Investigating Differentiation: A Role for Organelle Inheritance in Epidermal Growth

Balanced growth and differentiation is essential to tissue morphogenesis and homeostasis. How imbalances arise in disease states such as cancers is poorly understood. Loss of differentiation is associated with poorer prognosis in human patients and increasing malignancy in animal models. Here we explore this intersection between growth and differentiation in the context of epidermal development, where populations of stem cells are maintained in careful equilibrium and induced to proliferate and differentiate in response to stimuli such as injury or cyclical growth signals. During development, a naïve epidermis undergoes rapid proliferation and differentiation to form skin containing hair follicles, neurons, immune surveillance populations and melanocytes. The epidermis maintains homeostasis through many phases of development including rapid cytoskeletal dynamics and temporally coupled inductions of protein synthesis. However, it is still poorly understood how this maintenance is coordinated and sustained. After creating a quantitative differentiation assay to fractionate proliferative and differentiating cells from the embryonic mouse epidermis, I used transcriptional profiling to gain a deeper understanding of novel aspects of this process in vivo. By profiling time points across development spanning the naïve and fully competent epidermis I dissected mechanisms essential for establishing and maintaining the differentiated interfollicular epidermis. This map of the transcriptional landscape served as a tool for forming hypotheses about the association between a pathway or molecule of interest and epidermal development. Probing these profiles allowed me to directly assess the correlation between expression levels and known key regulators of epidermal differentiation. With this new understanding of signatures associated with key differentiation steps in the normal epidermis, I explored how genes commonly dysregulated in epithelial tumors may be involved in this developmental differentiation process. I mined my transcriptional profiles to identify overlap with genes reported to be dysregulated in a range of epithelial tumors. I then devised in vivo epidermal RNAi screen to identify which of these genes were candidate regulators of normal epidermal development. In utero lentiviral injection allows for direct manipulation of the developing epidermis and continued embryo development. The goal of the screen was to assess whether when a particular gene is lost during early epidermal development, the resulting epidermal clone is formed normally with respect to differentiation. Using a quantitative differentiation assay in combination with barcoded high throughput sequencing, I revealed how each gene altered differentiation. My screen identified a number of novel target genes likely to regulate individual steps of differentiation or differentiation more globally. The use of a tumor prone TGFbRII conditional knockout mouse line allowed for comparison of differentiation behavior in a more disease relevant setting. The use of wild type embryos implicated surprising new genes as potential regulators of differentiation. Focusing on one unexpected hit, peroxisome-associated protein PEX11b, I found that Pex11b-deficient epidermis fails to differentiate and form a barrier essential for life. Further study revealed mitotic changes associated with Pex11b-deficient basal progenitors including a mitotic delay, during which spindles rotate uncontrollably, perturbing polarized divisions and skewing daughter fates. Probing deeper, we discovered that without PEX11b, peroxisomes function, but fail to segregate properly. Intriguingly, peroxisome localization is directly coupled to mitotic progression, and when peroxisomes are ectopically mis-localized, mitotic abnormalities occur. Together, our findings unveil a hitherto unforeseen role for organelle inheritance in mitosis and spindle alignment, in the choice of daughter progenitors to differentiate or remain stem-like, and in maintaining proper tissue architecture

Replicative stress, stem cells and aging

Cilj ovog rada je analizirati mogućnosti razvoja specifičnih oblika turizma kao mjeru održivog razvoja turizma na području Europske unije. Europa kao vodeća turistička regija svijeta ima dobre temelje i tradiciju turizma te u skladu s time i olakšane mogućnosti daljnjeg razvoja raznih oblika turizma. U radu se analizira razvoj turističke politike EU te posebno mjere i aktivnosti usmjerene u cilju održivog razvoja turizma. Poseban osvrt se daje na mogućnosti razvoja pojedinih specifičnih oblika turizma kao što su ekoturizam, sportsko-rekreacijski turizam s posebnim osvrtom na cikloturizam, zatim omladinski turizam, kulturni turizam, enogastronomski te ukratko na ostale specifične oblike turizma u odabranim zemljama članicama EU s posebnim osvrtom na Republiku Hrvatsku. U radu su korišteni sekundarni izvori podataka pri čemu je proučena relevantna znanstvena i stručna literatura te razni internetski izvori o temi istraživanja.The purpose of this graduate thesis is to analyze the possibilites of special interest tourism development as a measure of sustainable tourism development in the European Union and in Republic of Croatia. Europe is the leading tourism region of the world; it has its background and tradition of tourism and facilitated possibilities od further development of numerous forms of tourism. The dissertation analyzes development of tourism policy of the EU, particularly measures and activities aimed at the sustainable development of tourism. Special attention is given to the development of ecotourism, sports and recreational tourism with emphasis on cyclotourism, youth tourism, cultural tourism, enogastronomic tourism and, shortly, on other special interest tourism in selected EU member states with emphasis on the Republic of Croatia. The dissertation is based on secondary sources that were studied with all the relevant professional and scientific literature on the topic of research; also numerous online sources have been studied in addition to those sources

