Anabolic Steroids Activate the NF-κB Pathway in Porcine Ovarian Putative Stem Cells Independently of the ZIP-9 Receptor.

Boldenone (Bdn) and nandrolone (Ndn) are anabolic androgenic steroids (AASs) that, as our previous studies have shown, may increase the risk of neoplastic transformation of porcine ovarian putative stem cells (poPSCs). The NF-κB pathway may be important in the processes of carcinogenesis and tumour progression. Therefore, in this work, we decided to test the hypothesis of whether Bdn and Ndn can activate the NF-κB pathway by acting through the membrane androgen receptor ZIP-9. For this purpose, the expression profiles of both genes involved in the NF-κB pathway and the gene coding for the ZIP-9 receptor were checked. The expression and localization of proteins of this pathway in poPSCs were also examined. Additionally, the expression of the ZIP-9 receptor and the concentration of the NF-κB1 and 2 protein complex were determined. Activation of the NF-κB pathway was primarily confirmed by an increase in the relative abundances of phosphorylated forms of RelA protein and IκBα inhibitor. Reduced quantitative profiles pinpointed not only for genes representing this pathway but also for unphosphorylated proteins, and, simultaneously, decreased concentration of the NF-κB1 and 2 complex may indicate post-activation silencing by negative feedback. However, the remarkably and sustainably diminished expression levels noticed for the SLC39A9 gene and ZIP-9 protein suggest that this receptor does not play an important role in the regulation of the NF-κB pathway.

Assessment of Stress, Depressive and Anxiety Symptoms in Patients with COPD during In-Hospital Pulmonary Rehabilitation: An Observational Cohort Study.

Background and Objectives: The relationship between physical health and mental health has been considered for years. A number of studies have shown a correlation between depressive states and the progress of somatic diseases. It seems that the proper cooperation of specialists may result in the improvement of the patient's well-being and a positive effect on the course of the rehabilitation process. The aim of this study was to assess the symptoms of depression, anxiety, and stress in patients with chronic obstructive pulmonary disease (COPD) as well as the assessment of the relationship of psychological symptoms with sociodemographic factors and physical condition. Materials and Methods: The study enrolled 51 COPD patients who underwent a three-week pulmonary rehabilitation program. After admission to the rehabilitation department, the subjects were asked to complete the Hospital Anxiety and Depression Scale (HADS) questionnaire, the Perception of Stress Questionnaire (PSQ), and a sociodemographic questionnaire. Results: Anxiety states were diagnosed in 70% of respondents and depressive states were diagnosed in 54% of patients. Some of the respondents (14%) also showed a tendency to experience various grounded stresses. Additionally, there were correlations between the mental state and the results of fitness and respiratory tests. Conclusions: Patients with COPD are at risk for mental disorders, which may adversely affect their general health and significantly limit their physical and respiratory efficiencies. The development of widely available therapeutic solutions to reduce symptoms associated with depression, anxiety, and stress seems to be an important challenge for the management of patients with COPD.

The human core exosome interacts with differentially localized processive RNases: hDIS3 and hDIS3L.

The eukaryotic RNA exosome is a ribonucleolytic complex involved in RNA processing and turnover. It consists of a nine-subunit catalytically inert core that serves a structural function and participates in substrate recognition. Best defined in Saccharomyces cerevisiae, enzymatic activity comes from the associated subunits Dis3p (Rrp44p) and Rrp6p. The former is a nuclear and cytoplasmic RNase II/R-like enzyme, which possesses both processive exo- and endonuclease activities, whereas the latter is a distributive RNase D-like nuclear exonuclease. Although the exosome core is highly conserved, identity and arrangements of its catalytic subunits in different vertebrates remain elusive. Here, we demonstrate the association of two different Dis3p homologs--hDIS3 and hDIS3L--with the human exosome core. Interestingly, these factors display markedly different intracellular localizations: hDIS3 is mainly nuclear, whereas hDIS3L is strictly cytoplasmic. This compartmental distribution reflects the substrate preferences of the complex in vivo. Both hDIS3 and hDIS3L are active exonucleases; however, only hDIS3 has retained endonucleolytic activity. Our data suggest that three different ribonucleases can serve as catalytic subunits for the exosome in human cells.

Loss of DRO1/CCDC80 in the tumor microenvironment promotes carcinogenesis.

