Diaphragmatic muscle reconstruction with an aligned electrospun poly(ε-caprolactone)/collagen hybrid scaffold

Large diaphragmatic muscle defects in congenital diaphragmatic hernia (CDH) are reconstructed by prosthetic materials or autologous grafts, which are associated with high complications and reherniation. In this study we examined the feasibility of using aligned electrospun poly(ε-caprolactone) (PCL)/collagen hybrid scaffolds for diaphragmatic muscle reconstruction. The hybrid scaffolds were implanted into a central left hemi-diaphragmatic defect (approximately 70% of the diaphragmatic tissue on the left side) in rats. Radiographic and magnetic resonance imaging (MRI) analyses showed no evidence of herniation or retraction up to 6 months after implantation. Histological and immunohistochemical evaluations revealed ingrowth of muscle tissue into the scaffolds. The mechanical properties of the retrieved diaphragmatic scaffolds were similar to those of normal diaphragm at the designated time points. Our results show that the aligned electrospun hybrid scaffolds allowed muscle cell migration and tissue formation. The aligned scaffolds may provide implantable functional muscle tissues for patients with diaphragmatic muscle defects.     

Biomimetic l-aspartic acid-derived functional poly(ester amide)s for vascular tissue engineering

Functionalization of polymeric biomaterials permits the conjugation of cell signaling molecules capable of directing cell function. In this study, l-phenylalanine and l-aspartic acid were used to synthesize poly(ester amide)s (PEAs) with pendant carboxylic acid groups through an interfacial polycondensation approach. Human coronary artery smooth muscle cell (HCASMC) attachment, spreading and proliferation was observed on all PEA films. Vinculin expression at the cell periphery suggested that HCASMCs formed focal adhesions on the functional PEAs, while the absence of smooth muscle α-actin (SMαA) expression implied the cells adopted a proliferative phenotype. The PEAs were also electrospun to yield nanoscale three-dimensional (3-D) scaffolds with average fiber diameters ranging from 130 to 294 nm. Immunoblotting studies suggested a potential increase in SMαA and calponin expression from HCASMCs cultured on 3-D fibrous scaffolds when compared to 2-D films. X-ray photoelectron spectroscopy and immunofluorescence demonstrated the conjugation of transforming growth factor-β1 to the surface of the functional PEA through the pendant carboxylic acid groups. Taken together, this study demonstrates that PEAs containing aspartic acid are viable biomaterials for further investigation in vascular tissue engineering.     

Small diameter vascular graft with fibroblast cells and electrospun poly (L-lactide-co-ε-caprolactone) scaffolds: Cell Matrix Engineering

Abstract

Electrospun scaffolds have been widely used in tissue engineering due to their similar structure to native extracellular matrices (ECM). However, one of the obstacles limiting the application of electrospun scaffolds for tissue engineering is the nano-sized pores, which inhibit cell infiltration into the scaffolds. To overcome this limitation, we approached to make layers which are consisted of cells onto the electrospun sheet and then tubular structure was constructed by rolling. We called this as ‘Cell Matrix Engineering’ because the electrospun sheets were combined with the cells to form one matrix. They maintained 3-D tubular structures well and their diameters were 4.1 mm (±0.1 mm). We compared the mechanical and biological properties of various vascular grafts with the electrospun PLCL sheets of different thickness. In these experiments, the vascular graft made with thin sheets showed a better cell proliferation and attachment than the grafts made with thick sheets because the thin layer allowed for more efficient mass transfer and better permeability than the thick layer. Culturing under physiological pulsatile flow condition was demonstrated in this work. These dynamic conditions provided the improved mass transport and aerobic cell metabolism. Therefore, the Cell Matrix Engineered vascular graft holds a great promise for clinical applications by overcoming the limitations associated with conventional scaffolds.     

