Synthesis and characterization of three covalently linked porphyrin-phthalocyanine pentamers with nucleophilic substitution

In this paper the synthetic methods of three covalently linked porphyrin-phthalocyanine heteropentamers, containing four units of porphyrin linked to a central phthalocyanine (Mpor-LPc; M = 2H, Fe, L = Zn, Fe), are described. The synthetic strategy is on the basis of nucleophilic substitution reactions between [1,8,15,22-tetra nitro phthalocyanines] and [5-(4-hydroxy phenyl)-10,15,20-triphenyl porphyrins] as the phenolic alcohols. Porphyrins are linked with oxygen as spacer through meso position of phenyl group to phthalocyanines. These macromolecules were characterized by ¹H NMR, UV-Vis, IR, fluorescence and mass spectroscopy. The electronic absorption spectrums of the hetero-dyad systems changed significantly upon coupling and showed a great red shift in the phthalocyanine Q-bands. These changes confirm the electron-donating effects of the porphyrin units and the extension of conjugated п-systems. The emission spectra of the products supports intramolecular energy and charge transfer between the sub-units.     

Alternative metabolic fates of thymine nucleotides in human cells.

Three types of experiments have been used to study the metabolism of thymine nucleotides by human cells. (1) Cells were labelled continuously with [3H]thymidine and the incorporation of label into DNA compared with the specific radioactivities of pools of individual thymine nucleotides separated by chromatography on polyethylene-imine-cellulose. (2) Cellular thymine nucleotides were labelled with [3H]thymidine at 13 degrees C, followed by incubation at 37 degrees C in unlabelled medium. Incorporation of label into DNA and loss of label from the nucleotide pools were monitored during the 'chase' period at 37 degrees C. (3) The experiments described in (2) above were repeated in the presence of the DNA-synthesis inhibitor cytosine arabinoside, in order to demonstrate more clearly and to quantify degradative pathways for thymine nucleotides. In phytohaemagglutinin-stimulated lymphocytes and in bone-marrow cells, only a proportion (25-60%) of labelled thymine nucleotide was incorporated into DNA, the rest being rapidly degraded and lost from the cell. In contrast, an established cell line (HPB-ALL) from a patient with acute lymphoblastic leukaemia of thymic origin incorporated 100% of its exogenously labelled thymine nucleotides into DNA. These results indicated that alternative metabolic routes are open to thymine nucleotides in human cells. In lymphocytes from patients with megaloblastic anaemia and in normal lymphocytes treated with methotrexate, the utilization of labelled thymine nucleotides for DNA synthesis was more efficient than in controls. These results offer an explanation for the observation of a normal pool of thymidine triphosphate in the cells of patients with untreated megaloblastic anaemia even though the amount of this compound available for DNA synthesis appears to be decreased.     

The Interplay between Chondrocyte Redifferentiation Pellet Size and Oxygen Concentration

Chondrocytes dedifferentiate during ex vivo expansion on 2-dimensional surfaces. Aggregation of the expanded cells into 3-dimensional pellets, in the presence of induction factors, facilitates their redifferentiation and restoration of the chondrogenic phenotype. Typically 1×10⁵–5×10⁵ chondrocytes are aggregated, resulting in “macro” pellets having diameters ranging from 1–2 mm. These macropellets are commonly used to study redifferentiation, and recently macropellets of autologous chondrocytes have been implanted directly into articular cartilage defects to facilitate their repair. However, diffusion of metabolites over the 1–2 mm pellet length-scales is inefficient, resulting in radial tissue heterogeneity. Herein we demonstrate that the aggregation of 2×10⁵ human chondrocytes into micropellets of 166 cells each, rather than into larger single macropellets, enhances chondrogenic redifferentiation. In this study, we describe the development of a cost effective fabrication strategy to manufacture a microwell surface for the large-scale production of micropellets. The thousands of micropellets were manufactured using the microwell platform, which is an array of 360×360 µm microwells cast into polydimethylsiloxane (PDMS), that has been surface modified with an electrostatic multilayer of hyaluronic acid and chitosan to enhance micropellet formation. Such surface modification was essential to prevent chondrocyte spreading on the PDMS. Sulfated glycosaminoglycan (sGAG) production and collagen II gene expression in chondrocyte micropellets increased significantly relative to macropellet controls, and redifferentiation was enhanced in both macro and micropellets with the provision of a hypoxic atmosphere (2% O₂). Once micropellet formation had been optimized, we demonstrated that micropellets could be assembled into larger cartilage tissues. Our results indicate that micropellet amalgamation efficiency is inversely related to the time cultured as discreet microtissues. In summary, we describe a micropellet production platform that represents an efficient tool for studying chondrocyte redifferentiation and demonstrate that the micropellets could be assembled into larger tissues, potentially useful in cartilage defect repair.     

