Isolation and culture of umbilical vein mesenchymal stem cells

Bone marrow contains a population of stem cells that can support hematopoiesis and can differentiate into different cell lines including adipocytes, osteocytes, chondrocytes, myocytes, astrocytes, and tenocytes. These cells have been denoted mesenchymal stem cells. In the present study we isolated a cell population derived from the endothelium and subendothelium of the umbilical cord vein which possesses morphological, immunophenotypical and cell differentiation characteristics similar to those of mesenchymal stem cells isolated from bone marrow. The cells were isolated from three umbilical cords after treatment of the umbilical vein lumen with collagenase. The cell population isolated consisted of adherent cells with fibroblastoid morphology which, when properly stimulated, gave origin to adipocytes and osteocytes in culture. Immunophenotypically, this cell population was found to be positive for the CD29, CD13, CD44, CD49e, CD54, CD90 and HLA-class 1 markers and negative for CD45, CD14, glycophorin A, HLA-DR, CD51/61, CD106, and CD49d. The characteristics described are the same as those presented by bone marrow mesenchymal stem cells. Taken together, these findings indicate that the umbilical cord obtained from term deliveries is an important source of mesenchymal stem cells that could be used in cell therapy protocols.

Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3

Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithelial cells, while endothelin-3 was added to stimulate their growth. By adding endothelin-3, the achievement ratio (viable cell cultures/total cultures) was enhanced to 60% of a total of 10 cultures (initiated from 8 distinct fetal small intestines), allowing the generation of viable epithelial cell cultures. Western blot, real-time PCR and immunofluorescent staining showed that cytokeratins 8, 18 and mouse intestinal mucosa-1/39 had high expression levels in human intestinal epithelial cells. Differentiated markers such as sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV also showed high expression levels in human intestinal epithelial cells. Differentiated human intestinal epithelial cells, with the expression of surface markers (cytokeratins 8, 18 and mouse intestinal mucosa-1/39) and secretion of cytokines (sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV), may be cultured by the thermolysin and endothelin-3 method and maintained for at least 20 passages. This is relatively simple, requiring no sophisticated techniques or instruments, and may have a number of varied applications.

Co-culture of chondrocytes and bone marrow mesenchymal stem cells in vitro enhances the expression of cartilaginous extracellular matrix components

Chondrocytes and bone marrow mesenchymal stem cells (BMSCs) are frequently used as seed cells in cartilage tissue engineering. In the present study, we determined if the co-culture of rabbit articular chondrocytes and BMSCs in vitro promotes the expression of cartilaginous extracellular matrix and, if so, what is the optimal ratio of the two cell types. Cultures of rabbit articular chondrocytes and BMSCs were expanded in vitro and then cultured individually or at a chondrocyte:BMSC ratio of 4:1, 2:1, 1:1, 1:2, 1:4 for 21 days and cultured in DMEM/F12. BMSCs were cultured in chondrogenic induction medium. Quantitative real-time RT-PCR and Western blot were used to evaluate gene expression. In the co-cultures, type II collagen and aggrecan expression increased on days 14 and 21. At the mRNA level, the expression of type II collagen and aggrecan on day 21 was much higher in the 4:1, 2:1, and 1:1 groups than in either the articular chondrocyte group or the induced BMSC group, and the best ratio of co-culture groups seems to be 2:1. Also on day 21, the expression of type II collagen and aggrecan proteins in the 2:1 group was much higher than in all other groups. The results demonstrate that the co-culture of rabbit chondrocytes and rabbit BMSCs at defined ratios can promote the expression of cartilaginous extracellular matrix. The optimal cell ratio appears to be 2:1 (chondrocytes:BMSCs). This approach has potential applications in cartilage tissue engineering since it provides a protocol for maintaining and promoting seed-cell differentiation and function.

