Partial meniscectomy changes fluid pressurization in articular cartilage in human knees

Partial meniscectomy is believed to change the biomechanics of the knee joint through alterations in the contact of articular cartilages and menisci. Although fluid pressure plays an important role in the load support mechanism of the knee, the fluid pressurization in the cartilages and menisci has been ignored in the finite element studies of the mechanics of meniscectomy. In the present study, a 3D fibril-reinforced poromechanical model of the knee joint was used to explore the fluid flow dependent changes in articular cartilage following partial medial and lateral meniscectomies. Six partial longitudinal meniscectomies were considered under relaxation, simple creep, and combined creep loading conditions. In comparison to the intact knee, partial meniscectomy not only caused a substantial increase in the maximum fluid pressure but also shifted the location of this pressure in the femoral cartilage. Furthermore, these changes were positively correlated to the size of meniscal resection. While in the intact joint, the location of the maximum fluid pressure was dependent on the loading conditions, in the meniscectomized joint the location was predominantly determined by the site of meniscal resection. The partial meniscectomy also reduced the rate of the pressure dissipation, resulting in even larger difference between creep and relaxation times as compared to the case of the intact knee. The knee joint became stiffer after meniscectomy because of higher fluid pressure at knee compression followed by slower pressure dissipation. The present study indicated the role of fluid pressurization in the altered mechanics of meniscectomized knees.     

Electrochemical studies of DNA immobilization onto the azide-terminated monolayers and its interaction with taxol

A surface modification procedure for the creation of self-assembled monolayers (SAMs) that can be used as a scaffold for double-stranded DNA (dsDNA) incorporation onto the gold surfaces is described. The SAMs of an azidohexane thiol derivative were prepared on the Au electrode and then used for the immobilization of dsDNA. The electrochemical characteristics of dsDNA onto the SAM-modified gold electrode were investigated by cyclic voltammetry and electrochemical impedance spectroscopy, and the surface concentration of dsDNA onto the SAMs surface was estimated. The interaction of dsDNA with the anticancer drug, taxol (paclitaxel), was also studied on the surface of DNA/SAM/Au electrode. The observed decrease in the guanine oxidation peak current was used to monitor the interaction of taxol with DNA. The resulting Langmuir isotherm for taxol binding to DNA at the modified electrode was used to evaluate the binding constant of taxol-DNA. The results obtained supported the groove binding interaction of taxol with DNA. The modified electrode was used as a sensitive sensor for quantification of taxol in human serum sample.     

Establishment and in vitro differentiation of a new embryonic stem cell line from human blastocyst

Embryonic stem cells have the ability to remain undifferentiated and proliferate indefinitely in vitro while maintaining the potential to differentiate into derivatives of all three embryonic germ layers. These cells have, therefore, potential for in vitro differentiation studies, gene function, and so on. The aim of this study was to produce a human embryonic stem cell line. An inner cell mass of a human blastocyst was separated and cultured on mouse embryonic fibroblasts in embryonic stem cell medium with related additives. The established line was evaluated by morphology; passaging; freezing and thawing; alkaline phosphatase; Oct-4 expression; anti-surface markers including Tra-1-60 and Tra-1-81; and karyotype and spontaneous differentiation. Differentiated cardiomyocytes and neurons were evaluated by transmission electron microscopy and immunocytochemistry. Here, we report the derivation of a new embryonic stem cell line (Royan H1) from a human blastocyst that remains undifferentiated in morphology during continuous passaging for more than 30 passages, maintains a normal XX karyotype, is viable after freezing and thawing, and expresses alkaline phosphatase, Oct-4, Tra-1-60, and Tra-1-81. These cells remain undifferentiated when grown on mouse embryonic fibroblast feeder layers in the presence or absence of recombinant human leukemia inhibitory factor. Royan H1 cells can differentiate in vitro in the absence of feeder cells and can produce embryoid bodies that can further differentiate into beating cardiomyocytes as well as neurons. These results define Royan H1 cells as a new human embryonic stem cell line.     

