TMEM45A, SERPINB5 and p16INK4A transcript levels are predictive for development of high-grade cervical lesions

Women persistently infected with human papillomavirus (HPV) type 16 are at high risk for development of cervical intraepithelial neoplasia grade 3 or cervical cancer (CIN3+). We aimed to identify biomarkers for progression to CIN3+ in women with persistent HPV16 infection. In this prospective study, 11,088 women aged 20-29 years were enrolled during 1991-1993, and re-invited for a second visit two years later. Cervical cytology samples obtained at both visits were tested for HPV DNA by Hybrid Capture 2 (HC2), and HC2-positive samples were genotyped by INNO-LiPA. The cohort was followed for up to 19 years via a national pathology register. To identify markers for progression to CIN3+, we performed microarray analysis on RNA extracted from cervical swabs of 30 women with persistent HPV16-infection and 11 HPV-negative women. Six genes were selected and validated by quantitative PCR. Three genes were subsequently validated within a different and large group of women from the same cohort. Secondly, Kaplan-Meier and Cox-regression analyses were used to investigate whether expression levels of those three genes predict progression to CIN3+. We found that high transcript levels of TMEM45A, SERPINB5 and p16INK4a at baseline were associated with increased risk of CIN3+ during follow-up. The hazard ratios of CIN3+ per 10-fold increase in baseline expression level were 1.6 (95% CI: 1.1-2.3) for TMEM45A, 1.6 (95% CI: 1.1-2.5) for p16INK4a, and 1.8 (95% CI: 1.2-2.7) for SERPINB5. In conclusion, high mRNA expression levels of TMEM45A, SERPINB5 and p16INK4a were associated with increased risk of CIN3+ in persistently HPV16-infected women

Ezrin interacts with the SARS coronavirus spike protein and restrains infection at the entry stage

© 2012 Millet et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Background: Entry of Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) and its envelope fusion with host cell membrane are controlled by a series of complex molecular mechanisms, largely dependent on the viral envelope glycoprotein Spike (S). There are still many unknowns on the implication of cellular factors that regulate the entry process. Methodology/Principal Findings: We performed a yeast two-hybrid screen using as bait the carboxy-terminal endodomain of S, which faces the cytosol during and after opening of the fusion pore at early stages of the virus life cycle. Here we show that the ezrin membrane-actin linker interacts with S endodomain through the F1 lobe of its FERM domain and that both the eight carboxy-terminal amino-acids and a membrane-proximal cysteine cluster of S endodomain are important for this interaction in vitro. Interestingly, we found that ezrin is present at the site of entry of S-pseudotyped lentiviral particles in Vero E6 cells. Targeting ezrin function by small interfering RNA increased S-mediated entry of pseudotyped particles in epithelial cells. Furthermore, deletion of the eight carboxy-terminal amino acids of S enhanced S-pseudotyped particles infection. Expression of the ezrin dominant negative FERM domain enhanced cell susceptibility to infection by SARS-CoV and S pseudotyped particles and potentiated S-dependent membrane fusion. Conclusions/Significance: Ezrin interacts with SARS-CoV S endodomain and limits virus entry and fusion. Our data present a novel mechanism involving a cellular factor in the regulation of S-dependent early events of infection.This work was supported by the Research Grant Council of Hong Kong (RGC#760208)and the RESPARI project of the International Network of Pasteur Institutes

Three-dimensional optoacoustic imaging of nailfold capillaries in systemic sclerosis and its potential for disease differentiation using deep learning

From Springer Nature via Jisc Publications RouterHistory: received 2020-05-07, accepted 2020-09-09, registration 2020-09-16, pub-electronic 2020-10-05, online 2020-10-05, collection 2020-12Publication status: PublishedFunder: Helmholtz Association; doi: http://dx.doi.org/10.13039/501100009318; Grant(s): i3 (ExNet-0022-Phase2-3)Funder: University of Manchester; doi: http://dx.doi.org/10.13039/501100000770; Grant(s): MC_PC_16053Funder: Manchester Biomedical Research Centre; doi: http://dx.doi.org/10.13039/100014653Funder: H2020 European Research Council; doi: http://dx.doi.org/10.13039/100010663; Grant(s): No 694968 (PREMSOT)Funder: Arthritis Research UK,United Kingdom; Grant(s): 19465Abstract: The autoimmune disease systemic sclerosis (SSc) causes microvascular changes that can be easily observed cutaneously at the finger nailfold. Optoacoustic imaging (OAI), a combination of optical and ultrasound imaging, specifically raster-scanning optoacoustic mesoscopy (RSOM), offers a non-invasive high-resolution 3D visualization of capillaries allowing for a better view of microvascular changes and an extraction of volumetric measures. In this study, nailfold capillaries of patients with SSc and healthy controls are imaged and compared with each other for the first time using OAI. The nailfolds of 23 patients with SSc and 19 controls were imaged using RSOM. The acquired images were qualitatively compared to images from state-of-the-art imaging tools for SSc, dermoscopy and high magnification capillaroscopy. The vascular volume in the nailfold capillaries were computed from the RSOM images. The vascular volumes differ significantly between both cohorts (0.216 ± 0.085 mm3 and 0.337 ± 0.110 mm3; p < 0.0005). In addition, an artificial neural network was trained to automatically differentiate nailfold images from both cohorts to further assess whether OAI is sensitive enough to visualize anatomical differences in the capillaries between the two cohorts. Using transfer learning, the model classifies images with an area under the ROC curve of 0.897, and a sensitivity of 0.783 and specificity of 0.895. In conclusion, this study demonstrates the capabilities of RSOM as an imaging tool for SSc and establishes it as a modality that facilitates more in-depth studies into the disease mechanisms and progression

