Functional recovery of denervated muscle by neurotization using nerve guidance channels

Tissue‐engineered muscle has been proposed as a means of repairing volumetric muscle defects to restore anatomical and functional recovery. We have previously demonstrated that denervated muscle, which is analogous to engineered muscle construct, can be reinnervated by direct transplantation of host nerve (neurotization) in a rat model. However, the use of this approach is not possible if the length of host nerve is inadequate and cannot be mobilized to the insertion site of the engineered muscle. In this study we investigated whether neurotization coupled with nerve guidance channels would increase the regeneration of neuromuscular junctions (NMJs) in completely denervated muscle and encourage neurofunctional recovery. Seventy‐two Lewis rats were evaluated in three groups, a normal control group (n = 8), a denervated group (n = 32) and a neurotization coupled with nerve guidance group (n = 32). Neurofunctional behaviour and histological evaluations were performed at 4, 8, 12 and 20 weeks postoperatively. Extensor postural thrust (EPT) and compound muscle action potential (CMAP) amplitude were significantly improved in the nerve guidance group when compared with the denervated group, even though these values were different from those of the normal control group at 20 weeks postoperation. Regeneration of axons and NMJs was demonstrated histologically in the nerve guidance group. Neurotization coupled with nerve guidance channels leads to regeneration of axons and NMJs in completely denervated muscle. To our knowledge, this is the first report to show that nerve guidance can allow re‐innervation in denervated muscle containing long‐gap nerve injuries. Copyright © 2013 John Wiley & Sons, Ltd.     

PTEN loss and KRAS activation leads to the formation of serrated adenomas and metastatic carcinoma in the mouse intestine

Mutation or loss of the genes PTEN and KRAS have been implicated in human colorectal cancer (CRC), and have been shown to co‐occur despite both playing a role in the PI3' kinase (PI3'K) pathway. We investigated the role of these genes in intestinal tumour progression in vivo, using genetically engineered mouse models, with the aim of generating more representative models of human CRC. Intestinal‐specific deletion of Pten and activation of an oncogenic allele of Kras was induced in wild‐type (WT) mice and mice with a predisposition to adenoma development (Apc^fl/+). The animals were euthanized when they became symptomatic of a high tumour burden. Histopathological examination of the tissues was carried out, and immunohistochemistry used to characterize signalling pathway activation. Mutation of Pten and Kras resulted in a significant life‐span reduction of mice predisposed to adenomas. Invasive adenocarcinoma was observed in these animals, with evidence of activation of the PI3'K pathway but no metastasis. However, mutation of Pten and Kras in WT animals not predisposed to adenomas led to perturbed homeostasis of the intestinal epithelium and the development of hyperplastic polyps, dysplastic sessile serrated adenomas and metastasizing adenocarcinomas with serrated features. These studies demonstrate synergism between Pten and Kras mutations in intestinal tumour progression, in an autochthonous and immunocompetent murine model, with potential application to preclinical drug testing. In particular, they show that Pten and Kras mutations alone predispose mice to the spectrum of serrated lesions that reflect the serrated pathway of CRC progression in humans. Published by John Wiley & Sons, Ltd. www.pathsoc.org.uk     

海洋蓝藻 Okeania hirsute 中的次级代谢产物及其细胞毒活性

目的 研究从海洋蓝藻 Okeania hirsute 中分离鉴定次级代谢产物并进行细胞毒活性评价。方法 三个天然产物(1~3)的结构确定是通过广泛的波谱数据,包括一维和二维核磁及质谱等。结果 化合物3对人大细胞肺癌细胞NCI-H460显示出强的细胞毒活性,IC₅₀值为0.384 μg/mL。结论 这是首次从Okeania属蓝细菌中分离获得化合物1~3。这些天然产物的发现表明Okeania属蓝细菌可以作为发现新颖化合物的重要来源。     

