The effect of different platelet-rich plasma concentrations on proliferation and differentiation of human periodontal ligament cells in vitro

OBJECTIVES: The use of platelets and platelet products has become increasingly popular clinically as a means of accelerating endosseous wound healing. It is likely that growth factors released by activated platelets at the site of injury play a role in periodontal regeneration by regulating cellular activity. The purpose of this study was to evaluate the biological effects of platelet-rich plasma (PRP) on human periodontal ligament cells (hPDLCs) in vitro. MATERIALS AND METHODS: Primary cultures of hPDLCs were obtained from healthy premolars. PRP was isolated by two-step centrifugation. Two main growth factors present in the thrombin-activated PRP (platelet-derived growth factor [PDGF-AB] and transforming growth factor-beta1 [TGF-beta1]) were evaluated using ELISA assay. Activated PRP or the combination of recombined human TGF-beta1 (rhTGF-beta1) and PDGF-AB (rhPDGF-AB) were added to hPDLCs in different concentrations to assess cell proliferation and osteogenic differentiation. RESULTS: PRP contained high levels of TGF-beta1 and PDGF-AB. Cell attachment, proliferation and ALP activity were enhanced by addition of PRP or rhTGF-beta1 and rhPDGF-AB combination to the cell cultures, while the stimulatory potency of PRP was much greater than the latter. These stimulatory effects presented in a dose-dependant manner, it seemed that PRP with 50~100 ng/ml TGF-beta1 was an ideal concentration. CONCLUSIONS: PRP can enhance hPDLC adhesion, proliferation and induce the differentiation of hPDLC into mineralized tissue formation cell; thereby contribute to the main processes of periodontal tissue regeneration. For economical and biological reasons, PRP has more clinical beneficial than analogous growth factors.     

通过染色体外同源重组建立乳腺中表达外源基因的转基因小鼠

为研究染色体外重组方法在创建转基因动物中的应用,选取牛asl酪蛋白基因的5′及3′侧翼区和氯霉素乙酰化酶编码区,构建了两个具有3 kb相互重叠的融合基因.将这两个DNA片段末端脱磷酸后,以摩尔比1:1的比例混合,通过显微注射导入小鼠受精原核,最后获得了11个品系的转基因小鼠.对其中10个品系的小鼠的整合分析表明,所注射的两个DNA片段均发生了染色体外同源重组,而且除了 1个品系的小鼠丢失了大约1kb的序列外,其余品系小鼠的重组产物与预想的结构符合.在当代和后代的转基因小鼠乳汁中均可测到氯霉素乙酰化酶的活性.这表明融合基因在转基因小鼠乳腺中得到表达和分泌,也说明显微共注射两个相互重叠的基因片段是建立转基因动物的一个可行途径.     

转基因红花中角质细胞生长因子KGF-1的表达

通过构建重组表达质粒载体p139035S-KGF1和根癌农杆菌介导在红花(Carthamus tinctorius)中表达角质细胞生长因子(KGF-1)。从侵染到诱导生根共需要14周, 转化率达0.1%。红花子叶在潮霉素筛选培养基上培养4–5周后便可获得丛生芽, 再生芽移入含潮霉素的伸长生根培养基, 培养4–8周可诱导生根。通过PCR、Southern blot、RT-PCR及Western blot检测证明目的基因KGF-1已经整合到红花细胞的染色体中, 实现了KGF-1外源蛋白在红花中的成功表达, 为开发KGF-1蛋白新的生产途径奠定了基础。     

Acylation-stimulating protein/C5L2-neutralizing antibodies alter triglyceride metabolism in vitro and in vivo