Reviewing the role of cardiac exosomes in myocardial repair at a glance

Exosome-based therapy is an emerging novel approach for myocardial infarction (MI) treatment. Exosomes are identified as extracellular vesicles that are produced within multivesicular bodies in the cells' cytosols and then are secreted from the cells. Exosomes are 30–100 nm in diameter that are released from viable cells and are different from other secreted vesicles such as apoptotic bodies and microvesicles in their origin and contents such as RNAs, proteins, and nucleic acid. The recent advances in exosome research have demonstrated the role of these bionanovesicles in the physiological, pathological, and molecular aspects of the heart. The results of in vitro and preclinical models have shown that exosomes from different cardiac cells can improve cardiac function following MI. For example, mesenchymal stem cells (MSCs) and cardiac progenitor cells (CPCs) containing exosomes can affect the proliferation, survival, and differentiation of cardiac fibroblasts and cardiomyocytes. Moreover, MSCs- and CPCs-derived exosomes can enhance the migration of endothelial cells. Exosome-based therapy approaches augment the cardiac function by multiple means, such as reducing fibrosis, stimulation of vascular angiogenesis, and proliferation of cardiomyocytes that result in replacing damaged heart tissue with newly generated functional myocytes. This review article aims to briefly discuss the recent advancements in the role of secreted exosomes in myocardial repair by focusing on cardiac cells-derived exosomes.     

Comparing the frequency of CD33+ pSTAT3+ myeloid-derived suppressor cells and IL-17+ lymphocytes in patients with prostate cancer and benign prostatic hyperplasia

Prostate cancer (PCa) is one of the most epidemic types of cancer in men. The tumor microenvironment (TME) of PCa is involved in the emergence of immunosuppressive factors such as myeloid-derived suppressor cells (MDSC), which regulate the immune system by several mechanisms, including interleukin (IL)-10 production. On the other hand, IL-17⁺ helper T cells (Th17) induce MDSCs and chronic inflammation in TME by producing IL-17. This study demonstrated that the frequency of CD33⁺ pSTAT3⁺ MDSC and IL-17⁺ lymphocyte as well as IL-10 messenger RNA (mRNA) expression were significantly higher in the PCa patients than in the benign prostatic hyperplasia (BPH) group. Moreover, there was no significant relationship between the frequency of CD33⁺ pSTAT3⁺ MDSC, and IL-17⁺ lymphocyte with Gleason scores in the PCa group. We suggested that the higher frequency of CD33⁺ pSTAT3⁺ MDSC and IL-17⁺ lymphocyte and the more frequent expression of IL-10 mRNA in PCa patients may play roles in tumor progression from BPH to PCa.     

Genotyping of –374A/T, –429A/G,And 63 bp Ins/Del Polymorphisms of Rage By Rapid One-Step Hexaprimer Amplification Refractory Mutation System Polymerase Chain Reaction in Breast Cancer Patients

Several studies have focused on the RAGE genetic background and have demonstrated that its polymorphisms affect the receptor's activity, expression, and downstream signaling. However, there is only little information regarding RAGE polymorphism in breast cancer. In the present study, the authors studied RAGE polymorphisms in 71 patients with breast cancer and 93 healthy women. RAGE –374T/A, –429T/C, and 63 bp Ins/del polymorphisms were analyzed using a hexaprimer amplification refractory mutation system PCR (H-ARMS-PCR). The results showed that RAGE polymorphisms are not associated with breast cancer in the current study population. Larger studies are required to confirm these data in other populations.     

Suitability indices and habitat suitability index model of Caspian kutum (Rutilus frisii kutum) in the southern Caspian Sea

The relationship between species and habitat is important in ecosystem-based fisheries management. Habitat suitability index (HSI) modeling is a valuable tool in ecology and can be used to describe the relationship between fish abundance and ecological variables in order to estimate the suitability of specific habitats. In the present study, an HSI model was applied to determine suitable habitats for the Caspian kutum (Rutilus frisii kutum), an important commercial species in the southern Caspian Sea. An arithmetic mean model (AMM) was found to be the most appropriate model for describing the relationship between two of the environmental variables investigated (depth and benthos biomass). However, a geometric mean model explained the evident relationship when all four environmental variables were used (depth, benthos biomass, photosynthetically active radiation and sea surface temperature). The areas with an HSI > 0.5 had over 85 % of the total catch indicating the reliability of the prediction of the Caspian kutum habitat using the AMM. The present study showed that depth and substrate structure are the most important environmental variables for the Caspian kutum to select its habitats, and between remotely sensed data, chlorophyll a, photosynthetically active radiation and sea surface temperature are the most critical parameters for near real-time prediction of the Caspian kutum habitat.     

