Electrospun gelatin scaffolds incorporating rat decellularized brain extracellular matrix for neural tissue engineering

The fabrication of an instructive bioabsorbable scaffold is one of the main goals for tissue engineering applications. In this regard, genipin cross-linked gelatin scaffolds, produced by electrospinning, were tested as a platform to include decellularized rat brain extracellular matrix as an active agent to provide fundamental biochemical cues to the seeded cells. This approach is expected to furnish a suitable natural-based polymeric scaffold with sufficient temporal stability to support cell attachment and spreading, also providing tissue-specific signals that can contribute to the expression of the requested cellular phenotype. We first demonstrated the effectiveness of the proposed decellularization protocol and the cytocompatibility of the resulting brain matrix. Then, the in vitro biological assays of the conditioned electrospun scaffolds, using rat allogeneic mesenchymal stromal cells, confirmed their biocompatibility and showed a differentiative potential in presence of just 1% w/w decellularized rat brain extracellular matrix.     

灌注法制备离体大鼠心脏脱细胞基质

目的 探索制备离体大鼠心脏脱细胞生物支架材料的新方法,为心脏组织工程研究提供三维立体天然支架.方法 取30只成年SD大鼠心脏,运用冻融加化学萃取的组织工程学方法 (胰蛋白酶、十二烷基硫酸钠和曲拉通X-100)处理离体大鼠心脏,同时观察心脏大体形态及颜色变化,并对脱细胞支架进行基因组DNA分析;HE染色,免疫荧光法,扫描和透射电镜进一步检测鉴定脱细胞支架的生物学特征.结果 心脏脱细胞支架外观透明,包膜完整,维持心脏三维立体结构,肉眼可见心脏内脉管系统;脱细胞支架DNA残留量不及对照组的3%;HE染色、扫描和透射电镜结果 显示,心脏脱细胞生物支架去细胞彻底,细胞外基质网状结构保留完整;免疫荧光结果 表明,胶原、弹性蛋白等细胞外支架成分保留较完整,未见明显细胞核成分残留.结论 运用冻融加化学萃取法所制备的离体心脏脱细胞生物支架去细胞彻底,细胞外基质保留较完整,是较为理想的心脏三维立体生物支架材料.     

不同交联密度甲基丙烯酰酯明胶/脱细胞半月板细胞外基质复合水凝胶的性能

文题释义：脱细胞半月板细胞外基质：由新鲜半月板组织通过湿法差速离心方法脱细胞制备而来,在去除了异体半月板的免疫原性的同时,仍保留了组织的大部分成分和组织本身的生物学性能,例如影响细胞的活性、调控细胞的增殖和分化、促进组织再生和修复等。 甲基丙烯酰酯明胶：是一种明胶的衍生物,由甲基丙烯酸酐与明胶合成而来。在加入光引发剂的基础上经紫外线照射,明胶链上的甲基丙烯酰胺和甲基丙烯酸甲酯侧基聚合形成聚合物链,聚合物链交错纵横,最终形成网状结构。 交联密度：通常用来表征交联聚合物里交联键的数量,主要受交联时间和交联光强度的影响,其可明显改变聚合物链的形成情况。 背景：交联后的聚合物链对水凝胶的基本性质和细胞相容性存在显著影响,而交联密度可明显改变聚合物链的形成情况,有关交联密度对水凝胶性能影响的针对性研究较少。 目的：制备一种细胞相容良好性的复合水凝胶,探究交联密度对该水凝胶性能的影响。 方法：配制甲基丙烯酰酯明胶溶液,然后加入脱细胞半月板细胞外基质溶液与LAP溶液,制备预凝胶溶液,采用蓝光对溶液进行交联,交联时间分别为10,30,60 s,检测3种水凝胶的压缩弹性模量、溶胀倍率与降解性能。将半月板纤维软骨细胞加入预凝胶溶液中,采用蓝光对溶液进行交联,交联时间分别为10,30,60 s,检测培养对应时间的细胞活性、形态与聚集。 结果与结论：①交联60 s水凝胶的压缩模明显高于交联10,30 s的水凝胶(P < 0.05);②交联10 s水凝胶的溶胀倍率明显高于交联30,60 s的水凝胶(P < 0.05),交联30,60 s的水凝胶的溶胀倍率比较差异无显著性意义(P > 0.05);③随着交联时间的延长,水凝胶的降解时间延长,交联60 s的水凝胶需要80 min完全降解,交联10 s的水凝胶50 min就可以完全降解;④培养24 h后,3组水凝胶中的细胞活性均在95%以上,各组之间无差异(P > 0.05);⑤培养1 d时,3组水凝胶中的细胞均呈球形且均匀分布;4 d时,3组细胞均开始伸展,交联10 s水凝胶中有较小的细胞团;7 d时,3组细胞树突状伸展更为明显,其中交联10 s水凝胶中形成了较为明显的细胞团;⑥培养1,7,14 d时,交联10 s水凝胶中的细胞活性均在85%以上。培养1 d时,交联10 s水凝胶中的细胞呈球形分布,且分布均匀;培养28 d时,细胞呈树突状伸展,并聚集形成网状结构;⑦结果表明,甲基丙烯酰酯明胶/脱细胞半月板细胞外基质复合水凝胶的性能可通过调整交联密度进行优化。 ORCID: 0000-0002-3606-1097(周建) 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

