Molecular factors regulating E-cadherin expression in urothelial bladder cancer and their correlations with the clinicopathological features

This study aimed to assess the expression of S100A4, Twist and E-cadherin (mRNA and protein) in urothelial bladder cancer, investigate the correlation between them and evaluate their association with the clinicopathological features of the disease. The study included 54 patients diagnosed as urothelial bladder cancer of different stages and grades. The expression levels of S100A4, Twist and E-cadherin (mRNA and protein) in tissue samples were determined by quantitative RT-PCR and immunohistochemistry. The expression of S100A4 and Twist was significantly upregulated while E- cadherin was significantly downregulated in urothelial bladder cancer tissues compared to the adjacent surrounding normal bladder tissues at both mRNA and protein levels (p?<?0.001). Expression levels of S100A4 and Twist were significantly higher in recurrent tumor than in non-recurrent tumors (p?<?0.001) while the expression level of E-cadherin was significantly lower in recurrent tumors than in non-recurrent tumors at both mRNA and protein levels (p?<?0.001). There was a significant positive correlation between S100A4 and Twist expressions (r?=?0.875, p?<?0.001) while significant negative correlations were found between E- cadherin and S100A4 expressions(r=- 0.803, p?<?0.001) and between E-cadherin and Twist (r?=??0.809, p?<?0.001). Up-regulation of S100A4 and Twist and down-regulation of E-cadherin in urothelial bladder cancer tissues compared to adjacent normal tissues were observed. There was a significant negative correlation between S100A4 and E- cadherin and between E- cadherin and Twist expression. However, there was a significant positive correlation between S100A4 and Twist expressions. Furthermore, the alterations in the gene expression were associated with disease stage and grade.     

Evaluation of 64Cu-labeled bifunctional chelate-bombesin conjugates

Several bifunctional chelates (BFCs) were investigated as carriers of (64)Cu for PET imaging. The most widely used chelator for (64)Cu labeling of BFCs is DOTA (1,4,7,10-tetraazacyclododecane-N,N',N″,N'-tretraacetic acid), even though this complex exhibits only moderate in vivo stability. In this study, we prepared a series of alternative chelator-peptide conjugates labeled with (64)Cu, measured in vitro receptor binding affinities in human breast cancer T47D cells expressing the gastrin-releasing peptide receptor (GRPR) and compared their in vivo stability in mice. DOTA-, NOTA-(1,4,7-triazacyclononane-1,4,7-triacetic acid), PCTA-(3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid), and Oxo-DO3A-(1-oxa-4,7,10-triazacyclododecane-4,7,10-triacetic acid) peptide conjugates were prepared using H(2)N-Aoc-[d-Tyr(6),βAla(11),Thi(13),Nle(14)]bombesin(6-14) (BBN) as a peptide template. The BBN moiety was selected since it binds with high affinity to the GRPR, which is overexpressed on human breast cancer cells. A convenient synthetic approach for the attachment of aniline-BFC to peptides on solid support is also presented. To facilitate the attachment of the aniline-PCTA and aniline-Oxo-DO3A to the peptide via an amide bond, a succinyl spacer was introduced at the N-terminus of BBN. The partially protected aniline-BFC (p-H(2)N-Bn-PCTA(Ot-Bu)(3) or p-H(2)N-Bn-DO3A(Ot-Bu)(3)) was then coupled to the resulting N-terminal carboxylic acid preactivated with DEPBT/ClHOBt on resin. After cleavage and purification, the peptide-conjugates were labeled with (64)Cu using [(64)Cu]Cu(OAc)(2) in 0.1 M ammonium acetate buffer at 100 °C for 15 min. Labeling efficacy was >90% for all peptides; Oxo-DO3A-BBN was incubated an additional 150 min at 100 °C to achieve this high yield. Specific activities varied from 76 to 101 TBq/mmol. Competition assays on T47D cells showed that all BFC-BBN complexes retained high affinity for the GRPR. All BFC-BBN (64)Cu-conjugates were stable for over 20 h when incubated at 37 °C in mouse plasma samples. However, in vivo, only 37% of the (64)Cu/Oxo-DO3A complex remained intact after 20 h while the (64)Cu/DOTA-BBN complex was completely demetalated. In contrast, both (64)Cu/NOTA- and (64)Cu/PCTA-BBN conjugates remained stable during the 20 h time period. Our results indicate that it is possible to successfully conjugate aniline-BFC with peptide on solid support. Our data also show that (64)Cu-labeled NOTA- and PCTA-BBN peptide conjugates are promising radiotracers for PET imaging of many human cancers overexpressing the GRP receptor.     

The effects of extracellular matrix (ECM) proteins on the attachment of Pneumocystis carinii to lung cell lines in vitro.