Tumors are composed of the tumor cells and the surrounding microenvironment. Both are closely interwoven and interact by a complex and multifaceted cross-talk which plays an integral part in tumor initiation, growth, and progression. Dro1/Ccdc80 has been shown to be a potent suppressor of colorectal cancer and ubiquitous inactivation of Dro1/Ccdc80 strongly promoted colorectal carcinogenesis in ApcMin/+ mice and in a chemically-induced colorectal cancer model. The aim of the present study was to investigate whether Dro1/Ccdc80's tumor suppressive function is tumor-cell-autonomous. Expression of Dro1/Ccdc80 in cancer cells had no effect on both colon tumor development in ApcMin/+ mice and formation of xenograft tumors. In contrast, DRO1/CCDC80 loss in the microenvironment strongly increased tumor growth in xenograft models, inhibited cancer cell apoptosis, and promoted intestinal epithelial cell migration. Moreover, stromal Dro1/Ccdc80 inactivation facilitated formation of intestinal epithelial organoids. Expression analyses showed Dro1/Ccdc80 to be significantly down-regulated in murine gastric cancer associated fibroblasts, in ApcMin/+ colon tumor primary stromal cells and in microdissected stroma from human colorectal cancer compared to normal, non-tumor stroma. Our results demonstrate epithelial derived DRO1/CCDC80 to be dispensable for intestinal tissue homeostasis and identify Dro1/Ccdc80 as tumor suppressor in the tumor microenvironment.

Myeloid-derived suppressor cells: a double-edged sword?

Myeloid-derived suppressor cells are important cell population with an immunoregulatory potential in both adaptive and innate immunity. Their immunosuppressive activity is widely accepted. However, emerging evidence suggests that this heterogeneous cell population can be, under some circumstances, immunostimulatory rather than suppressive. This finding can shed a new light on antitumour immunity which is believed to be impaired in immunosuppressive environments.

Induction of Epithelial-mesenchymal Transition in MDCK II Cells.

Epithelial-mesenchymal transition (EMT) is a reversible process of epithelial cell transdifferentiation into a mesenchymal cell, that enables initiation of cell migration. EMT plays an important role in embryonic development, tissue repair and cancer metastasis. Better understanding of cellular and molecular events during EMT will not only provide novel insights on how mammalian organism develops and how epithelial tissues regenerate, but also can identify novel therapeutic targets for cancer therapy. Here we aim to provide a detailed protocol on how to induce EMT in Madin-Darby Canine Kidney (MDCK) II epithelial cell line and perform immunofluorescent staining on EMT-induced cells.

Training Using a Commercial Immersive Virtual Reality System on Hand-Eye Coordination and Reaction Time in Young Musicians: A Pilot Study.

The implementation of virtual reality (VR) opens up a wide range of possibilities for the development of dexterity, speed and precision of movements. The aim of this study was to investigate whether immersive VR training affected the hand-eye coordination and reaction time in students of the state music school. This study implemented a single-group pre-post study design. This study enrolled 14 individuals, submitted to a 15 min training session of the immersive music game "Beat Saber", once a day for 5 consecutive days. The plate-tapping test (PTT) and the ruler-drop test (Ditrich's test) were used to assess the reaction time. Trial-making test (TMT) A and TMT B were used to assess coordination and visual attention. Analysis of the results showed a statistically significant improvement in hand-eye coordination and reaction time of music school students using the TMT-A (p < 0.002), TMT-B (p < 0.001), Ditrich's test for the non-dominant hand (0.025) and PTT (0.0001) after applying a week-long training period in immersive VR. The results obtained in the present study show that the VR system, along with the immersive music game, has the potential to improve hand-eye coordination and reaction time in young musicians, which may lead to the faster mastering of a musical instrument.

Cellular Interactions in the Intestinal Stem Cell Niche.

Epithelial cells are one of the most actively cycling cells in a mammalian organism and therefore are prone to malignant transformation. Already during organogenesis, the connective tissue (mesenchyme) provides instructive signals for the epithelium. In an adult organism, the mesenchyme is believed to provide crucial regulatory signals for the maintenance and regeneration of epithelial cells. Here, we discuss the role of intestinal myofibroblasts, α-smooth muscle actin-positive stromal (mesenchymal) cells, as an important regulatory part of the intestinal stem cell niche. Better understanding of the cross-talk between myofibroblasts and the epithelium in the intestine has implications for advances in regenerative medicine, and improved therapeutic strategies for inflammatory bowel disease, intestinal fibrosis and colorectal cancer.

Goblet Cell Ratio in Combination with Differentiation and Stem Cell Markers in Barrett Esophagus Allow Distinction of Patients with and without Esophageal Adenocarcinoma.

