Hybrid chitosan–ß‐glycerol phosphate–gelatin nano‐/micro fibrous scaffolds with suitable mechanical and biological properties for tissue engineering

Scaffold‐based tissue engineering is considered as a promising approach in the regenerative medicine. Graft instability of collagen, by causing poor mechanical properties and rapid degradation, and their hard handling remains major challenges to be addressed. In this research, a composite structured nano‐/microfibrous scaffold, made from a mixture of chitosan–ß‐glycerol phosphate–gelatin (chitosan–GP–gelatin) using a standard electrospinning set‐up was developed. Gelatin–acid acetic and chitosan ß‐glycerol phosphate–HCL solutions were prepared at ratios of 30/70, 50/50, 70/30 (w/w) and their mechanical and biological properties were engineered. Furthermore, the pore structure of the fabricated nanofibrous scaffolds was investigated and predicted using a theoretical model. Higher gelatin concentrations in the polymer blend resulted in significant increase in mean pore size and its distribution. Interaction between the scaffold and the contained cells was also monitored and compared in the test and control groups. Scaffolds with higher chitosan concentrations showed higher rate of cell attachment with better proliferation property, compared with gelatin‐only scaffolds. The fabricated scaffolds, unlike many other natural polymers, also exhibit non‐toxic and biodegradable properties in the grafted tissues. In conclusion, the data clearly showed that the fabricated biomaterial is a biologically compatible scaffold with potential to serve as a proper platform for retaining the cultured cells for further application in cell‐based tissue engineering, especially in wound healing practices. These results suggested the potential of using mesoporous composite chitosan–GP–gelatin fibrous scaffolds for engineering three‐dimensional tissues with different inherent cell characteristics. © 2015 Wiley Periodicals, Inc. Biopolymers 105: 163–175, 2016.     

All-Optical Tunable Plasmonic Biosensor Made of Graphene and Metamaterial

Plasmonics - We demonstrate a novel, label-free and real-time tunable infrared biosensor by employing surface-plasmon polaritons in asymmetric Mach–Zehnder interferometer. The waveguides...     

Adipose tissue‐derived mesenchymal stem cells and keratinocytes co‐culture on gelatin/chitosan/β‐glycerol phosphate nanoscaffold in skin regeneration

Using cell‐based engineered skin is an emerging strategy for treating difficult‐to‐heal wounds. To date, much endeavor has been devoted to the fabrication of appropriate scaffolds with suitable biomechanical properties to support cell viability and growth in the microenvironment of a wound. The aim of this research was to assess the impact of adipose tissue‐derived mesenchymal stem cells (AD‐MSCs) and keratinocytes on gelatin/chitosan/β‐glycerol phosphate (GCGP) nanoscaffold in full‐thickness excisional skin wound healing of rats. For this purpose, AD‐MSCs and keratinocytes were isolated from rats and GCGP nanoscaffolds were electrospun. Through an in vivo study, the percentage of wound closure was assessed on days 7, 14, and 21 after wound induction. Samples were taken from the wound sites in order to evaluate the density of collagen fibers and vessels at 7 and 14 days. Moreover, sampling was done on days 7 and 14 from wound sites to assess the density of collagen fibers and vessels. The wound closure rate was significantly increased in the keratinocytes‐AD‐MSCs‐scaffold (KMS) group compared with other groups. The expressions of vascular endothelial growth factor, collagen type 1, and CD34 were also significantly higher in the KMS group compared with the other groups. These results suggest that the combination of AD‐MSCs and keratinocytes seeded onto GCGP nanoscaffold provides a promising treatment for wound healing.     

ERK1/2 inactivation and p38 MAPK-dependent caspase activation during guanosine 5'-triphosphate-mediated terminal erythroid differentiation of K562 cells

Since differentiation therapy is one of the promising strategies for treatment of leukemia, universal efforts have been focused on finding new differentiating agents. In that respect, it was recently shown that guanosine 5'-triphosphate (GTP) induced the differentiation of K562 cells, suggesting its possible efficiency in treatment of chronic myelogenous leukemia (CML). However, further investigations are required to verify this possibility. Here, the effects of GTP on activation of mitogen-activated protein kinases (MAPKs) and caspases in K562 cells were examined. Exposure of K562 cells to 100muM GTP markedly inhibited growth (4-70%) and increased percent glycophorin A positive cells after 1-6 days. GTP-induced terminal erythroid differentiation of K562 cells was accompanied with activation of three key caspases, i.e., caspase-3, -6 and -9. More detailed studies revealed that mitochondrial pathway is activated along with down-regulation of Bcl-xL and releasing of cytochrome c into cytosol. Among MAPKs, ERK1/2and p38 were modulated after GTP treatment. Western blot analyses showed that sustained phosphorylation of p38 MAPK was accompanied by a decrease in ERK1/2 activation. These modulatory effects of GTP were observed at early exposure times before the onset of differentiation (3h), and followed for 24-96h. Interestingly, inhibition of p38 MAPK pathway by SB202190 impeded GTP-mediated caspases activation and differentiation in K562 cells, suggesting that p38 MAPK may act upstream of caspases in our system. These results point to a pivotal role for p38 MAPK pathway during GTP-mediated erythroid differentiation of K562 cells and will hopefully have important impact on pharmaceutical evaluation of GTP for CML treatment in differentiation therapy approaches.     

