藻酸钙凝胶-成骨细胞-骨粉构建工程化骨修复兔颅骨缺损的实验研究

目的探讨藻酸钙凝胶、成骨细胞、骨粉复合构建的可塑形组织工程骨修补兔颅骨缺损后的形态学变化和成骨效果.方法28只日本大耳白兔,随机分为A(n=20)、B(n=8)两组,手术在兔颅顶骨矢状缝两侧分别各建立一个直径1cm的圆形全层缺损,用两种方法藻酸钙凝胶、成骨细胞、骨粉和藻酸钙凝胶、骨粉分别构建组成可塑形的组织工程骨复合材料,分别填补修复A组兔颅骨左右两侧的缺损,B组为空白对照组,通过大体、组织学、X线观察材料的形态变化、成骨情况,并对X线片和组织学切片进行评分.结果材料植入后,局部未见红肿、积液、渗出等异常反应.①藻酸钙凝胶-成骨细胞-骨粉组修补后12周颅骨缺损基本被硬性组织所修复,镜下见修复材料大多被骨组织替代,骨粉基本被吸收,有块状凝胶残留其中,组织学评分为(5.50±1.00).X线片见兔颅骨缺损处有高密度骨痂影存在,布满整个缺损区,X线片评分为(3.25±0.95).②藻酸钙凝胶-骨粉组修补后12周部分颅骨缺损被硬性组织所修复,镜下见修复材料部分转变成骨组织,骨粉基本被吸收,有凝胶残留其中,组织学评分为(3.25±1.50).X线片见兔颅骨缺损处有高密度骨痂影存在,主要分布在缺损区的边缘部位,X线片评分(2.25±0.25).③空白对照组骨缺损主要被膜样纤维组织修复,在紧邻骨缺损边缘处有硬性组织形成,镜下见修复组织边缘有骨组织存在,中央大部为膜状致密纤维组织,组织学评分为(1.50±0.50),X线片仅见靠近骨缺损边缘的部位存在致密骨痂影,X线片评分为(1.00±0.57).结论藻酸钙凝胶、成骨细胞、骨粉构建的可塑形组织工程骨可根据颅骨缺损的形态进行塑形填补,在体内有良好的成骨能力,可达到对兔颅骨缺损的骨性修复,但部分藻酸盐凝胶吸收缓慢,手术后12周仍不能满意吸收.     

红霉素治疗前后下呼吸道绿脓杆菌藻酸盐含量的变化

目的　比较红霉素治疗前后绿脓杆菌菌落中藻酸盐含量的变化 ,了解红霉素对绿脓杆菌生物被膜的作用。方法　利用藻酸盐经强酸处理后生成的糠醛或其衍生物 ,再与酚类、胺类化合物作用生成有色物质的原理 ,采用比色法测定红霉素治疗前后绿脓杆菌菌落中的藻酸盐含量 ,并测定比色法的准确性和重复性。结果　菌落中的藻酸盐含量与反应物的吸光度值 (A)呈正相关 (r =0 .98,P <0 .0 1) ;比色法的回收率为 98.18% ,变异系数小于 5 % ;红霉素治疗前后绿脓杆菌菌落的藻酸盐含量分别为 2 3.5 4和 18.0 2 μg/ml,差异有显著性 (P <0 .0 5 )。结论　比色法的准确性和重复性均较高 ;采用比色法可测得红霉素治疗后绿脓杆菌菌落中的藻酸盐含量降低     

可注射原位交联海藻酸钙骨修复材料小鼠体内异位成骨研究

目的对复合了人重组骨形态发生蛋白2(rhBMP-2)的可注射原位交联海藻酸钙骨修复材料在小鼠体内异位成骨进行评估。方法实验组为含0．1mg rhBMP-2的材料0．1ml,空白对照组为不含rhBMP-2 的材料0．1ml,分别注射到小鼠左后肢大腿肌陷窝中,对照组植入含0．1mg rh BMP-2的骨优导,21天后, 影像学检查成骨情况,解剖称骨湿重,同时在注射后7天,14天,21天解剖实验组动物各三只,取注射部位进行HE染色组织学检查。结果 21天影像学检查显示实验组小鼠左后肢肌陷窝处有骨痂生成,解剖取异位成骨称湿重实验组为239．2±59．7mg,对照组为225．5±56．9mg,无显著性差异,2周后组织学检查有大量骨髓细胞及骨小梁生成。结论该材料有良好的诱导新骨生成的能力。     