Co-culture of apoptotic breast cancer cells with immature dendritic cells: a novel approach for DC-based vaccination in breast cancer

A dendritic cell (DC)-based vaccine strategy could reduce the risk of recurrence and improve the survival of breast cancer patients. However, while therapy-induced apoptosis of hepatocellular and colorectal carcinoma cells can enhance maturation and antigen presentation of DCs, whether this effect occurs in breast cancer is currently unknown. In the present study, we investigated the effect of doxorubicin (ADM)-induced apoptotic MCF-7 breast cancer cells on the activation of DCs. ADM-induced apoptotic MCF-7 cells could effectively induce immature DC (iDC) maturation. The mean fluorescence intensity (MFI) of DC maturity marker CD83 was 23.3 in the ADM-induced apoptotic MCF-7 cell group compared with 8.5 in the MCF-7 cell group. The MFI of DC co-stimulatory marker CD86 and HLA-DR were also increased after iDCs were treated with ADM-induced apoptotic MCF-7 cells. Furthermore, the proliferating autologous T-lymphocytes increased from 14.2 to 40.3% after incubated with DCs induced by apoptotic MCF-7 cells. The secretion of interferon-γ by these T-lymphocytes was also increased. In addition, cell-cell interaction between apoptotic MCF-7 cells and iDCs, but not soluble factors released by apoptotic MCF-7 cells, was crucial for the maturation of iDCs. These findings constitute a novel in vitro DC-based vaccine strategy for the treatment of breast cancer by ADM-induced apoptotic MCF-7 cells.

Culture of bovine ovarian follicle wall sections maintained the highly estrogenic profile under basal and chemically defined conditions

Follicle cultures reproduce in vitro the functional features observed in vivo. In a search for an ideal model, we cultured bovine antral follicle wall sections (FWS) in a serum-free defined medium (DM) known to induce 17β-estradiol (E2) production, and in a nondefined medium (NDM) containing serum. Follicles were sectioned and cultured in NDM or DM for 24 or 48 h. Morphological features were determined by light microscopy. Gene expression of steroidogenic enzymes and follicle-stimulating hormone (FSH) receptor were determined by RT-PCR; progesterone (P4) and E2 concentrations in the media were measured by radioimmunoassay. DM, but not NDM, maintained an FWS morphology in vitro that was similar to fresh tissue. DM also induced an increase in the expression of all steroidogenic enzymes, except FSH receptor, but NDM did not. In both DM and NDM, there was a gradual increase in P4 throughout the culture period; however, P4 concentration was significantly higher in NDM. In both media, E2 concentration was increased at 24 h, followed by a decrease at 48 h. The E2:P4 ratio was higher in DM than in NDM. These results suggest that DM maintains morphological structure, upregulates the expression of steroidogenic enzyme genes, and maintains steroid production with a high E2:P4 ratio in FWS cultures.

Managing Risk in a Culture of Rights: Providing Support and Treatment in Community-Based Settings for Persons with Intellectual Disabilities who Sexually Offend

People with intellectual disability who sexually offend commonly live in community-based settings since the closing of all institutions across the province of Ontario. Nine (n=9) front line staff who provide support to these individuals in three different settings (treatment setting, transitional setting, residential setting) were interviewed. Participants responded to 47 questions to explore how sex offenders with intellectual disability can be supported in the community to prevent re-offenses. Questions encompassed variables that included staff attitudes, various factors impacting support, structural components of the setting, quality of life and the good life, staff training, staff perspectives on treatment, and understanding of risk management. Three overlapping models that have been supported in the literature were used collectively for the basis of this research: The Good Lives Model (Ward & Gannon, 2006; Ward et al., 2007), the quality of life model (Felce & Perry, 1995), and variables associated with risk management. Results of this research showed how this population is being supported in the community with an emphasis on the following elements: positive and objective staff attitude, teamwork, clear rules and protocols, ongoing supervision, consistency, highly trained staff, and environments that promote quality of life. New concepts arose which suggested that all settings display an unequal balance of upholding human rights and managing risks when supporting this high-risk population. This highlights the need for comprehensive assessments in order to match the offender to the proper setting and supports, using an integration of a Risk, Need, Responsivity model and the Good Lives model for offender rehabilitation and to reduce the likelihood of re-offenses.