脱细胞基质水凝胶在再生医学中的研究进展

脱细胞基质(dECM)由于其高仿生性和优异的生物相容性,已被广泛用作再生医学的支架材料。水凝胶(hydrogel)作为一种高含水量、可控流动性的功能高分子材料,非常适合用于临床中的部分微创手术。近年来,随着水凝胶理论和技术的快速发展,dECM水凝胶逐渐成为再生医学领域的研究热点。本文从dECM水凝胶的制备及其临床前应用两个方面对近年来的相关研究进行综述,并展望其未来的临床前景。     

Synthetic biodegradable hydrogel delivery of demineralized bone matrix for bone augmentation in a rat model

There exists a strong clinical need for a more capable and robust method to achieve bone augmentation, and a system with fine-tuned delivery of demineralized bone matrix (DBM) has the potential to meet that need. As such, the objective of the present study was to investigate a synthetic biodegradable hydrogel for the delivery of DBM for bone augmentation in a rat model. Oligo(poly(ethylene glycol) fumarate) (OPF) constructs were designed and fabricated by varying the content of rat-derived DBM particles (either 1:3, 1:1 or 3:1 DBM:OPF weight ratio on a dry basis) and using two DBM particle size ranges (50–150 or 150–250 μm). The physical properties of the constructs and the bioactivity of the DBM were evaluated. Selected formulations (1:1 and 3:1 with 50–150 μm DBM) were evaluated in vivo compared to an empty control to investigate the effect of DBM dose and construct properties on bone augmentation. Overall, 3:1 constructs with higher DBM content achieved the greatest volume of bone augmentation, exceeding 1:1 constructs and empty implants by 3- and 5-fold, respectively. As such, we have established that a synthetic, biodegradable hydrogel can function as a carrier for DBM, and that the volume of bone augmentation achieved by the constructs correlates directly to the DBM dose.     