新型mucA基因突变的铜绿假单胞菌生物被膜的形成和藻酸盐相关基因表达的研究

目的 确定黏液型铜绿假单胞菌PA17的mum基因突变位点，研究藻酸盐合成相关基因在其生物被膜形成过程中的表达，并观察PAl7生物被膜形成过程和形态。方法 PCR方法扩增铜绿假单胞菌PA17的mueA基因全长并测序；改良平板培养法建立PA17的生物被膜模型，半定量RT-PCR测定生物被膜形成24h、3d．6d时藻酸盐合成相关基因,algD、algU和algR的表达，并进行统计学分析；扫描电镜观察不同时间点的生物被膜形态。结果 PA17的mucA基因第166～333位核苷酸片段缺失，第342位A→G；其藻酸盐相关基因algD和algU均在生物被膜形成过程的第6天表达水平最高，algR在24h表达最高，单因素方差分析显示，上述基因在生物被膜形成过程不同时间点表达的差异有统计学意义；PA17于第6天形成成熟生物被膜，形态为薄膜状。结论 PA17是一株含新型mucA突变基因的黏液型铜绿假单胞菌，其藻酸盐相关基因在生物被膜形成的不同时间点的表达差异具有统计学意义，其生物被膜形态为薄膜状。     

可注射矿化胶原水凝胶修复大鼠颅骨临界性骨缺损的研究

目的 基于矿化胶原（MC）与海藻酸盐（ALG）,制备一种新型可注射矿化胶原水凝胶（MCA）,研究MCA对颅骨临界性骨缺损的修复作用。方法 将MC粉末以200 mg/mL的比例加入到ALG（C_ALG=15 mg/mL）中制备出矿化胶原水凝胶（MCA）。采用扫描电子显微镜对MCA的交联状态和元素分析情况进行观察。采用L929细胞增殖实验评估MCA的生物安全性,体内降解实验观察MCA降解性和生物相容性。通过大鼠颅骨临界骨缺损模型来验证MCA的骨修复效果。结果 MCA水凝胶经过扫描电子显微镜（SEM）观察后发现水凝胶表面粗糙,其内部Ca/P信号均匀且密集。为了评估MCA的生物安全性,使用MCA浸提液与L929细胞共同孵育后,发现200 mg/mL浸提液实验组的L929细胞增值率为99.95%,毒性评级为1级。同时,将MCA水凝胶注射至大鼠皮下,术后60 d注射部位仍见残留的MCA植入物,而在MCA水凝胶注射到颅骨缺损处的第60天时,MCA则全部降解。最后,观察MCA在颅骨临界骨缺损动物模型中的骨缺损修复效果,实验结果表明MCA水凝胶组的颅骨缺损修复效果优于ALG水凝胶组和对照组。结论 MCA水凝胶具有良好的生物安全性,降解性和促进骨再生的能力,是一种安全可靠的骨修复材料。     

天然多糖材料微囊肾动脉栓塞对肾功能和形态的影响

目的: 研究海藻酸钠-壳聚糖微囊肾动脉栓塞对肾功能和形态的影响.方法: 以天然多糖材料海藻酸钠与壳聚糖制备的海藻酸钠-壳聚糖微囊(ACM)为栓塞剂,采用经导管介入治疗技术进行兔肾动脉栓塞,于术前及术后1,2,4及8周检测血常规、肾功能及肾脏病理变化.结果: 手术前后,动物的血常规各项指标均在正常范围内波动;术后初期肾功能有所下降,1周后基本恢复正常.从血管造影与病理组织学检查结果看,健侧肾未发生任何变化,栓侧肾随栓塞时间延长出现大片弥漫性凝固性坏死,体积与重量不断减小,未出现血管再通且无侧支循环形成.结论: ACM具有良好的血管栓塞作用,使用方便,是一种较理想的血管栓塞剂.     

Pharmacokinetics,biocompatibility and bioavailability of a controlled release monoclonal antibody formulation

The sustained and localized delivery of monoclonal antibodies has become highly relevant, because of the increasing number of investigated local delivery applications in recent years. As the local delivery of antibodies is associated with high technological hurdles, very few successful approaches have been reported in the literature so far. Alginate-based delivery systems were previously described as promising sustained release formulations for monoclonal antibodies (mAbs). In order to further investigate their applicability, a single-dose animal study was conducted to compare the biocompatibility, the pharmacokinetics and the bioavailability of a human monoclonal antibody liquid formulation with two alginate-based sustained delivery systems after subcutaneous administration in rats. 28 days after injection, the depot systems were still found in the subcutis of the animals. A calcium cross-linked alginate formulation, which was injected as a hydrogel, was present as multiple compartments separated by subcutaneous tissue. An in situ forming alginate formulation was recovered as a single compact and cohesive structure. It can be assumed that the multiple compartments of the hydrogel formulation led to almost identical pharmacokinetic profiles for all tested animals, whereas the compact nature of the in situ forming system resulted in large interindividual variations in pharmacokinetics. As compared to the liquid formulation the hydrogel formulations led to lower mAb serum levels, and the in situ forming system to a shift in the time to reach the maximum mAb serum concentration (T_max) from 2 to 4 days. Importantly, it was shown that after 28 days only marginal amounts of residual mAb were present in the alginate matrix and in the tissue at the injection site indicating nearly complete release. In line with this finding, systemic drug bioavailability was not affected by using the controlled release systems. This study successfully demonstrates the suitability and underlines the potential of polyanionic systems for local and controlled mAb delivery.     

