlled trial of minimally invasive surgery using acellular dermal matrix for complex anorectal fistula

AIM: To compare the efficacy and safety of acellular dermal matrix (ADM) bioprosthetic material and endorectal advancement flap (ERAF) in treatment of complex anorectal fistula. METHODS: Ninety consecutive patients with complex anorectal fistulae admitted to Anorectal Surgical Department of First Affi liated Hospital, Xinjiang Medical University from March 2008 to July 2009, were enrolled in this study. Complex anorectal fistula was diagnosed following its clinical, radiographic, or endoscopic diagnostic criteria. Under spinal anesthesia, patients underwent identification and irrigation of the fistula tracts using hydrogen peroxide. ADM was securely sutured at the secondary opening to the primary opening using absorbable suture. Outcomes of ADM and ERAF closure werecompared in terms of success rate, fecal incontinence rate, anorectal deformity rate, postoperative pain time, closure time and life quality score. Success was defined as closure of all external openings, absence of drainage without further intervention, and absence of abscess formation. Follow-up examination was performed 2 d, 2, 4, 6, 12 wk, and 5 mo after surgery, respectively. RESULTS: No patient was lost to follow-up. The overall success rate was 82.22% (37/45) 5.7 mo after surgery. ADM dislodgement occured in 5 patients (11.11%), abscess formation was found in 1 patient, and fistula recurred in 2 patients. Of the 13 patients with recurrent fistula using ERAF, 5 (11.11%) received surgical drainage because of abscess formation. The success rate, postoperative pain time and closure time of ADM were significantly higher than those of ERAF (P < 0.05). However, no difference was observed in fecal incontinence rate and anorectal deformity rate after treatment with ADM and ERAF. CONCLUSION: Closure of fistula tract opening with ADM is an effective procedure for complex anorectal fistula. ADM should be considered a first line treatment for patients with complex anorectal fistula.

Risk factors for pertussis among children hospitalized for pertussis during 2016-2017, in Guizhou Province of China: a case-control study

Aim:Pertussis is a respiratory tract infection,the vaccine for which was introduced in the Expanded Programme on Immunization(EPI)in Guizhou Province of China(hereafter referred as Guizhou)in the 1980s.This vaccine rapidly decreased incidence rates of pertussis in the province,however,despite the wide high coverage of the diphtheria,tetanus and acellular pertussis(DTaP)combined vaccine,there has been a resurgence of pertussis since 2014.Even with this recent increase in disease transmission,risk factors for pertussis infection have not been evaluated in Guizhou.We aimed to provide information on pertussis risk factors and insight on designing targeted pertussis control policies and measures through this study.Methods:A 1:2 matched retrospective case-control study was conducted between 2016 and 2017,involving infants and children younger than 6 years old and parents of the participants.The enrolled cases included clinical and laboratory confirmed pertussis cases according to the WHO-recommended pertussis definition.Controls were selected from children in the same neighborhood who were not diagnosed with pertussis prior to our investigation and did not exhibit any clinical manifestations of pertussis.Results:The household size[Matched odds ratio(OR_(m)=1.4,95%confidence interval(Cl):1.1-1.7]and household members with antecedent cough(OR_(m)=3.6,95%CI:1.8-7.2)were significantly associated with their child’s pertussis onset.The parents’occupations were significantly associated with their child’s pertussis onset(ORm=9.4,95%CI:1.6-54.8,for mother side;ORm=4.5,95%CI:1.2-16.5,for father side),when they worked in the business and/or service industry.Having family members with a history of cough was an independent risk factor for pertussis[Adjusted odds ratio(AOR)=43.6,95%CI:2.7-694.0].Besides,the parents’demographic characteristics and DTaP doses were not found to be independent factors.Conclusion:Household exposure is an important risk factor for pertussis infection in infants and young children and should therefore be considered as a major factor during formulation of pertussis control policies and measures.