LLA/GA/ε-CL摩尔比对三元共聚物性能的影响

辛酸亚锡为催化剂,采用本体开环聚合方法,合成了一组不同摩尔比的L-丙交酯/乙交酯/ε-己内酯(LLA/GA/CL)三元共聚物。通过GPC、1H-NMR、DSC和XRD测试,研究了摩尔比对共聚物分子量及其分布、热性能和结晶性能的影响。结果表明共聚物组成摩尔比与投料摩尔比基本一致,共聚物玻璃化转变温度(Tg)随LLA和CL含量增加分别呈现升高和降低趋势,并且当LLA含量增加至77 mol%,即LLA/GA摩尔比85∶15时,共聚物表现出明显的部分结晶性。     

Controlling the migration behaviors of vascular smooth muscle cells by methoxy poly(ethylene glycol) brushes of different molecular weight and density

Cell migration is an important biological activity. Regulating the migration of vascular smooth muscle cells (VSMCs) is critical in tissue engineering and therapy of cardiovascular disease. In this work, methoxy poly(ethylene glycol) (mPEG) brushes of different molecular weight (Mw 2 kDa, 5 kDa and 10 kDa) and grafting mass (0-859 ng/cm²) were prepared on aldehyde-activated glass slides, and were characterized by X-ray photoelectron spectrometer (XPS) and quartz crystal microbalance with dissipation (QCM-d). Adhesion and migration processes of VSMCs were studied as a function of different mPEG Mw and grafting density. We found that these events were mainly regulated by the grafting mass of mPEG regardless of mPEG Mw and grafting density. The VSMCs migrated on the surfaces randomly without a preferential direction. Their migration rates increased initially and then decreased along with the increase of mPEG grafting mass. The fastest rates (∼24 μm/h) appeared on the mPEG brushes with grafting mass of 300-500 ng/cm² depending on the Mw. Cell adhesion strength, arrangement of cytoskeleton, and gene and protein expression levels of adhesion related proteins were studied to unveil the intrinsic mechanism. It was found that the cell-substrate interaction controlled the cell mobility, and the highest migration rate was achieved on the surfaces with appropriate adhesion force.     

Adsorption of albumin,collagen, and fibronectin on the surface of poly(hydroxybutyrate-hydroxyvalerate) (PHB/HV) and of poly(ε-caprolactone) (PCL) films modified by an alkaline hydrolysis and of poly(ethylene terephtalate) (PET) track-etched membranes

The effect of alkaline hydrolysis on several surface properties of poly(hydroxybutyratehydroxyvalerate) (92/8) (PHB/HV) and poly(ε-caprolactone) (PCL) films and of poly(ethylene terephtalate) (PET) track-etched membranes have been characterized, as well as the adsorption of three proteins normally encountered by mammalian cells in vivo, namely albumin, collagen, and fibronectin. The water contact angle decreases and the number of -COOH functions accessible to a chemical reaction at the surface of PCL increases with alkaline hydrolysis. Analysis by atomic force microscopy pictures reveals a change in surface morphology. The modifications of surface properties are correlated with a two times increase of the adsorption of three radiolabelled proteins. The hydrolysis results in a slight increase in the water contact angle of one face of the PHB/HV film and a sharp increase in the number of -COOH functions. Important morphology changes are also induced. The adsorption of the radiolabelled proteins is almost 100 times higher on the hydrolyzed polymer than on the native surface. The increase in hydrophilicity of different PET batches correlates to an increase in the number of -COOH functions. Nevertheless, the surface chemical composition and rugosity are constant and no significant difference in the amount of radiolabelled fibronectin adsorbed on the different surfaces is detectable. In conclusion, the effect of hydrolysis on the surface properties of each of the polyesters studied as well as the proteins adsorption on the different surfaces are different. The results strongly support the hypothesis that, in the system studied, parameters other than hydrophilicity influence protein adsorp     

