A High-Content Small Molecule Screen Identifies Sensitivity of Glioblastoma Stem Cells to Inhibition of Polo-Like Kinase 1

Glioblastoma multiforme (GBM) is the most common primary brain cancer in adults and there are few effective treatments. GBMs contain cells with molecular and cellular characteristics of neural stem cells that drive tumour growth. Here we compare responses of human glioblastoma-derived neural stem (GNS) cells and genetically normal neural stem (NS) cells to a panel of 160 small molecule kinase inhibitors. We used live-cell imaging and high content image analysis tools and identified JNJ-10198409 (J101) as an agent that induces mitotic arrest at prometaphase in GNS cells but not NS cells. Antibody microarrays and kinase profiling suggested that J101 responses are triggered by suppression of the active phosphorylated form of polo-like kinase 1 (Plk1) (phospho T210), with resultant spindle defects and arrest at prometaphase. We found that potent and specific Plk1 inhibitors already in clinical development (BI 2536, BI 6727 and GSK 461364) phenocopied J101 and were selective against GNS cells. Using a porcine brain endothelial cell blood-brain barrier model we also observed that these compounds exhibited greater blood-brain barrier permeability in vitro than J101. Our analysis of mouse mutant NS cells (INK4a/ARF^−/−, or p53^−/−), as well as the acute genetic deletion of p53 from a conditional p53 floxed NS cell line, suggests that the sensitivity of GNS cells to BI 2536 or J101 may be explained by the lack of a p53-mediated compensatory pathway. Together these data indicate that GBM stem cells are acutely susceptible to proliferative disruption by Plk1 inhibitors and that such agents may have immediate therapeutic value.     

Urinary excretion of a novel hexasaccharide and glycopeptide analogue in fucosidosis.

Repeated Biogel P6 chromatography of the urine from a patient with fucosidosis yielded several fractions containing fucosyloligosaccharides and glycopeptides. Two of these were shown by ¹H nuclear magnetic resonance (¹H-n.m.r.) spectroscopy and permethylation analysis to have the following structures respectively: (I) αfuc (1→3) [βgal (1→4)] βglcNAc (1→2) αman ($\text{1→}\text{3}\text{6}$) βman (1→4) glcNAc and (II) αfuc (1→3) [βgal (1→4)] βglcNAc (1→2) αman ($\text{1→}\text{3}\text{6}$) βman (1→4) βglcNAc (1→4) [αfuc ($\text{1→}\text{3}\text{6}$)] βglcNAc-Asn.     

Cthrc1 is a positive regulator of osteoblastic bone formation

Background

Bone mass is maintained by continuous remodeling through repeated cycles of bone resorption by osteoclasts and bone formation by osteoblasts. This remodeling process is regulated by many systemic and local factors.

Methodology/Principal Findings

We identified collagen triple helix repeat containing-1 (Cthrc1) as a downstream target of bone morphogenetic protein-2 (BMP2) in osteochondroprogenitor-like cells by PCR-based suppression subtractive hybridization followed by differential hybridization, and found that Cthrc1 was expressed in bone tissues in vivo. To investigate the role of Cthrc1 in bone, we generated Cthrc1-null mice and transgenic mice which overexpress Cthrc1 in osteoblasts (Cthrc1 transgenic mice). Microcomputed tomography (micro-CT) and bone histomorphometry analyses showed that Cthrc1-null mice displayed low bone mass as a result of decreased osteoblastic bone formation, whereas Cthrc1 transgenic mice displayed high bone mass by increase in osteoblastic bone formation. Osteoblast number was decreased in Cthrc1-null mice, and increased in Cthrc1 transgenic mice, respectively, while osteoclast number had no change in both mutant mice. In vitro, colony-forming unit (CFU) assays in bone marrow cells harvested from Cthrc1-null mice or Cthrc1 transgenic mice revealed that Cthrc1 stimulated differentiation and mineralization of osteoprogenitor cells. Expression levels of osteoblast specific genes, ALP, Col1a1, and Osteocalcin, in primary osteoblasts were decreased in Cthrc1-null mice and increased in Cthrc1 transgenic mice, respectively. Furthermore, BrdU incorporation assays showed that Cthrc1 accelerated osteoblast proliferation in vitro and in vivo. In addition, overexpression of Cthrc1 in the transgenic mice attenuated ovariectomy-induced bone loss.

Conclusions/Significance

Our results indicate that Cthrc1 increases bone mass as a positive regulator of osteoblastic bone formation and offers an anabolic approach for the treatment of osteoporosis.     

Testing hypotheses driving genetic structure in the cooperatively breeding Brown‐headed Nuthatch Sitta pusilla

Habitat fragmentation has often been implicated in the decline of many species. For habitat specialists and/or sedentary species, loss of habitat can result in population isolation and lead to negative genetic effects. However, factors other than fragmentation can often be important and also need to be considered when assessing the genetic structure of a species. We genotyped individuals from 13 populations of the cooperatively breeding Brown‐headed Nuthatch Sitta pusilla in Florida to test three alternative hypotheses regarding the effects that habitat fragmentation might have on genetic structure. A map of potential habitat developed from recent satellite imagery suggested that Brown‐headed Nuthatch populations in southern Florida occupied smaller and more isolated habitat patches (i.e. were more fragmented) than populations in northern Florida. We also genotyped individuals from a small, isolated Brown‐headed Nuthatch population on Grand Bahama Island. We found that populations associated with more fragmented habitat in southern Florida had lower allelic richness than populations in northern Florida (P = 0.02), although there were no differences in heterozygosity. Although pairwise estimates of F_ST were low overall, values among southern populations were generally higher than northern populations. Population assignment tests identified K = 3 clusters corresponding to a northern cluster, a southern cluster and a unique population in southeast Florida; using sampling localities as prior information revealed K = 7 clusters, with greater structure only among southern Florida populations. The Bahamas population showed moderate to high differentiation compared with Florida populations. Overall, our results suggest that fragmentation could affect gene flow in Brown‐headed Nuthatch populations and is likely to become more pronounced over time.     

