Tissue Harvester with Functional Valve (THFV): Shidham's device for reproducibly higher specimen yield by fine needle aspiration biopsy with easy to perform steps

BACKGROUND: Fine needle aspiration biopsy (FNAB) cytology has been a highly effective methodology for tissue diagnosis and for various ancillary studies including molecular tests. In addition to other benefits, FNAB predominantly retrieves the diagnostic loosely cohesive cells in the lesion as compared to the adjacent supporting stroma with relatively higher cohesiveness. However, FNAB procedure performed with currently available resources is highly skill dependent with inter-performer variability, which compromises its full potential as a diagnostic tool. In this study we report a device overcoming these limitations. METHODS: 'Tissue Harvester with Functional Valve' (THFV) was evaluated as part of a phase 1 National Institute of Health (NIH) research grant under Small Business Technology Transfer (STTR) Program. Working prototypes of the device were prepared. Each of the four cytopathologists with previous cytopathology fellowship training and experience in performing FNAB evaluated 5 THFV and 5 hypodermic needles resulting in 40 specimens (20 with THFV, 20 with hypodermic needles). A piece of fresh cattle liver stuffed in latex glove was used as the specimen. Based on these results a finished design was finalized. RESULTS: The smears and cell blocks prepared from the specimens obtained by THFV were superior in terms of cellularity to specimens obtained with hypodermic needles. The tissuecrit of specimens obtained with THFV ranged from 70 to 100 μl (mean 87, SD 10), compared to 17 to 30 μl (mean 24, SD 4) with conventional hypodermic needles (p < .0001, Student t-test). The technical ease [on a scale of 1 (easy) to 5 (difficult)] with THFV ranged from 1 to 2 as compared to 2 to 3 with hypodermic needles. CONCLUSION: The specimen yield with the new THFV was significantly higher when compared to hypodermic needles. Also, the FNAB procedure with THFV was relatively easier in comparison with hypodermic needles. The final version of Shidham's THFV device would improve the FNAB specimen yield by eliminating the skill factor. The increased specimen yield by this device would also facilitate wider application of FNAB specimens for various ancillary tests, including molecular tests

Recovery of a US Endangered Fish

BACKGROUND: More fish have been afforded US Endangered Species Act protection than any other vertebrate taxonomic group, and none has been designated as recovered. Shortnose sturgeon (Acipenser brevirostrum) occupy large rivers and estuaries along the Atlantic coast of North America, and the species has been protected by the US Endangered Species Act since its enactment. METHODOLOGY/PRINCIPAL FINDINGS: Data on the shortnose sturgeon in the Hudson River (New York to Albany, NY, USA) were obtained from a 1970s population study, a population and fish distribution study we conducted in the late 1990s, and a fish monitoring program during the 1980s and 1990s. Population estimates indicate a late 1990s abundance of about 60,000 fish, dominated by adults. The Hudson River population has increased by more than 400% since the 1970s, appears healthy, and has attributes typical for a long-lived species. Our population estimates exceed the government and scientific population recovery criteria by more than 500%, we found a positive trend in population abundance, and key habitats have remained intact despite heavy human river use. CONCLUSIONS/SIGNIFICANCE: Scientists and legislators have called for changes in the US Endangered Species Act, the Act is being debated in the US Congress, and the Act has been characterized as failing to recover species. Recovery of the Hudson River population of shortnose sturgeon suggests the combination of species and habitat protection with patience can yield successful species recovery, even near one of the world's largest human population centers

A Genetic Screen for Dihydropyridine (DHP)-Resistant Worms Reveals New Residues Required for DHP-Blockage of Mammalian Calcium Channels

Dihydropyridines (DHPs) are L-type calcium channel (Cav1) blockers prescribed to treat several diseases including hypertension. Cav1 channels normally exist in three states: a resting closed state, an open state that is triggered by membrane depolarization, followed by a non-conducting inactivated state that is triggered by the influx of calcium ions, and a rapid change in voltage. DHP binding is thought to alter the conformation of the channel, possibly by engaging a mechanism similar to voltage dependent inactivation, and locking a calcium ion in the pore, thereby blocking channel conductance. As a Cav1 channel crystal structure is lacking, the current model of DHP action has largely been achieved by investigating the role of candidate Cav1 residues in mediating DHP-sensitivity. To better understand DHP-block and identify additional Cav1 residues important for DHP-sensitivity, we screened 440,000 randomly mutated Caenorhabditis elegans genomes for worms resistant to DHP-induced growth defects. We identified 30 missense mutations in the worm Cav1 pore-forming (α1) subunit, including eleven in conserved residues known to be necessary for DHP-binding. The remaining polymorphisms are in eight conserved residues not previously associated with DHP-sensitivity. Intriguingly, all of the worm mutants that we analyzed phenotypically exhibited increased channel activity. We also created orthologous mutations in the rat α1C subunit and examined the DHP-block of current through the mutant channels in culture. Six of the seven mutant channels examined either decreased the DHP-sensitivity of the channel and/or exhibited significant residual current at DHP concentrations sufficient to block wild-type channels. Our results further support the idea that DHP-block is intimately associated with voltage dependent inactivation and underscores the utility of C. elegans as a screening tool to identify residues important for DHP interaction with mammalian Cav1 channels

