Correlation of tumor-associated macrophages and clinicopathological factors in Wilms tumor

Background/purpose: Despite high long-term survival rates in patients with Wilms tumor, there is a need to develop better prognostic biomarkers in order to maximize cure while avoiding treatment-associated morbidities. Tumor-associated macrophages have been recently associated with poorer prognosis and increased disease progression in a number of adult cancers. We investigated the relationship between macrophages and clinicopathological fators in this pediatric solid tumor. Methods: Tissue microarray sections of 124 Wilms tumor cases obtained from the Cooperative Human Tissue Network were stained with CD68, a macrophage marker using standard immunohistochemical techniques and quantified using digital image processing techniques. Macrophage densities were correlated by tumor stage, and survival analyses were conducted with available clinical data. Immunohistochemistry was performed on 25 additional Wilms tumor cases obtained from the tumor bank at Columbia University Medical Center and correlated with presence of tumor microvascular invasion. Results: Mean macrophage count densities in stage IV tumors were significantly greater than densities in stage I and III tumors (p=.021, .036). Although the overall and disease-free survival did not differ between high and low macrophage presence groups across all stages, increased macrophage presence was associated with decreased disease-free survival in patients with stage II tumors (p=0.035). Increased macrophage presence may have also correlated with decreased disease-free survival in stage IV patients, but the sample size was too small to allow detection of this difference with significance (p=0.575). Increased macrophage presence was associated with tumor microvascular invasion (p=0.0004). Conclusion: Our results suggest that macrophage recruitment may be associated with disease progression in Wilms tumor. Quantitation of macrophage presence may therefore be a useful adjunct in refining staging algorithms for patients with stage II Wilms tumor. Such data might be useful in the effort to reduce the risk of adverse effects associated with under- or overtreatment of this neoplasm

Mapping the Hsp90 Genetic Interaction Network in Candida albicans Reveals Environmental Contingency and Rewired Circuitry

The molecular chaperone Hsp90 regulates the folding of diverse signal transducers in all eukaryotes, profoundly affecting cellular circuitry. In fungi, Hsp90 influences development, drug resistance, and evolution. Hsp90 interacts with ∼10% of the proteome in the model yeast Saccharomyces cerevisiae, while only two interactions have been identified in Candida albicans, the leading fungal pathogen of humans. Utilizing a chemical genomic approach, we mapped the C. albicans Hsp90 interaction network under diverse stress conditions. The chaperone network is environmentally contingent, and most of the 226 genetic interactors are important for growth only under specific conditions, suggesting that they operate downstream of Hsp90, as with the MAPK Hog1. Few interactors are important for growth in many environments, and these are poised to operate upstream of Hsp90, as with the protein kinase CK2 and the transcription factor Ahr1. We establish environmental contingency in the first chaperone network of a fungal pathogen, novel effectors upstream and downstream of Hsp90, and network rewiring over evolutionary time

Generational distribution of a Candida glabrata population: Resilient old cells prevail, while younger cells dominate in the vulnerable host.

Similar to other yeasts, the human pathogen Candida glabrata ages when it undergoes asymmetric, finite cell divisions, which determines its replicative lifespan. We sought to investigate if and how aging changes resilience of C. glabrata populations in the host environment. Our data demonstrate that old C. glabrata are more resistant to hydrogen peroxide and neutrophil killing, whereas young cells adhere better to epithelial cell layers. Consequently, virulence of old compared to younger C. glabrata cells is enhanced in the Galleria mellonella infection model. Electron microscopy images of old C. glabrata cells indicate a marked increase in cell wall thickness. Comparison of transcriptomes of old and young C. glabrata cells reveals differential regulation of ergosterol and Hog pathway associated genes as well as adhesion proteins, and suggests that aging is accompanied by remodeling of the fungal cell wall. Biochemical analysis supports this conclusion as older cells exhibit a qualitatively different lipid composition, leading to the observed increased emergence of fluconazole resistance when grown in the presence of fluconazole selection pressure. Older C. glabrata cells accumulate during murine and human infection, which is statistically unlikely without very strong selection. Therefore, we tested the hypothesis that neutrophils constitute the predominant selection pressure in vivo. When we altered experimentally the selection pressure by antibody-mediated removal of neutrophils, we observed a significantly younger pathogen population in mice. Mathematical modeling confirmed that differential selection of older cells is sufficient to cause the observed demographic shift in the fungal population. Hence our data support the concept that pathogenesis is affected by the generational age distribution of the infecting C. glabrata population in a host. We conclude that replicative aging constitutes an emerging trait, which is selected by the host and may even play an unanticipated role in the transition from a commensal to a pathogen state.post-print10768 K

Genetic Determinants of Intrinsic Antibiotic Tolerance in Mycobacterium avium.

The Mycobacterium avium complex (MAC) is one of the most prevalent causes of nontuberculous mycobacteria pulmonary infection in the United States, and yet it remains understudied. Current MAC treatment requires more than a year of intermittent to daily combination antibiotic therapy, depending on disease severity. In order to shorten and simplify curative regimens, it is important to identify the innate bacterial factors contributing to reduced antibiotic susceptibility, namely, antibiotic tolerance genes. In this study, we performed a genome-wide transposon screen to elucidate M. avium genes that play a role in the bacterium\u27s tolerance to first- and second-line antibiotics. We identified a total of 193 unique M. avium mutants with significantly altered susceptibility to at least one of the four clinically used antibiotics we tested, including two mutants (in DFS55_00905 and DFS55_12730) with panhypersusceptibility. The products of the antibiotic tolerance genes we have identified may represent novel targets for future drug development studies aimed at shortening the duration of therapy for MAC infections

The protein kinase CK2 regulatory subunits regulate function of the Hsp90/Cdc37 protein complex.