不同比例软骨细胞与间充质干细胞对构建组织工程软骨的影响

目的以不同比例将大鼠软骨细胞及骨髓间充质干细胞（bonemarrowstromalceu,BMSCs）依次体外及体内混合培养,探讨二者构建组织工程软骨的最佳比例。方法分别取1个月及1d龄SD大鼠的BMSCs及关节软骨,原代软骨细胞与第2代BMSCs混合比例为全软骨细胞组、3：1、1：1、1：3、全BMSCs组,共五组。以终浓度4×10^7／mI．种于聚乳酸一聚羟基乙酸支架上,体外培养4周后,植入裸鼠皮下继续培养,8周后对标本进行大体观察、组织学切片、Ⅱ型胶原免疫荧光染色及检测氨基糖胺多糖的含量。结果软骨细胞与BMSCs混合比例3：1组较其他各组所构建出的组织工程软骨体积大,厚度厚,有光泽;组织染色见软骨细胞增殖较旺盛,被大量细胞外基质包围,分布均匀,并可见软骨陷窝;Ⅱ型胶原免疫荧光染色见细胞内和细胞外基质发出的红色荧光较明亮;3：1组的组织块氨基糖胺多糖含量（470．38±29．78）μg高于其他各组（P〈O．05）。结论软骨细胞与BMSCs混合培养有助于节省软骨细胞,二者混合浓度比例以3：1最好。     

骨髓间充质干细胞体外构建组织工程化半月板

目的探索以骨髓间充质干细胞（BMSCs）为种子细胞,在体外构建具有完整内侧半月板形态的软骨样组织的方法。方法运用模具制备内侧半月板形的PGA/PLA支架。抽取犬骨髓,分离培养BMSCs,将其接种于支架材料上,5 d后使用软骨诱导液培养。体外培养6周后,行大体观察、组织学检测及生物力学检测。结果细胞材料复合物能够较好地维持半月板三维立体结构,形成了表面光滑、触之有弹性的瓷白色软骨样组织。 HE染色可见典型的软骨陷窝出现,说明成熟软骨组织的形成。Safranine O染色证实有蛋白聚糖基质产生。生物力学检测显示,新生组织弹性模量达正常半月板组织的12.7%。结论 BMSCs通过体外诱导,可在体外分化为较成熟的软骨组织,并构建出组织工程化半月板。     

靶向抑制巨噬细胞Act1表达对溃疡性结肠炎的作用

目的 探讨巨噬细胞NF-κB活化因子1（nuclear factor kappa B activator 1,Act1）在炎症性肠病中的作用。方法 以巨噬细胞Act1表达被靶向抑制的小鼠（anti-Act1）为研究对象,利用葡聚糖硫酸钠（dextran sodium sulfate,DSS）诱导溃疡性结肠炎模型,通过检测小鼠体重变化、便血情况、粪便黏稠度等进行疾病活动指数评分,对结直肠组织进行HE染色评价炎症严重程度,免疫组化方法分析白细胞和巨噬细胞等的浸润情况并用RT-qPCR法检测相关细胞因子的表达变化。结果 在DSS诱导7 d后与C57小鼠相比,anti-Act1小鼠结直肠炎症指标评分较低,巨噬细胞等浸润数量较少,TNF-α、IL-1β和IL-6 mRNA表达较低。结论 靶向抑制巨噬细胞Act1表达能够减轻DSS诱导的溃疡性结肠炎。     

Performance Test and Microstructure of Modified PVC Aggregate-Hybrid Fiber Reinforced Engineering Cementitious Composite (ECC)