Acellular ostrich corneal stroma used as scaffold for construction of tissue-engineered cornea

AIM： To assess acellular ostrich corneal matrix used as a scaffold to reconstruct a damaged cornea. METHODS： A hypertonic saline solution combined with a digestion method was used to decellularize the ostrich cornea. The microstructure of the acellular corneal matrix was observed by transmission electron microscopy （TEM） and hematoxylin and eosin （H＆E） staining. The mechanical properties were detected by a rheometer and a tension machine. The acellular corneal matrix was also transplanted into a rabbit cornea and cytokeratin 3 was used to check the immune phenotype, RESULTS： The microstructure and mechanical properties of the ostrich cornea were well preserved after the decellularization process, in vitro, the methyl thiazolyl tetrazoUum results revealed that extracts of the acellular ostrich corneas （AOCs） had no inhibitory effects on the proliferation of the corneal epithelial or endothelial cells or on the keratocytes, The rabbit lamellar keratoplasty showed that the transplanted AOCs were transparent and completely incorporated into the host cornea while corneal turbidity and graft dissolution occurred in the acellular porcine cornea （APC） transplantation, The phenotype of the reconstructed cornea was similar to a normal rabbit cornea with a high expression of cytokeratin 3 in the superficial epithelial cell layer, CONCLUSION： We first used AOCs as scaffolds to reconstruct damaged corneas. Compared with porcine corneas, the anatomical structures of ostrich corneas are closer to those of human corneas. In accordance with the principle that structure determines function, a xenograft lamellar keratoplasty also confirmed that the AOC transplantation generated a superior outcome compared to that of the APC graft.

Biomechanical properties of acellular sciatic nerves treated with a modified chemical method

Nerve grafts are able to adapt to surrounding biomechanical environments if the nerve graft itself exhibits appropriate biomechanical properties （load, elastic modulus, etc.）. The present study was designed to determine the differences in biomechanical properties between fresh and chemically acellularized sciatic nerve grafts. Two different chemical methods were used to establish acellular nerve grafts. The nerve was chemically extracted in the Sondell method with a combination of Triton X-100 （nonionic detergent） and sodium deoxycholate （anionic detergent）, and in the modified method with a combination of Triton X-200 （anionic detergent）, sulfobetaine-10 （SB-10, amphoteric detergents）, and sulfobetaine-16 （SB-16, amphoteric detergents）. Following acellularization, hematoxylin-eosin staining and scanning electron microscopy demonstrated that the effect of acellularization via the modified method was similar to the traditional Sondell method. However, effects of demyelination and nerve fiber tube integrity were superior to the traditional Sondell method. Biomechanical testing showed that peripheral nerve graft treated using the chemical method resulted in decreased biomechanical properties （ultimate load, ultimate stress, ultimate strain, and mechanical work to fracture） compared with fresh nerves, but the differences had no statistical significance （P 〉 0.05）. These results demonstrated no significant effect on biomechanical properties of nerves treated using the chemical method. In conclusion, nerve grafts treated via the modified method removed Schwann cells, preserved neural structures, and ensured biomechanical properties of the nerve graft, which could be more appropriate for implantation studies.

A novel porcine acellular dermal matrix scaffold used in periodontal regeneration

Regeneration of periodontal tissue is the most promising method for restoring periodontal structures.To find a suitable bioactive three- dimensional scaffold promoting cell proliferation and differentiation is critical in periodontal tissue engineering.The objective of this study was to evaluate the biocompatibility of a novel porcine acellular dermal matrix as periodontal tissue scaffolds both in vitro and in vivo.The scaffolds in this study were purified porcine acellular dermal matrix(PADM) and hydroxyapatite-treated PADM(HA-PADM). The biodegradation patterns of the scaffolds were evaluated in vitro.The biocompatibility of the scaffolds in vivo was assessed by implanting them into the sacrospinal muscle of 20 New Zealand white rabbits.The hPDL cells were cultured with PADM or HA-PADM scaffolds for 3,7,14,21 and 28 days.Cell viability assay,scanning electron microscopy(SEM),hematoxylin and eosin(H&E) staining, immunohistochemistry and confocal microscopy were used to evaluate the biocompatibility of the scaffolds.In vitro,both PADM and HA-PADM scaffolds displayed appropriate biodegradation pattern,and also,demonstrated favorable tissue compatibility without tissue necrosis,fibrosis and other abnormal response.The absorbance readings of the WST-1 assay were increased with the time course, suggesting the cell proliferation in the scaffolds.The hPDL cells attaching,spreading and morphology on the surface of the scaffold were visualized by SEM,H&E staining,immnuohjstochemistry and confocal microscopy,demonstrated that hPDL cells were able to grow into the HA-PADM scaffolds and the amount of cells were growing up in the course of time.This study proved that HA-PADM scaffold had good biocompatibility in animals in vivo and appropriate biodegrading characteristics in vitro.The hPDL cells were able to proliferate and migrate into the scaffold.These observations may suggest that HA-PADM scaffold is a potential cell carrier for periodontal tissue regeneration.