Brock Essay - The Political Culture of Ontario

An essay submitted by Sean O'Sullivan to Professor W.A. Matheson, 25 April 1977. The focus of the essay is stable government, "As one of the chief, if not the predominant, force in giving Confederation its political shape, Ontario helped bring about a central government designed to promote, and dedicated to preserve, stability. In the governing of their own province, the people of Ontario have been faithful to that same goal of stability. Perhaps that steadfast attitude says more than anything else about the political culture of Ontario."

Innovations pédagogiques et culture disciplinaire en enseignement supérieur

Cette recherche qualitative porte sur les innovations pédagogiques utilisées à l’Université de Montréal en contexte de cultures disciplinaires. La recherche visait à étudier les innovations pédagogiques, entendues comme tout enseignement dispensé de manières différentes de la pratique traditionnelle du cours magistral, proposées par les professeurs dans une université fortement engagée en recherche. ll nous a paru utile de contribuer à compléter les savoirs existants dans ce domaine peu exploré, particulièrement lorsque nous savons que les innovations pédagogiques en contexte de cultures disciplinaires n’ont encore pas été étudiées à l’Université de Montréal. D’un point de vue social, la pertinence de cette recherche réside dans le cadre de la valorisation de l’enseignement universitaire souhaitée tant par les politiques, les institutions et la société que les professeurs et, au-delà, elle invite au rééquilibrage des deux piliers indispensables à l’université centrée sur la recherche qui laisse paraître une relation dichotomique marquée entre enseignement et recherche. Deux modes de cueillettes de données ont été privilégiés : les entrevues individuelles semi-structurées et un entretien de groupe auprès de trente-deux professeurs lauréats du Prix d’excellence en enseignement de l’Université de Montréal. Pour cette recherche, nous avons employé la théorisation ancrée comme méthode d’analyse de données recueillies selon d’autres approches. En d’autres mots, nous avons souhaité utiliser la théorisation ancrée comme un « processus » (Paillé, 1994, p. 149) d’analyse des données avec pour objectif d’approfondir l’objet de notre recherche par-delà la simple analyse descriptive sans pour autant prétendre à une théorisation avancée. En premier lieu, nos résultats nous ont permis de connaître les innovations pédagogiques utilisées dans l’enceinte de notre terrain de recherche, l’Université de Montréal et de dresser un portrait actualisé de leurs innovateurs. Nous avons aussi exploré les raisons qui amènent les professeurs à innover, décrit le processus nécessaire à l’innovation pédagogique et expliqué les freins s’opposant à cette dernière. En second lieu, nos résultats énumèrent les différences liées à la matière d’enseignement, au champ de recherche et soulignent le rapport de la discipline à l’innovation comme critère déterminant à son implication. En dernier lieu, nos résultats révèlent l’existence d’une expertise pédagogique partagée au sein de l’Université de Montréal qui permet de rompre la solitude des professeurs par un soutien entre pairs et favorise la transférabilité des innovations pédagogiques d’une culture disciplinaire à une autre. Finalement, nous présentons une théorisation actualisée des paramètres constituant une innovation pédagogique et détaillons les nouveaux paramètres qui influent sur la construction d’une innovation pédagogique. Nous concluons cette thèse par des recommandations et des pistes de recherches.

Telling the truth about HIV? Testing and disclosure in a culture of stigma

Étude de cas / Case study

Studies on the macrobenthic community of Cochin backwaters with special reference to culture of Eriopisa chilkensis (Gammaridae-Amphipoda)