Acylation-stimulating protein (ASP), a lipogenic hormone, stimulates triglyceride (TG) synthesis and glucose transport upon activation of C5L2, a G protein-coupled receptor. ASP-deficient mice have reduced adipose tissue mass due to increased energy expenditure despite increased food intake. The objective of this study was to evaluate the blocking of ASP-C5L2 interaction via neutralizing antibodies (anti-ASP and anti-C5L2-L1 against C5L2 extracellular loop 1). In vitro, anti-ASP and anti-C5L2-L1 blocked ASP binding to C5L2 and efficiently inhibited ASP stimulation of TG synthesis and glucose transport. In vivo, neither anti-ASP nor anti-C5L2-L1 altered body weight, adipose tissue mass, food intake, or hormone levels (insulin, leptin, and adiponectin), but they did induce a significant delay in TG clearance [P < 0.0001, 2-way repeated-measures (RM) ANOVA] and NEFA clearance (P < 0.0001, 2-way RM ANOVA) after a fat load. After treatment with either anti-ASP or anti-C5L2-L1 antibody there was no change in adipose tissue AMPK activity, but neutralizing antibodies decreased perirenal TG mass (-38.4% anti-ASP, -18.8% anti-C5L2, P < 0.01-0.001) and perirenal LPL activity (-75.6% anti-ASP, -72.5% anti-C5L2, P < 0.05). In liver, anti-C5L2-L1 decreased TG mass (-42.8%, P < 0.05), whereas anti-ASP increased AMPK activity (+34.6%, P < 0.001). In the muscle, anti-C5L2-L1 significantly increased TG mass (+128.0%, P < 0.05), LPL activity (+226.1%, P < 0.001), and AMPK activity (+71.1%, P < 0.01). In addition, anti-ASP increased LPL activity (+164.4, P < 0.05) and AMPK activity (+53.9%, P < 0.05) in muscle. ASP/C5L2-neutralizing antibodies effectively block ASP-C5L2 interaction, altering lipid distribution and energy utilization.     

Chaperone-mediated refolding of recombinant prochymosin

It has been verified that prochymosin is characterized by a two-stage refolding: dilution of unfolded protein into pH 11 buffer followed by neutralization at pH 8; the high-pH step is indispensable. Here we demonstrate that one-stage refolding around pH 8 can be achieved when GroE or 10-fold molar excess (rather than catalytic concentration) of protein disulfide isomerase (PDI) over prochymosin is present. The helping effect varies with the oxidation states of prochymosin. GroE and PDI increase the reactivation of the unfolded, partially reduced and the unfolded, oxidized prochymosin from 5% to 40% and from 50% to 100%, respectively. For the unfolded and fully reduced prochymosin, GroE does not have a positive effect, whereas PDI promotes renaturation from 2% to 28%. Based on our previous and present observations, we propose that at pH 8 there may be two kinds of incorrect interactions within and between prochymosin polypeptides leading to unproductive pathways: one prevents disulfide rearrangement, which can be avoided by high pH; the other interferes with acquisition of native conformation, which can be relieved by GroE and PDI.     

A Passive Anti-icing Strategy Based on a Superhydrophobic Mesh with Extremely Low Ice Adhesion Strength

Although superhydrophobic materials have attracted much research interest in anti-icing,some controversy still exists.In this research,we report a cost-effective method used to verify the contribution of area fraction to ice adhesion strength.We tried to partially-embed siliea nanopnarticles into microscale fabrics of a commercial polyamide mesh.Then,the area fraction could be determined by altering the mesh size.Generally,the ice adhesion strength decreases as the area fraction decreases.An ice adhesion strength of~1.9 kPa and a delayed freezing time of~1048 s can be obtained.We attribute the low ice adhesion strength to the combination of superhydro-phobicity and stress concentration.The superhydrophobicity prohibits the water from penetrating into the voids of the meshes,and the small actual contact area leads to stress concentration which promotes interfacial crack propagation.Moreover,our superhydrophobic mesh simultaneously exhibis a micro-nano hierarchical structure and a partally-cmbedded structure.Therefore,the as-prepared superhydrophobic mesh retained the ieephobicity after 20 icingldeicing cycles,and maintained its superhydrophobicity even afier 60 sandpaper-abrasion cycles and a 220"C thermal treatment.