Effects of protein binding on a lipid bilayer containing local anesthetic articaine, and the potential of mean force calculation: a molecular dynamics simulation approach

Articaine, as a local anesthetic drug has been simulated in neutral and charged forms, and its interaction with the dimyristoylphosphatidylcholine (DMPC) lipid bilayer membrane is investigated by molecular dynamics simulation using GROMACS software. In order to obtain the optimum location of the drug molecules, as they penetrate into the membrane, umbrella sampling is applied and the free energy is calculated. The effect of protein binding to DMPC membrane on the process of drug diffusion through the membrane is considered. Five simulation systems are designed and by applying the potential of mean force, the molecular dynamics simulation on the system is performed. In light of the obtained results, the electrostatic potential, variation of lipid bilayer’s order parameter and the diffusion coefficient of drug are discussed.

Spectrophotometric study of ketoconazole binding with citrate capped silver nanoparticles and its monitoring in human plasma samples

Ketoconazole or Nizoral is an antifungal medication used to treat various type of fungal infections. It has been reported that this antifungal agent is able to induce a variety of heart function side effects, such as long‐QT syndrome, and ventricular arrhythmias. Hence, prescribing, identifying and controlling the side effects of medications such as ketoconazole is essential for human safety. In this study, a distinct and fast colorimetric probe based on citrate capped silver nanoparticles (Cit‐AgNPs) was introduced for determination of trace amounts of ketoconazole. Cit‐AgNPs were synthesized trough a novel method and applied for the quantification of ketoconazole in human plasma samples. Ultraviolet‐visible spectroscopy, transmission electron microcopy, dynamic light scattering, and energy dispersive X‐ray spectroscopy analysis, have been utilized for characterization of Cit‐AgNPs. It was revealed that in the presence of ketoconazole, the absorption intensity of Cit‐AgNPs was decreased by increasing the ketoconazole concentration. Moreover, this reaction is accompanied by a color change from shiny yellow to brown/red in acidic pH. Under acidic pH and optimum condition, the calibration graph of the assay was linear in the range of 0.1 to 0.8 μM. According to the obtained results Cit‐AgNPs can be used as a novel probe for the sensitive and specific detection and determination of similar drugs in clinical samples. Also, this method open a new way to biomedical analyses of pharmaceutical samples which is necessary in TDM.     

Significant differences between mouse and human trophinins are revealed by their expression patterns and targeted disruption of mouse trophinin gene.

Trophinin has been identified as a membrane protein mediating apical cell adhesion between two human cell lines: trophoblastic HT-H cells, and endometrial epithelial SNG-M cells. Expression patterns of trophinin in humans suggested its involvement in embryo implantation and early placental development. The mouse trophinin gene maps to the distal part of the X chromosome and corresponds to human chromosome Xp11.21-22, the locus where the human trophinin gene maps. Western blot analysis indicates that the molecular weight of mouse trophinin is 110 kDa, which is consistent with the calculated value of 107 kDa. Positive signals for trophinin proteins were detected in preimplantation mouse embryos at the morula and blastocyst stages. Implanting blastocysts do not show detectable levels of trophinin protein, demonstrating that trophinin is not involved in blastocyst adhesion to the uterus in the mouse. Mouse embryo strongly expressed trophinin in the epiblast 1 day after implantation. Trophinin protein was not found in the mouse uteri and placenta after 5.5 days postcoitus (dpc). Targeted disruption of the trophinin gene in the mouse showed a partial embryonic lethality in a 129/SvJ background, but the cause of this lethality remains undetermined. The present study indicates significant differences between mouse and human trophinins in their expression patterns, and it suggests that trophinin is not involved in embryo implantation and placental development in the mouse.     

Zonula occludens-1 (ZO-1) is involved in morula to blastocyst transformation in the mouse

It is unknown whether or not tight junction formation plays any role in morula to blastocyst transformation that is associated with development of polarized trophoblast cells and fluid accumulation. Tight junctions are a hallmark of polarized epithelial cells and zonula occludens-1 (ZO-1) is a known key regulator of tight junction formation. Here we show that ZO-1 protein is first expressed during compaction of 8-cell embryos. This stage-specific appearance of ZO-1 suggests its participation in morula to blastocyst transition. Consistent with this idea, we demonstrate that ZO-1 siRNA delivery inside the blastomeres of zona-weakened embryos using electroporation not only knocks down ZO-1 gene and protein expressions, but also inhibits morula to blastocyst transformation in a concentration-dependent manner. In addition, ZO-1 inactivation reduced the expression of Cdx2 and Oct-4, but not ZO-2 and F-actin. These results provide the first evidence that ZO-1 is involved in blastocyst formation from the morula by regulating accumulation of fluid and differentiation of nonpolar blastomeres to polar trophoblast cells.     

