organ culture model of liver for the study of cancer treatment for hepatocellular carcinoma

The liver, the largest organ of the human body, is a multifunctional organ with various metabolic activities that plays a fundamental role in maintaining the body and in sustaining life. Although the liver has great regenerative capacity and recovery, the damage caused by chronic diseases such as cancer or viral infections can lead to permanent loss of liver function. Studies on the mechanism of liver disease, have focused on the selection of cell and tissue culture techniques, including strategies based on in vitro models. The organ culture is a promising tool for the study of liver diseases, because it can mimic the complex of the microenvironment in vivo using a three-dimensional model of human liver tissue. These models allow a better study of the specific functions of the liver. In this context, we have analyzed the development of a hepatocarcinoma, obtained by inoculating a murine hepatocarcinoma cell line, Hepa 1/A1s, in the liver of 10 mice of the strain C57BL / 6. After 20 days from the inoculation, the portion of liver invaded by the tumor was removed from the animals and cultured. A group of 5 liver explants were used as a control and other 5 explants were cultured for 4 weeks in a complete medium containing 10% Citozym , a food supplement with reported antioxidant properties. The cancer-invaded hepatic lobes, treated with Citozym , showed a clear reduction of the weight and the volume of the hepatic tumors, when compared with the control explants

Structure and Mechanism of Dimer-Monomer Transition of a Plant Poly(A)-Binding Protein upon RNA Interaction: Insights into Its Poly(A) Tail Assembly

Poly(A)-binding proteins (PABPs) play crucial roles in mRNA biogenesis, stability, transport and translational control in most eukaryotic cells. Although animal PABPs are well-studied proteins, the biological role, three-dimensional structure and RNA-binding mode of plant PABPs remain largely uncharacterized. Here, we report the structural features and RNA-binding mode of a Citrus sinensis PABP (CsPABPN1). CsPABPN1 has a domain architecture of nuclear PABPs (PABPNs) with a single RNA recognition motif (RRM) flanked by an acidic N-terminus and a GRPF-rich C-terminus. The RRM domain of CsPABPN1 displays virtually the same three-dimensional structure and poly(A)-binding mode of animal PABPNs. However, while the CsPABPN1 RRM domain specifically binds poly(A), the full-length protein also binds poly(U). CsPABPN1 localizes to the nucleus of plant cells and undergoes a dimer–monomer transition upon poly(A) interaction. We show that poly(A) binding by CsPABPN1 begins with the recognition of the RNA-binding sites RNP1 and RNP2, followed by interactions with residues of the β2 strands, which stabilize the dimer, thus leading to dimer dissociation. Like human PABPN1, CsPABPN1 also seems to form filaments in the presence of poly(A). Based on these data, we propose a structural model in which contiguous CsPABPN1 RRM monomers wrap around the RNA molecule creating a superhelical structure that could not only shield the poly(A) tail but also serve as a scaffold for the assembly of additional mRNA processing factors

Endometrial receptivity and human embryo implantation : in vivo and in vitro studies