This thesis Entitled studies on the macrobenthic community of cochin backwaters with special reference to culture of eriopisa chilkensis (Gammaridae- amphipoda).Benthic organisms are usually studied for environmental impact assessment, pollution control and resource conservation. The benthic monitoring component has three major objectives: 1) characterize the benthic communities to assess the estuarine health, 2) determine seasonal and spatial variability in benthic communities, and 3) detect changes in the estuarine community through examination of changes in abundances of specific indicator taxa and other standard benthic indices.Cochin backwaters situated at the tip of the northern Vembanad lake is a tropical positive estuarine system. The backwaters of Kerala support as much biological productivity and diversity as tropical rain forest and are responsible for the rich fishery potential of Kerala. Backwaters also act as nursery grounds for commercially important prawns and fishes.The thesis has been subdivided into seven chapters. The first chapter gives a general introduction about the topic and also highlights the scope and purpose of the study. The second chapter covers the methodology adopted for the collection and analysis of water quality parameters, sediment and the macrobenthic fauna.Chapter 3 deals with hydrographic features, sediment characteristics and the spatial variation and abundance of macrobenthic fauna in the Cochin estuary.Chapter 4 explains the impact of organic enrichment on macrobenthic popUlation in the Cochin estuary and includes the comparison of the present data with the earlier work in this region.Chapter 5 deals with seasonal variability in abundance of macrobenthic species in the estuary. The study was conducted from 9 stations during three seasons (pre-monsoon, monsoon and post-monsoon) in 2003.Chapter 6 deals with Life history and Population Dynamics of Eriopisa chilkensis Chilton (Gammaridae-Amphipoda). The life cycle of the gammarid amphipod Eriopisa chilkensis from the Cochin estuary, south west coast of India was studied for the first time under laboratory conditions.

Development of plant tissue culture of Plumbago rosea Linn. for enhanced production of secondary metabolites

P rosea syn. Indica belong to the family of plumbaginaceae, is an important medicinal plant, cultivated widely in India. The roots of these plant are generally used for medicinal purposes mainly as diuretic, germicidal, vessicant, and abortifacient. It is also used for anaemia, diarrhea, leprosy and common wart. The bark of the root contains orange yellow pigment named plumbagin, a crystalline substance, belongs to the class of naphthoquinone. Its chemical structure is 5-hydroxy 2-methyl 1,4naphthoquinone. Apart from P rosea, P zeylanica, P europea, Drosera and Drosophyllum also contains plumbagin. The most exploited source of plumbagin is, of course, P. rosea roots. The roots contain O.9mg/ g D.Wt. of plumbagin in the roots. These plants grow very slowly and the roots suitable for plumbagin extraction can be obtained only after several years of growth. The productivity of the plant is also rather poor. The focus of the present study was to develop alternative strategies to obtain plumbagin. The tissue culture of P rosea for micropropagation has been studied

Primary hemocyte culture of Penaeus monodon as an in vitro model for white spot syndrome virus titration, viral and immune related gene expression and cytotoxicity assays

Immortal cell lines have not yet been reported from Penaeus monodon, which delimits the prospects of investigating the associated viral pathogens especially white spot syndrome virus (WSSV). In this context, a method of developing primary hemocyte culture from this crustacean has been standardized by employing modified double strength Leibovitz-15 (L-15) growth medium supplemented with 2% glucose, MEM vitamins (1 ), tryptose phosphate broth (2.95 g l 1), 20% FBS, N-phenylthiourea (0.2 mM), 0.06 lgml 1 chloramphenicol, 100 lgml 1 streptomycin and 100 IU ml 1 penicillin and hemolymph drawn from shrimp grown under a bio-secured recirculating aquaculture system (RAS). In this medium the hemocytes remained viable up to 8 days. 5-Bromo-20-deoxyuridine (BrdU) labeling assay revealed its incorporation in 22 ± 7% of cells at 24 h. Susceptibility of the cells to WSSV was confirmed by immunofluoresence assay using a monoclonal antibody against 28 kDa envelope protein of WSSV. A convenient method for determining virus titer as MTT50/ml was standardized employing the primary hemocyte culture. Expression of viral genes and cellular immune genes were also investigated. The cell culture could be demonstrated for determining toxicity of a management chemical (benzalkonium chloride) by determining its IC50. The primary hemocyte culture could serve as a model for WSSV titration and viral and cellular immune related gene expression and also for investigations on cytotoxicity of aquaculture drugs and chemicals

Application of primary haemocyte culture of Penaeus monodon in the assessment of cytotoxicity and genotoxicity of heavy metals and pesticides