This study aims to solve the problems of the high cost, heavy pollution and poor performance of traditional engineered cementitious composites (ECC) by adding modified Polyvinyl chloride (PVC) aggregate, Polypropylene (PP)–Polyvinyl alcohol (PVA) hybrid fiber and large amount of fly ash. The PVC aggregate is modified by pre-coating silica fume with a PP fiber volume content of 0.5%, PVA fiber volume contents of 1%, 1.5%, and 2%, PVC aggregate contents of 10%, 20%, and 30%, and fly ash volume content of 69%. Different properties and microstructures were studied by carrying out cube compression tests, splitting tensile tests, water absorption tests, drop hammer impact tests, scanning electron microscopy and nuclear magnetic resonance tests. According to the test results, under the same content of PVC aggregate, the use of modified PVC aggregate can, not only effectively avoid the decrease in strength and increase of water absorption, but also improve brittleness and impact failure energy. Regardless of the kind and content of fiber, the compressive strength and brittleness will decrease, while the splitting tensile strength, water absorption, and impact failure energy will increase. After adding 0.5% PP and 1.5% PVA fiber, the performance is ordinary and a negative mixing effect occurs. As more modified PVC aggregate is added, the strength of the ECC concrete with PP–PVA hybrid fiber and modified PVC aggregate added slowly decrease, while the water absorption and impact failure energy increase. Based on a comprehensive analysis of the test data, the reinforcement method of adding 1.5% PVA-0.5% PP hybrid fiber-30% modified PVC aggregate is superior to adding 1.5% PVA fiber, but slightly inferior to adding 2% PVA fiber. This study argues that the reinforcement method is of great significance for the promotion and application of ECC.     

Involvement of Nax sodium channel in peripheral nerve regeneration via lactate signaling

Na_x, a sodium concentration‐sensitive sodium channel, is expressed in non‐myelinating Schwann cells of the adult peripheral nervous system, but the pathophysiological role remains unclear. We found that functional recovery of the hind paw responses from the sciatic nerve transection was delayed in Na_x knockout ( ) mice. Histological analyses showed a decrease in the number of regenerated myelinated axons in sciatic nerves. The delay in the recovery in mice was improved by lactate and inhibited by a monocarboxylate transporter inhibitor. In vitro experiments using cultured Schwann cells showed that lactate release was enhanced by endothelin (ET)‐1 and blocked by an ET receptor type B antagonist. Here, it is conceivable that Na_x was activated by ET‐1. The amount of lactate release by ET‐1 was lower in mice than in wild‐type mice. These results indicated that Na_x is functionally coupled to ET for lactate release via ET receptor type B and is involved in peripheral nerve regeneration.     

Neural tissue engineering options for peripheral nerve regeneration

Tissue engineered nerve grafts (TENGs) have emerged as a potential alternative to autologous nerve grafts, the gold standard for peripheral nerve repair. Typically, TENGs are composed of a biomaterial-based template that incorporates biochemical cues. A number of TENGs have been used experimentally to bridge long peripheral nerve gaps in various animal models, where the desired outcome is nerve tissue regeneration and functional recovery. So far, the translation of TENGs to the clinic for use in humans has met with a certain degree of success. In order to optimize the TENG design and further approach the matching of TENGs with autologous nerve grafts, many new cues, beyond the traditional ones, will have to be integrated into TENGs. Furthermore, there is a strong requirement for monitoring the real-time dynamic information related to the construction of TENGs. The aim of this opinion paper is to specifically and critically describe the latest advances in the field of neural tissue engineering for peripheral nerve regeneration. Here we delineate new attempts in the design of template (or scaffold) materials, especially in the context of biocompatibility, the choice and handling of support cells, and growth factor release systems. We further discuss the significance of RNAi for peripheral nerve regeneration, anticipate the potential application of RNAi reagents for TENGs, and speculate on the possible contributions of additional elements, including angiogenesis, electrical stimulation, molecular inflammatory mediators, bioactive peptides, antioxidant reagents, and cultured biological constructs, to TENGs. Finally, we consider that a diverse array of physicochemical and biological cues must be orchestrated within a TENG to create a self-consistent coordinated system with a close proximity to the regenerative microenvironment of the peripheral nervous system.