Pneumocystis carinii cells labeled with fluorescein isothiocyanate were co-cultured with tissue culture cells. Measurements of attachment was determined by the tissue culture cell fluorescence after washing out the P. carinii organisms. The effects of the extracellular matrix proteins, laminin and fibronectin, on the binding of P. carinii onto the monolayer of cultured cells were investigated for better understanding of organism-cell interactions. The internalization of P. carinii by MRC5 cells was observed.     

Fleshing Out Vulnerability

In the literature on medical ethics, it is generally admitted that vulnerable persons or groups deserve special attention, care or protection. One can define vulnerable persons as those having a greater likelihood of being wronged – that is, of being denied adequate satisfaction of certain legitimate claims. The conjunction of these two points entails what we call the Special Protection Thesis. It asserts that persons with a greater likelihood of being denied adequate satisfaction of their legitimate claims deserve special attention, care or protection. Such a thesis remains vague, however, as long as we do not know what legitimate claims are. This article aims at dispelling this vagueness by exploring what claims we have in relation to health care – thus fleshing out a claim‐based conception of vulnerability. We argue that the Special Protection Thesis must be enriched as follows: If individual or group X has a greater likelihood of being denied adequate satisfaction of some of their legitimate claims to (i) physical integrity, (ii) autonomy, (iii) freedom, (iv) social provision, (v) impartial quality of government, (vi) social bases of self‐respect or (vii) communal belonging, then X deserves special attention, care or protection. With this improved understanding of vulnerability, vulnerability talk in healthcare ethics can escape vagueness and serve as an adequate basis for practice.     

Stem cells: cross-talk and developmental programs

The thesis advanced in this essay is that stem cells-particularly those in the nervous system-are components in a series of inborn 'programs' that not only ensure normal development, but persist throughout life so as to maintain homeostasis in the face of perturbations-both small and great. These programs encode what has come to be called 'plasticity'. The stem cell is one of the repositories of this plasticity. This review examines the evidence that interaction between the neural stem cell (as a prototypical somatic stem cell) and the developing or injured brain is a dynamic, complex, ongoing reciprocal set of interactions where both entities are constantly in flux. We suggest that this interaction can be viewed almost from a 'systems biology' vantage point. We further advance the notion that clones of exogenous stem cells in transplantation paradigms may not only be viewed for their therapeutic potential, but also as biological tools for 'interrogating' the normal or abnormal central nervous system environment, indicating what salient cues (among the many present) are actually guiding the expression of these 'programs'; in other words, using the stem cell as a 'reporter cell'. Based on this type of analysis, we suggest some of the relevant molecular pathways responsible for this 'cross-talk' which, in turn, lead to proliferation, migration, cell genesis, trophic support, protection, guidance, detoxification, rescue, etc. This type of developmental insight, we propose, is required for the development of therapeutic strategies for neurodegenerative disease and other nervous system afflictions in humans. Understanding the relevant molecular pathways of stem cell repair phenotype should be a priority, in our view, for the entire stem cell field.     

Effect of Controlled Crossing on the Triglyceride and Fatty Acid Composition of Virgin Olive Oils

Our study reports the triglyceride (triacylglycerol, TAG) composition of five new Tunisian virgin olive oil cultivars obtained through controlled crossings of the cultivar (cv.) Chemlali Sfax. These cultivars have been selected among a progeny of 500 olive descendants, based on an evaluation of the fatty‐acid (FA) composition of their oils. Among these samples, two were derived from the crossing with the cv. Sigoise as pollinator (SM634) or pollen acceptor (SM1110) and the others from the crossing with the cv. Meski as pollen acceptor (SM513, SM514, and SM517). The five descendants were characterized by a good fat value, a balanced FA composition, and a high content of triolein, varying between 26.9 (SM514) and 45.46% (SM1110). They had an improved FA composition as compared to that of the cv. Chemlali Sfax and their fruits were slightly bigger. The principal component analysis suggested that the TAG variables were more suitable than the total FAs for an optimum classification of the cultivar samples analyzed. The cultivars obtained through the crossing with the cv. Sigoise (as pollen acceptor or pollinator) had a more favorable composition of FAs and TAG than those obtained through the crossing with the cv. Meski, which indicated that genetic factors had the most important influence on the quality of the virgin olive oils.     