紫杉醇聚合物纳米囊泡的制备和体外释放研究

目的 以聚己内酯-b-聚乙二醇-6-聚己内酯（PCEP）两亲性三嵌段共聚物为载体研制紫杉醇聚合物纳米囊泡.方法 以不同分子量的聚乙二醇（PEG）引发合成不同亲水段、疏水段链长的PCEP并进行FT-IR、1H NMR和GPC表征,以合成的嵌段聚合物PCEP为载体,通过薄膜-超声分散法制备紫杉醇聚合物纳米囊泡,用透射电子显微镜（TEM）表征其形态和构造,用粒度分析仪测定其粒径及分布,用高效液相色谱（HPLC）法测定其载药量及包封率,用透析袋法研究药物体外释放;同时,研究不同亲水链长、疏水链长对紫杉醇聚合物囊泡载药量、包封率、粒径及体外释放紫杉醇药物的影响.结果 研制的紫杉醇聚合物囊泡呈核-壳结构球形,粒径为纳米级,随着PCEP共聚物相对分子质量的增加而增大;紫杉醇聚合物囊泡体外释放无突释现象,能稳定缓慢释放紫杉醇,且释放速率随共聚物中亲水段PEG含量增加而增大,随疏水段PCL含量增大而减小.结论 以PCEP两亲性三嵌段共聚物为载体制备的紫杉醇聚合物纳米囊泡,其粒径小且分布均匀,包封率较高,有望成为一种用于提高紫杉醇的药效且降低不良反应的新的紫杉醇缓控释剂型.     

Rab28相关信号通路在低切应力诱导 血管平滑肌细胞迁移中的作用

目的 研究低切应力（low shear stress, LowSS）诱导的血管平滑肌细胞（vascular smooth muscle cells, VSMCs）迁移功能异常在动脉粥样硬化血管重建病理过程中的作用及其分子机制。 方法 应用双向凝胶电泳结合质谱分析的差异蛋白质组学方法,研究1.5 Pa正常切应力（normal shear stress, NSS）与0.5 Pa LowSS条件下培养血管组织的蛋白质差异表达谱。应用血管内皮细胞（endothelial cells, ECs）与VSMCs联合培养的平行平板流动腔系统,分别施加NSS和LowSS,Western blot检测ECs与VSMCs的Rab28表达水平以及VSMCs的磷酸化ERK表达水平;Transwell法检测VSMCs的迁移能力;RNA干扰和PD98059分别特异性抑制VSMCs的Rab28表达和ERK磷酸化,再观察VSMCs迁移能力变化。结果 血管差异蛋白质组学的结果发现,与NSS组相比,Rab28在LowSS组血管组织的表达水平明显升高。细胞实验结果显示,LowSS加载明显上调VSMCs的Rab28蛋白表达、VSMCs迁移和ERK磷酸化。静态条件下RNA干扰抑制单独培养VSMCs的Rab28表达,VSMCs迁移能力明显降低,但ERK磷酸化水平无明显变化;应用PD98059特异性抑制VSMCs的ERK磷酸化,VSMCs迁移能力和Rab28表达水平均明显降低。结论 LowSS可能通过上调VSMCs的ERK磷酸化水平引起Rab28表达水平增加,从而诱导VSMCs迁移。探讨Rab28及其相关信号通路在切应力调控VSMCs功能中的作用及其机制,可能为深入理解动脉粥样硬化血管重建疾病发病机制和寻找新的药物治疗靶点提供力学生物学依据。     

紫杉醇聚己内酯／泊洛沙姆188共混载药微球及其抗肿瘤活性

目的本研究首次尝试利用聚己内酯（PCL）与亲水性添加剂泊洛沙姆188（Pluronic F68,F68）共混物作为载体材料与抗癌药物紫杉醇组成微球缓释载药系统。方法采用乳化,溶剂挥发法制备紫杉醇PCL/F68共混微球;考察紫杉醇PCL／F68共混微球的表面形态、平均粒径、包埋率及体外释放性能：利用DSC法分析紫杉醇在PCL／F68共混徽球中的分散状态;考察紫杉醇PCL/F68共混微球在小鼠肝癌H22腹水瘤模型中的抗肿瘤活性。结果表明载体材料中的亲水性添加剂F68可在微球表面形成孔状结构,F68的加入提高了紫杉醇从PCL／F68共混载药微球的释放并获得了接近恒定的释放性能;在小鼠肝癌H22腹水瘤模型中。紫杉醇PCL／F68共混载药微球对肿瘤生长具有抑制作用,荷瘤小鼠生存期明显延长。结论以PCL/F68共混物为载体制备的紫杉醇控释微球具有较高的释放能力和明显的控释效果．     