The increasing incidence of esophageal adenocarcinoma (EAC) is mirrored by the increasing prevalence of Barrett esophagus, a precursor lesion resulting in a large number of individuals "at risk" for this lethal malignancy. Among patients with Barrett esophagus, only about 0.3% annually will develop EAC. Because large numbers of patients are followed in endoscopic surveillance, there is a need for risk prediction among a growing population of patients with Barrett esophagus. We identified four potential biomarkers from an inflammation (IL1ß)-dependent mouse model of Barrett esophagus and tested them in 189 patients with Barrett esophagus with and without high-grade dysplasia (HGD)/early cancer (T1). The primary goal was to distinguish patients with Barrett esophagus with no evidence of dysplasia from those with dysplasia. Increasing stem cell marker LGR5 and niche cell marker DCLK1 and decreasing differentiation marker (secretory mucus cells, TFF2+ cells) correlated with elevated tumor score in the mouse. Having outlined the origin of those markers in the Barrett esophagus mouse model, we showed the applicability for human Barrett esophagus. We compared 94 patients with nondysplastic Barrett esophagus tissue with 95 patients with Barrett esophagus and HGD or early cancer. Low levels of TFF2 (AUC 87.2%) provided the best discrimination between nondysplastic Barrett esophagus and Barrett esophagus with cancer, followed by high levels of DCLK1 (AUC 83.4%), low goblet cell ratio (AUC 79.4%), and high LGR5 (AUC 71.4%). The goblet cell ratio, rather than the presence of goblet cells per se, was found to be an important discriminator. These findings may be useful in developing future risk prediction models for patients with Barrett esophagus and ultimately to improve EAC surveillance. Cancer Prev Res; 10(1); 55-66. ©2016 AACR.

Three-Dimensional Gastrointestinal Organoid Culture in Combination with Nerves or Fibroblasts: A Method to Characterize the Gastrointestinal Stem Cell Niche.

The gastrointestinal epithelium is characterized by a high turnover of cells and intestinal stem cells predominantly reside at the bottom of crypts and their progeny serve to maintain normal intestinal homeostasis. Accumulating evidence demonstrates the pivotal role of a niche surrounding intestinal stem cells in crypts, which consists of cellular and soluble components and creates an environment constantly influencing the fate of stem cells. Here we describe different 3D culture systems to culture gastrointestinal epithelium that should enable us to study the stem cell niche in vitro in the future: organoid culture and multilayered systems such as organotypic cell culture and culture of intestinal tissue fragments ex vivo. These methods mimic the in vivo situation in vitro by creating 3D culture conditions that reflect the physiological situation of intestinal crypts. Modifications of the composition of the culture media as well as coculturing epithelial organoids with previously described cellular components such as myofibroblasts, collagen, and neurons show the impact of the methods applied to investigate niche interactions in vitro. We further present a novel method to isolate labeled nerves from the enteric nervous system using Dclk1-CreGFP mice.

Both activating and inhibitory Fc gamma receptors mediate rituximab-induced trogocytosis of CD20 in mice.

Antigenic modulation by trogocytosis during anti-CD20 mAb treatment with rituximab (RTX) leads to loss of CD20 and therefore can compromise therapy. During trogocytosis, effector cells, such as macrophages, remove CD20 from the surface of antibody-coated cells in an Fc receptor-dependent manner. Importantly, Fcγ receptors (FcγRs) are also crucial in the anti-tumor effects of RTX by inducing antibody dependent cell-mediated cytotoxicity (ADCC). Here we studied the role of FcγR during RTX-induced trogocytosis of CD20 in an intraperitoneal tumor model with EL4-CD20 cells. We found marked RTX-induced trogocytosis of CD20 in FcγRI- or FcγRIII-deficient mice, similar to wild type mice, demonstrating a redundancy for activating FcγR in trogocytosis. Interestingly, in FcRγ-chain-deficient mice, trogocytosis was still apparent, indicating that the inhibitory receptor FcγRIIB alone can also mediate trogocytosis. These data were confirmed by in vitro analysis with blocking antibodies. Decreasing the amount of RTX in vivo resulted in less trogocytosis of CD20, supporting clinical studies with lower RTX dose. Importantly, we show that cells which undergo in vivo trogocytosis can still be killed ex vivo by ADCC but not by complement-mediated cytotoxicity (CDC), underscoring the clinical relevance of trogocytosis. Taken together, our study provides more insights into the mechanism and consequences of RTX-induced trogocytosis of CD20.