Chemically extracted acellular allogeneic nerve graft combined with ciliary neurotrophic factor promotes sciatic nerve repair

A chemically extracted acellular allogeneic nerve graft can reduce postoperative immune rejection, similar to an autologous nerve graft, and can guide neural regeneration. However, it remains poorly understood whether a chemically extracted acellular allogeneic nerve graft combined with neurotrophic factors provides a good local environment for neural regeneration. This study investigated the repair of injured rat sciatic nerve using a chemically extracted acellular allogeneic nerve graft combined with ciliary neurotrophic factor. An autologous nerve anastomosis group and a chemical acellular allogeneic nerve bridging group were prepared as controls. At 8 weeks after repair, sciatic functional index, evoked potential amplitude of the soleus muscle, triceps wet weight recovery rate, total number of myelinated nerve fibers and myelin sheath thickness were measured. For these indices, values in the three groups showed the autologous nerve anastomosis group 〉 chemically extracted acellular nerve graft ＋ ciliary neurotrophic factor group 〉 chemical acellular allogeneic nerve bridging group. These results suggest that chemically extracted acellular nerve grafts combined with ciliary neurotrophic factor can repair sciatic nerve defects, and that this repair is inferior to autologous nerve anastomosis, but superior to chemically extracted acellular allogeneic nerve bridging alone.

Acellular porcine corneal matrix as a carrier scaffold for cultivating human corneal epithelial cells and fibroblasts in vitro

AIM：To investigate the feasibility of corneal anterior lamellar reconstruction with human corneal epithelial cells and fibroblasts,and an acellular porcine cornea matrix（APCM） in vitro.·METHODS：The scaffold was prepared from fresh porcine corneas which were treated with 0.5%sodium dodecyl sulfate（SDS）solution and the complete removal of corneal cells was confirmed by hematoxylin-eosin（HE）staining and 4’,6-diamidino-2-phenylindole（DAPI）staining.Human corneal fibroblasts and epithelial cells were cultured with leaching liquid extracted from APCM,and then cell proliferative ability was evaluated by MTT assay.To construct a human corneal anterior lamellar replacement,corneal fibroblasts were injected into the APCM and cultured for 3d,followed by culturing corneal epithelial cells on the stroma construction surface for another 10d.The corneal replacement was analyzed by HE staining,and immunofluorescence staining.·R ESULTS：Histological examination indicated that there were no cells in the APCM by HE staining,and DAPI staining did not detect any residual DNA.The leaching liquid from APCM had little influence on the proliferation ability of human corneal fibroblasts and epithelial cells.At 10d,a continuous 3 to 5 layers of human corneal epithelial cells covering the surface of the APCM was observed,and the injected corneal fibroblasts distributed within the scaffold.The phenotype of the construction was similar to normal human corneas,with high expression of cytokeratin 12 in the epithelial cell layer and high expression of Vimentin in the stroma.·CONCLUSION：Corneal anterior lamellar replacement can be reconstructed in vitro by cultivating human corneal epithelial cells and fibroblasts with an acellular porcine cornea matrix.This laid the foundation for the further transplantation in vitro.

Establishment of an untransfected human corneal stromal cell line and its biocompatibility to acellular porcine corneal stroma

AIM: To establish an untransfected human corneal stromal (HCS) cell line and characterize its biocompatibility to acellular porcine corneal stoma (aPCS). METHODS: Primary culture was initiated with a pure population of HCS cells in DMEM/F12 media (pH 7.2) containing 20% fetal bovine serum and various necessary growth factors. The established cell line was characterized by growth property, chromosome analysis, tumorigenicity assay, expression of marker proteins and functional proteins. Furthermore, the biocompatibility of HCS cells with aPCS was examined through histological and immunocytochemistry analyses and with light, electron microscopies. RESULTS: HCS cells proliferated to confluence 2 weeks later in primary culture and have been subcultured to passage 140 so far. A continuous untransfected HCS cell line with a population doubling time of 41.44 hours at passage 80 has been determined. Results of chromosome analysis, morphology, combined with the results of expression of marker protein and functional proteins suggested that the cells retained HCS cell properties. Furthermore, HCS cells have no tumorigenicity, and with excellent biocompatibility to aPCS. CONCLUSION: An untransfected and non-tumorigenic HCS cell line has been established, and the cells maintained positive expression of marker proteins and functional proteins. The cell line, with excellent biocompatibility to aPCS, might be used for in vitroreconstruction of tissue-engineered HCS.