Esophageal cancer alters the expression of nuclear pore complex binding protein Hsc70 and eIF5A-1

Nuclear pore complex (NPC) is the only corridor for macromolecules exchange between nucleus and cytoplasm. NPC and its components, nucleoporins, play important role in the diverse physiological processes including macromolecule exchange, chromosome segregation, apoptosis and gene expression. Recent reports also suggest involvement of nucleoporins in carcinogenesis. Applying proteomics, we analyzed expression pattern of the NPC components in a newly established esophageal cancer cell line from Persia (Iran), the high-risk region for esophageal cancer. Our results indicate overexpression of Hsc70 and downregulation of subunit alpha type-3 of proteasome, calpain small subunit 1, and eIF5A-1. Among these proteins, Hsc70 and eIF5A-1 are in direct interaction with NPC and involved in the nucleocytoplasmic exchange. Hsc70 plays a critical role as a chaperone in the formation of a cargo–receptor complex in nucleocytoplasmic transport. On the other hand, it is an NPC-associated protein that binds to nucleoporins and contributes in recycling of the nucleocytoplasmic transport receptors in mammals and affects transport of proteins between nucleus and cytoplasm. The other nuclear pore interacting protein: eIF5A-1 binds to the several nucleoporins and participates in nucleocytoplasmic transport. Altered expression of Hsc70 and eIF5A-1 may cause defects in nucleocytoplasmic transport and play a role in esophageal carcinogenesis.     

Development of Novel Prime-Boost Strategies Based on a Tri-Gene Fusion Recombinant L. tarentolae Vaccine against Experimental Murine Visceral Leishmaniasis

Visceral leishmaniasis (VL) is a vector-borne disease affecting humans and domestic animals that constitutes a serious public health problem in many countries. Although many antigens have been examined so far as protein- or DNA-based vaccines, none of them conferred complete long-term protection. The use of the lizard non-pathogenic to humans Leishmania (L.) tarentolae species as a live vaccine vector to deliver specific Leishmania antigens is a recent approach that needs to be explored further. In this study, we evaluated the effectiveness of live vaccination in protecting BALB/c mice against L. infantum infection using prime-boost regimens, namely Live/Live and DNA/Live. As a live vaccine, we used recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinases (CPA and CPB without its unusual C-terminal extension (CPB^-CTE)) as a tri-fusion gene. For DNA priming, the tri-fusion gene was encoded in pcDNA formulated with cationic solid lipid nanoparticles (cSLN) acting as an adjuvant. At different time points post-challenge, parasite burden and histopathological changes as well as humoral and cellular immune responses were assessed. Our results showed that immunization with both prime-boost A2-CPA-CPB^-CTE-recombinant L. tarentolae protects BALB/c mice against L. infantum challenge. This protective immunity is associated with a Th1-type immune response due to high levels of IFN-γ production prior and after challenge and with lower levels of IL-10 production after challenge, leading to a significantly higher IFN-γ/IL-10 ratio compared to the control groups. Moreover, this immunization elicited high IgG1 and IgG2a humoral immune responses. Protection in mice was also correlated with a high nitric oxide production and low parasite burden. Altogether, these results indicate the promise of the A2-CPA-CPB^-CTE-recombinant L. tarentolae as a safe live vaccine candidate against VL.     

An Investigation into the Strength of the Association and Agreement Levels between Subjective and Objective Sleep Duration in Adolescents

Study Objectives

The majority of adolescent sleep research has utilized self-reported sleep duration and some have based information on a solitary question. Whilst some have claimed to have validated sleep survey data with objective actigraphy measures in adolescents, the statistical approach applied only demonstrates the strength of the association between subjective and objective sleep duration data and does not reflect if these different methods actually agree.

Methods

Data were collected as part of the Midlands Adolescents Schools Sleep Education Study (MASSES). Adolescents (n=225) aged 11-13 years provided estimates for weekday, weekend and combined sleep duration based on self-reported survey data, a 7-day sleep diary, and wrist-worn actigraphy.

Results

We assessed the strength of the relationship as well as agreement levels between subjective and objectively determined sleep duration (weekday, weekend and combined). Subjective diary sleep duration was significantly correlated with actigraphy estimates for weekday and weekend sleep duration r=0.30, p≤0.001 and r=0.31, p≤0.001 respectively. Pitman’s test demonstrated no significant difference in the variance between weekend sleep duration (r=0.09, p=0.16) and combined sleep duration (r=0.12, p=0.08) indicating acceptable agreement between actigraphy and sleep diary sleep duration only. Self-reported sleep duration estimates (weekday, weekend and combined) did not agree with actigraphy determined sleep duration.

Conclusions

Sleep diaries are a cost-effective alternative to survey/questionnaire data. Self-reported measures of sleep duration in adolescents do not agree with actigraphy measures and should be avoided where possible. Previous adolescent sleep studies that have utilized self-reported survey data may not provide a complete representation of sleep on the outcome measure of interest.