Bacterial behavior on coated porous titanium substrates for biomedical applications

In this work, bacterial behavior on dense and porous titanium substrates is discussed. Porous titanium was fabricated by a space holder technique using 50 vol , NH4HCO3 with particle sizes between 250 and 355 amp; 956;m . These substrates were coated by sulfonated PEEK termed SPEEK . Characterization of the porous substrate was carried out using the Archimedes Method, Image Analysis, and three dimensional X ray Micro Computed Tomography including total and interconnected porosity, equivalent diameter, and pore shape factor , as well as mechanical characterization specifically stiffness and yield strength . A detailed study was performed here to investigate the influence of substrate porosity on the adhesion and proliferation of E. coli, MRSA, and P. aeruginosa common causes of orthopedic device associated infections . Bacterial colonization was examined in terms of the initial bacterial concentration, as well as bacterial adherence to and growth on the surface and inside the pores. Results suggest that fully dense titanium supported the least bacterial colonization, while the porous titanium promoted bacterial growth in the medium and inside the cavities. Furthermore, the SPEEK coating deposited onto the samples inhibited bacteria growth inside the porous materials. In this manner, this study showed for the first time that SPEEK could have potential antibacterial properties to offset the increase in bacteria growth commonly observed in porous material

Therapeutic Potential of a Novel Vitamin D3 Oxime Analogue, VD1-6, with CYP24A1 Enzyme Inhibitory Activity and Negligible Vitamin D Receptor Binding

Abstract: The regulation of vitamin D3 actions in humans occurs mainly through the Cytochrome P450 24-hydroxylase (CYP24A1) enzyme activity. CYP24A1 hydroxylates both 25-hydroxycholecalciferol (25(OH)D3) and 1,25-dihydroxycholecalciferol (1,25(OH)2D3), which is the first step of vitamin D catabolism. An abnormal status of the upregulation of CYP24A1 occurs in many diseases, including chronic kidney disease (CKD). CYP24A1 upregulation in CKD and diminished activation of vitamin D3 contribute to secondary hyperparathyroidism (SHPT), progressive bone deterioration, and soft tissue and cardiovascular calcification. Previous studies have indicated that CYP24A1 inhibition may be an effective strategy to increase endogenous vitamin D activity and decrease SHPT. This study has designed and synthesized a novel C-24 O-methyloxime analogue of vitamin D3 (VD1-6) to have specific CYP24A1 inhibitory properties. VD1-6 did not bind to the vitamin D receptor (VDR) in concentrations up to 10-7 M, assessed by a VDR binding assay. The absence of VDR binding by VD1-6 was confirmed in human embryonic kidney HEK293T cultures through the lack of CYP24A1 induction. However, in silico docking experiments demonstrated that VD1-6 was predicted to have superior binding to CYP24A1, when compared to that of 1,25(OH)2D3. The inhibition of CYP24A1 by VD1-6 was also evident by the synergistic potentiation of 1,25(OH)2D3-mediated transcription and reduced 1,25(OH)2D3 catabolism over 24 h. A further indication of CYP24A1 inhibition by VD1-6 was the reduced accumulation of the 24,25(OH)D3, the first metabolite of 25(OH)D catabolism by CYP24A1. Our findings suggest the potent CYP24A1 inhibitory properties of VD1-6 and its potential for testing as an alternative therapeutic candidate for treating SHPT.Ali K. Alshabrawy, Yingjie Cui, Cyan Sylvester, Dongqing Yang, Emilio S. Petito, Kate R. Barratt, Rebecca K. Sawyer, Jessica K. Heatlie, Ruhi Polara, Matthew J. Sykes, Gerald J. Atkins, Shane M. Hickey, Michael D. Wiese, Andrea M. Stringer, Zhaopeng Liu, and Paul H. Anderso

Prediction of Prostate Cancer Biochemical and Clinical Recurrence Is Improved by IHC-Assisted Grading Using Appl1, Sortilin and Syndecan-1.

Gleason scoring is used within a five-tier risk stratification system to guide therapeutic decisions for patients with prostate cancer. This study aimed to compare the predictive performance of routine H&E or biomarker-assisted ISUP (International Society of Urological Pathology) grade grouping for assessing the risk of biochemical recurrence (BCR) and clinical recurrence (CR) in patients with prostate cancer. This retrospective study was an assessment of 114 men with prostate cancer who provided radical prostatectomy samples to the Australian Prostate Cancer Bioresource between 2006 and 2014. The prediction of CR was the primary outcome (median time to CR 79.8 months), and BCR was assessed as a secondary outcome (median time to BCR 41.7 months). The associations of (1) H&E ISUP grade groups and (2) modified ISUP grade groups informed by the Appl1, Sortilin and Syndecan-1 immunohistochemistry (IHC) labelling were modelled with BCR and CR using Cox proportional hazard approaches. IHC-assisted grading was more predictive than H&E for BCR (C-statistic 0.63 vs. 0.59) and CR (C-statistic 0.71 vs. 0.66). On adjusted analysis, IHC-assisted ISUP grading was independently associated with both outcome measures. IHC-assisted ISUP grading using the biomarker panel was an independent predictor of individual BCR and CR. Prospective studies are needed to further validate this biomarker technology and to define BCR and CR associations in real-world cohorts.Jessica M. Logan ... Lisa M. Butler ... Douglas A. Brooks ... et al

