Raman Spectral Signatures of Cervical Exfoliated Cells from Liquid-Based Cytology Samples

It is widely accepted that cervical screening has significantly reduced the incidence of cervical cancer worldwide. The primary screening test for cervical cancer is the Papanicolaou (Pap) test, which has extremely variable specificity and sensitivity. There is an unmet clinical need for methods to aid clinicians in the early detection of cervical precancer. Raman spectroscopy is a label-free objective method that can provide a biochemical fingerprint of a given sample. Compared with studies on infrared spectroscopy, relatively few Raman spectroscopy studies have been carried out to date on cervical cytology. The aim of this study was to define the Raman spectral signatures of cervical exfoliated cells present in liquid-based cytology Pap test specimens and to compare the signature of high-grade dysplastic cells to each of the normal cell types. Raman spectra were recorded from single exfoliated cells and subjected to multivariate statistical analysis. The study demonstrated that Raman spectroscopy can identify biochemical signatures associated with the most common cell types seen in liquid-based cytology samples; superficial, intermediate, and parabasal cells. In addition, biochemical changes associated with high-grade dysplasia could be identified suggesting that Raman spectroscopy could be used to aid current cervical screening tests

Raman Spectroscopy of Liquid-Based Cervical Smear Samples as a Triage to Stratify Women Who Are HPV-Positive on Screening

Persistent high-risk human papillomavirus (HPV) infection can lead to cervical precancer and cancer. Recently, HPV testing has been introduced for primary cervical screening, but the HPV DNA test cannot distinguish between transient and persistent HPV infection. Thus, there is an unmet clinical need to develop a new test to identify women with a high-risk persistent HPV infection. Raman spectra were recorded from cervical smear samples (n = 60) and, on the basis of HPV DNA and HPV mRNA test results, a classifier was developed to identify persistent HPV infection. A further blinded independent test set (n = 14) was used to validate the model, and sensitivity of 90% and specificity of 100% were achieved. Improved triage would allow women with a high-risk persistent HPV infection to be referred for immediate treatment, while women with a low-risk transient infection could avoid overtreatment

Phenotyping dividing cells in mouse models of neurodegenerative basal ganglia diseases

BACKGROUND: Mice generated by a Cre/LoxP transgenic paradigm were used to model neurodegenerative basal ganglia disease of which Huntington disease (HD) is the prototypical example. In HD, death occurs in striatal projection neurons as well as cortical neurons. Cortical and striatal neurons that express the D1 dopamine receptor (Drd1a) degenerate in HD. The contribution that death of specific neuronal cell populations makes to the HD disease phenotype and the response of the brain to loss of defined cell subtypes is largely unknown. METHODS: Drd1a-expressing cells were targeted for cell death and three independent lines generated; a striatal-restricted line, a cortical-restricted line and a global line in which Drd1a cells were deleted from both the striatum and cortex. Two independent experimental approaches were used. In the first, the proliferative marker Ki-67 was used to identify proliferating cells in eighty-week-old mice belonging to a generic global line, a global in which Drd1a cells express green fluorescent protein (GFP-global) and in eighty-week-old mice of a cortical line. In the second experiment, the proliferative response of four-week-old mice belonging to GFP-global and striatal lines was assessed using the thymidine analogue BrdU. The phenotype of proliferating cells was ascertained by double staining for BrdU and Olig2 (an oligodendrocyte marker), Iba1 (a microglial cell marker), S100β (an astroglial cell marker), or NeuN (a neuronal cell marker). RESULTS: In the first study, we found that Ki-67-expressing cells were restricted to the striatal side of the lateral ventricles. Control mice had a greater number of Ki-67+ cells than mutant mice. There was no overlap between Ki-67 and GFP staining in control or mutant mice, suggesting that cells did not undergo cell division once they acquired a Drd1a phenotype. In contrast, in the second study we found that BrdU+ cells were identified throughout the cortex, striatum and periventricular region of control and mutant mice. Mutant mice from the GFP-global line showed increased BrdU+ cells in the cortex, striatum and periventricular region relative to control. Striatal line mutant mice had an increased number of BrdU+ cells in the striatum and periventricular region, but not the cortex. The number of microglia, astrocytes, oligodendrocytes and neurons generated from dividing progenitors was increased relative to control mice in most brain regions in mutant mice from the GFP-global line. In contrast, striatal line mutant mice displayed an increase only in the number of dividing microglia in striatal and periventricular regions. CONCLUSIONS: Genetically programmed post-natal ablation of Drd1a-expressing neurons is associated with an extensive proliferative response involving multiple cell lineages. The nature of the tissue response has the potential not only to remove cellular debris but also to forge physiologically meaningful brain repair. Age related deficits in proliferation are seen in mutant lines. A blunted endogenous reparative response may underlie the cumulative deficits characteristic of age related neurodegeneration

