Twenty-one novel tri- and tetranucleotide microsatellite loci for the Amur tiger (Panthera tigris altaica)

Amur tiger is the largest subspecies of tiger in the world and his conservation has also received much attention. In this study, we isolated and characterized twenty-one tri- and tetranucleotide microsatellite markers from this species. The number of alleles for each locus ranged from two to nine in a group of 60 individuals and the observed and expected heterozygosities were 0.333–0.917 and 0.302–0.822, respectively. The overall discrimination power and exclusion probabilities in parentage and paternity testing for these markers were 1.00, 0.9947 and 0.9999, respectively, indicating high-resolution power of microsatellite markers.     

Non-peptide calcitonin gene-related peptide receptor antagonists from a benzodiazepinone lead

High-throughput screening of the Merck sample collection identified benzodiazepinone tetralin-spirohydantoin 1 as a CGRP receptor antagonist with micromolar activity. Comparing the structure of 1 with those of earlier peptide-based antagonists such as BIBN 4096 BS, a key hydrogen bond donor-acceptor pharmacophore was hypothesized. Subsequent structure activity studies supported this hypothesis and led to benzodiazepinone piperidinyldihydroquinazolinone 7, CGRP receptor K(i)=44nM and IC(50)=38nM. Compound 7 was orally bioavailabile in rats and is a lead in the development of orally bioavailable CGRP antagonists for the treatment of migraine.     

Optimization of CCR4 antagonists: side-chain exploration

The design, synthesis, and activity of novel and selective small molecule antagonists of the CC chemokine receptor-4 (CCR4) are presented. Compound 8c was efficacious in a murine allergic inflammation model (ED(50) 30 mg/kg).     

Bicyclic and tricyclic thiophenes as protein tyrosine phosphatase 1B inhibitors

A novel pyridothiophene inhibitor of PTP1B was discovered by rational screening of phosphotyrosine mimics at high micromolar concentrations. The potency of this lead compound has been improved significantly by medicinal chemistry guided by X-ray crystallography and molecular modeling. Excellent consistency has been observed between structure-activity relationships and structural information from PTP1B-inhibitor complexes.     

Chelerythrine and sanguinarine dock at distinct sites on BclXL that are not the classic BH3 binding cleft

The ratio of the levels of pro-survival and pro-apoptotic members of the Bcl-2 protein family is thought to be an important regulatory factor for determining the sensitivity of the mammalian cells to apoptotic stimuli. High levels of expression of pro-survival members such as Bcl(XL) in human cancers were frequently found to be a good prognostic indicator predicting poor response to chemotherapy. The pro-survival members of the Bcl-2 family mediate their effects through heterodimerization with the BH3 region of the pro-apoptotic members. Structural analyses of the binding complex of the BH3 peptide and Bcl(XL) showed that a hydrophobic groove termed the BH3 binding cleft is the docking site for the BH3 region. Chemical mimetics of the BH3 region such as BH3I-1 that target the BH3 binding cleft indeed exhibit pro-apoptotic activities. Chelerythrine (CHE) and sanguinarine (SAN) are natural benzophenanthridine alkaloids that are structurally homologous to each other. CHE was previously identified as an inhibitor of Bcl(XL) function from a high-throughput screen of natural products, but its mode of interaction with Bcl(XL) is not known. By determining the effect of site-directed mutagenesis on ligand binding and using saturation transfer difference (STD) NMR experiments, we have verified locations of these docked ligands. Surprisingly, CHE and SAN bind separately at the BH groove and BH1 region of Bcl(XL) respectively, different from the BH3 binding cleft where other known inhibitors of Bcl(XL) target. Interestingly, certain residues on the flexible loop between helices alpha1 and alpha2 of Bcl(XL) are also perturbed upon CHE, but not SAN or BH3I-1 binding. Although CHE and SAN are similarly effective as BH3I-1 in displacing bound BH3 peptide, they are much more effective in inducing apoptosis, raising the possibility that CHE and SAN might be able to antagonize other pro-survival mechanisms in addition to the one that involves BH3 region binding.     