A draft human pangenome reference

Here the Human Pangenome Reference Consortium presents a first draft of the human pangenome reference. The pangenome contains 47 phased, diploid assemblies from a cohort of genetically diverse individuals. These assemblies cover more than 99% of the expected sequence in each genome and are more than 99% accurate at the structural and base pair levels. Based on alignments of the assemblies, we generate a draft pangenome that captures known variants and haplotypes and reveals new alleles at structurally complex loci. We also add 119 million base pairs of euchromatic polymorphic sequences and 1,115 gene duplications relative to the existing reference GRCh38. Roughly 90 million of the additional base pairs are derived from structural variation. Using our draft pangenome to analyse short-read data reduced small variant discovery errors by 34% and increased the number of structural variants detected per haplotype by 104% compared with GRCh38-based workflows, which enabled the typing of the vast majority of structural variant alleles per sample

Understanding the genetic complexity of puberty timing across the allele frequency spectrum

Pubertal timing varies considerably and is associated with later health outcomes. We performed multi-ancestry genetic analyses on ~800,000 women, identifying 1,080 signals for age at menarche. Collectively, these explained 11% of trait variance in an independent sample. Women at the top and bottom 1% of polygenic risk exhibited ~11 and ~14-fold higher risks of delayed and precocious puberty, respectively. We identified several genes harboring rare loss-of-function variants in ~200,000 women, including variants in ZNF483, which abolished the impact of polygenic risk. Variant-to-gene mapping approaches and mouse gonadotropin-releasing hormone neuron RNA sequencing implicated 665 genes, including an uncharacterized G-protein-coupled receptor, GPR83, which amplified the signaling of MC3R, a key nutritional sensor. Shared signals with menopause timing at genes involved in DNA damage response suggest that the ovarian reserve might signal centrally to trigger puberty. We also highlight body size-dependent and independent mechanisms that potentially link reproductive timing to later life disease

The combined actions of DHT and MPA lead to altered AR signaling in normal and malignant post-menopausal breast epithelial cells

Abstract Consistent with several observational studies examining combined hormone replacement therapy (cHRT: conjugated equine estrogen in conjunction with the synthetic progestin medroxyprogesterone acetate, MPA) in postmenopausal women, a re-analysis of the placebo-controlled randomized Women's Health Initiative clinical trial demonstrated a markedly increased breast cancer risk in newly menopausal women following ≥ 5 years of cHRT (Hazard Ratio, 3.05; 95% Confidence Interval, 1.62-5.70) [1]. We have previously demonstrated that androgen receptor (AR)-mediated effects of MPA impede 5α-dihydrotestosterone (DHT)-induced AR signaling in normal and malignant breast epithelial cells (AACR abstract 2010). The current study aimed to further investigate the biological actions of DHT and/or MPA on steroid receptor expression and cancer-related intracellular signaling pathways. Immunohistochemical analysis of estrogen receptor alpha (ERα), progesterone receptor (PR) and AR expression was performed on histologically normal human post-menopausal breast tissues and measured by image analysis in tissues cultured ex vivo with vehicle (0.1% ethanol control), DHT (1nM), MPA (1nM) or the AR antagonist, bicalutamide (Bic;1uM) for 48 hr, either alone or in combination. Microarray analysis and qRT-PCR validation were performed using the ERα positive breast cancer cell line, ZR-75-1 to determine changes in gene expression in key intracellular signaling pathways. The microarray data was analyzed with Ingenuity Gene Pathway Analysis and Gene Ontology software. Statistical tests included both Wilcoxon matched pairs test and one-way ANOVA (p<0.05). DHT treatment increased AR expression in cultured breast tissues compared to vehicle control (p<0.05), and co-treatment with either MPA or Bic impeded this effect. No change in ERα or PR protein levels was induced by the hormone treatments. Microarray studies revealed that DHT or MPA treatment for 6 hr altered the expression of 439 and 858 genes, respectively, whereas co-treatment altered 1494 genes (p<0.05). Only 114 genes were uniquely regulated by DHT, and the expression of 32% (41% induced and 27% repressed) of these genes was abrogated by MPA. Similarly, the expression of 38% (51% induced and 24% repressed) of the 690 genes uniquely regulated by co-treatment with DHT and MPA was altered compared to DHT alone (p<0.05). Examples of genes that were regulated by DHT (p<0.05) and this effect of DHT was antagonised by co-treatment with DHT and MPA (p<0.05) are FGFR2, OLR1 and C1ORF116. Co-treatment with DHT and MPA altered the expression of genes involved in cell growth, cell cycle, cell death, cancer and intracellular signaling pathways compared to individual treatments (p<0.05). Collectively, these findings suggest an AR-mediated mechanism for the action of MPA in breast cancer. 1.Prentice, R.L., et al. Am J Epidemiol, 2009 170(1): 12-23. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 274. doi:1538-7445.AM2012-274Aleksandra M. Ochnik, Nicole L. Moore, Stephen N. Birrell, Lisa M. Butler, Shalini Jindal, Luke Selth, Wayne D. Tilley and Theresa E. Hicke

Pangenome graph construction from genome alignments with Minigraph-Cactus

Pangenome references address biases of reference genomes by storing a representative set of diverse haplotypes and their alignment, usually as a graph. Alternate alleles determined by variant callers can be used to construct pangenome graphs, but advances in long-read sequencing are leading to widely available, high-quality phased assemblies. Constructing a pangenome graph directly from assemblies, as opposed to variant calls, leverages the graph’s ability to represent variation at different scales. Here we present the Minigraph-Cactus pangenome pipeline, which creates pangenomes directly from whole-genome alignments, and demonstrate its ability to scale to 90 human haplotypes from the Human Pangenome Reference Consortium. The method builds graphs containing all forms of genetic variation while allowing use of current mapping and genotyping tools. We measure the effect of the quality and completeness of reference genomes used for analysis within the pangenomes and show that using the CHM13 reference from the Telomere-to-Telomere Consortium improves the accuracy of our methods. We also demonstrate construction of a Drosophila melanogaster pangenome