骨髓基质干细胞与骨基质明胶复合培养体内异位成骨的实验研究

目的：探讨骨髓基质干细胞(MSC)与骨基质明胶复合培养体内异位成骨的可行性。方法：将SD大鼠来源的MSC与同种异体的骨基质明胶复合培养后植入SD大鼠背部竖脊肌肌膜内，分别于术后不同的时间点处死大鼠，标本进行碱性磷酸酶(ALP)活性测定及组织形态学观察：结果：实验组标本自术后第3周ALP活性起就表现出阳性，第4周MSC与骨基质明胶复合体内大量成骨。结论：MSC与骨基质明胶复合培养体内异位成骨完全可行的。     

SD大鼠骨基质明胶吸附骨髓间充质干细胞修复骨缺损的实验研究

目的:探讨大鼠骨基质明胶吸附骨髓间充质干细胞修复骨缺损的可行性。方法:采用第5代(P5)SD大鼠骨髓间充质干细胞(MSCs),经Hoeschst 33344(sigma)荧光杂料标记后,调配制成的1×106个／ml细胞浓度后与骨基质明胶(BMG)共同培养6 h,然后植入SD大鼠双侧胫骨的实验性骨缺损中(A组),同时作胫骨骨缺损单纯BMG植入(B组),无植入物(C组)两组对照。术后8周处死,取骨缺损区组织进行组织学观察,其中A组行荧光染料标记测定以确认骨缺损区的骨痂是否来源于MSCS。结果:①A组:胫骨骨缺损区可见大量新生不规则骨纤维组织、软骨及纤维骨痂填充,可见骨细胞、骨组织和骨小梁,已形成骨髓腔。②B组:胫骨骨缺损区可见大量纤维组织、少量新生不规则骨纤维组织及骨骼肌组织,伴有多核巨细胞和少量炎性细胞,缺损区边缘带有骨痂组织。③C组:胫骨骨缺损区可见大量纤维组织及骨骼肌组织填充生长,伴有多核巨细胞和少量炎性细胞,缺损区边缘带有少量骨痂组织。荧光染色鉴定确认胫骨骨缺损区的骨痂来源于MSCs。结论:大鼠骨基质明胶吸附骨髓间充质干细胞修复骨缺损具有安全性、可行性及有效性。     

The extracellular matrix during neural crest formation and migration in rat embryos

Summary Changes in the distribution of extracellular matrix components have been investigated immunohistochemically during neural crest development in the rat. Inside the ectodermal epithelium basal lamina components are formed resulting in a separation of neurectoderm and epidermal ectoderm. Within the presumptive neural crest area fibronectin, hyaluronan and chondroitin sulphate become apparent. Upon subsequent neural crest migration the basal lamina becomes disrupted. As the neural crest cells take part in mesectoderm formation, fragments of the basal lamina remain attached to their surface, as is demonstrated with antibodies against laminin and collagen type IV. The extracellular matrix is therefore active both in the separation of neuroectoderm from epidermal ectoderm and in mesectoderm formation.     

同种与异种骨基质明胶修复颅骨缺损的实验研究

以家兔颅顶骨直径１０ｍｍ园形骨缺损作为动物模型，分别植入同种（兔）、异种（人、猪、羊）的骨基质明胶。植入后４、８、１２、１６ｗ进行ｘ线摄片和组织学检查。结果显示，同种骨基质明胶无免疫排斥反应，具有良好的骨诱导作用，术后１２ｗ骨缺损完全修复；异种骨基质明胶植入早期，存在着不同程度的排斥反应，植骨后期（１２～１６ｗ），随着排斥反应的减轻，亦出现了诱导成骨。结果说明，如能改进异种骨基质明胶的制作方法，降低其抗原性，将具有重要的临床应用价值。     

离体大鼠胰腺去细胞生物支架的制备与鉴定

目的 通过离体灌注加浸泡的方法制备大鼠胰腺去细胞生物支架(PDBC),为胰岛和胰腺的组织工程研究提供新型天然生物支架. 方法 健康成年大鼠20只,以物理冻融及灌注洗脱法(脱氧胆酸钠+ DNAase),采用胆管、胰管联合逆向在体灌流法获取胰腺去细胞生物支架.并通过基因组DNA定量定性分析,组织化学染色、免疫荧光组织化学染色、荧光积分吸光度分析、透射电镜观察等进一步测定细胞残留以及观察去细胞支架,如胶原IV、纤维连接蛋白和层黏连蛋白等成分的保留. 结果 DNA定性分析显示,实验组未见明显DNA条带,对照组DNA条带可达100bp,定量检测实验组DNA残留不足对照组的5％;组织化学染色和透射电镜观察均表明,去细胞支架无细胞残留,细胞外支架连续性完好,脉管支架保存完整;免疫荧光结果表明,胶原蛋白、弹性蛋白等细胞外支架成分保留较完整,未见明显细胞核成分残留. 结论 运用冻融加浸泡灌注制备的胰腺去细胞生物支架,细胞去除彻底,细胞外支架保留完好.     