Background: Infertility, one of the common gynaecological disorders, affects 10-15% women in their reproductive years. Despite the advances in assisted reproduction techniques and the best efforts of infertility specialists, implantation rates do not exceed more than 28-30%. Implantation failure is one of the major causes of infertility of couples with unexplained infertility. Understanding the mechanisms of human embryo implantation is important in the attempt to alleviate infertility, improve pregnancy rates and foetal health. This knowledge could also be further explored to develop new modalities for fertility regulation. Objective: To understand endometrial receptivity and the human embryo implantation process through experimental and translational research using in vivo and in vitro approaches. In Paper I, an in vitro three-dimensional embryo-endometrial cell culture model expressing receptivity markers ER-α, ER-β, PR-(A+B), PR-B, VEGF, LIF, IL-1β and COX-2 and αVβ3 and MUC1 was developed and tested for its progesterone regulation using anti-progestin mifepristone and gestagen levonorgestrel on the human embryo implantation process. It was found that none of the 15 embryos in the cultures exposed to mifepristone attached to the endometrial construct, whereas 10/17 in control and 6/14 in levonorgestrel groups did attach. This model was further utilised in Paper II to study the role of Leukemia inhibitory factor (LIF) in human embryo implantation by using a potent LIF inhibitor, PEGLA (PEGylated LIF inhibitor). Inhibition of LIF by PEGLA, inhibited blastocyst attachment to the in vitro model, down-regulated its expression of AKT and triggered apoptosis in the inner cell mass, as studied by immunofluorescence and real-time PCR. Papers III and IV explored large-scale progesterone regulated human endometrial receptivity markers during the implantation window in stromal and epithelial compartments using a laser capture microdissection and microarray analysis. Interestingly, the expression of both the mRNA and the protein for ectonucleoside pyrophosphatase/phosphodiestrase 3 (ENPP3) in the epithelial compartment were not detectable in the progesterone-depleted group, as studied by microarray, real-time PCR and immunohistochemistry. The major canonical pathways altered in epithelial compartment were oxidative phosphorylation and the mitochondrial dysfunction pathway. In the stromal compartment, 101 genes were potentially differentially regulated (FC > 2; p < 0.05). Realtime PCR analysis showed significant differences in the expression of SFRP4 (6.73; p=0.005), CTSC (2.3; p=0.04), SMARCA 1 (1.66; p=0.02), CPM (16.37; p=0.03), MGN5 (1.82; p=0.03), MT1G (-333; p= 0.003) and MT2A (-4.67; p=0.03) with progesterone. The major canonical pathways differentially regulated with progesterone in stromal cells, as analysed by IPA, were EIF2 signalling and the mitochondrial dysfunction pathway. Conclusion: In this study, an in vitro-three dimensional embryo-endometrial cell culture model to investigate the human embryo implantation process was established and this shed more light onto the role of LIF in the embryo implantation process. This model could be used to examine the embryoendometrial interaction that leads to successful implantation, as well as to develop new contraceptive agents. A novel endometrial epithelial progesterone-regulated receptivity marker, ENPP3, was identified and its potential application in ART needs to be explored further. The knowledge gained from the expression of progesterone-regulated genes in the endometrial, glandular and stromal compartments could assist in understanding the molecular mechanisms involved in endometrial receptivity and would be beneficial for improving fertility rates in women, as well as paving way to the development of new endometrium-based fertility regulating agents

Interleukin 6 plays a role in the migration of magnetically levitated mesenchymal stem cells spheroids

Mesenchymal stem cells (MSCs) reside quiescently within a specialised ‘niche’ environment in the bone marrow. However, following appropriate signalling cues, MSCs mobilise and migrate out from the niche, typically toward either sites of injury (a regenerative response) or toward primary tumours (an intrinsic homing response, which promotes MSCs as cellular vectors for therapeutic delivery). To date, very little is known about MSC mobilisation. By adopting a 3D MSC niche model, whereby MSC spheroids are cultured within a type I collagen gel, recent studies have highlighted interleukin-6 (IL-6) as a key cytokine involved in MSC migration. Herein, the ability of IL-6 to induce MSC migration was further investigated, and the key matrix metalloproteinases used to effect cell mobilisation were identified. Briefly, the impact of IL-6 on the MSC migration in a two-dimensional model systems was characterised—both visually using an Ibidi chemotaxis plate array (assessing for directional migration) and then via a standard 2D monolayer experiment, where cultured cells were challenged with IL-6 and extracted media tested using an Abcam Human MMP membrane antibody array. The 2D assay displayed a strong migratory response toward IL-6 and analysis of the membrane arrays data showed significant increases of several key MMPs. Both data sets indicated that IL-6 is important in MSC mobilisation and migration. We also investigated the impact of IL-6 induction on MSCs in 3D spheroid culture, serving as a simplistic model of the bone marrow niche, characterised by fluorescently tagged magnetic nanoparticles and identical membrane antibody arrays. An increase in MMP levels secreted by cells treated with 1 ng/mL IL-6 versus control conditions was noted in addition to migration of cells away from the central spheroid mass

Managing Growth: Best Practices of Family-Owned Businesses

Family-owned businesses represent the majority of business in the United States. As consumers and employees, we are compelled to their sense of trustworthiness that all too often disappears in the business world. Our economy depends on the success of family-owned businesses, but only one third of these organizations successfully transition to the second generation and only one in ten survive to the third generation. While a series of best practices attempt to prescribe solutions their challenges, these practices fail to account for the various types of family-owned businesses. More specifically, many types of family-owned businesses exist as evident by specific transitions in terms of ownership, family and business. Therefore, the study of best practices in family firms must consider the timing of implementation. This study analyzes three family-owned businesses that successfully transitioned from start-up businesses owned by a single controlling owner ready to give up control to an expanding business owned by a sibling partnership with young children. From this analysis of a specific type of family-owned business, six common practices emerged