A New Role for TIMP-1 in Modulating Neurite Outgrowth and Morphology of Cortical Neurons

Background

Tissue inhibitor of metalloproteinases-1 (TIMP-1) displays pleiotropic activities, both dependent and independent of its inhibitory activity on matrix metalloproteinases (MMPs). In the central nervous system (CNS), TIMP-1 is strongly upregulated in reactive astrocytes and cortical neurons following excitotoxic/inflammatory stimuli, but no information exists on its effects on growth and morphology of cortical neurons.

Principal Findings

We found that 24 h incubation with recombinant TIMP-1 induced a 35% reduction in neurite length and significantly increased growth cones size and the number of F-actin rich microprocesses. TIMP-1 mediated reduction in neurite length affected both dendrites and axons after 48 h treatment. The effects on neurite length and morphology were not elicited by a mutated form of TIMP-1 inactive against MMP-1, -2 and -3, and still inhibitory for MMP-9, but were mimicked by a broad spectrum MMP inhibitor. MMP-9 was poorly expressed in developing cortical neurons, unlike MMP-2 which was present in growth cones and whose selective inhibition caused neurite length reductions similar to those induced by TIMP-1. Moreover, TIMP-1 mediated changes in cytoskeleton reorganisation were not accompanied by modifications in the expression levels of actin, βIII-tubulin, or microtubule assembly regulatory protein MAP2c. Transfection-mediated overexpression of TIMP-1 dramatically reduced neuritic arbour extension in the absence of detectable levels of released extracellular TIMP-1.

Conclusions

Altogether, TIMP-1 emerges as a modulator of neuronal outgrowth and morphology in a paracrine and autrocrine manner through the inhibition, at least in part, of MMP-2 and not MMP-9. These findings may help us understand the role of the MMP/TIMP system in post-lesion pre-scarring conditions.     

Seasonal dynamics of zooplankton community in four Mediterranean reservoirs in humid area (Beni Mtir: north of Tunisia) and semi arid area (Lakhmes,Nabhana and Sidi Saâd: center of Tunisia)

The zooplankton community was studied in four Mediterranean reservoirs to assess the relative importance of environmental factors as determinants of zooplankton dynamic in the four seasons. The water temperature and hydrology variations affect the distribution of zooplankton. A positive correlation was established between the total zooplankton and the water temperature (r=0.9, n=9, p<0.05). Fourteen zooplankton species were identified. Seasonal changes in the density (ANOVA, F=3.7, d.f=36, p<0.01) and the biomass of total zooplankton (ANOVA, F=4.4, d.f=36, p<0.001) were observed. Our results suggest that planktivorous fish may not modify the zooplankton dynamics in Beni Mtir reservoir (oligotrophic). On the contrary, in Sidi Saâd reservoir (mesotrophic), fish predation has major effects on seasonal zooplankton dynamics.     

Cell cycle dependent regulation of deoxycytidine kinase, deoxyguanosine kinase, and cytosolic 5'-nucleotidase I activity in MOLT-4 cells

Activation of nucleoside analogues is dependent on kinases and 5'-nucleotidases and the balance between the activity of these enzymes. The purpose of this study was to analyze deoxycytidine kinase, deoxyguanosine kinase, and 4 different 5'-nucleotidases during cell cycle progression in MOLT-4 cells. The activity of both kinases was cell cycle dependent and increased during proliferation while the activity of cytosolic 5'-nucleotidase I decreased. We could show that the kinase activity was higher than the total nucleotidase activity, which was unchanged or decreased during cell cycle progression. These data may be important in designing modern combination therapy with nucleoside analogues.     

Production of haploid and doubled haploid plants of melon (<Emphasis Type="Italic">Cucumis melo</Emphasis> L.) for use in breeding for multiple virus resistance

We have developed improved procedures for recovery of haploid and doubled haploid (DH) melon plants, using hybrids derived from crosses of lines with multiple virus resistance. Seeds formed after pollination with irradiated pollen were cultured in liquid medium for 10 days before excision of the embryos for further culture. This made it easier to identify the seeds containing parthenogenetic embryos, thereby reducing the effort required and increasing the percentage of plants recovered. The plants obtained (approximately 175) were transferred to a greenhouse for evaluation. Three fertile lines were identified, and selfed seeds were obtained for evaluating virus resistance. Flow cytometry of leaf tissues showed that two of these lines were spontaneous DH and the third was a mixoploid containing haploid and diploid cells. The other plants remained sterile through the flowering stage. Flow cytometry of 20 sterile plants showed that all were haploid. Attempts to induce chromosome doubling by applying colchicine to greenhouse-grown plants were unsuccessful. Shoot tips from the haploid plants were used to establish new in vitro cultures. In vitro treatment of 167 micropropagated haploid shoots with colchicine produced 10 diploid plants as well as 100 mixoploid plants. Pollen from male flowers that formed in vitro on the colchicine-treated plants was examined. High percentages of viable pollen that stained with acetocarmine were found not only in the diploids but also in >60% of the plants scored as mixoploid or haploid by flow cytometry. Efficient recovery of DH from hybrid melon lines carrying combinations of important horticultural traits will be a valuable tool for melon breeders.