Synthesis of a folate functionalized PEGylated poly(propylene imine) dendrimer as prospective targeted drug delivery system

Based on fourth generation diaminobutane poly(propylene imine) dendrimer, a novel targeted drug nanocarrier was prepared, bearing protective PEG chains and a folate targeting ligand. As a control a PEGylated derivative without folate was also synthesized. The encapsulation and release properties of these PEGylated derivatives were investigated employing etoposide, an anticancer hydrophobic drug. Enhanced solubility of etoposide was achieved inside the dendrimeric scaffold which was subsequently released in a controlled manner. These properties coupled with specificity towards the folate receptor and the low toxicity render folate functionalized PEGylated poly(propylene imine) dendrimer promising candidate for targeted drug delivery.     

The ex vivo toll-like receptor 7 tolerance induction in donor lymphocytes prevents murine acute graft-versus-host disease

Background aims

Acute graft-versus-host disease (aGVHD) remains a major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation, mediated by alloreactive donor T cells. Toll-like receptors (TLRs), a family of conserved pattern-recognition receptors (PRRs), represent key players in donors' T-cell activation during aGVHD; however, a regulatory, tolerogenic role for certain TLRs has been recognized in a different context. We investigated whether the ex vivo–induced TLR-2,-4,-7 tolerance in donor cells could prevent alloreactivity in a mismatched transplantation model.

Vitamin D3 ameliorates podocyte injury through the nephrin signalling pathway

Renal podocytes form the main filtration barrier possessing unique phenotype maintained by proteins including podocalyxin and nephrin, which are modulated in pathological conditions. In diabetic nephropathy (DN), podocytes become structurally and functionally compromised. Nephrin, a structural backbone protein of the slit diaphragm, acts as regulator of podocyte intracellular signalling with renoprotective role. Vitamin D₃ through its receptor, VDR, provides renal protection in DN but limited data exist about its effect on podocytes. In this study, we used isolated rat glomeruli to assess podocalyxin and nephrin expression after treatment with the 1,25‐dihydroxyvitamin D₃ analogue paricalcitol in the presence of normal and diabetic glucose levels. The role of 1,25‐dihydroxyvitamin D₃ (calcitriol) and its analogue, paricalcitol, on podocyte morphology and survival was also investigated in the streptozotocin (STZ)‐diabetic animal model. In our ex vivo model, glomeruli exhibited high glucose‐mediated down‐regulation of podocalyxin, and nephrin, while paricalcitol reversed the high glucose‐induced decrease of nephrin and podocalyxin expression. Paricalcitol treatment enhanced VDR expression and promoted VDR and RXR co‐localization in the nucleus. Our data also indicated that hyperglycaemia impaired survival of cultured glomeruli and suggested that the implemented nephrin down‐regulation was reversed by paricalcitol treatment, initiating Akt signal transduction which may be involved in glomerular survival. Our findings were further verified in vivo, as in the STZ‐diabetic animal model, calcitriol and paricalcitol treatment resulted in significant amelioration of hyperglycaemia and restoration of nephrin signalling, suggesting that calcitriol and paricalcitol may provide molecular bases for protection against loss of the permselective renal barrier in DN.     

Expression of ptc and gli genes in talpid3 suggests bifurcation in Shh pathway

talpid3 is an embryonic-lethal chicken mutation in a molecularly un-characterised autosomal gene. The recessive, pleiotropic phenotype includes polydactylous limbs with morphologically similar digits. Previous analysis established that hox-D and bmp genes, that are normally expressed posteriorly in the limb bud in response to a localised, posterior source of Sonic Hedgehog (Shh) are expressed symmetrically across the entire anteroposterior axis in talpid3 limb buds. In contrast, Shh expression itself is unaffected. Here we examine expression of patched (ptc), which encodes a component of the Shh receptor, and is probably itself a direct target of Shh signalling, to establish whether talpid3 acts in the Shh pathway. We find that ptc expression is significantly reduced in talpid3 embryos. We also demonstrate that talpid3 function is not required for Shh signal production but is required for normal response to Shh signals, implicating talpid3 in transduction of Shh signals in responding cells. Our analysis of expression of putative components of the Shh pathway, gli1, gli3 and coupTFII shows that genes regulated by Shh are either ectopically expressed or no longer responsive to Shh signals in talpid3 limbs, suggesting possible bifurcation in the Shh pathway. We also describe genetic mapping of gli1, ptc, shh and smoothened in chickens and confirm by co-segregation analysis that none of these genes correspond to talpid3.     

