Characterization and expression of senescence marker in prolonged passages of rat bone marrow-derived mesenchymal stem cells

The present study is aimed at optimizing the in vitro culture protocol for generation of rat bone marrow- (BM-) derived mesenchymal stem cells (MSCs) and characterizing the culture-mediated cellular senescence. The initial phase of generation and characterization was conducted using the adherent cells from Sprague Dawley (SD) rat’s BM via morphological analysis, growth kinetics, colony forming unit capacity, immunophenotyping, and mesodermal lineage differentiation. Mesenchymal stem cells were successfully generated and characterized as delineated by the expressions of CD90.1, CD44H, CD29, and CD71 and lack of CD11b/c and CD45 markers. Upon induction, rBM-MSCs differentiated into osteocytes and adipocytes and expressed osteocytes and adipocytes genes. However, a decline in cell growth was observed at passage 4 onwards and it was further deciphered through apoptosis, cell cycle, and senescence assays. Despite the enhanced cell viability at later passages (P4-5), the expression of senescence marker, β-galactosidase, was significantly increased at passage 5. Furthermore, the cell cycle analysis has confirmed the in vitro culture-mediated cellular senescence where cells were arrested at the G0/G1 phase of cell cycle. Although the currently optimized protocols had successfully yielded rBM-MSCs, the culture-mediated cellular senescence limits the growth of rBM-MSCs and its potential use in rat-based MSC research

An Investigation of Object Shadows Utilization In 3D Shape Re-Construction Using Inexpensive Equipment

An approach for automatic 3D object re-construction using its shadow ispresented. The approach investigates the use of information inherited by thegenerated object shadows to re-construct the object geometry. An algorithm isdeveloped that make use of object height information for the directions associatedwith the incident light and the generated object shadows, hence, acquired heightfeatures represents the object features that have actually obstructed the incidentlight. The technique is tested using objects of different shapes. Close to realmeasurements are gained and the overall accuracy of the system is found to bewithin 0.75 mm using the adopted imaging hardware and setup. Obtained resultsconfirmed the validity of the proposed approach

Production of lentiviral vector with polycistronic transcripts for reprogramming of mouse fibroblast cells

iPS cells were originally generated using monocistronic retroviral vectors carrying the Yamanaka factors 'OSKM'. The development of a polycistronic viral vector with OSKM linked by 2A peptides has simplified reprogramming procedure and reduced the risk of multiple proviral integrations and insertional mutagenesis. In this study, we demonstrated the production of the polycistronic lentiviral vector encoding OSKM in a single cassette without a reporter gene or drug-based selection system. Syncytia formations were clearly seen following the co-transfection of a lentiviral plasmid construct with the structural and packaging plasmids. The virion was collected at 48 hours post-transfection. Afterwards, the viral titers were measured by the expression of Sox2 protein from transduced HT1080 cells. Subsequently, Oct4 expression was successfully detected in mouse fibroblasts in the range of 5, 10 and 20 MOIs with expression of 90.7%, 97.5% and 98%, respectively. The results obtained from this study could be used as a model for the production of OSKM lentiviral vector for newcomers to cellular reprogramming research

Generation of induced pluripotent stem cells by a polycistronic lentiviral vector in feeder- and serum- free defined culture

Induced pluripotent stem cells (iPSCs) have great potentials for regenerative medicine. However, serious concerns such as the use of the viral-mediated reprogramming strategies and exposure of iPSCs to animal products from feeder cells and serum-containing medium have restricted the application of iPSCs in the clinics. Therefore, the generation of iPSCs with minimal viral integrations and in non-animal sourced and serum-free medium is necessary. In this report, a polycistronic lentiviral vector carrying Yamanaka's factors was used to reprogram mouse fibroblasts into iPSCs in feeder- and xeno-free culture environment. The generated iPSCs exhibited morphology and self-renewal properties of embryonic stem cells (ESCs), expression of specific pluripotent markers, and potentials to differentiate into the three-major distinct specialized germ layers in vitro. The iPSCs were also shown to have the potential to differentiate into neural precursor and neurons in culture, with greater than 95% expression of nestin, Pax6 and βIII-tubulin. This body of work describes an alternative method of generating iPSCs by using polycistronic lentiviral vector that may minimize the risks associated with viral vector-mediated reprogramming and animal derived products in the culture media