Scaffold stiffness affects the contractile function of three-dimensional engineered cardiac constructs

We investigated the effects of the initial stiffness of a three-dimensional elastomer scaffold--highly porous poly(glycerol sebacate)--on functional assembly of cardiomyocytes cultured with perfusion for 8 days. The polymer elasticity varied with the extent of polymer cross-links, resulting in three different stiffness groups, with compressive modulus of 2.35 ± 0.03 (low), 5.28 ± 0.36 (medium), and 5.99 ± 0.40 (high) kPa. Laminin coating improved the efficiency of cell seeding (from 59 ± 15 to 90 ± 21%), resulting in markedly increased final cell density, construct contractility, and matrix deposition, likely because of enhanced cell interaction and spreading on scaffold surfaces. Compact tissue was formed in the low and medium stiffness groups, but not in the high stiffness group. In particular, the low stiffness group exhibited the greatest contraction amplitude in response to electric field pacing, and had the highest compressive modulus at the end of culture. A mathematical model was developed to establish a correlation between the contractile amplitude and the cell distribution within the scaffold. Taken together, our findings suggest that the contractile function of engineered cardiac constructs positively correlates with low compressive stiffness of the scaffold.     

Importance of the stem cell microenvironment for ophthalmological cell-based therapy

Cell therapy is a promising treatment for diseases that are caused by cell degeneration or death. The cells for clinical transplantation are usually obtained by culturing healthy allogeneic or exogenous tissue invitro. However, for diseases of the eye, obtaining the adequate number of cells for clinical transplantation is difficult due to the small size of tissue donors and the frequent needs of long-term amplification of cells in vitro, which results in low cell viability after transplantation. In addition, the transplanted cells often develop fibrosis or degrade and have very low survival. Embryonic stem cells(ESCs) and induced pluripotent stem cells(i PS) are also promising candidates for cell therapy. Unfortunately, the differentiation of ESCs can bring immune rejection, tumorigenicity and undesired differentiated cells, limiting its clinical application. Although i PS cells can avoid the risk of immune rejection caused by ES cell differentiation post-transplantation, the low conversion rate, the risk of tumor formation and the potentially unpredictable biological changes that could occur through genetic manipulation hinder its clinical application. Thus, the desired clinical effect of cell therapy is impaired by these factors. Recent research findings recognize that the reason for low survival of the implanted cells not only depends on the seeded cells, but also on the cell microenvironment, which determines the cell survival, proliferation and even reverse differentiation. When used for cell therapy, the transplanted cells need a specific three-dimensional structure to anchor and specific extra cellular matrix components in addition to relevant cytokine signaling to transfer the required information to support their growth. These structures present in the matrix in which the stem cells reside are known as the stem cell microenvironment. The microenvironment interaction with the stem cells provides the necessary homeostasis for cell maintenance and growth. A large number of studies suggest that to explore how to reconstruct the stem cell microenvironment and strengthen its combination with the transplanted cells are key steps to successful cell therapy. In this review, we will describe the interactions of the stem cell microenvironment with the stem cells, discuss the importance of the stem cell microenvironment for cell-based therapy in ocular diseases, and introduce the progress of stem cell-basedtherapy for ocular diseases.     

Ultrastructure changes of seedlings of <Emphasis Type="Italic">Kosteletzkya virginica</Emphasis> under waterlogging conditions

The changes in ultrastructure of leaf cell in waterlogged Kosteletzkya virginica seedlings were observed by transmission electron microscope. During waterlogging, the chloroplasts turned into a round shape and their volume was gradually reduced. Furthermore, the lamellae of thylakoids swelled, and the inclusions in chloroplast decreased. The shape of starch grains slightly changed, but their number and volume decreased, and they even disappeared in the end. On the other hand, plastoglobules enlarged and their amount enhanced gradually. The membrane and inner cristea of mitochondria gradually became unclear, while the mitochondria firstly enlarged but disappeared at the end. Similar to chloroplast, cell nucleus dwindled gradually, concentrated and tended to a round shape. Additionally, the annulate lamellae and multivesicular body occurred on the 20th day of experiment, and the cell wall distorted and twisted at the late stage of waterlogging. Consequently, these changes of organelles are the typical characteristics of the complete cell disintegration or death for Kosteletzkya virginica under long-term waterlogging.