A Transdisciplinary Approach to Determining the Provenience of a Distorted, Pre-Columbian Skull Recovered in Rural Idaho

Transdisciplinary research involves cooperation, exchange of information, sharing of resources and integration of disciplines to achieve a common scientific goal. In this study, collaborators utilized tools and knowledge of materials science, anthropology, archaeology, geosciences and biology in an attempt to determine the provenience of skeletal remains of unknown origin. The exchange of ideas and skills along with the crossing of disciplines in this study sucessfully allowed the incorporation of expertise from many team members. This transdisciplinary approach to research provided a more comprehensive and detailed analysis than any one field alone could provide. An archaeological assessment of a human skull recovered in rural Idaho recognized cranial deformation and post-mortem application of a red pigment. A combination of scanning electron microscopy (SEM), x-ray fluorescence (XRF) and energy-dispersive x-ray spectroscopy (EDS) identified the major and trace elements present in the red post-mortem pigment as cinnabar and rare earth metals. Analysis via carbon and oxygen stable isotopes from teeth and bone to provided insight into the diet and habitat for distinct segments of the individual’s life, indicating a regional separation in early life versus late adulthood. Radiocarbon dating determined the approximate age of the skull to be between 600-700 years old and a forensic mtDNA assessmentcategorized a mitochondrial haplogroup for the remains as originating from the East African or Arabian Peninsula

"It's a can of worms": understanding primary care practitioners' behaviours in relation to HPV using the Theoretical Domains Framework

Background: The relationship between infection with high-risk human papillomavirus (HPV) and cervical cancer is transforming cervical cancer prevention. HPV tests and vaccinations have recently become available. In Ireland, as elsewhere, primary care practitioners play a key role in prevention. ATHENS (A Trial of HPV Education and Support) aims to develop a theorybased intervention to support primary care practitioners in their HPV-related practice. This study, the first step in the intervention development process, aimed to: identify HPV-related clinical behaviours that the intervention will target; clarify general practitioners’ (GPs’) and practice nurses’ roles and responsibilities; and determine factors that potentially influence clinical behaviour. A secondary objective was to informally assess the utility of the Theoretical Domains Framework (TDF) in understanding clinical behaviours in an area with an evolving evidence-base. Methods: In-depth semi-structured telephone interviews were conducted with GPs and practice nurses. The topic guide, which contained open questions and HPV-related clinical scenarios, was developed through literature review and clinical experience. Interview transcripts were content-analysed using the TDF as the coding framework. Results: 19 GPs and 14 practice nurses were interviewed. The major HPV-related clinical behaviours were: initiating a discussion about HPV infection with female patients; offering/recommending HPV vaccination to appropriate patients; and answering patients’ questions about HPV testing. While the responsibility for taking smears was considered a female role, both male and female practitioners dealt with HPV-related issues. All 12 theoretical domains arose in relation to HPV infection; the domains judged to be most important were: knowledge, emotion, social influences, beliefs about capabilities and beliefs about consequences. Eleven domains emerged in relation to HPV vaccination, with beliefs about consequences, social influences, knowledge and environmental context and resources judged to be the most important. Nine domains were relevant to HPV testing, with knowledge and beliefs about capabilities judged to be the most important. Conclusions: The findings confirm the need for an intervention to support primary care practitioners around HPV and suggest it should target a range of theoretical domains. The TDF proved valuable in analysing qualitative data collected using a topic guide not specifically designed to capture TDF domains and understanding clinical behaviours in an area with an evolving evidence-base