<p>(A) Hsp90 serine and threonine phosphorylation is severely reduced in the <i>ckb1</i>Δ/<i>ckb1</i>Δ mutant, and Cdc37 serine and threonine phosphorylation is severely reduced in both the <i>ckb1</i>Δ/<i>ckb1</i>Δ and <i>ckb2</i>Δ/<i>ckb2</i>Δ mutants. Hsp90 or Cdc37 were immunoprecipitated and Western blots were hybridized with CaHsp90, TAP (to detect Cdc37-TAP), phosphothreonine, or phosphoserine antibodies. The ratio of phosphorylated to unphosphorylated Hsp90 or Cdc37 in each CK2 mutant was quantified relative to the wild type. (B) Western analysis demonstrates that Cdc37 levels are severely reduced (>50%, red box) in the mutant lacking the regulatory subunit Ckb2 (<i>ckb2</i>Δ/<i>ckb2</i>Δ) in the absence of external stress compared to the wild type (WT, BWP17); Hsp90 and Hog1 levels, however, are reduced (>25%, yellow box) in strains that lack the regulatory subunits (<i>ckb1</i>Δ/<i>ckb1</i>Δ or <i>ckb2</i>Δ/<i>ckb2</i>Δ) in response to oxidative stress in the form of a 10 minute treatment with 1 mM hydrogen peroxide. Actin served as loading control. (C) Deletion of CK2 regulatory subunits, <i>CKB1</i> or <i>CKB2</i>, phenocopies deletion of <i>HOG1</i> in terms of hypersensitivity to high osmolarity stress exerted by sorbitol. Growth is quantitatively displayed with color as indicated with the color bar. (D) Our results support a model in which the regulatory subunits of CK2 (Ckb1 and Ckb2) affect phosphorylation of Hsp90 and Cdc37, protein levels of the Hsp90-Cdc37 complex under basal or stress conditions, and levels of the target kinase Hog1.</p

Hsp90 genetic interactions identified in six screens.

<p>Hsp90 genetic interactions identified in six screens.</p

Hsp90 kinase genetic interaction network.

<p>Of the 226 interactions, 34 are with kinases. Kinases are color-coded depending on their degree of connectivity, ranging from grey for one connection to orange for five connections. Kinases and test conditions (black squares) are connected with each other via edges. While the caspofungin screen shared only one of its kinase interactors (<i>CKB2</i>) with another screen, every other screen shared half or more of its interactors with another screen. For six kinases (diamonds), protein levels were measured upon Hsp90 depletion.</p

The high-connectivity interactor Ahr1 influences <i>HSP90</i> expression and morphogenesis.

<p>(A) <i>HSP90</i> transcript levels are reduced in the <i>ahr1</i>Δ/<i>ahr1</i>Δ mutant. <i>HSP90</i> transcript levels were measured in the wild type (SN152), the <i>ahr1</i>Δ/<i>ahr1</i>Δ mutant, and the <i>AHR1</i> complemented strain by quantitative RT-PCR and normalized to <i>GPD1</i>. Data are means ± standard deviation for triplicate samples. (B) The <i>ahr1</i>Δ/<i>ahr1</i>Δ mutant filaments in rich medium at 30°C, consistent with the effects of compromised Hsp90 function. Differential Interference Contrast microscopy of strains incubated in rich medium at 30°C for 24 hours with or without 10 µM geldanamycin (GdA). Scale bar is 10 µm.</p

Model of high- and low-connectivity interactors identified in this screen that either modify the Hsp90/Cdc37 complex or are affected by it.

<p>The high connectivity interactors (red) modulate gene expression, protein levels, or phosphorylation of Hsp90 and Cdc37, while the chaperone complex regulates gene expression, protein levels or activation of low-connectivity interactors (blue).</p

Early adolescent gender diversity and mental health in the Adolescent Brain Cognitive Development study

BackgroundThere are known associations between mental health symptoms and transgender identity among adults. Whether this relationship extends to early adolescents and to gender domains other than identity is unclear. This study measured dimensions of gender in a large, diverse, sample of youth, and examined associations between diverse gender experiences and mental health.MethodsThe ABCD study is an ongoing, longitudinal, US cohort study. Baseline data (release 2.0) include 11,873 youth age 9/10 (48% female); and the 4,951 1-year follow-up visits (age 10/11; 48% female) completed prior to data release. A novel gender survey at the 1-year visit assessed felt-gender, gender noncontentedness, and gender nonconformity using a 5-point scale. Mental health measures included youth- and parent-reports.ResultsRoughly half a percent of 9/10-year-olds (n&nbsp;=&nbsp;58) responded 'yes' or 'maybe' when asked, 'Are you transgender' at baseline. Recurrent thoughts of death were more prevalent among these youth compared to the rest of the cohort (19.6% vs. 6.4%, χ2 &nbsp;=&nbsp;16.0, p&nbsp;&lt;&nbsp;.001). At the 1-year visit, when asked about the three dimensions of gender on a 5-point scale, 33.2% (n&nbsp;=&nbsp;1,605) provided responses that were not exclusively and totally aligned with one gender. Significant relationships were observed between mental health symptoms and gender diversity for all dimensions assessed.ConclusionsSimilar to adult studies, early adolescents identifying as transgender reported increased mental health symptoms. Results also point to considerable diversity in other dimensions of gender (felt-gender, gender noncontentedness, gender nonconformity) among 10/11-year-olds, and find this diversity to be related to critical mental health symptoms. These findings add to our limited understanding of the relationship between dimensions of gender and wellness for youth