在高血压动脉重建中microRNA-21对血管平滑肌细胞细胞外基质表达的调控作用

目的探讨在高血压动脉重建中microRNA-21(miR-21)对血管平滑肌细胞(vascular smooth muscle cells,VSMCs)细胞外基质(extracellular matrix,ECM)的调控作用及其机制。方法建立腹主动脉缩窄型大鼠高血压模型,大鼠分为假手术对照组、高血压2周组和高血压4周组;对体外培养的大鼠主动脉VSMCs施加频率为1.25 Hz周期性张应变,加载幅度分别为0%(静态对照组)、5%(正常张应变组)、15%(模拟高血压状态的高张应变组),加载持续时间均为12 h。采用Western blotting和Real time RT-PCR技术,分别检测动脉和细胞样品ECM以及miR-21的表达。用miR-21特异干扰片段抑制培养的VSMCs miR-21表达,然后检测VSMCs的ECM、miR-21和Smad 7表达变化。结果与假手术对照组相比,高血压2周组胸主动脉ECM和miR-21的表达显著上升;高血压4周组胸主动脉的I型胶原、III型胶原和miR-21表达显著上升。与静态对照组和5%张应变组相比,15%张应变组VSMCs的I型胶原表达无显著变化,而III型胶原表达显著升高,Smad 7表达显著下降,周期性张应变增强VSMCs的miR-21表达。干扰miR-21降低周期性张应变状态下VSMCs的miR-21表达以及III型胶原蛋白水平表达,上调VSMCs的Smad 7表达。结论高血压血管重建导致大鼠胸主动脉ECM和miR-21高表达。周期性高张应变可诱导VSMCs的miR-21高表达,再通过其调节Smad 7蛋白,进而调控VSMCs的ECM,尤其是III型胶原的表达,参与高血压血管重建。     

microRNA-34a在低切应力诱导血管平滑肌细胞增殖中的作用

目的探讨micro RNA-34a(mi R-34a)在低切应力诱导血管平滑肌细胞(vascular smooth muscle cells,VSMCs)增殖中的作用。方法应用细胞联合培养平行平板流动腔系统对与VSMCs联合培养的内皮细胞(endothelial cells,ECs)施加1.5 Pa正常切应力(normal shear stress,NSS)和0.5 Pa低切应力(low shear stress,Low SS),加载时间为12 h。Western blotting检测联合培养VSMCs的增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)蛋白表达,以此反映VSMCs的增殖能力。实时PCR检测联合培养VSMCs的mi R-34a表达变化。通过Target Scan、mi RWalk等网站预测mi R-34a的下游靶蛋白。Western blotting检测联合培养VSMCs的mi R-34a靶蛋白Forkhead转录因子J2(forkhead box j2,Foxj2)表达。通过mimics和inhibitor转染技术,分别上调和抑制VSMCs的mi R-34a表达,Western blotting检测Foxj2及PCNA的表达变化,验证mi R-34a和Foxj2之间的调控关系。结果与NSS相比,Low SS促进联合培养VSMCs的PCNA表达,并显著上调联合培养VSMCs的mi R-34a表达。通过Target Scan、mi RWalk等网站预测mi R-34a的下游靶蛋白为Foxj2。Low SS加载下联合培养VSMCs的mi R-34a靶蛋白Foxj2表达明显降低。静态条件下上调VSMCs的mi R-34a表达,靶蛋白Foxj2表达明显降低,PCNA表达显著升高;抑制VSMCs的mi R-34a表达,靶蛋白Foxj2表达明显上调,PCNA表达显著降低。结论 Low SS通过调控联合培养VSMCs的mi R-34a和靶蛋白Foxj2,促进VSMCs增殖。研究结果为进一步阐明动脉粥样硬化疾病的发病机制及药物治疗靶标提供新的力学生物学实验依据。     

Mussel-inspired protein-mediated surface functionalization of electrospun nanofibers for pH-responsive drug delivery

pH-responsive drug delivery systems could mediate drug releasing rate by changing the pH values at specific times as per the pathophysiological need of the disease. This paper demonstrates that a mussel-inspired protein polydopamine coating can tune the loading and releasing rate of charged molecules from electrospun poly(ε-caprolactone) (PCL) nanofibers in solutions with different pH values. In vitro release profiles show that the positive charged molecules release significantly faster in acidic than those in neutral and basic environments within the same incubation time. The results of fluorescein diacetate staining and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays show the viability of cancer cells after treatment with doxorubicin-released media at different pH values qualitatively and quantitatively, indicating that the media containing doxorubicin that were released in solutions at low pH values could kill a significantly higher number of cells than those released in solutions at high pH values. Together, the pH-responsive drug delivery systems based on polydopamine-coated PCL nanofibers could have potential application in the oral delivery of anticancer drugs for treating gastric cancer and in vaginal delivery of anti-viral drugs or anti-inflammatory drugs, which could raise their efficacy, deliver them to the specific target and minimize their toxic side effects.     