A model for anteroposterior patterning of the vertebrate limb based on sequential long- and short-range Shh signalling and Bmp signalling

It has been proposed that digit identity in chick limb bud is specified in a dose-dependent fashion by a long-range morphogen, produced by the polarising region. One candidate is Sonic hedgehog (Shh) protein, but it is not clear whether Shh acts long or short range or via Bmps. Here we dissect the relationship between Shh and Bmp signalling. We show that Shh is necessary not only for initiating bmp2 expression but also for sustaining its expression during the period when additional digits are being specified. We also show that we can reproduce much of the effect of Shh during this period by applying only Bmp2. We further demonstrate that it is Bmps that are responsible for digit specification by transiently adding Noggin or Bmp antibodies to limbs treated with Shh. In such limbs, multiple additional digits still form but they all have the same identity. We also explored time dependency and range of Shh signalling by examining ptc expression. We show that high-level ptc expression is induced rapidly when either Shh beads or polarising regions are grafted to a host limb. Furthermore, we find that high-level ptc expression is first widespread but later more restricted. All these data lead us to propose a new model for digit patterning. We suggest that Shh initially acts long range to prime the region of the limb competent to form digits and thus control digit number. Then later, Shh acts short range to induce expression of Bmps, whose morphogenetic action specifies digit identity.     

Glucose-induced gradual phenotypic modulation of cultured human glomerular epithelial cells may be independent of Wilms’ tumor 1 (WT1)

Background

Melatonin, a hormone-like substance involved in the regulation of the circadian rhythm, has been demonstrated to protect cells against oxidative DNA damage and to inhibit tumorigenesis.

Results

In the current study, we investigated the effect of melatonin on DNA strand breaks using the alkaline DNA comet assay in breast cancer (MCF-7) and colon cancer (HCT-15) cell lines. Our results demonstrated that cells pretreated with melatonin had significantly shorter Olive tail moments compared to non-melatonin treated cells upon mutagen (methyl methanesulfonate, MMS) exposure, indicating an increased DNA repair capacity after melatonin treatment. We further examined the genome-wide gene expression in melatonin pretreated MCF-7 cells upon carcinogen exposure and detected altered expression of many genes involved in multiple DNA damage responsive pathways. Genes exhibiting altered expression were further analyzed for functional interrelatedness using network- and pathway-based bioinformatics analysis. The top functional network was defined as having relevance for “DNA Replication, Recombination, and Repair, Gene Expression, [and] Cancer”.

Conclusions

These findings suggest that melatonin may enhance DNA repair capacity by affecting several key genes involved in DNA damage responsive pathways.     

Grain number determination in durum wheat as affected by drought stress: An analysis at spike and spikelet level

Yield studies show that increases in grain yield are always accompanied by an increase in grain number and, hence, further increases in yield potential may require additional improvements in grain number. The improvement of modern durum wheat was mainly based on the introduction of semidwarf genes. A 2‐year field drought stress experiment, concerning two different genotype groups (landraces vs modern cultivars), was carried out under a rainout shelter in order (a) to assess the effect of water deficit on floret dynamics and grain number determination, (b) to explore the relationship between plant water status with grain number per spike and its components (i.e., spikelets per spike, fertile florets per spikelet and grain set) and (c) to quantify the importance of several plant traits in determining the final number of grains per spike and fertile florets per spike when the main source of variation is water availability. Compared to control (well irrigated), grain number per spike was reduced, depending on year, genotype and water availability level, by 12.4–58.7% and this reduction was evident almost in all spikelet positions along the spike. Although there were some doubts in the past about the increased sensitivity of semidwarf cultivars to drought stress, they were not confirmed from our results. In most of the cases, the variation in plant water status (by means of water potential index [WPI]) during floret primordia phase (FPP) explained most of the variance in grain number per spike, fertile florets per spikelet, grain set and fertile spikelets per spike. In general, increasing water stress intensity decreased grain number per spike by an average rate of 13.5 and 9.4 grains per 0.2 MPa decrease in WPI, in modern cultivars and landraces, respectively. However, seasonal and genotypic effects were evident by modifying the slopes of the linear regressions between WPI and the studied plant traits. Commonality analysis revealed that the number of fertile florets per spikelet was the best predictor of grain number per spike, indicating that there is still much space for further improvement for this trait in landraces. However, this trait has been clearly improved in modern cultivars, especially at the basal and central spikelets. Although the number of spikelets per spike was the best unique predictor of the number of grains per spike in modern cultivars, grain set presented the highest total effect.