Acellular allogeneic nerve grafting combined with bone marrow mesenchymal stem cell transplantation for the repair of long-segment sciatic nerve defects:biomechanics and validation of mathematical models

We hypothesized that a chemically extracted acellular allogeneic nerve graft used in combination with bone marrow mesenchymal stem cell transplantation would be an effective treatment for long-segment sciatic nerve defects.To test this,we established rabbit models of 30 mm sciatic nerve defects,and treated them using either an autograft or a chemically decellularized allogeneic nerve graft with or without simultaneous transplantation of bone marrow mesenchymal stem cells.We compared the tensile properties,electrophysiological function and morphology of the damaged nerve in each group.Sciatic nerves repaired by the allogeneic nerve graft combined with stem cell transplantation showed better recovery than those repaired by the acellular allogeneic nerve graft alone,and produced similar results to those observed with the autograft.These findings confirm that a chemically extracted acellular allogeneic nerve graft combined with transplantation of bone marrow mesenchymal stem cells is an effective method of repairing long-segment sciatic nerve defects.

Acellular nerve allograft promotes selective regeneration

Acellular nerve ailograft preserves the basilar membrane tube and extracellular matrix, which promotes selective regeneration of neural defects via bridging. In the present study, a Sprague Dawley rat sciatic nerve was utilized to prepare acellular nerve allografts through the use of the chemical extraction method. Subsequently, the allograft was transplanted into a 10-mm sciatic nerve defect in Wistar rats, while autologous nerve grafts from Wistar rats served as controls. Compared with autologous nerve grafts, the acellular nerve allografts induced a greater number of degenerated nerve fibers from sural nerves, as well as a reduced misconnect rate in motor fibers, fewer acetylcholine esterase-positive sural nerves, and a greater number of carbonic anhydrase-positive sensory nerve fibers. Results demonstrated that the acellular nerve allograft exhibited significant neural selective regeneration in the process of bridging nerve defects.

Repair of peripheral nerve defects with chemically extracted acellular nerve allografts loaded with neurotrophic factors-transfected bone marrow mesenchymal stem cells

Chemically extracted acellular nerve allografts loaded with brain-derived neurotrophic fac- tor-transfected or ciliary neurotrophic factor-transfected bone marrow mesenchymal stem cells have been shown to repair sciatic nerve injury better than chemically extracted acellular nerve allografts alone, or chemically extracted acellular nerve allografts loaded with bone marrow mesenchymal stem cells. We hypothesized that these allografts compounded with both brain-derived neurotrophic factor- and ciliary neurotrophic factor-transfected bone marrow mesenchymal stem cells may demonstrate even better effects in the repair of peripheral nerve injury. We cultured bone marrow mesenchymal stem cells expressing brain-derived neuro- trophic factor and/or ciliary neurotrophic factor and used them to treat sciatic nerve injury in rats. We observed an increase in sciatic functional index, triceps wet weight recovery rate, myelin thickness, number of myelinated nerve fibers, amplitude of motor-evoked potentials and nerve conduction velocity, and a shortened latency of motor-evoked potentials when al- lografts loaded with both neurotrophic factors were used, compared with allografts loaded with just one factor. Thus, the combination of both brain-derived neurotrophic factor and cili- ary neurotrophic factor-transfected bone marrow mesenchymal stem cells can greatly improve nerve injury.

Combining acellular nerve allografts with brainderived neurotrophic factor transfected bone marrow mesenchymal stem cells restores sciatic nerve injury better than either intervention alone

In this study, we chemically extracted acellular nerve allografts from bilateral sciatic nerves, and repaired 10-mm sciatic nerve defects in rats using these grafts and brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells. Experiments were performed in three groups： the acellular nerve allograft bridging group, acellular nerve allograft ＋ bone marrow mesenchymal stem cells group, and the acellular nerve allograft ＋ brain-derived neurotrophic factor transfected bone marrow mesenchyrnal stem cells group. Results showed that at 8 weeks after bridging, sciatic functional index, triceps wet weight recovery rate, myelin thickness, and number of myelinated nerve fibers were significantly changed in the three groups. Variations were the largest in the acellular nerve allograft ＋ brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells group compared with the other two groups. Experimental findings suggest that chemically extracted acellular nerve allograft combined nerve factor and mesenchymal stem cells can promote the restoration of sciatic nerve defects. The repair effect seen is better than the single application of acellular nerve allograft or acellular nerve allograft combined mesenchymal stem cell transplantation.