大鼠单叶肝去细胞生物支架的制备与鉴定

目的通过灌注法制备大鼠单叶肝去细胞生物支架,并对其进行鉴定。方法健康成年SD大鼠20只,随机分为去细胞组和正常对照组,每组10只。去细胞组经门静脉灌胃针插管,恒温37℃依次灌注肝素化PBS溶液,1%Triton X-100(p H 7.5~8.0)及PBS溶液。HE、Masson染色及扫描电子显微镜观察组织学及超微结构改变;免疫荧光结合4’,6-二脒基-2-苯基吲哚(DAPI)观察2组胶原蛋白Ⅳ和Ⅰ、层黏连蛋白和纤维连接蛋白观察细胞外基质的主要成分;DNA定性和定量分析组织中残留DNA浓度和片段长度;聚甲基丙烯酸甲酯铸型观察肝脏内血管分布情况。结果 Triton X-100灌注4h左右即可制备单叶肝脏去细胞生物支架。在灌注过程中,肝内细胞和细胞碎片逐渐被清洗,最终变成半透明状。HE、Masson、免疫荧光染色及扫描电子显微镜显示,去细胞组肝生物支架较完整地保留了细胞外支架的成分,未见明显细胞及细胞核成分残留;去细胞组支架DNA残留量较正常对照组下降了97.32%,琼脂糖凝胶电泳未见明显的DNA条带。血管铸型标本显示,去细胞组血管分布与正常对照组相仿,其分支完整、清晰。结论运用Triton X-100灌注法所制备的大鼠单叶肝生物支架去细胞彻底,较完整地保留细胞外基质和血管网络结构,是一种简单易行且较为理想的制备实验用单叶肝生物支架的方法。     

Micro- and nanotopography with extracellular matrix coating modulate human corneal endothelial cell behavior

The human corneal endothelium plays an important role in maintaining corneal transparency. Human corneal endothelial cells have limited regenerative capability in vivo. Consequently, endothelial dysfunction can occur following corneal endothelial trauma or inherited diseases. To restore endothelial function, corneal transplantation is needed. However, there is a worldwide shortage of donor corneas, motivating the development of a tissue-engineered graft alternative using cultivated endothelial cells. To induce in vitro cell proliferation, much effort has been made to improve culture conditions and to mimic the native extracellular microenvironment. We incorporated topographical and biochemical cues in our in vitro culture of human corneal endothelial cell line B4G12 (HCEC-B4G12) and hypothesized that manipulation of the extracellular environment can modulate cell proliferation, morphometry and phenotype. The topographies tested were nanopillars, microwells and micropillars on polydimethylsiloxane, while the biochemical factors were extracellular matrix protein coatings of fibronectin-collagen I (FC), FNC® coating mix (FNC) and laminin-chondroitin sulfate (LC). Cellular morphometry, Na⁺/K⁺-ATPase and zona occludens 1 (ZO-1) gene and protein expression were analyzed 3 days after cells had formed a confluent monolayer. The cell circularity on all patterns and coatings was above 0.78. On all coatings, cell area was the lowest on micropillars. The coefficient of variation (CV) of the cell area was the lowest on nanopillars with an LC coating. With an FC coating, micropillars induced a better cellular outcome as the cells had the greatest circularity, smallest cell area and highest Na⁺/K⁺-ATPase and ZO-1 gene and protein expression. With the LC coating, HCECs grown on nanopillars resulted in the lowest CV of the cell area and the highest ZO-1 gene expression. Thus, HCEC-B4G12 morphometry and phenotype can be improved using different topographical and biochemical cues.     

