CFP and YFP, but Not GFP, Provide Stable Fluorescent Marking of Rat Hepatic Adult Stem Cells

The stable expression of reporter genes in adult stem cells (ASCs) has important applications in stem cell biology. The ability to integrate a noncytotoxic, fluorescent reporter gene into the genome of ASCs with the capability to track ASCs and their progeny is particularly desirable for transplantation studies. The use of fluorescent proteins has greatly aided the investigations of protein and cell function on short-time scales. In contrast, the obtainment of stably expressing cell strains with low variability in expression for studies on longer-time scales is often problematic. We show that this difficulty is partly due to the cytotoxicity of a commonly used reporter, green fluorescent protein (GFP). To avoid GFP-specific toxicity effects during attempts to stably mark a rat hepatic ASC strain and, therefore, obtain stable, long-term fluorescent ASCs, we evaluated cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP), in addition to GFP. Although we were unable to derive stable GFP-expressing strains, stable fluorescent clones (up to 140 doublings) expressing either CFP or YFP were established. When fluorescently marked ASCs were induced to produce differentiated progeny cells, stable fluorescence expression was maintained. This property is essential for studies that track fluorescently marked ASCs and their differentiated progeny in transplantation studies

CXCR6, a Newly Defined Biomarker of Tissue-Specific Stem Cell Asymmetric Self-Renewal, Identifies More Aggressive Human Melanoma Cancer Stem Cells

Background: A fundamental problem in cancer research is identifying the cell type that is capable of sustaining neoplastic growth and its origin from normal tissue cells. Recent investigations of a variety of tumor types have shown that phenotypically identifiable and isolable subfractions of cells possess the tumor-forming ability. In the present paper, using two lineage-related human melanoma cell lines, primary melanoma line IGR39 and its metastatic derivative line IGR37, two main observations are reported. The first one is the first phenotypic evidence to support the origin of melanoma cancer stem cells (CSCs) from mutated tissue-specific stem cells; and the second one is the identification of a more aggressive subpopulation of CSCs in melanoma that are CXCR6+. Conclusions/Significance: The association of a more aggressive tumor phenotype with asymmetric self-renewal phenotype reveals a previously unrecognized aspect of tumor cell physiology. Namely, the retention of some tissue-specific stem cell attributes, like the ability to asymmetrically self-renew, impacts the natural history of human tumor development. Knowledge of this new aspect of tumor development and progression may provide new targets for cancer prevention and treatment

Assessing the Potential of Hydroponic Farming to Reduce Food Imports: The Case of Lettuce Production in Sweden

Many studies have investigated the issue of feeding the world’s growing population from different perspectives. Conventional agricultural methods usually have small production yields while requiring large amounts of scarce and unevenly distributed resources such as farmland and water. Furthermore, although produced food may meet the demands, it is still inefficiently delivered among different regions. Urban agriculture has been proposed to produce food inside urban areas with higher yields and less resource consumption. Hydroponics is one of the urban farming methods that needs further research before being applied on large scales. This study aims to investigate the potential lettuce production using hydroponic systems to grow lettuce domestically in urban areas in Sweden to lower its lettuce import and motivate local food production to become self-sufficient. The study is performed using a literature review, theoretical design of a system, and scenario-based assessment of hydroponic lettuce production. The detailed analysis is performed via a case study of lettuce production in Uppsala municipality and two scenarios considering the reduction of lettuce import to Sweden and related challenges and opportunities of the designed system. The analyses demonstrate that the system paves the way to decrease lettuce import and dependence on trade. Therefore, applying the system increases self-sufficiency and decreases vulnerability to shocks. Nevertheless, the study does not necessarily address issues related to transportation and food miles. Moreover, finding suitable places to establish the system needs further studies.

Assessing the Potential of Hydroponic Farming to Reduce Food Imports: The Case of Lettuce Production in Sweden

Many studies have investigated the issue of feeding the world’s growing population from different perspectives. Conventional agricultural methods usually have small production yields while requiring large amounts of scarce and unevenly distributed resources such as farmland and water. Furthermore, although produced food may meet the demands, it is still inefficiently delivered among different regions. Urban agriculture has been proposed to produce food inside urban areas with higher yields and less resource consumption. Hydroponics is one of the urban farming methods that needs further research before being applied on large scales. This study aims to investigate the potential lettuce production using hydroponic systems to grow lettuce domestically in urban areas in Sweden to lower its lettuce import and motivate local food production to become self-sufficient. The study is performed using a literature review, theoretical design of a system, and scenario-based assessment of hydroponic lettuce production. The detailed analysis is performed via a case study of lettuce production in Uppsala municipality and two scenarios considering the reduction of lettuce import to Sweden and related challenges and opportunities of the designed system. The analyses demonstrate that the system paves the way to decrease lettuce import and dependence on trade. Therefore, applying the system increases self-sufficiency and decreases vulnerability to shocks. Nevertheless, the study does not necessarily address issues related to transportation and food miles. Moreover, finding suitable places to establish the system needs further studies.

