Emerging Imagery: the Great Famine in Nineteenth Century Irish Lit

The critical debate surrounding the Great Famine in Irish Literature centers on the notion of a perceived silence. While some scholars claim that there is a literary void in Irish Literature following this cataclysmic event, others wonder whether language is even capable of describing the extreme physical, emotional, and psychological suffering that is inflicted upon the victims when such tragedies occur. Centuries of imperialism and colonialism had created a class divide so wide and an Irish economy so fragile that when a calamity such as famine occurred, it was the poverty-stricken, predominately Irish-Catholic peasantry that suffered most. Poor and illiterate, powerless and voiceless, they also lacked the ability -- and oftentimes, the willingness -- to articulate the horrors that they experienced, witnessed, and sometimes even committed in order to survive. Trauma, too, would add to their muteness as feelings of guilt, shame, blame, and immeasurable grief followed in its wake. Anguished by the question of why the Famine had occurred and feeling somehow responsible for it, the Irish peasantry were at a loss as to how to cope with it, how to address it, and how to articulate it to those who had not experienced it first-hand. What must also be kept in mind when discussing the Great Famine is that there were two stories to be told in regards to location: those who remained behind in Ireland and those who departed to other shores. Each had their own experiences to share and their own trauma with which to cope. While the horrific images and memories brought on by the Famine still plagued this sector of the Irish population, they also experienced a haunting sense of melancholy and nostalgia for the homeland that they had left behind oftentimes as a last resort in order to survive. While there was a silence of sorts immediately following the Famine that echoed the silence that reverberated across the decimated land, a torrent of imagery would emerge to express the inexpressible and describe the indescriba

Pasireotide versus pituitary surgery: a retrospective analysis of 12 months of treatment in patients with Cushing's disease

Pituitary surgery represents the first-line treatment for most patients with Cushing\u2019s disease (CD). In the case of surgery failure, additional treatment options are required. Pasireotide has shown favourable results in the first-line treatment of patients with CD, who are not candidates for surgery or in the second-line when surgery has failed. The aim of the current study is to compare the effects of surgery and pasireotide treatment in a cohort of patients with CD, and to evaluate the differences in response rate in terms of hormonal and clinical control, and improvement of metabolic complication

PFN1 and integrin-β1/mTOR axis involvement in cornea differentiation of fibroblast limbal stem cells

Ex vivo limbal stem cell transplantation is the main therapeutic approach to address a complete and functional re-epithelialization in corneal blindness, the second most common eye disorder. Although important key points were defined, the molecular mechanisms involved in the epithelial phenotype determination are unclear. Our previous studies have demonstrated the pluripotency and immune-modulatory of fibroblast limbal stem cells (f-LSCs), isolated from the corneal limbus. We defined a proteomic profile especially enriched in wound healing and cytoskeleton-remodelling proteins, including Profilin-1 (PFN1). In this study we postulate that pfn-1 knock down promotes epithelial lineage by inhibiting the integrin-β1(CD29)/mTOR pathway and subsequent NANOG down-expression. We showed that it is possible modulate pfn1 expression levels by treating f-LSCs with Resveratrol (RSV), a natural compound: pfn1 decline is accompanied with up-regulation of the specific differentiation epithelial genes pax6 (paired-box 6), sox17 (sex determining region Y-box 17) and ΔNp63-α (p63 splice variant), consistent with drop-down of the principle stem gene levels. These results contribute to understand the molecular biology of corneal epithelium development and suggest that pfn1 is a potential molecular target for the treatment of corneal blindness based on epithelial cell dysfunction

OpenSPIM - an open access platform for light sheet microscopy

Light sheet microscopy promises to revolutionize developmental biology by enabling live in toto imaging of entire embryos with minimal phototoxicity. We present detailed instructions for building a compact and customizable Selective Plane Illumination Microscopy (SPIM) system. The integrated OpenSPIM hardware and software platform is shared with the scientific community through a public website, thereby making light sheet microscopy accessible for widespread use and optimization to various applications.Comment: 7 pages, 3 figures, 6 supplementary videos, submitted to Nature Methods, associated public website http://openspim.or