聚己二酸丁二醇酯-对苯二甲酸丁二醇酯/Ⅰ型胶原取向纤维促进前交叉韧带断裂后腱-骨愈合

背景:材料表面的微/纳米结构对细胞的行为具有调控作用,肌腱组织主要由平行的胶原纤维组成,因此取向纤维结构对腱-骨愈合具有一定的促进作用.目的:探索聚己二酸丁二醇酯-对苯二甲酸丁二醇酯[poly(butylene adipate-co-terephthalate),PBAT]/Ⅰ型胶原取向纤维支架的生物相容性和成骨诱导活...     

Angiotensin-(1-7)对血管平滑肌细胞钙化的影响及信号转导通路

目的： 在钙化大鼠主动脉血管平滑肌细胞上观察血管紧张素-(1-7)［Angiotensin-(1-7)］对钙化的影响及其信号通道。方法: 用β-磷酸甘油制备钙化的大鼠血管平滑肌细胞，再以血管紧张素-(1-7)、血管紧张素Ⅱ、血管紧张素Ⅱ +血管紧张素-(1-7)、选择性蛋白激酶A（PKA）或蛋白激酶C（PKC）抑制剂等干预，通过Von Kossa染色及检测钙含量、碱性磷酸酶活性、骨钙素浓度和Cbfa1 mRNA表达来探讨血管紧张素Ⅱ对钙化的影响及其信号通道。结果: 血管紧张素-(1-7)抑制钙化大鼠血管平滑肌细胞的钙含量、碱性磷酸酶活性（P>0.05）、骨钙素浓度和Cbfa1 mRNA表达（P<0.05），也抑制血管紧张素Ⅱ对血管平滑肌细胞的钙含量、碱性磷酸酶活性、骨钙素浓度和Cbfa1 mRNA表达的促进作用（P<0.05）；血管紧张素-(1-7)提高血管平滑肌细胞内cAMP浓度（P<0.05），PKA抑制剂可阻断血管紧张素-(1-7)对钙化血管平滑肌细胞的钙含量、碱性磷酸酶活性、骨钙素浓度和Cbfa1 mRNA表达的抑制作用（P<0.05）。结论: 血管紧张素-(1-7)可抑制β-磷酸甘油诱导的血管平滑肌细胞钙化，并拮抗血管紧张素Ⅱ促进的血管平滑肌细胞钙化；这些效应与cAMP-PKA-Cbfa1信号途径有关。     

Mesenchymal stromal cells form vascular tubes when placed in fibrin sealant and accelerate wound healing in vivo

Non-healing, chronic wounds are a growing public health problem and may stem from insufficient angiogenesis in affected sites. Here, we have developed a fibrin formulation that allows adipose-derived mesenchymal stromal cells (ADSCs) to form tubular structures in vitro. The tubular structures express markers of endothelium, including CD31 and VE-Cadherin, as well as the pericyte marker NG2. The ability for the MSCs to form tubular structures within the fibrin gels was directly dependent on the stoichiometric ratios of thrombin and fibrinogen and the resulting gel concentration, as well as on the presence of bFGF. Fibrin gel formulations that varied in stiffness were tested. ADSCs that are embedded in a stiff fibrin formulation express VE-cadherin and CD31 as shown by PCR, FACS and immunostaining. Confocal imaging analysis demonstrated that tubular structures formed, containing visible lumens, in the stiff fibrin gels in vitro. There was also a difference in the amounts of bFGF secreted by ADSCs grown in the stiffer gels as compared to softer gels. Additionally, hAT-MSCs gave rise to perfusable vessels that were VE-cadherin positive after subcutaneous injection into mice, whereas the softer fibrin formulation containing ADSCs did not. The application of ADSCs delivered in the stiff fibrin gels allowed for the wounds to heal more quickly, as assessed by wound size, amount of granulation tissue and collagen content. Interestingly, following 5 days of healing, the ADSCs remained within the fibrin gel and did not integrate into the granulation tissue of healing wounds in vivo. These data show that ADSCs are able to form tubular structures within fibrin gels, and may also contribute to faster wound healing, as compared with no treatment or to wounds treated with fibrin gels devoid of ADSCs.     

