Probes for INS

Epstein-Barr virus (EBV), a common pathogen in humans, is suspected as the cause of multiple pregnancy-related pathologies including depression, preeclampsia, and stillbirth. Moreover, transmission of EBV through the placenta has been reported. However, the focus of EBV infection within the placenta has remained unknown to date. In this study, we proved the expression of latent EBV genes in the endometrial glandular epithelial cells of the placenta and investigated the cytological characteristics of these cells. Sixty-eight placentas were obtained from pregnant women. Tissue microarray was constructed. EBV latent genes including EBV-encoding RNA-1 (EBER1), Epstein-Barr virusnuclear antigen 1 (EBNA1), late membrane antigen (LMP1), and RPMS1 were detected with silver in situ hybridization and/or mRNA in situ hybridization. Nuclear features of EBV-positive cells in EBV-infected placenta were compared with those of EBV-negative cells via image analysis. Sixteen placentas (23.5%) showed positive expression of all 4 EBV latent genes; only the glandular epithelial cells of the decidua showed EBV gene expression. EBV infection status was not significantly correlated with maternal, fetal, or placental factors. The nuclei of EBV-positive cells were significantly larger, longer, and round-shaped than those of EBV-negative cells regardless of EBV-infection status of the placenta. For the first time, evidence of EBV gene expression has been shown in placental tissues. Furthermore, we have characterized its cytological features, allowing screening of EBV infection through microscopic examination.

The lack of representative nasopharyngeal carcinoma (NPC) models has seriously hampered research on EBV carcinogenesis and preclinical studies in NPC. Here we report the successful growth of five NPC patient-derived xenografts (PDXs) from fifty-eight attempts of transplantation of NPC specimens into NOD/SCID mice. The take rates for primary and recurrent NPC are 4.9% and 17.6%, respectively. Successful establishment of a new EBV-positive NPC cell line, NPC43, is achieved directly from patient NPC tissues by including Rho-associated coiled-coil containing kinases inhibitor (Y-27632) in culture medium. Spontaneous lytic reactivation of EBV can be observed in NPC43 upon withdrawal of Y-27632. Whole-exome sequencing (WES) reveals a close similarity in mutational profiles of these NPC PDXs with their corresponding patient NPC. Whole-genome sequencing (WGS) further delineates the genomic landscape and sequences of EBV genomes in these newly established NPC models, which supports their potential use in future studies of NPC.

Regardless of host, pathogenic mycobacteria of the Mycobacterium tuberculosiscomplex such as Mycobacterium bovis, induce a characteristic lesion known as agranuloma, tubercle or tuberculoid granuloma. Granulomas represent a distinct host response to chronic antigenic stimuli, such as foreign bodies, certain bacterial components, or persistent pathogens such as M. bovis. Granulomas are composed of specific cell types including epithelioid macrophages, lymphocytes and a morphologically distinctive cell type, the multinucleated giant cell. Multinucleated giant cells are formed by the fusion of multiple macrophages; however, their function remains unclear. In humans, giant cells in tuberculous granulomas have been shown to express various cytokines, chemokines and enzymes important to the formation and maintenance of the granuloma. The objective of this study was to quantitatively assess multinucleated giant cell cytokine expression in bovine tuberculoid granulomas; focusing on cytokines of suspected relevance to bovine tuberculosis. Using calves experimentally infected with M. bovis, in situ cytokine expression was quantitatively assessed using RNAScope^® for the following cytokines TNF-α, IFN-γ, TGF-β, IL-17A and IL-10. Multinucleated giant cells in bovine tuberculoid granulomas expressed all examined cytokines to varying degrees, with differential expression of TGF-β, IL-17A and IL-10 in giant cells from early versus late stage granulomas. There was a modest, positive correlation between the level of cytokine expression and cell size or number of nuclei. These results suggest that multinucleated giant cells are active participants within bovine tuberculoid granulomas, contributing to the cytokine milieu necessary to form and maintain granulomas.

The hallmark lesion of tuberculosis in humans and animals is the granuloma. The granuloma represents a distinct host cellular immune response composed of epithelioid macrophages, lymphocytes, and multinucleated giant cells, often surrounding a caseous necrotic core. Within the granuloma, host-pathogen interactions determine disease outcome. Factors within the granulomas such as cytokines and chemokines drive cell recruitment, activity, function and ultimately the success or failure of the host's ability to control infection. Hence, an understanding of the granuloma-level cytokine response is necessary to understand tuberculosis pathogenesis. In-situ cytokine expression patterns were measured using a novel in situ hybridization assay, known as RNAScope^® in granulomas of the lungs, tracheobronchial lymph nodes and caudal mediastinal lymph nodes of cattle experimentally infected with Mycobacterium bovis via aerosol exposure. In spite of microscopic morphological similarities, significant differences were seen between late stage granulomas of the lung compared to those of the tracheobronchial lymph nodes for IL-17A, IFN-γ, TGF-β, IL10 and IL-22 but not for TNF-α. Additionally, significant differences were noted between granulomas from two different thoracic lymph nodes that both receive afferent lymphatics from the lungs (i.e., tracheobronchial and caudal mediastinal lymph nodes) for TNF-α, IL-17A, IFN-γ, TGF-β and IL-10 but not for IL-22. These findings show that granuloma morphology alone is not a reliable indicator of granuloma function as granulomas of similar morphologies can have disparate cytokine expression patterns. Moreover, anatomically distinct lymph nodes (tracheobronchial vs caudal mediastinal) differ in cytokine expression patterns even when both receive afferent lymphatics from a lung containing tuberculoid granulomas. These findings show that selection of tissue and anatomic location are critical factors in assessing host immune response to M. bovis and should be considered carefully.