Human limbal fibroblast-like stem cells induce immune-tolerance in autoreactive T lymphocytes from female patients with Hashimoto\u2019s thyroiditis

Abstract Background: Due to their \u201cnatural immune privilege\u201d and immunoregulatory properties human fibroblast-like limbal stem cells (f-LSCs) have acquired great interest as a potential tool for achieving immunotolerance. Hashimoto\u2019s thyroiditis (HT) is the most common thyroid autoimmune disease and cause of hypothyroidism. To date, conventional hormone replacement therapy and unspecific immunosuppressive regimens cannot provide a definitive cure for HT subjects. We explored the immunosuppressant potential of human f-LSCs on circulating lymphomonocytes (PBMCs) collected from healthy donors and female HT patients. Methods: We assessed the immunophenotyping of f-LSCs, both untreated and after 48 h of proinflammatory cytokine exposure, by means of quantitative reverse-transcription polymerase chain reaction (qRT-PCR) and flow cytometry. The immunosuppressant effects of f-LSCs on healthy activated PBMCs were investigated in cell-cell contact and transwell settings through cell cycle assay, acridine orange staining, and caspase-3 detection. We also studied T-cell responses and possible Treg conversion by means of flow cytometry. Functional assays were conducted in activated HT lymphocytes cocultured with f-LSCs after carboxyfluorescein succinimidyl ester labeling and intracellular detection of pro- and antiinflammatory cytokines. Results: The hypo-immunogenicity of the f-LSC population depended on both cell contact and soluble factors produced, as well as the undetectable expression of all those molecules required to fully activate T lymphocytes. Following exposure to Th1 cytokines, f-LSCs augmented expression of programmed death-ligand 1 and 2 (PDL-1 and -2), indoleamine-pyrrole-2,3-dioxygenase (IDO), interleukin (IL)-6, and monocyte chemotactic protein 1 (MCP-1) while maintaining their negative phenotype for major histocompatibility (MHC) class II and costimulatory molecules. During coculture, f-LSCs suppressed up to 40% of proliferation in healthy activated PBMCs, arrested them in the G0/G1 cell cycle phase without inducing apoptosis cascade, inverted the CD4/CD8 ratio, and promoted sustained expression of the immunomodulator marker CD69. Under coculture conditions the Th imbalance of autoreactive T cells from female HT patients was fully restored. Conclusions: Our study describes an in vitro coculture system able to prevent inappropriate activation of autoreactive T lymphocytes of female HT patients and to generate a tolerogenic environment even in an inflammatory background. Further investigations are necessary to establish whether this stem cell-based therapy approach in HT could avoid lifetime hormone replacement therapy by inducing T-cell education

Mesenchymal stem cells derived from inflamed dental pulpal and gingival tissue: a potential application for bone formation