MaxHiC: A robust background correction model to identify biologically relevant chromatin interactions in Hi-C and capture Hi-C experiments

Hi-C is a genome-wide chromosome conformation capture technology that detects interactions between pairs of genomic regions and exploits higher order chromatin structures. Conceptually Hi-C data counts interaction frequencies between every position in the genome and every other position. Biologically functional interactions are expected to occur more frequently than transient background and artefactual interactions. To identify biologically relevant interactions, several background models that take biases such as distance, GC content and mappability into account have been proposed. Here we introduce MaxHiC, a background correction tool that deals with these complex biases and robustly identifies statistically significant interactions in both Hi-C and capture Hi-C experiments. MaxHiC uses a negative binomial distribution model and a maximum likelihood technique to correct biases in both Hi-C and capture Hi-C libraries. We systematically benchmark MaxHiC against major Hi-C background correction tools including Hi-C significant interaction callers (SIC) and Hi-C loop callers using published Hi-C, capture Hi-C, and Micro-C datasets. Our results demonstrate that 1) Interacting regions identified by MaxHiC have significantly greater levels of overlap with known regulatory features (e.g. active chromatin histone marks, CTCF binding sites, DNase sensitivity) and also disease-associated genome-wide association SNPs than those identified by currently existing models, 2) the pairs of interacting regions are more likely to be linked by eQTL pairs and 3) more likely to link known regulatory features including known functional enhancer-promoter pairs validated by CRISPRi than any of the existing methods. We also demonstrate that interactions between different genomic region types have distinct distance distributions only revealed by MaxHiC. MaxHiC is publicly available as a python package for the analysis of Hi-C, capture Hi-C and Micro-C data.     

Polyphyly of Asian Tree Toads,Genus Pedostibes Günther, 1876 (Anura: Bufonidae), and the Description of a New Genus from Southeast Asia

The Asian Tree Toad genus Pedostibes, as currently understood, exhibits a conspicuously disjunct distribution, posing several immediate questions relating to the biogeography and taxonomy of this poorly known group. The type species, P. tuberculosus and P. kempi, are known only from India, whereas P. hosii, P. rugosus, and P. everetti are restricted to Southeast Asia. Several studies have shown that these allopatric groups are polyphyletic, with the Indian Pedostibes embedded within a primarily South Asian clade of toads, containing the genera Adenomus, Xanthophryne, and Duttaphrynus. Southeast Asian Pedostibes on the other hand, are nested within a Southeast Asian clade, which is the sister lineage to the Southeast Asian river toad genus Phrynoidis. We demonstrate that Indian and Southeast Asian Pedostibes are not only allopatric and polyphyletic, but also exhibit significant differences in morphology and reproductive mode, indicating that the Southeast Asian species’ are not congeneric with the true Pedostibes of India. As a taxonomic solution, we describe a new genus, Rentapiagen. nov. to accommodate the Southeast Asian species.     

Noise Exposure and Hearing Impairment among Chinese Restaurant Workers and Entertainment Employees in Hong Kong

Background

Noise-induced hearing loss (NIHL) is a major concern in the non-manufacturing industries. This study aimed to investigate the occupational noise exposure and the NIHL among Chinese restaurant workers and entertainment employees working in the service industry in Hong Kong.

Methods

This cross-sectional survey involved a total of 1,670 participants. Among them, 937 were randomly selected from the workers of Chinese restaurants and 733 were selected from workers in three entertainment sectors: radio and television stations; cultural performance halls or auditoria of the Leisure and Cultural Services Department (LCSD); and karaoke bars. Noise exposure levels were measured in the sampled restaurants and entertainment sectors. Each participant received an audiometric screening test. Those who were found to have abnormalities were required to take another diagnostic test in the health center. The “Klockhoff digit” method was used to classify NIHL in the present study.

Results

The main source of noise inside restaurants was the stoves. The mean hearing thresholds showed a typical dip at 3 to 6 KHz and a substantial proportion (23.7%) of the workers fulfilled the criteria for presumptive NIHL. For entertainment sectors, employees in radio and television stations generally had higher exposure levels than those in the halls or auditoria of the LCSD and karaoke bars. The mean hearing thresholds showed a typical dip at 6 KHz and a substantial proportion of the employees fulfilled the criteria for presumptive NIHL (38.6%, 95%CI: 35.1–42.1%). Being male, older, and having longer service and daily alcohol consumption were associated with noise-induced hearing impairment both in restaurant workers and entertainment employees.

Conclusion

Excessive noise exposure is common in the Chinese restaurant and entertainment industries and a substantial proportion of restaurant workers and entertainment employees suffer from NIHL. Comprehensive hearing conservation programs should be introduced to the service industry in Hong Kong.