Tunable hydrogel composite with two-step processing in combination with innovative hardware upgrade for cell-based three-dimensional bioprinting

Three-dimensional (3-D) bioprinting is the layer-by-layer deposition of biological material with the aim of achieving stable 3-D constructs for application in tissue engineering. It is a powerful tool for the spatially directed placement of multiple materials and/or cells within the 3-D sample. Encapsulated cells are protected by the bioink during the printing process. Very few materials are available that fulfill requirements for bioprinting as well as provide adequate properties for cell encapsulation during and after the printing process. A hydrogel composite including alginate and gelatin precursors was tuned with different concentrations of hydroxyapatite (HA) and characterized in terms of rheology, swelling behavior and mechanical properties to assess the versatility of the system. Instantaneous as well as long-term structural integrity of the printed hydrogel was achieved with a two-step mechanism combining the thermosensitive properties of gelatin with chemical crosslinking of alginate. Novel syringe tip heaters were developed for improved temperature control of the bioink to avoid clogging. Human mesenchymal stem cells mixed into the hydrogel precursor survived the printing process and showed high cell viability of 85% living cells after 3 days of subsequent in vitro culture. HA enabled the visualization of the printed structures with micro-computed tomography. The inclusion of HA also favors the use of the bioink for bone tissue engineering applications. By adding factors other than HA, the composite could be used as a bioink for applications in drug delivery, microsphere deposition or soft tissue engineering.     

Material design for an artificial extracellular matrix: Cell entrapment in poly (N-isopropylacrylamide) (PNIPAM)-grafted gelatin hydrogel

As a thermoresponsive extracellular matrix, PNIPAM-derivatized gelatin (PNIPAM-gelatin) was synthesized by an iniferter-based graft polymerization of NIPAM on side chains of gelatin (molecular weight, ca. 9.5 ×10⁴ g/mol). The degree of grafting was 22.6 groups per molecule, and the estimated molecular weight of PNIPAM was ca. 1.2×10⁴ g/mol. The phase transition of dissolution/precipitation of PNIPAM-gelatin occurred at around 35°C. At concentrations above 15 w/v% over about 35°C, the solution was converted to hydrogel. The mechanical strength of the produced hydrogel increased with the concentration of PNIPAM-gelatin. The apparent elastic modulus of the hydrogel at a concentration of 20 w/v% was 1.2×10⁴Pa, which is nearly equal to that of collagen gel prepared at 0.15w/v%. When a culture medium containing the PNIPAM-gelatin (concentration, 20 w/v%) and bovine smooth muscle cells was incubated at 37°C, the cells were entrapped into a hydrogel. The entrapped cells apparently died in hydrogel with a thickness of 1 mm. However, the use of thinner hydrogel (thickness, 0.1 mm) or comixing with a small amount of PNIPAM-derivatized hyaluronic acid (PNIPAM-HA), even at 1 mm thickness, appeared to increase the survival of entrapped cells.     