An integrated analysis of the SOX2 microRNA response program in human pluripotent and nullipotent stem cell lines

Abstract Background SOX2 is a core component of the transcriptional network responsible for maintaining embryonal carcinoma cells (ECCs) in a pluripotent, undifferentiated state of self-renewal. As such, SOX2 is an oncogenic transcription factor and crucial cancer stem cell (CSC) biomarker in embryonal carcinoma and, as more recently found, in the stem-like cancer cell component of many other malignancies. SOX2 is furthermore a crucial factor in the maintenance of adult stem cell phenotypes and has additional roles in cell fate determination. The SOX2-linked microRNA (miRNA) transcriptome and regulome has not yet been fully defined in human pluripotent cells or CSCs. To improve our understanding of the SOX2-linked miRNA regulatory network as a contribution to the phenotype of these cell types, we used high-throughput differential miRNA and gene expression analysis combined with existing genome-wide SOX2 chromatin immunoprecipitation (ChIP) data to map the SOX2 miRNA transcriptome in two human embryonal carcinoma cell (hECC) lines. Results Whole-microRNAome and genome analysis of SOX2-silenced hECCs revealed many miRNAs regulated by SOX2, including several with highly characterised functions in both cancer and embryonic stem cell (ESC) biology. We subsequently performed genome-wide differential expression analysis and applied a Monte Carlo simulation algorithm and target prediction to identify a SOX2-linked miRNA regulome, which was strongly enriched with epithelial-to-mesenchymal transition (EMT) markers. Additionally, several deregulated miRNAs important to EMT processes had SOX2 binding sites in their promoter regions. Conclusion In ESC-like CSCs, SOX2 regulates a large miRNA network that regulates and interlinks the expression of crucial genes involved in EMT

TLR7 promotes chronic airway disease in RSV-infected mice

Respiratory syncytial virus (RSV) commonly infects the upper respiratory tract (URT) of humans, manifesting with mild cold or flu-like symptoms. However, in infants and the elderly, severe disease of the lower respiratory tract (LRT) often occurs and can develop into chronic airway disease. A better understanding of how an acute RSV infection transitions to a LRT chronic inflammatory disease is critically important to improve patient care and long-term health outcomes. To model acute and chronic phases of the disease, we infected wild-type C57BL/6 and toll-like receptor 7 knockout (TLR7 KO) mice with RSV and temporally assessed nasal, airway and lung inflammation for up to 42 days post-infection. We show that TLR7 reduced viral titers in the URT during acute infection but promoted pronounced pathogenic and chronic airway inflammation and hyperreactivity in the LRT. This study defines a hitherto unappreciated molecular mechanism of lower respiratory pathogenesis to RSV, highlighting the potential of TLR7 modulation to constrain RSV pathology to the URT

Epithelioid trophoblastic tumour : a case with genetic linkage to a child born over seventeen years prior, successfully treated with surgery and pembrolizumab

Epithelioid trophoblastic tumours are rare neoplasms showing differentiation towards the chorion leave-type intermediate cytotrophoblast, with only a handful of cases being reported in the literature. These tumours are slow-growing and are typically confined to the uterus for extended periods of time. While the pathogenesis is unclear, they are thought to arise from a remnant intermediate trophoblast originating from prior normal pregnancies or, less frequently, gestational trophoblastic tumours. A protracted time period between the gestational event and tumour development is typical. This case describes a 49-year-old previously healthy female who presented with a completely asymptomatic uterine mass, discovered incidentally during a routine gynaecological assessment. The pathological analysis of the hysterectomy specimen confirmed an epithelioid trophoblastic tumour, involving the uterus and cervix. This is a rare gynaecological tumour. A comparative short tandem repeat analysis revealed genetic similarities to a previous healthy gestation seventeen years prior. She was successful treated with adjuvant pembrolizumab, with no evidence of disease recurrence to date.peer-reviewe