Non-invasive endotracheal delivery of paclitaxel-loaded alginate microparticles

Aerosolized chemotherapeutics leads to higher, localized and continuous concentrations of active agents in lung tissue with lower side effects for other organs. The present study was performed on jugular vein cannulated rats which endothracheally received 4 mg/kg of free paclitaxel powder (Free-PTX), paclitaxel-loaded alginate microparticles (PTX-ALG-MPs) and i.v. paclitaxel (Anzatax^®). Pharmacokinetic parameters for Free-PTX and PTX-ALG-MPs contain higher AUC, mean residence time (MRT),half-life and bioavailability, with lower elimination constant (k_e). Statistical analysis showed that the amount of paclitaxel per gram of lung tissue after 0.5, 6 and 24 h after administration of Free-PTX was lower than PTX-ALG-MPs. Lung tissue AUC for Free-PTX was lower than PTX-ALG-MPs. According to the obvious advantages obtained, such as dose lowering and increasing paclitaxel residence time and half-life. It should be noted that cell cytotoxicity test on normal airway cell lines was not examined in this study but due to previous reports on safety of inhaled paclitaxel, it can be suggested that pulmonary delivery of paclitaxel can be a useful non-invasive route of administration compared with i.v administration.     

Homeostasis maintenance of encapsulated cells

Cell niche homeostasis plays a critical role in many bodily functions including tissue functionality, stem cell maintenance and differentiation, wound healing, cancer development and propagation, and many others. Many tissue engineering approaches overlook the importance of engineered constructs homeostasis, in particular for transplantation purposes. Here, we present a study of the effect of encapsulation duration on engineered tissue maturation and provide a protocol for the evaluation of critical conditions for transplantation purposes. In brief, SHSY5Y human neuroblastoma cells were encapsulated in 2% alginate by electrohydrodynamic jetting method for up to 4 weeks. We evaluated extracellular matrix niche formation and tissue maturation in situ through COL1A1 expression. In in vitro conditions, we studied the ability of cells to maintain their critical functions after being released from alginate beads. Cellular viability was evaluated via an apoptosis/necrosis detection kit and AlamarBlue assay, and functionality via immunocytochemistry. We proved the importance of engineered tissue homeostasis stabilization for future cell recovery, in particular, for our system cells encapsulated for 28 days met all critical requirements for successful tissue transplantation. Maturation of engineered tissue constructs could be accelerated by enriching alginate with growth factors or extracellular matrix molecules.     

降脂通脉胶囊联合藻酸双酯钠治疗高脂血症的临床研究

目的探讨降脂通脉胶囊联合藻酸双酯钠片治疗高脂血症的临床疗效。方法选择2018年3月-2019年3月在陕西省荣誉军人康复医院治疗的高脂血症患者86例,根据用药的差别分为对照组(43例)和治疗组(43例)。对照组口服藻酸双酯钠片,50mg/次,3次/d;治疗组在对照组基础上口服降脂通脉胶囊,2.0g/次,3次/d。两组患者经4周治疗。观察两组患者临床疗效,同时比较治疗前后两组患者高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)、三酰甘油(TG)、胆固醇(TC)、嗜酸性粒细胞阳离子蛋白(ECP)、摄食抑制因子-1(NSF-1)、硫氧还蛋白相互作用蛋白(TXNIP)、肿瘤坏死因子-α(TFN-α)、白细胞介素-1β(IL-1β)、脂联素(ADP)、超氧化物歧化酶(SOD)、丙二醛(MDA)和谷胱甘肽过氧化物酶(GSH-Px)水平。结果治疗后,对照组临床有效率为81.40%,显著低于治疗组的97.67%,两组比较差异有统计学意义(P<0.05)。治疗后,两组患者TC、TG、LDL-C、ECP、IL-1β、TFN-α、TXNIP、NSF-1和MDA水平均显著降低(P<0.05),而HDL-C、ADP、SOD和GSH-Px水平均显著升高(P<0.05),且治疗组患者这些指标明显好于对照组(P<0.05)。结论降脂通脉胶囊联合藻酸双酯钠片治疗高脂血症可有效降低血脂指标,促进机体细胞因子和氧化应激指标的改善,具有一定的临床推广应用价值。