Cryopreservation of sour orange (Citrus aurantium L.) shoot tips

Summary The objective of this study was to establish a cryopreservation protocol for sour orange (Citrus aurantium L.). Cryopreservation was carried out via encapsulation-dehydration, vitrification, and encapsulation-vitrification on shoot tips excised from in vitro cultures. Results indicated that a maximum of 83% survival and 47% regrowth of encapsulated-dehydrated and cryopreserved shoot tips was obtained with 0.5M sucrose in the preculture medium and further dehydration for 6 h to attain 18% moisture content. Dehydration of encapsulated shoot tips with silica gel for 2h resulted in 93% survival but only 37% regrowth of cryopreserved shoot tips. After preculturing with 0.5M sucrose, 80% of the vitrified cryopreserved shoots survived when 2M sucrose plus 10% dimethyl sulfoxide (DMSO) was used as a cryoprotectant for 20 min at 25°C. Survival and regrowth of vitrified cryopreserved shoot tips were 67% and 43%, respectively, when 0.4M sucrose plus 2M glycerol was used as a loading solution followed by application of 100% plant vitrification solution (PVS2) for 20 min. Increased duration of exposure to the loading solution up to 60 min increased survival (83%) and regrowth (47%) of cryopreserved shoot tips. With encapsulation-vitrification, dehydration with 100% PVS2 for 2 or 3 h at 0°C resulted in 50 or 57% survival and 30 or 40% regrowth, respectively, of cryopreserved shoot tips.     

Phytochemical Compounds from the Crop Byproducts of Tunisian Globe Artichoke Cultivars

The phytochemical composition in two Tunisian globe artichoke cultivars (bracts, leaves, and floral stems) was evaluated in the plant byproducts. The results indicated that the bracts contain the highest levels of total phenols, o‐diphenols, and flavonoids, whereas tannins seem to be more abundant in the leaves. Bracts from the ‘Violet d'Hyères’ cultivar possessed more total phenols (160.8 mg/g DW), flavonoids (64.9 mg/g DW), and anthocyanins (15.3 μg/g DW) than the ‘Blanc d'Oran’ bracts (134.5 mg/g DW, 51.2 mg/g DW, and 8.3 μg/g DW, resp.). Sixty‐four volatile compounds were identified in the headspace of globe artichoke material, particularly in the bracts. The volatile profile showed that sesquiterpene hydrocarbons and non‐terpene derivatives were the main volatiles emitted by the bracts in both cultivars. These results suggest that globe artichoke byproducts might represent a potential source of natural compounds, which could be used as nutraceuticals or as ingredients in the design of functional foods.     

Expansion strategies for human mesenchymal stromal cells culture under xeno‐free conditions

Choosing the culture system and culture medium used to produce cells are key steps toward a safe, scalable, and cost‐effective expansion bioprocess for cell therapy purposes. The use of AB human serum (AB HS) as an alternative xeno‐free supplement for mesenchymal stromal cells (MSC) cultivation has increasingly gained relevance due to safety and efficiency aspects. Here we have evaluated different scalable culture systems to produce a meaningful number of umbilical cord matrix‐derived MSC (UCM MSC) using AB HS for culture medium supplementation during expansion and cryopreservation to enable a xeno‐free bioprocess. UCM MSC were cultured in a scalable planar (compact 10‐layer flasks and roller bottles) and 3‐D microcarrier‐based culture systems (spinner flasks and stirred tank bioreactor). Ten layer flasks and roller bottles enabled the production of 2.6 ± 0.6 × 10⁴ and 1.4 ± 0.3 × 10⁴ cells/cm². UCM MSC‐based microcarrier expansion in the stirred conditions has enabled the production of higher cell densities (5.5–23.0 × 10⁴ cells/cm²) when compared to planar systems. Nevertheless, due to the moderate harvesting efficiency attained, (80% for spinner flasks and 46.6% for bioreactor) the total cell number recovered was lower than expected. Cells maintained the functional properties after expansion in all the culture systems evaluated. The cryopreservation of cells (using AB HS) was also successfully carried out. Establishing scalable xeno‐free expansion processes represents an important step toward a GMP compliant large‐scale production platform for MSC‐based clinical applications. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1358–1367, 2017     

Complexes of vitamin B6, part VII: Ternary complexes of cobalt(II), nickel(II), and copper(II) involving pyridoxamine and some amino acids

The stability constants of the ternary Cu(II), Ni(II), and Co(II) complexes containing pyridoxamine (PM) and as a second ligand (L) glycine, DL-alanine, DL-valine, and β-phenylalnine were determined by pH-metric titration in 0.50 M KNO₃ at 30°C. The corresponding constants of the equilibrium, log X, are greater than would be expected for purely statistical reasons (log X = 0.6), except for few complex cases of Co(II). It has been also concluded that amino acids compete more than pyridoxamine for Ni(II) and Co(II) through the formation of 1:2:1:0 species rather than 2:1:1:0 of PM⁻:L⁻:M²⁺:H⁺.