Maternal Behavior is Impaired in Female Mice Lacking Type 3 Adenylyl Cyclase

Although chemosensory signals generated by mouse pups may trigger maternal behavior of females, the mechanism for detection of these signals has not been fully defined. As some odorant receptors are coupled to the type 3 adenylyl cyclase (AC3), we evaluated the role of AC3 for maternal behavior using AC3−/− female mice. Here, we report that maternal behavior is impaired in virgin and postpartum AC3−/− mice. Female AC3−/− mice failed the pup retrieval assay, did not construct well-defined nests, and did not exhibit maternal aggression. Furthermore, AC3−/− females could not detect odorants or pup urine in the odorant habituation test and were unable to detect pups by chemoreception. In contrast to wild-type mice, AC activity in main olfactory epithelium (MOE) preparations from AC3−/− female mice was not stimulated by odorants or pheromones. Moreover, odorants and pheromones did not evoke electro-olfactogram (EOG) responses in the MOE of AC3−/− female mice. We hypothesize that the detection of chemical signals that trigger maternal behavior in female mice depends upon AC3 in the MOE

Investigation of Exomic Variants Associated with Overall Survival in Ovarian Cancer

BACKGROUND: While numerous susceptibility loci for epithelial ovarian cancer (EOC) have been identified, few associations have been reported with overall survival. In the absence of common prognostic genetic markers, we hypothesize that rare coding variants may be associated with overall EOC survival and assessed their contribution in two exome-based genotyping projects of the Ovarian Cancer Association Consortium (OCAC). METHODS: The primary patient set (Set 1) included 14 independent EOC studies (4,293 patients) and 227,892 variants, and a secondary patient set (Set 2) included six additional EOC studies (1,744 patients) and 114,620 variants. Because power to detect rare variants individually is reduced, gene-level tests were conducted. Sets were analyzed separately at individual variants and by gene, and then combined with meta-analyses (73,203 variants and 13,163 genes overlapped). RESULTS: No individual variant reached genome-wide statistical significance. A SNP previously implicated to be associated with EOC risk and, to a lesser extent, survival, rs8170, showed the strongest evidence of association with survival and similar effect size estimates across sets (Pmeta = 1.1E-6, HRSet1 = 1.17, HRSet2 = 1.14). Rare variants in ATG2B, an autophagy gene important for apoptosis, were significantly associated with survival after multiple testing correction (Pmeta = 1.1E-6; Pcorrected = 0.01). CONCLUSIONS: Common variant rs8170 and rare variants in ATG2B may be associated with EOC overall survival, although further study is needed. IMPACT: This study represents the first exome-wide association study of EOC survival to include rare variant analyses, and suggests that complementary single variant and gene-level analyses in large studies are needed to identify rare variants that warrant follow-up study

The development of sex differences in ring-tailed lemur feeding ecology

Sex differences in feeding ecology may develop in response to fluctuations in physiological costs to females over their reproductive cycles, or to sexual size dimorphism, or function to minimize feeding competition within a group via resource partitioning. For most mammal species, it is unknown how these factors contribute to sex differences in feeding, or how the development of males and females reflects these intraspecific feeding differences. We show changes in dietary composition, diversity, overlap, and foraging behavior throughout development in ring-tailed lemurs (Lemur catta) and test how the development of sex differences in feeding is related to female costs of reproduction and year-round resource partitioning. Sex differences in dietary composition were only present when females were lactating, but sex differences in other aspects of feeding, including dietary diversity, and relative time spent feeding and foraging, developed at or near the time of weaning. Sex difference in juveniles and subadults, when present, were similar to the differences found in adults. The low year-round dietary overlap and early differences in dietary diversity indicate that some resource partitioning may begin with young individuals and fluctuate throughout development. The major differences between males and females in dietary composition suggest that these larger changes in diet are closely tied to female reproductive state when females must shift their diet to meet energetic and nutritional requirements

Network-Based Integration of GWAS and Gene Expression Identifies a HOX-Centric Network Associated with Serous Ovarian Cancer Risk