Background: Chronic periodontal disease is an infectious disease consisting of prolonged inflammation of the supporting tooth tissue and resulting in bone loss. Guided bone regeneration procedures have become common and safe treatments in dentistry, and in this context dental stem cells would represent the ideal solution as autologous cells. In this study, we verified the ability of dental pulp mesenchymal stem cells (DPSCs) and gingival mesenchymal stem cells (GMSCs) harvested from periodontally affected teeth to produce new mineralized bone tissue in vitro, and compared this to cells from healthy teeth. Methods: To characterize DPSCs and GMSCs, we assessed colony-forming assay, immunophenotyping, mesenchymal/stem cell phenotyping, stem gene profiling by means of flow cytometry, and quantitative polymerase chain reaction (qPCR). The effects of proinflammatory cytokines on mesenchymal stem cell (MSC) proliferation and differentiation potential were investigated. We also observed participation of several heat shock proteins (HSPs) and actin-depolymerizing factors (ADFs) during osteogenic differentiation. Results: DPSCs and GMSCs were successfully isolated both from periodontally affected dental tissue and controls. Periodontally affected dental MSCs proliferated faster, and the inflamed environment did not affect MSC marker expressions. The calcium deposition was higher in periodontally affected MSCs than in the control group. Proinflammatory cytokines activate a cytoskeleton remodeling, interacting with HSPs including HSP90 and HSPA9, thioredoxin-1, and ADFs such as as profilin-1, cofilin-1, and vinculin that probably mediate the increased acquisition in the inflamed environment. Conclusions: Our findings provide evidence that periodontally affected dental tissue (both pulp and gingiva) can be used as a source of MSCs with intact stem cell properties. Moreover, we demonstrated that the osteogenic capability of DPSCs and GMSCs in the test group was not only preserved but increased by the overexpression of several proinflammatory cytokine-dependent chaperones and stress response proteins

Light-sheet microscopy for everyone? Experience of building an OpenSPIM to study flatworm development.

Background: Selective plane illumination microscopy (SPIM a type of light-sheet microscopy) involves focusing a thin sheet of laser light through a specimen at right angles to the objective lens. As only the thin section of the specimen at the focal plane of the lens is illuminated, out of focus light is naturally absent and toxicity due to light (phototoxicity) is greatly reduced enabling longer term live imaging. OpenSPIM is an open access platform (Pitrone et al. 2013 and OpenSPIM.org) created to give new users step-by-step instructions on building a basic configuration of a SPIM microscope, which can in principle be adapted and upgraded to each laboratory’s own requirements and budget. Here we describe our own experience with the process of designing, building, configuring and using an OpenSPIM for our research into the early development of the polyclad flatworm Maritigrella crozieri – a non-model animal. Results: Our OpenSPIM builds on the standard design with the addition of two colour laser illumination for simultaneous detection of two probes/molecules and dual sided illumination, which provides more even signal intensity across a specimen. Our OpenSPIM provides high resolution 3d images and time lapse recordings, and we demonstrate the use of two colour lasers and the benefits of two color dual-sided imaging. We used our microscope to study the development of the embryo of the polyclad flatworm M. crozieri. The capabilities of our microscope are demonstrated by our ability to record the stereotypical spiral cleavage pattern of M. crozieri with high-speed multi-view time lapse imaging. 3D and 4D (3D + time) reconstruction of early development from these data is possible using image registration and deconvolution tools provided as part of the open source Fiji platform. We discuss our findings on the pros and cons of a self built microscope. Conclusions: We conclude that home-built microscopes, such as an OpenSPIM, together with the available open source software, such as MicroManager and Fiji, make SPIM accessible to anyone interested in having continuous access to their own light-sheet microscope. However, building an OpenSPIM is not without challenges and an open access microscope is a worthwhile, if significant, investment of time and money. Multi-view 4D microscopy is more challenging than we had expected. We hope that our experience gained during this project will help future OpenSPIM users with similar ambitions

In vitro generation of pancreatic endocrine cells from human adult fibroblast-like limbal stem cells