Angiotensin-(1-7)对血管平滑肌细胞钙化的影响及信号转导通路

目的:在钙化大鼠主动脉血管平滑肌细胞上观察血管紧张素-(1-7)[Angiotensin-(1-7)]对钙化的影响及其信号通道。方法:用β-磷酸甘油制备钙化的大鼠血管平滑肌细胞,再以血管紧张素-(1-7)、血管紧张素Ⅱ、血管紧张素Ⅱ 血管紧张素-(1-7)、选择性蛋白激酶A(PKA)或蛋白激酶C(PKC)抑制剂等干预,通过Von Kossa染色及检测钙含量、碱性磷酸酶活性、骨钙素浓度和Cbfa1 mRNA表达来探讨血管紧张素Ⅱ对钙化的影响及其信号通道。结果:血管紧张素-(1-7)抑制钙化大鼠血管平滑肌细胞的钙含量、碱性磷酸酶活性(P>0.05)、骨钙素浓度和Cbfa1 mRNA表达(P<0.05),也抑制血管紧张素Ⅱ对血管平滑肌细胞的钙含量、碱性磷酸酶活性、骨钙素浓度和Cbfa1 mRNA表达的促进作用(P<0.05);血管紧张素-(1-7)提高血管平滑肌细胞内cAMP浓度(P<0.05),PKA抑制剂可阻断血管紧张素-(1-7)对钙化血管平滑肌细胞的钙含量、碱性磷酸酶活性、骨钙素浓度和Cbfa1 mRNA表达的抑制作用(P<0.05)。结论:血管紧张素-(1-7)可抑制β-磷酸甘油诱导的血管平滑肌细胞钙化,并拮抗血管紧张素Ⅱ促进的血管平滑肌细胞钙化;这些效应与cAMP-PKA-Cbfa1信号途径有关。     

Protein release from poly(ε-caprolactone) microspheres prepared by melt encapsulation and solvent evaporation techniques: A comparative study

Poly(ε-caprolactone) (PCL) microspheres containing c. 3% bovine serum albumin (BSA) were prepared by melt encapsulation and solvent evaporation techniques. PCL, because of its low T_m, enabled the melt encapsulation of BSA at 75°C thereby avoiding potentially toxic organic solvents such as dichloromethane (DCM). Unlike the solvent evaporation method, melt encapsulation led to 100% incorporation efficiency which is a key factor in the microencapsulation of water-soluble drugs. Examination of the stability of the encapsulated protein by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that protein integrity was unaffected by both methods of encapsulation. In vitro release of the protein into phosphate buffer examined at 37 °C from microspheres prepared by both techniques showed that the release rate from melt-encapsulated microspheres was somewhat slower compared to the release from solvent-evaporated spheres. Both released around 20% of the incorporated protein in 2 weeks amounting to approximately 6.5 μgmg^-1 of microspheres. Although the diffusivity of macromolecules in PCL is rather low, it is shown that PCL microspheres are capable of delivering sufficient quantity of proteins by diffusion for prolonged periods to function as a carrier for many vaccines. Unlike poly(lactic acid) (PLA) and poly(glycolic acid) (PGA) polymers which generate extreme acid environments during their degradation, the delayed degradation characteristics of PCL do not generate an acid environment during protein release and, therefore, may be advantageous for sustained delivery of proteins and polypeptides.     