Coupled effects of local movement and global interaction on contagion

By incorporating segregated spatial domain and individual-based linkage into the SIS (susceptible-infected-susceptible) model, we investigate the coupled effects of random walk and intragroup interaction on contagion. Compared with the situation where only local movement or individual-based linkage exists, the coexistence of them leads to a wider spread of infectious disease. The roles of narrowing segregated spatial domain and reducing mobility in epidemic control are checked, these two measures are found to be conducive to curbing the spread of infectious disease. Considering heterogeneous time scales between local movement and global interaction, a log-log relation between the change in the number of infected individuals and the timescale $\tau$ is found. A theoretical analysis indicates that the evolutionary dynamics in the present model is related to the encounter probability and the encounter time. A functional relation between the epidemic threshold and the ratio of shortcuts, and a functional relation between the encounter time and the timescale $\tau$ are found

Modeling mammalian gastrulation with embryonic stem cells

Understanding cell fate patterning and morphogenesis in the mammalian embryo remains a formidable challenge. Recently, in vivo models based on embryonic stem cells (ESCs) have emerged as complementary methods to quantitatively dissect the physical and molecular processes that shape the embryo. Here we review recent developments in using embryonic stem cells to create both two and three-dimensional culture models that shed light on mammalian gastrulation.Comment: 18 pages, 1 figur

A model for providing emotion awareness and feedback using fuzzy logic in online learning

Monitoring users’ emotive states and using that information for providing feedback and scaffolding is crucial. In the learning context, emotions can be used to increase students’ attention as well as to improve memory and reasoning. In this context, tutors should be prepared to create affective learning situations and encourage collaborative knowledge construction as well as identify those students’ feelings which hinder learning process. In this paper, we propose a novel approach to label affective behavior in educational discourse based on fuzzy logic, which enables a human or virtual tutor to capture students’ emotions, make students aware of their own emotions, assess these emotions and provide appropriate affective feedback. To that end, we propose a fuzzy classifier that provides a priori qualitative assessment and fuzzy qualifiers bound to the amounts such as few, regular and many assigned by an affective dictionary to every word. The advantage of the statistical approach is to reduce the classical pollution problem of training and analyzing the scenario using the same dataset. Our approach has been tested in a real online learning environment and proved to have a very positive influence on students’ learning performance.Peer ReviewedPostprint (author's final draft

It’s not the model that doesn’t fit, it’s the controller! The role of cognitive skills in understanding the links between natural mapping, performance, and enjoyment of console video games

This study examines differences in performance, frustration, and game ratings of individuals playing first person shooter video games using two different controllers (motion controller and a traditional, pushbutton controller) in a within-subjects, randomized order design. Structural equation modeling was used to demonstrate that cognitive skills such as mental rotation ability and eye/hand coordination predicted performance for both controllers, but the motion control was significantly more frustrating. Moreover, increased performance was only related to game ratings for the traditional controller input. We interpret these data as evidence that, contrary to the assumption that motion controlled interfaces are more naturally mapped than traditional push-button controllers, the traditional controller was more naturally mapped as an interface for gameplay

Designing stem cell niches for differentiation and self-renewal

Mesenchymal stem cells, characterized by their ability to differentiate into skeletal tissues and self-renew, hold great promise for both regenerative medicine and novel therapeutic discovery. However, their regenerative capacity is retained only when in contact with their specialized microenvironment, termed the stem cell niche. Niches provide structural and functional cues that are both biochemical and biophysical, stem cells integrate this complex array of signals with intrinsic regulatory networks to meet physiological demands. Although, some of these regulatory mechanisms remain poorly understood or difficult to harness with traditional culture systems. Biomaterial strategies are being developed that aim to recapitulate stem cell niches, by engineering microenvironments with physiological-like niche properties that aim to elucidate stem cell-regulatory mechanisms, and to harness their regenerative capacity in vitro. In the future, engineered niches will prove important tools for both regenerative medicine and therapeutic discoveries