Lack of shrimp cell lines has hindered the study of pollutants which adversely affects shrimp health and its export value. In this context a primary haemocyte culture developed from Penaeus monodon was employed for assessing the cytotoxicity and genotoxicity of two heavy metal compounds, cadmium chloride and mercuric chloride and two organophosphate insecticides, malathion and monocrotophos. Using MTT assay 12 h IC50 values calculated were 31.09 16.27 mM and 5.52 1.16 mM for cadmium chloride and mercuric chloride and 59.94 52.30 mg l 1 and 186.76 77.00 mg l 1 for malathion and monocrotophos respectively. Employing Comet assay, DNA damage inflicted by these pollutants on haemocytes were evaluated and the pollutants induced DNA damage in >60% of the cells. The study suggested that haemocyte culture could be used as a tool for quantifying cytotoxicity and genotoxicity of aquaculture drugs, management chemicals and pollutants

Made in “Polish village” – modern ethno-design vs. traditional culture of Polish countryside

Tradycyjna polska wieś – kolorowa, wesoła, zintegrowana z naturą i rządząca się swoimi prawami to zbiorowy mit, odległy w czasie. To klasyczny temat wielu poszukiwań etnografi cznych – terenowych, naukowych opracowań i porównań. We współczesnym świecie mamy do czynienia jedynie z wybranymi elementami – tradycjami jeszcze żywymi lub przetworzonymi i dostosowanymi do wymogów współczesnego odbiorcy – turysty, ale także artysty i projektanta. Obecne zainteresowanie polską sztuką ludową i rzemiosłem to kolejna fala mody folkowej. Ten ogólnoeuropejski trend, poszukujący w kulturze egalitarnej natchnienia i inspiracji swój początek bierze w I połowie XIX wieku. W polskiej sztuce dekoracyjnej (wzornictwie przemysłowym) także nastąpił powrót do źródeł, a w ludowości i wiejskości doszukiwano się narodowego ducha, inspiracją dla polskich artystów był angielski Ruch Arts and Craft, a w ich projektach widoczne były echa fi lozofi i Johna Ruskina oraz Williama Morrisa m. in. odrzucających masową produkcję, mającą wg nich zły wpływ na jakość produkowanych przedmiotów, jednocześnie na piedestał wynoszących umiejętności rękodzielnicze i rzemieślnicze. Po okresie PRLu utożsamianym głównie z funkcjonowaniem Cepelii i sprowadzeniem produktów inspirowanych ludowością do roli masowych dekoracji, stanowiących często synonim kiczu, powróciła moda na polskie rękodzieło, tradycyjną sztukę ludową oraz na inspiracje wypływające z obserwowania wiejskiego stylu życia. Współczesne poszukiwania polskich twórców w obszarze tworzących w ramach nurtu zwanego etnodizajnem przebiegają dwutorowo. Z jednej strony to inspiracja tylko przejawami ludowości, jej konkretnymi wytworami, które stanowią głównie dekorację dla współczesnych rozwiązań formalnych i technologicznych. Jednak coraz silniejszy jest drugi nurt,starający się wniknąć głębiej, którego celem jest zrozumienie istoty tradycyjnej kultury ludowej polskiej wsi. To inspiracja wiejskim stylem życia, wraz z jego zaletami i wadami, to wreszcie inspiracja zmieniającym się krajobrazem kulturowym i poszukiwanie nowych rozwiązań, dzięki którym odbiorca zbliży się do natury. Projekty zaliczane do nurtu etnodizajnu cechuje przede wszystkim różnorodność wykorzystanych materiałów (tradycyjnych i nowoczesnych), sposób ich przetworzenia, zachowanie pierwotnej funkcji lub nadanie nowej. Większość tych projektów sytuuje się też w nurcie eko-artu, sztuki nawiązującej do natury, z wykorzystaniem ekologicznych materiałów. Jest więc etnodizajn XXI kontynuacją ideologii stworzonej już w wieku XIX – przeciwstawieniem dla przemysłu, technicyzacji i umasowienia. A dla współczesnego człowieka – miejskiego – może być etnodizajn powrotem do źródeł.