Coordinate Regulation of Matrix Metalloproteinase-1 and Tissue Inhibitor of Metalloproteinase-1 Expression in Human Vascular Smooth Muscle Cells

The expression of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) by human vascular smooth muscle cells (SMC) was monitored as a function of the phenotypic modulation in vitro. Cell phenotype was manipulated by varying serum concentration and cell density. Synthetic phenotype was characterized by a minimum expression of the contractile proteins and a maximal proliferation rate. Contractile phenotype was quiescent and expressed a maximal level of contractile proteins. Synthetic cells expressed the highest levels of both MMP-1 and TIMP-1 and displayed maximal collagenolytic activity. No significant change was detected in MMP-2 expression or catalytic activity. Enzyme immunoassays revealed that MMP-1 expression fell by 77 ± 2.4-95 ± 0.5%, and that of TIMP-1 by 34 ± 0.5-59 ± 1.9%, as the cells acquired a contractile phenotype. The level of the MMP-1/TIMP-1 complex was similarly reduced by 78 ± 2.9-85 ± 1.6%. These data demonstrate that the expression of MMP-1 and TIMP-1 are coordinately regulated with SMC phenotype.     

Three dimensionally printed pearl powder/poly-caprolactone composite scaffolds for bone regeneration**

Abstract

Pearl has great potential as a natural biomaterial for bone tissue engineering, but it suffers from low porosity, difficulty in molding, and poor anti-buckling property. In this study, we used the 3-D printing technique to fabricate original pearl powder and PCL composite scaffolds with different concentrations of pearl powder. The four groups of scaffolds were termed PCL, 30% Pearl/PCL, 50% Pearl/PCL and 80% Pearl/PCL scaffolds according to the proportion of pearl powder. The samples were systematically investigated by scanning electron microscopy (SEM), wide-angle XRD, liquid substitution, Zwick static materials testing, and energy dispersive X-ray analysis. Biological characterization included SEM, fluorescent staining using calcein-AM, cell counting kit-8 assay, alkaline phosphatase and qRT-PCR analysis. The results show that the pore size and the pore morphology of the scaffolds are closely controlled via 3-D printing. This is very beneficial for tissue growth and nutrition transmission. The regular and uniform square macropore structure ensured that the pearl powder/PCL scaffolds had favorable mechanical strength. As the concentration of pearl powder in the scaffolds increase, the compressive strength and apatite formation increase as well as cell adhesion, proliferation, and osteogenic differentiation. These results show that pearl powder/PCL scaffolds fit the requirements of bone tissue engineering. The structures as well as physicochemical and biological properties of pearl powder/PCL composite scaffolds are positively associated with pearl powder concentrations.     

血管紧张素Ⅱ和血管紧张素-(1-7)对大鼠血管平滑肌细胞肾素(原)受体表达的影响

目的: 研究血管紧张素Ⅱ(Ang II)和血管紧张素-(1-7) 对大鼠血管平滑肌细胞(VSMCs)肾素(原)受体 表达的影响。方法: 将VSMCs按以下分组:(1)对照组:不加干预因素;(2)不同浓度AngⅡ组:分别加入AngⅡ10、100、1 000 nmol/L;(3)不同浓度Ang-(1-7)组:分别加入Ang-(1-7) 10、100、1 000 nmol/L; (4)AngⅡ+ losartan(AT₁受体拮抗剂)组:losartan 10^-6 mol/L预处理30 min后,再加入AngⅡ100 nmol/L; (5)AngⅡ+ PD123319(AT₂受体拮抗剂)组: 先用10^-5 mol/LPD123319预处理30 min后,再用终浓度为100 nmol/L AngⅡ;(6)CGP42112A(AT₂受体激动剂)组:加入10^-7 mol/L CGP42112A。各组用real-time PCR法和Western blotting法检测(P)RR的表达情况。结果: 与对照组比,不同浓度AngⅡ可以促进(P)RR mRNA和蛋白的表达,并且呈浓度依赖性(均P<0.01);不同浓度Ang-(1-7)可抑制 (P)RR mRNA和蛋白表达(均P<0.01),各浓度之间比较无显著差异(均P>0.05);加入CGP42112A可以促进(P)RR mRNA和蛋白的表达(均P<0.01),与AngⅡ处理组比较,加入PD123319可抑制(P)RR mRNA和蛋白表达(均P<0.01),加入losartan不能抑制(P)RR mRNA和蛋白表达(P>0.05)。结论: AngⅡ可通过AT₂受体促进(P)RR mRNA和蛋白表达,而Ang-(1-7) 可抑制(P)RR mRNA和蛋白的表达。