Stem cells might provide unlimited supply of transplantable cells for β-cell replacement therapy in diabetes. The human limbus is a highly specialized region hosting a well-recognized population of epithelial stem cells, which sustain the continuous renewal of the cornea, and the recently identified stromal fibroblast-like stem cells (f-LSCs), with apparent broader plasticity. However, the lack of specific molecular markers for the identification of the multipotent limbal subpopulation has so far limited the investigation of their differentiation potential. In this study we show that the human limbus contains uncommitted cells that could be potentially harnessed for the treatment of diabetes. Fourteen limbal biopsies were obtained from patients undergoing surgery for ocular diseases not involving the conjunctiva or corneal surface. We identified a subpopulation of f-LSCs characterized by robust proliferative capacity, expressing several pluripotent stem cell markers and exhibiting self-renewal ability. We then demonstrated the potential of f-LSCs to differentiate in vitro into functional insulin-secreting cells by developing a four-step differentiation protocol that efficiently directed f-LSCs towards the pancreatic endocrine cell fate. The expression of specific endodermal, pancreatic, islet, and β-cell markers, as well as functional properties of f-LSC-derived insulin-producing cells, were evaluated during differentiation. With our stage-specific approach, up to 77% of f-LSCs eventually differentiated into cells expressing insulin (also assessed as C-peptide) and exhibited phenotypic features of mature β-cells, such as expression of critical transcription factors and presence of secretory granules. Although insulin content was about 160-fold lower than what observed in adult islets, differentiated cells processed ∼98% of their proinsulin content, similar to mature β-cells. Moreover, they responded in vitro in a regulated manner to multiple secretory stimuli, including glucose. In conclusion, f-LSCs represent a possible relevant source of autologous, transplantable, insulin-producing cells that could be tested for the reversal of diabetes

MULTIPLE PLURIPOTENT STEM CELL MARKERS IN HUMAN ANAPLASTIC THYROID CANCER: THE PUTATIVE UPSTREAM ROLE OF SOX-2

Background: Anaplastic Thyroid Carcinoma (ATC) is a rare and aggressive endocrine tumor, with highly undifferentiated morphology. It has been suggested that cancer stem cells (CSCs) might play a central role in ATC. The objectives of this study were the following: 1) to characterize CSCs from ex vivo ATC specimens by investigating the expression of several pluripotent stem cell markers; 2) to evaluate in vitro drug resistance modifications after specific CSC transcription factor switch off. Methods: Ex vivo: eight formalin-fixed, paraffin-embedded ATC specimens were analyzed by RT and qRT-PCR and immunohistochemistry. In vitro: in ATC SW1736 cells the expression levels of OCT-4, NANOG and ABCG2 and the sensitivity to either cisplatin or doxorubicin were evaluated after silencing. Results: OCT-4, KLF4 and SOX2 transcription factors and C-KIT and THY-1 stem surface antigens showed variable up-regulation in all ATC cases. The SW1736 cell line was characterized by a high percentage of stem population (10.4 \ub1 2.1 % of cells were aldehyde dehydrogenase positive) and a high expression of several CSC markers (SOX2, OCT4, NANOG, C-MYC, SSEA4). SOX2 silencing down-regulated OCT-4, NANOG and ABCG2. SOX2 silencing sensitized SW1736 cells, causing a significant cell death increase (1.8 fold) in comparison to control cells with 10 \ub5M cisplatin (93.9\ub13.4% vs. 52.6\ub19.4%, p<0.01) and 2.7 fold with 0.5\ub5M doxorubicin (45.8\ub19.9% vs. 17.1\ub13.4% p<0.01). ABCG2 silencing caused increased cell death with both cisplatin (74.9\ub11.4%) and doxorubicin treatment (74.1\ub10.1%) vs. no-target-treated cells (respectively, 45.8\ub11.0% and 48.6\ub11.0%, p<0.001). Conclusions: The characterization of CSCs in ATC through the analysis of multiple pluripotent stem cell markers might be useful in identifying cells with a stem-like phenotype capable of resisting conventional chemotherapy. In addition, our data demonstrate that SOX2 switch-off through ABCG2 transporter down-regulation has a major role in overcoming CSC chemotherapy resistance

Donor age and long-term culture do not negatively influence the stem potential of limbal fibroblast-like stem cells

In regenerative medicine the maintenance of stem cell properties is of crucial importance. Ageing is considered a cause of reduced stemness capability. The limbus is a stem niche of easy access and harbors two stem cell populations: epithelial stem cells and fibroblast-like stem cells. Our aim was to investigate whether donor age and/or long-term culture have any influence on stem cell marker expression and the profiles in the fibroblast-like stem cell population