Lack of EGF receptor contributes to drug sensitivity of human germline cells

Germline mutations have been associated with generation of various types of tumour. In this study, we investigated genetic alteration of germline tumours that affect the drug sensitivity of cells. Although all germline tumour cells we tested were hypersensitive to DNA-damaging drugs, no significant alteration was observed in their DNA repair activity or the expression of DNA repair proteins. In contrast, germline tumours expressed very low level of epidermal growth factor receptor (EGFR) compared to drug-resistant ovarian cancer cells. An immunohistochemical analysis indicated that most of the primary germline tumours we tested expressed very low level of EGFR. In accordance with this, overexpression of EGFR in germline tumour cells showed an increase in drug resistance, suggesting that a lack of EGFR, at least in part, contributes to the drug sensitivity of germline tumours

SUMO-Interacting Motifs of Human TRIM5α are Important for Antiviral Activity

Human TRIM5α potently restricts particular strains of murine leukemia viruses (the so-called N-tropic strains) but not others (the B- or NB-tropic strains) during early stages of infection. We show that overexpression of SUMO-1 in human 293T cells, but not in mouse MDTF cells, profoundly blocks N-MLV infection. This block is dependent on the tropism of the incoming virus, as neither B-, NB-, nor the mutant R110E of N-MLV CA (a B-tropic switch) are affected by SUMO-1 overexpression. The block occurred prior to reverse transcription and could be abrogated by large amounts of restricted virus. Knockdown of TRIM5α in 293T SUMO-1-overexpressing cells resulted in ablation of the SUMO-1 antiviral effects, and this loss of restriction could be restored by expression of a human TRIM5α shRNA-resistant plasmid. Amino acid sequence analysis of human TRIM5α revealed a consensus SUMO conjugation site at the N-terminus and three putative SUMO interacting motifs (SIMs) in the B30.2 domain. Mutations of the TRIM5α consensus SUMO conjugation site did not affect the antiviral activity of TRIM5α in any of the cell types tested. Mutation of the SIM consensus sequences, however, abolished TRIM5α antiviral activity against N-MLV. Mutation of lysines at a potential site of SUMOylation in the CA region of the Gag gene reduced the SUMO-1 block and the TRIM5α restriction of N-MLV. Our data suggest a novel aspect of TRIM5α-mediated restriction, in which the presence of intact SIMs in TRIM5α, and also the SUMO conjugation of CA, are required for restriction. We propose that at least a portion of the antiviral activity of TRIM5α is mediated through the binding of its SIMs to SUMO-conjugated CA

Sonophore labeled RGD: A targeted contrast agent for optoacoustic imaging.

Optoacoustic imaging is a rapidly expanding field for the diagnosis, characterization, and treatment evaluation of cancer. However, the availability of tumor specific exogenous contrast agents is still limited. Here, we report on a small targeted contrast agent for optoacoustic imaging using a black hole quencher&reg; (BHQ) dye. The sonophore BHQ-1 exhibited strong, concentration-dependent, optoacoustic signals in phantoms, demonstrating its ideal suitability for optoacoustic imaging. After labeling BHQ-1 with cyclic RGD-peptide, BHQ-1-cRGD specifically bound to &alpha;v&beta;3-integrin expressing glioblastoma cell spheroids in vitro. The excellent optoacoustic properties of BHQ-1-cRGD could furthermore be proven in vivo. Together with this emerging imaging modality, our sonophore labeled small peptide probe offers new possibilities for non-invasive detection of molecular structures with high resolution in vivo and furthers the specificity of optoacoustic imaging. Ultimately, the discovery of tailor-made sonophores might offer new avenues for various molecular optoacoustic imaging applications, similar to what we see with fluorescence imaging