利用罗曼鸡肌腱滑液鞘体内构建趾深屈肌腱的生物力学及组织学研究

目的 探索肌腱滑液鞘对肌腱体内再生的作用。 方法 将36只罗曼鸡随机平均分为A、B两组。A组把左趾深屈肌腱滑液鞘的上、下、右侧分离,不切断趾深屈肌腱。同种异体脱细胞肌腱用部分分离的腱滑液鞘包裹,并固定在趾深屈肌腱左侧。B组直接把同种异体脱细胞肌腱固定在去除肌腱滑液鞘的趾深屈肌腱左侧。另取右趾深屈肌腱作为正常对照组。分别在第4、8、12周取材进行趾深屈肌腱最大载荷、弹性模量的生物力学检测及HE染色的组织学检测。结果 术后第4、8、12周时,除第4周A、B组在弹性模量方面差异无统计学意义,其余时间点A组最大载荷和弹性模量都大于B组（P<0.05）。随时间延长,A组胶原纤维增多,排列紧密,方向一致,炎性细胞浸润及纤维组织增生程度较轻。而B组胶原纤维数量逐渐减少、排列逐渐散乱,炎性细胞浸润、纤维组织增生程度明显高于A组。结论腱滑液鞘有助于肌腱再生,说明合适的体内环境对肌腱构建具有重要作用,本研究结果对临床上肌腱缺损病人找到合适肌腱替代物有积极作用。     

失下牙槽神经支配与拔牙窝新骨形成及骨量保持

背景：牙齿拔除后拔牙窝内新骨形成及改建与神经支配密切相关。 目的：探讨失神经支配对拔牙窝新骨形成和骨量保持的可能调节作用。 方法：切除犬一侧下牙槽神经建立失神经支配动物模型,失神经支配侧为实验组,以未切除侧为对照组,组织学染色检测术后2,4,8,12周时间点拔牙窝新骨形成和骨量保持。 结果与结论：实验组拔牙窝骨缺损区的新生骨量面积百分比在术后2,4,8周明显低于对照组(P < 0.01);术后2,4,8,12周,实验组颊舌侧牙槽嵴高度差明显高于对照组(P < 0.05或P < 0.01)。说明失神经支配与拔牙窝新骨形成和骨量保持存在密切相关性。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

在体制备大鼠全肾去细胞支架的程序性分析

目的通过在体灌注Triton X-100、SDS溶液法制备大鼠全肾去细胞生物支架,并对支架制备过程中的关键步骤进行程序性检测、分析,为制备科学合理的实验用大鼠全肾去细胞生物支架提供基础。方法 SD大鼠40只,随机分为4组,每组10只。肝素化后直接切取肾脏作为对照组(control组);肝素PBS灌注组(H组);肝素PBS、Triton X-100灌注组(HT组);肝素PBS、Triton X-100、十二烷基硫酸钠(SDS)溶液灌注组(HTS组)。HE、Masson、PAS染色及透射电镜观察各组肾组织病理及超微结构改变,免疫荧光结合4,6-二脒基-2-苯基吲哚(DAPI)观察各组胶原蛋白Ⅳ(collagenⅣ)、层黏连蛋白(LN)、纤维连接蛋白(FN)、硫酸乙酰肝素蛋白多糖2(HSPG2)、弹性蛋白(elastin)及细胞核的表达情况,总DNA测定各组DNA残留浓度。结果 Triton X-100、SDS灌注6h左右制备大鼠肾脏去细胞生物支架。在灌注过程中,肾内细胞和细胞碎片逐渐被清洗,最终变成半透明状。HE、Masson、PAS染色及透射电镜显示连续分布、形态排列类似肾小球、肾小管轮廓结构的网状结构,基膜连续完整。免疫荧光结合DAPI染色结果表明,去细胞过程中细胞外基质中的重要蛋白collagen IV、LN、FN、HSPG2、elastin都较好的得到了保存,细胞核在灌注过程中逐渐减少,直至完全消失;最终残留组织的DNA为4.90μg/g。结论通过合适浓度和灌注比例的Triton X-100、SDS制备大鼠肾脏去细胞生物支架,并对其进行程序性分析,发现能有效清除大鼠肾内所有细胞成分,较完整的保留网络状肾及肾血管细胞外基质结构和成分,是一种简单易行且较为理想的制备实验用全肾生物支架的方法。     