Factors influencing eating behavior of Benghazi University students

Background: University students are more exposed to new individual and environmental influences. This transition period is considered as a risky life phase because it’s characterized by changing in physical and social status as well as changing in the lifestyle that will affect the eating behavior of students. Aims and Objectives: The current study aimed to determine the factors influencing the eating behaviors of Benghazi University students. Materials and Methods: A cross sectional study was undertaken for a period from January to May 2019 in Benghazi University. Samples of 300 students were requested to fill out a questionnaire. SPSS was used to analyze the data. Results: After starting university, (64%) of students stated that they had a change in eating behavior and (59%) of participants reported unhealthy eating pattern. About (67%) of students had a sedentary lifestyle with the majority of them were having unhealthy eating patterns (P value = 0.000). According to the BMI the majority of students had normal weight (62.6%). About (80%) of student reported that the lack of time to prepare a healthy meal during study period was effective and More than half of the students reported that inaccessibility of healthy food, student’s positive emotions, poor knowledge of healthy food, and stress associated with exams period were effective. There was a statistical difference between student’s eating patterns and poor knowledge, lack of time, stress, body weight concerns, negative emotions, peer pressure, lack of parental control, mass media and social life (P value < 0.05). Conclusion: This study concluded that the majority of students undergo a negative shift in their eating and lifestyle after starting university, and there is statistical difference between many factors and student’s eating pattern

ASSESSMENT OF MOLECULAR BIOMARKERS FOR BLADDER CANCER DIAGNOSIS, GRADING AND PROGNOSIS

Worldwide, bladder cancer is a very significant public health problem, in terms of prevalence, mortality and management for individuals and their families. Bladder cancer is the fourth most commonly diagnosed cancer in men and one of the heaviest cancers in terms of cost. Although transurethral resection of the bladder followed by intravesical instillation of live attenuated Bacillus Calmette–Guérin (BCG) is considered as the gold standard for patients with intermediate and high risk, only a small portion of patient responds to the BCG-therapy. Bladder cancer management is faced by the therapy failure, great side effects and some difficulties in histological classification. Accordingly, growing interest is given to the use of genetic, epigenetic and immunologic biomarkers for molecular signature characterization and tumor stratification. In Morocco, great efforts have been made to contribute to the improvement of management of bladder cancer and many studies were made to evaluate some genetic and epigenetic biomarkers. Generated data is of a great interest for characterization of bladder cancer tumors in Morocco and could be used for better management of this disease

Induced pluripotent stem cells: reprogramming platforms and applications in cell replacement therapy

The generation of induced pluripotent stem cells (iPSCs) from differentiated mature cells is one of the most promising technologies in the field of regenerative medicine. The ability to generate patient-specific iPSCs offers an invaluable reservoir of pluripotent cells, which could be genetically engineered and differentiated into target cells to treat various genetic and degenerative diseases once transplanted, hence counteracting the risk of graft versus host disease. In this context, we review the scientific research streams that lead to the emergence of iPSCs, the roles of reprogramming factors in reprogramming to pluripotency, and the reprogramming strategies. As iPSCs serve tremendous correction potentials for various diseases, we highlight the successes and challenges of iPSCs in cell replacement therapy and the synergy of iPSCs and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 gene editing tools in therapeutics research

Lack of methylation on transgene leads to high level and persistent transgene expression in induced pluripotent stem cells

Short-lived therapeutic gene expression in mammalian cells by DNA methylation is one of the major challenges in gene therapy. In this study, we assessed the implication of DNA methylation on the duration of GFP expression in mouse embryonic stem (ES) and mouse induced pluripotent stem (iPS) cells. The cells were transduced with lentivirus (LV) carrying green fluorescent protein (GFP) driven by either human elongation factor (EF1α) or cytomegalovirus (CMV) promoter. Transduced iPS cells exhibited higher percentage of GFP+ cells with persistent mean fluorescent intensity than transduced ES cells. Analysis on the integrated copy of transgene in the population of the transduced cells demonstrated similar copy number. However, significant increase in GFP intensity following 5-azaC treatment was observed in transduced ES cells only, suggesting the influence of DNA methylation in transgene silencing. Subsequent DNA methylation analysis showed that the promoter and the GFP region of the provirus in iPS cells had negligible methylation profile compared to transduced ES cells. Interestingly, sustained transgene expression was observed upon directed differentiation of transduced iPS cells towards CD34+ CD45+ cells. Hence, this study has shown that favourable transgene activity from lentiviral transduced iPS cells was due to the lack of methylation at the proviral regions

La investigación sobre el hidraulismo andalusí y los asentamientos localizados en el Alto Maestrazgo (Castellón)

Editada en la Fundación Empresa PúblicaLos asentamientos rurales andalusíes incluyen es sus áreas de influencia política zonas de residencia y espacios hidráulicos. Estos últimos, generalmente pequeños y medianos, son el resultado de un diseño previo, llevado a cabo por las comunidades campesinas a partir de la captación del agua y de las características físicas del territorio. En esta investigación, mediante diversas técnicas de trabajo de campo, junto con la documentación escrita y la toponimia, se analizan algunos sistemas hidráulicos localizados en el que fuera distrito administrativo andalusí de Culla (Castellón).Rural andalusí settlements include both residential and hydraulic zones in their areas the political influence. These hydraulic zones, usually small and medium-sized, are the result of a previous design carried out by the peasant communities and determined by water collecting and the features of the terrain. This paper focusses on some hydraulic systems located in the old andalusí administrative district of Culla in the Spanish province of Castellón. Different field work techniques along with toponyms, documents and other references Me used in this research.Publicad