Nonmalignant AR-positive prostate epithelial cells and cancer cells respond differently to androgen.

Prostate cancer research suffers from the lack of suitable models to study the role of normal cells in prostate carcinogenesis. To address this challenge, we developed a cell line model mimicking luminal prostate epithelial cells by modifying the immortalized prostate epithelial cell line RWPE-1 to constitutively express the androgen receptor (AR). RWPE-1-AR cells express known AR target genes, and exhibit coexpression of luminal and basal markers characteristic of transient amplifying cells, and an RNA signature resembling prostate luminal progenitor cells. Under unstimulated conditions, constitutive AR expression does not have a biologically significant effect on the proliferation of RWPE-1 cells, but when stimulated by androgens, growth is retarded. The transcriptional response of RWPE-1-AR cells to androgen stimulation involves suppression of the growth-related KRAS pathway and is thus markedly different from that of the prostate cancer cell line LNCaP and its derivative AR-overexpressing LNCaP-ARhi cells, in which growth- and cancer-related pathways are upregulated. Hence, the nonmalignant AR-positive RWPE-1-AR cell line model could be used to study the transformation of the prostate epithelium.

Herpesviruses and their genetic diversity in the blood virome of healthy individuals: effect of aging.

BACKGROUND: As we age, the functioning of the human immune system declines. The results of this are increases in morbidity and mortality associated with infectious diseases, cancer, cardiovascular disease, and neurodegenerative disease in elderly individuals, as well as a weakened vaccination response. The aging of the immune system is thought to affect and be affected by the human virome, the collection of all viruses present in an individual. Persistent viral infections, such as those caused by certain herpesviruses, can be present in an individual for long periods of time without any overt pathology, yet are associated with disease in states of compromised immune function. To better understand the effects on human health of such persistent viral infections, we must first understand how the human virome changes with age. We have now analyzed the composition of the whole blood virome of 317 individuals, 21-70 years old, using a metatranscriptomic approach. Use of RNA sequencing data allows for the unbiased detection of RNA viruses and active DNA viruses. RESULTS: The data obtained showed that Epstein-Barr virus (EBV) was the most frequently expressed virus, with other detected viruses being herpes simplex virus 1, human cytomegalovirus, torque teno viruses, and papillomaviruses. Of the 317 studied blood samples, 68 (21%) had EBV expression, whereas the other detected viruses were only detected in at most 6 samples (2%). We therefore focused on EBV in our further analyses. Frequency of EBV detection, relative EBV RNA abundance and the genetic diversity of EBV was not significantly different between age groups (21-59 and 60-70 years old). No significant correlation was seen between EBV RNA abundance and age. Deconvolution analysis revealed a significant difference in proportions of activated dendritic cells, macrophages M1, and activated mast cells between EBV expression positive and negative individuals. CONCLUSIONS: As it is likely that the EBV RNA quantified in this work is derived from reactivation of the latent EBV virus, these data suggest that age does not affect the rate of reactivation nor the genetic landscape of EBV. These findings offer new insight on the genetic diversity of a persistent EBV infection in the long-term.

Genetic Adaptation of Coxsackievirus B1 during Persistent Infection in Pancreatic Cells.

Coxsackie B (CVB) viruses have been associated with type 1 diabetes. We have recently observed that CVB1 was linked to the initiation of the autoimmune process leading to type 1 diabetes in Finnish children. Viral persistency in the pancreas is currently considered as one possible mechanism. In the current study persistent infection was established in pancreatic ductal and beta cell lines (PANC-1 and 1.1B4) using four different CVB1 strains, including the prototype strain and three clinical isolates. We sequenced 5' untranslated region (UTR) and regions coding for structural and non-structural proteins and the second single open reading frame (ORF) protein of all persisting CVB1 strains using next generation sequencing to identify mutations that are common for all of these strains. One mutation, K257R in VP1, was found from all persisting CVB1 strains. The mutations were mainly accumulated in viral structural proteins, especially at BC, DE, EF loops and C-terminus of viral capsid protein 1 (VP1), the puff region of VP2, the knob region of VP3 and infection-enhancing epitope of VP4. This showed that the capsid region of the viruses sustains various changes during persistency some of which could be hallmark(s) of persistency.

Bioinformatics Assembling and Assessment of Novel Coxsackievirus B1 Genome.

The human microbiome project via application of metagenomic next-generation sequencing techniques has found surprising large and diverse amounts of microbial sequences across different body sites. There is a wave of investigators studying autoimmune related diseases designing from birth case and control studies to elucidate microbial associations and potential direct triggers. Sequencing analysis, considered big data as it typically includes millions of reads, is challenging but particularly demanding and complex is virome profiling due to its lack of pan-viral genomic signature. Impressively thousands of virus complete genomes have been deposited and these high-quality references are core components of virus profiling pipelines and databases. Still it is commonly known that most viral sequences do not map to known viruses. Moreover human viruses, particularly RNA groups, are notoriously heterogeneous due to high mutation rates. Here, we present the related assembling challenges and a series of bioinformatics steps that were applied in the construction of the complete consensus genome of a novel clinical isolate of Coxsackievirus B1. We further demonstrate our effort in calling mutations between prototype Coxsackievirus B1 sequence from GenBank and serial clinical isolate genome grown in cell culture.