Pluronic F-127水凝胶复合骨髓间充质干细胞构建组织工程化脂肪的体外研究

本实验应用Pluronic F-127水凝胶三维培养大鼠骨髓间充质干细胞(BMSCs),观察其生物相容性,并研究其对BMSCs诱导分化成脂肪细胞的影响。原代培养SD大鼠BMSCs,经诱导分化成脂、成骨和成神经鉴定后,用MTT法检测Pluronic F-127水凝胶对BMSCs的细胞毒性;将P4BMSCs均匀混合于水凝胶中,复合BMSCs的水凝胶经成胶后分别用成脂诱导液和完全培养基培养,观察并记录脂滴、脂肪细胞及脂肪组织形成情况,培养2周后油红O染色鉴定。结果表明BMSCs在Pluronic F-127水凝胶中能较好的黏附、生长及增殖。成脂诱导的水凝胶中BMSCs可分化为脂肪细胞,其中少部分聚集成脂肪细胞团,形成脂肪样组织,并经油红O染色鉴定证实。而仅加入完全培养基的水凝胶内BMSCs则未发生分化。提示BMSCs可在Pluronic F-127水凝胶三维支架中培养并被成功诱导分化为脂肪细胞,利用水凝胶负载BMSCs向脂肪细胞诱导分化可望为组织工程化脂肪提供新的思路,Plu-ronic F-127水凝胶可作为BMSCs治疗体外培养的良好支架。     

异基因组织工程骨修复猪胫骨缺损术后外周血T淋巴细胞亚群的检测

目的 通过检测异基因组织工程骨移植修复猪胫骨缺损术后T淋巴细胞亚群CD4 和CD8 细胞的变化,探索异基因组织工程骨的免疫原性.方法 骨髓间充质干细胞(MSCs)体外培养,扩增,诱导成骨后与磷酸三钙(TCP)复合为组织工程骨,分自体和同种异体组,植入构建好的猪胫骨中段2.0 cm的缺损处,对照组为纯TCP组,流式检测术前,术后3、7、14、28、56 d外周血T淋巴细胞亚群变化.结果 小型猪外周血T淋巴细胞亚群分别为CD4 (9.37±1.65)%,CD8 (38.43±1.62)%,CD4 CD8 (7.23±1.24)%;术后组内不同时间点CD4 和CD8 细胞测定值无显著变化,CD4 CD8 双阳性细胞在3 d和7 d增加明显(P＜0.05);组间各时间点CD4 、CD8 和CD4 CD8 细胞无显著变化.结论 术后T细胞亚群检测结果显示无明显的免疫排斥,推论异基因组织工程骨免疫原性低.     

自体脂肪干细胞复合胶原海绵多孔支架构建大鼠乳腺脂肪组织的初步研究

目的 探讨自体脂肪干细胞复合胶原海绵多孔支架构建大鼠乳腺脂肪组织的可行性.方法 采用胶原酶消化、离心方法分离并培养6只雌性SD大鼠的脂肪干细胞.对脂肪干细胞进行成骨、成脂诱导及鉴定.将自体脂肪干细胞复合Ⅰ型胶原海绵多孔支架进行体外培养.将自体脂肪干细胞复合Ⅰ型胶原海绵多孔支架移植到大鼠右上侧乳腺(实验组),将Ⅰ型胶原海绵多孔支架移植到大鼠左上侧乳腺(对照组),共移植6例.12周后观测有无新生脂肪形成;测量新生脂肪组织湿质量;组织切片行HE及油红0染色.结果 SD大鼠脂肪干细胞增殖能力强;具有成骨及成脂分化能力;体外培养2周,胶原支架内可见ASC生长.实验组可见乳腺与胸肌之间有脂肪样新生组织形成,平均湿质量为(121±9) mg.对照组见乳腺与胸肌之间有胶原海绵样组织形成,平均湿质量为(77±6) mg,两组间湿质量存在统计学差异,P＜0.05.HE及油红O染色证实,实验组新生组织主要为成熟脂肪组织;而对照组主要为胶原纤维.结论 自体脂肪干细胞复合Ⅰ型胶原海绵多孔支架可在大鼠乳腺区域形成脂肪组织.