ATR and H2AX Cooperate in Maintaining Genome Stability under Replication Stress*S⃞

Chromosomal abnormalities are frequently caused by problems encountered during DNA replication. Although the ATR-Chk1 pathway has previously been implicated in preventing the collapse of stalled replication forks into double-strand breaks (DSB), the importance of the response to fork collapse in ATR-deficient cells has not been well characterized. Herein, we demonstrate that, upon stalled replication, ATR deficiency leads to the phosphorylation of H2AX by ATM and DNA-PKcs and to the focal accumulation of Rad51, a marker of homologous recombination and fork restart. Because H2AX has been shown to play a facilitative role in homologous recombination, we hypothesized that H2AX participates in Rad51-mediated suppression of DSBs generated in the absence of ATR. Consistent with this model, increased Rad51 focal accumulation in ATR-deficient cells is largely dependent on H2AX, and dual deficiencies in ATR and H2AX lead to synergistic increases in chromatid breaks and translocations. Importantly, the ATM and DNA-PK phosphorylation site on H2AX (Ser139) is required for genome stabilization in the absence of ATR; therefore, phosphorylation of H2AX by ATM and DNA-PKcs plays a pivotal role in suppressing DSBs during DNA synthesis in instances of ATR pathway failure. These results imply that ATR-dependent fork stabilization and H2AX/ATM/DNA-PKcs-dependent restart pathways cooperatively suppress double-strand breaks as a layered response network when replication stalls

Burden of Sickle Cell Disease in Ghana: The Korle-Bu Experience

In Africa, sickle cell disease (SCD) is a major public health problem with over 200,000 babies born per year. In Ghana, approximately 15,000 (2%) of Ghanaian newborns are diagnosed with SCD annually. A retrospective review of medical records of all SCD patients aged 13 years and above, who presented to the sickle cell clinic at Ghana Institute of Clinical Genetics (GICG), Korle-Bu, from 1st January 2013 to 31st December 2014, was carried out, using a data abstraction instrument to document their phenotypes, demographics, attendance/clinic visits, pattern of attendance, and common complications seen. During the period under review 5,451 patients were seen at the GICG, with 20,788 clinic visits. The phenotypes were HbSS (55.7%) and HbSC (39.6%) with other sickle cell phenotypes (4.7%). Out of the 20,788 clinic visits, outpatient visits were 15,802 (76%), and urgent care visits were 4,986 (24%), out of which 128 (2.6%) patients were admitted to the Teaching Hospital for further management of their acute complications. There were 904 patient referrals (out of 5,451 patients) for specialist care; the 3 specialties that had the most referrals were Obstetrics and Gynaecology (168 patients), Orthopaedics (150 patients), and Ophthalmology (143 patients). In 2014, complications seen at KBTH included 53 patients with avascular necrosis (AVN) and 61 patients with chronic leg ulcers. Our centre has a large number of patients living with sickle cell disease. From our experience, early recognition and referral of sickle cell related complications can reduce morbidity and mortality associated with this disease. A multidisciplinary approach to care of SCD patients is therefore important

Impact of socio-demographic and economic factors on intimate partner violence justification among women in union in Papua New Guinea

Peer reviewedPublisher PD

Panâ cancer clinical and molecular analysis of racial disparities

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/153667/1/cncr32598_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/153667/2/cncr32598.pd

Isolation and enrichment of newborn and adult skin stem cells of the interfollicular epidermis

The interfollicular epidermis regenerates from a heterogeneous population of basal cells undergoing either self-renewal or terminal differentiation, thereby balancing cell loss in tissue turnover or in wound repair. In this chapter, we describe a reliable and simple method for isolating interfollicular epithelial stem cells from the skin of newborn mice or from tail and ear skin of adult mice using fluorescence-activated cell sorting (FACS). We also provide a detailed protocol for culturing interfollicular epidermal stem cells and to assess their proliferative potential and self-renewing ability. These techniques are useful for directly evaluating epidermal stem cell function in normal mice under different conditions or in genetically modified mouse models