BACKGROUND: Genome-wide association studies (GWAS) have so far reported 12 loci associated with serous epithelial ovarian cancer (EOC) risk. We hypothesized that some of these loci function through nearby transcription factor (TF) genes and that putative target genes of these TFs as identified by coexpression may also be enriched for additional EOC risk associations. METHODS: We selected TF genes within 1 Mb of the top signal at the 12 genome-wide significant risk loci. Mutual information, a form of correlation, was used to build networks of genes strongly coexpressed with each selected TF gene in the unified microarray dataset of 489 serous EOC tumors from The Cancer Genome Atlas. Genes represented in this dataset were subsequently ranked using a gene-level test based on results for germline SNPs from a serous EOC GWAS meta-analysis (2,196 cases/4,396 controls). RESULTS: Gene set enrichment analysis identified six networks centered on TF genes (HOXB2, HOXB5, HOXB6, HOXB7 at 17q21.32 and HOXD1, HOXD3 at 2q31) that were significantly enriched for genes from the risk-associated end of the ranked list (P < 0.05 and FDR < 0.05). These results were replicated (P < 0.05) using an independent association study (7,035 cases/21,693 controls). Genes underlying enrichment in the six networks were pooled into a combined network. CONCLUSION: We identified a HOX-centric network associated with serous EOC risk containing several genes with known or emerging roles in serous EOC development. IMPACT: Network analysis integrating large, context-specific datasets has the potential to offer mechanistic insights into cancer susceptibility and prioritize genes for experimental characterization

Assessment of variation in immunosuppressive pathway genes reveals TGFBR2 to be associated with risk of clear cell ovarian cancer

BACKGROUND: Regulatory T (Treg) cells, a subset of CD4+ T lymphocytes, are mediators of immunosuppression in cancer, and, thus, variants in genes encoding Treg cell immune molecules could be associated with ovarian cancer. METHODS: In a population of 15,596 epithelial ovarian cancer (EOC) cases and 23,236 controls, we measured genetic associations of 1,351 SNPs in Treg cell pathway genes with odds of ovarian cancer and tested pathway and gene-level associations, overall and by histotype, for the 25 genes, using the admixture likelihood (AML) method. The most significant single SNP associations were tested for correlation with expression levels in 44 ovarian cancer patients. RESULTS: The most significant global associations for all genes in the pathway were seen in endometrioid (p = 0.082) and clear cell (p = 0.083), with the most significant gene level association seen with TGFBR2 (p = 0.001) and clear cell EOC. Gene associations with histotypes at p < 0.05 included: IL12 (p = 0.005 and p = 0.008, serous and high-grade serous, respectively), IL8RA (p = 0.035, endometrioid and mucinous), LGALS1 (p = 0.03, mucinous), STAT5B (p = 0.022, clear cell), TGFBR1 (p = 0.021 endometrioid) and TGFBR2 (p = 0.017 and p = 0.025, endometrioid and mucinous, respectively). CONCLUSIONS: Common inherited gene variation in Treg cell pathways shows some evidence of germline genetic contribution to odds of EOC that varies by histologic subtype and may be associated with mRNA expression of immune-complex receptor in EOC patients

Variants in genes encoding small GTPases and association with epithelial ovarian cancer susceptibility

Epithelial ovarian cancer (EOC) is the fifth leading cause of cancer mortality in American women. Normal ovarian physiology is intricately connected to small GTP binding proteins of the Ras superfamily (Ras, Rho, Rab, Arf, and Ran) which govern processes such as signal transduction, cell proliferation, cell motility, and vesicle transport. We hypothesized that common germline variation in genes encoding small GTPases is associated with EOC risk. We investigated 322 variants in 88 small GTPase genes in germline DNA of 18,736 EOC patients and 26,138 controls of European ancestry using a custom genotype array and logistic regression fitting log-additive models. Functional annotation was used to identify biofeatures and expression quantitative trait loci that intersect with risk variants. One variant, ARHGEF10L (Rho guanine nucleotide exchange factor 10 like) rs2256787, was associated with increased endometrioid EOC risk (OR=1.33, p=4.46 x 10-6). Other variants of interest included another in ARHGEF10L, rs10788679, which was associated with invasive serous EOC risk (OR=1.07, p=0.00026) and two variants in AKAP6 (A-kinase anchoring protein 6) which were associated with risk of invasive EOC (rs1955513, OR=0.90, p=0.00033; rs927062, OR =0.94, p=0.00059). Functional annotation revealed that the two ARHGEF10L variants were located in super-enhancer regions and that AKAP6 rs927062 was associated with expression of GTPase gene ARHGAP5 (Rho GTPase activating protein 5). Inherited variants in ARHGEF10L and AKAP6, with potential transcriptional regulatory function and association with EOC risk, warrant investigation in independent EOC study populations