Investigation of a SACK approach for ex vivo expansion of human HSCs

Thesis (Ph. D.)--Massachusetts Institute of Technology, Biological Engineering Division, 2006.Includes bibliographical references.Ex vivo expansion of hematopoietic stem cells (HSCs) is a long-standing challenge faced by both researchers and clinicians. To date, no robust, efficient method for the pure, ex vivo expansion of human HSCs has been demonstrated. Previous methods primarily induced the expansion of committed hematopoietic progenitor cells (HPCs), yielding even less pure populations of HSCs. This research was based on the hypothesis that, like for other adult stem cells (ASCs), the major barrier to expanding HSCs ex vivo is in preferentially regulating the asymmetric self-renewal of HSCs without loss in their ability to produce differentiated committed HPCs. This laboratory has shown that a p53-dependent pathway specifically controls the self-renewal pattern of several types of ASCs and thereby provides an effective means for expansion of ASCs in culture. The method, which involves the use of purine metabolites to achieve suppression of asymmetric cell kinetics, is referred to as SACK. The utility of the p53-dependent pathway was investigated for directing expansion of human HSCs. In order to support this investigation, the proliferation of HPCs in in vitro cultures was repressed by culturing cells without hematopoietic growth factors and cytokines.(cont.) This allowed the in vitro detection of SACK-effects on a small sub-population of cells, predicted to include HSCs. In order to determine the self-renewal capacity and multilineage potential of SACK- cultured cells, they were transplanted into non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice. In vivo transplantation investigations exhibited 1.9- fold to 4.5-fold increased engraftment efficiency with SACK-agents compared to SACK-free controls, suitable for clinical applications. This result suggests that SACK-culture expands a population of SCID-repopulating cells (SRCs) that yields self-renewal and multilineage engraftment in NOD/SCID mice. Accordingly, increased engraftment efficiency for successful clinical applications may be achieved after additional optimization of HSC expansion. To obtain the full therapeutic potential of expanded HSCs, development of methods for independently marking putative ASCs for future analyses and gene therapy was explored. This early success with human HSCs supports the basic hypothesis that the SACK approach may be applicable to expansion of many types of ASCs.by Rouzbeh R. Taghizadeh.Ph.D

Phylogenetic Comparison of Influenza Virus Isolates from Three Medical Centers in Tehran with the Vaccine Strains during 2008-2009

Background: Influenza virus is a major infectious pathogen of the respiratory system causing a high degree of morbidity and mortality annually. The worldwide vaccines are decided and produced annually by World Health Organization and licensed companies based on the samples collected from all over the world. The aim of this study was to determine phylogenecity and heterogenecity of the circulating influenza isolates during 2008-2009 outbreaks in Tehran, compare them with the vaccine strains that were recommended by WHO for the same period. Methods: Nasopharyngeal swabs (n=142) were collected from patients with influenza and influenza-like illness. Typing and subtyping of the isolates were performed using multiplex RT-PCR and phylogenetic analysis was carried out for hemagglutinin genes of the isolates. Results: Fifty out of 142 samples were positive for influenza A virus, and no influenza B virus was detected. Phylogenetic analyses revealed that the A/H1N1 isolates were related closely to A/Brisbane/59/2007, and the A/H3N2 isolates were close to A/Brisbane/10/2007 vaccine strains. Conclusion: The findings of the present study demonstrate that the A/H1N1 was the predominant subtype of human influenza virus among the patients studied in Tehran during 2008-2009 winter seasons. In addition, some amino acid variation was found in Tehran/2008/H1N1 isolates from the 2008-2009 vaccine strain, but the H3N2 isolates showed higher genetic resemblance to the vaccine strai

Primers and thermal protocols utilized for the RT-PCR evaluation of CTA and differentiation antigens expression in human melanoma cells.

<p>Primers and thermal protocols utilized for the RT-PCR evaluation of CTA and differentiation antigens expression in human melanoma cells.</p

Evaluation of the effect of the CXCR6 ligand CXCL16 on melanoma cell line growth.

<p>Twenty thousand cells were seeded overnight in individual wells of 12 multi-well plates in complete medium growth condition. Recombinant sCXCL16 (100 ng/ml) was added the next day and replaced every 48 hours. On the 4th and 7th day of culture, the cells were trypsinized, and viable cell numbers determined. The bar graph represents the mean ± S.D. of three independent experiments each carried out in triplicate. <b>*</b>, p<0.01 <i>vs</i>. untreated cells (black bars); <b>**</b>, p<0.001 <i>vs</i>. untreated cells (black bars).</p

Tumor xenografts derived from and CXCR6- IGR37 cells.

<p>5×10⁵ CXCR6- sorted cells were injected subcutaneously in five-week-old NOD-SCID mice (3 mice for each experimental condition). CXCR6- cells did not yield tumors.</p