Electrospun poly-l-lactic acid/polyvinyl alcohol nanofibers improved insulin-producing cell differentiation potential of human adipose-derived mesenchymal stem cells

Combination of adipose-derived mesenchymal stem cells (ADSCs) and synthetic materials in terms of pancreatic tissue engineering can be considered as a treatment of diabetes. This study aimed to evaluate the differentiation of human ADSCs to pancreatic cells on poly-l -lactic acid/polyvinyl alcohol (PLLA/PVA) nanofibers as a three-dimensional (3D) scaffold. Mesenchymal stem cells (MSCs) were characterized for mesenchymal surface markers by flow cytometry. Then ADSCs were seeded on 3D scaffolds and treated with pancreatic differentiation medium. Immunostaining assay showed that ADSCs were very efficiently differentiated into a relatively homogeneous population of insulin-producing cells. Moreover, real-time RT-PCR results revealed that pancreas-specific markers were highly expressed in 3D scaffolds compared with their expression in tissue culture plates and this difference in expression level was significant. In addition, insulin and C-peptide secreted in response to varying concentrations of glucose in the 3D scaffold group was significantly higher than that in 2D culture. The results of the present study confirmed that PLLA/PVA scaffold seeded with ADSCs could be a suitable option in pancreatic tissue engineering.     

Characterisation and expression of phospholipases B from the opportunistic fungus Aspergillus fumigatus

The phospholipase B family (PLB) are enzymes sharing phospholipase (PL), lysophospholipase (LPL) and lysophospholipase-transacylase (LPTA) activities. They have been shown to be important virulence factors in several human fungal pathogens including Candida albicans and Cryptococcus neoformans. Aspergillus fumigatus, a human opportunistic fungal pathogen leading to a high rate of mortality in immunosuppressed patients is known to possess an extracellular phospholipase B activity. In this paper, we report the molecular characterisation of three PLB genes from A. fumigatus (afplb) using degenerate primers in PCR amplification and data from the A. fumigatus genome project. They are expressed at 37 degrees C, and two of them (afplb1 and afplb3) are induced by lecithin. They encode proteins of 633, 588 and 630 amino acids, respectively, presenting together a T-Coffee score of 81. They also possess the amino acid triad responsible for enzymatic activity in the mammalian cytosolic PLA2 and other fungal PLBs. AfPLB1 and afPLB3 are secreted with a cleaved signal peptide. The complete cDNA sequences were obtained by RACE-PCR for the two secreted afPLBs and probably account for the extracellular phospholipase activity previously reported in the culture media of A. fumigatus.     

Targeted co-delivery of paclitaxel and anti P-gp shRNA by low molecular weight PEI decorated with L-3,4-dihydroxyphenylalanine

Co-delivery of small chemotherapeutic molecules and nucleic acid materials via targeted carriers has attracted great attention for treatment of resistant tumors and reducing adverse effects. In this study, a targeted carrier for co-delivery was prepared based on low-molecular weight polyethylenimine (LMW PEI). Paclitaxel (PTX) was covalently conjugated onto PEI via a succinate linker. The PEI conjugate was decorated with L-DOPA in order to target large neutral amino acid transporter-1 (LAT-1) that is over-expressed on various cancer cells. This PEI conjugate was complexed with human ABCB1 shRNA plasmid to down-regulate the expression of P-glycoprotein, as one of the major efflux pumps inducing resistance against chemotherapeutics. The formation of PEI conjugate enhanced the solubility of PTX and resulted in the condensation and protection of plasmid DNA in nanosized polyplexes. The results of targeted delivery into the cells demonstrated that PEI conjugate transferred the payloads to the cells over-expressing LAT-1 transporter, while the biological effects on the cells lacking the transporter was negligible. Also, shRNA-mediated down-regulation of P-gp led to the increase of toxic effects on the cells over-expressing P-gp. This study suggests a promising approach for co-delivery of small molecules and nucleic acid materials in a targeted manner for cancer therapy.