Potential applications of equine genomics in dissecting diseases and fertility

Following the recent development of high-resolution gene maps and generation of several basic tools and resources to use them in analyzing traits that are economically important to horse owners, genome analysis in horses is witnessing a shift towards developing an ability to analyze complex traits. The likelihood of this happening in the very near future is great, mainly because of the recent availability of the whole genome sequence in the horse. The latter has triggered the development of novel tools like SNP-chip and expression arrays that will permit rapid genome-wide analysis. While these tools will be used for a range of multi-factorial disease traits, attempts are underway to develop focused tools that can target reproduction, fertility and sex determination. For this, a catalog of sex and reproduction related (SRR) genes is being developed in horses. A recently developed dense map of the horse Y chromosome will provide genes that are expressed exclusively in males and, therefore, have an impact on stallion fertility. Overall, these advances in equine genome analysis hold promise for improved diagnosis and treatment of various conditions in horses.     

Prostaglandin E2 is a product of induced prostaglandin-endoperoxide synthase 2 and microsomal-type prostaglandin E synthase at the implantation site of the hamster

Certain uterine prostaglandins (PGs) are elevated at implantation sites and are needed to trigger the events of blastocyst implantation that include blastocyst-uterine attachment and stromal decidualization with vascular permeability changes. Several decades of investigations showed that treatment with PG synthesis inhibitors, prior to or during the time of implantation, resulted in either complete inhibition or a delay in implantation or reduction in the number of implantation sites with diminished decidual tissue. Consistent with these findings, we observed that whereas a selective PG endoperoxide synthase (Ptgs) 1 inhibitor SC-560 failed to inhibit implantation, a selective Ptgs2 inhibitor SC-236 showed significantly reduced number and size of implantation sites in progesterone-treated ovariectomized pregnant hamsters. It is known that Ptgs2 expression and Ptgs2-derived prostacyclin (PGI2) synthesis at implantation sites are needed for implantation in the mouse (a rodent that needs ovarian estrogen for implantation). However, it is unknown which Ptgs and PG synthases produce which PGs at implantation sites of the hamster (a rodent that does not need ovarian estrogen for implantation). Here we demonstrate that as blastocyst implantation proceeds, a reduction in Ptgs1 expression from uterine luminal epithelial cells and a gradual induction in Ptgs2 expression exclusively in luminal epithelial and adjacent decidual cells occurred at implantation sites of hamsters. Results also reveal that PGE2, but not PGI2, is the major PG at implantation sites where Ptgs2 and microsomal type PGE synthases but not PGI synthases are co-expressed. This elevated uterine PGE2 at implantation sites may serve to initiate or amplify physiological signals required for specific aspects of the implantation process in hamsters.     

HB-EGF directs stromal cell polyploidy and decidualization via cyclin D3 during implantation

Stromal cell polyploidy is a unique phenomenon that occurs during uterine decidualization following embryo implantation, although the developmental mechanism still remains elusive. The general consensus is that the aberrant expression and altered functional activity of cell cycle regulatory molecules at two particular checkpoints G1 to S and G2 to M in the cell cycle play an important role in the development of cellular polyploidy. Despite the compelling evidence of intrinsic cell cycle alteration, it has been implicated that the development of cellular polyploidy may be controlled by specific actions of extracellular growth regulators. Here we show a novel role for heparin-binding EGF-like growth factor (HB-EGF) in the developmental process of stromal cell polyploidy in mice. HB-EGF, which is one of the earliest known molecular mediators of implantation in mice and humans, promotes stromal cell polyploidy via upregulation of cyclin D3. Adenoviral delivery of antisense cyclin D3 attenuates cyclin D3 expression and abrogates HB-EGF-induced stromal cell polyploidy in vitro and in vivo. Collectively, the results demonstrate that the regulation of stromal cell polyploidy and decidualization induced by HB-EGF depend on cyclin D3 induction.     

Prediction of protein secondary structure based on residue pair types and conformational states using dynamic programming algorithm

We have used a statistical approach for protein secondary structure prediction based on information theory and simultaneously taking into consideration pairwise residue types and conformational states. Since the prediction of residue secondary structure by one residue window sliding make ambiguity in state prediction, we used a dynamic programming algorithm to find the path with maximum score. A score system for residue pairs in particular conformations is derived for adjacent neighbors up to ten residue apart in sequence. The three state overall per-residue accuracy, Q3, of this method in a jackknife test with dataset created from PDBSELECT is more than 70%.