Salt stress responses in pear and quince: physiological and molecular aspects

The productivity of agricultural crops is seriously limited by salinity. This problem is rapidly increasing, particularly in irrigated lands. Like almost all the fruit tree species, Pyrus communis is generally considered a salt sensitive species, but only little information is available on its behavior under saline conditions. Previous studies, carried out in the Department of Fruit Tree and Woody Plant Science (University of Bologna), focused their attention on pear and quince salt stress responses to understand which rootstock would be the most suitable for pear in order to tolerate a salt stress condition. It has been reported that pear and quince have different ability in the uptake, translocation and accumulation of chloride (Cl-) and sodium (Na+) ions, when plants were irrigated for one season with saline water (5 dS/m). The aim of the present work was to deepen these aspects and investigate salt stress responses in pear and quince. Two different experiments have been performed: a “short-term” trial in a growth chamber and a “long-term” experiment in the open field. In the short-term experiment, three different genotypes usually adopted as pear rootstocks (MC, BA29 and Farold®40) and the pear variety Abbé Fétel own rooted have been compared under salt stress conditions. The trial was performed in a hydroponic culture system, applying a 90 mM NaCl stress to half of the plants, after five weeks of normal growth in Hoagland’s solution. During the three-weeks of salt stress treatment, physiological, mineral and molecular analyses were performed in order to monitor, for each genotype, the development of the salt stress responses in comparison with the corresponding “unstressed” plants. Farold®40 and Abbé Fétel own rooted showed the onset of leaf necrosis, due to salt toxicity, one week before quinces. Moreover, quinces displayed a significant delay in premature senescence of old leaves, while pears emerged for their ability to regenerate new leaves from apparently dead foliage with the salt stress still running. Physiological measurements, such as shoots length, chlorophyll (Chl) content, and photosynthesis, have been carried out and revealed that pears exhibited a significant reduction in water content and a wilting aspect, while for quinces a decrease in Chl content and a growth slowdown were observed. At the end of the trial, all plants were collected and organs separated for dry weight estimation and mineral analyses (Cu, Fe, Mn, Zn Mg, Ca, K, Na and Cl). Mineral contents have been affected by salinity; same macro/micro nutrients were altered in some organs or relocated within the plant. This plant response could have partially contributed to face the salt stress. Leaves and roots have been harvested for molecular analyses at four different times during stress conditions. Molecular analyses consisted of the gene expression study of three main ion transporters, well known in Arabidopsis thaliana as salt-tolerance determinants in the “SOS” pathway: NHX1 (tonoplast Na+/H+ antiporter), SOS1 (plasmalemma Na+/H+ antiporter) and HKT1 (K+ high-affinity and Na+ low-affinity transporter). These studies showed that two quince rootstocks adopted different responsive mechanisms to NaCl stress. BA29 increased its Na+ sequestration activity into leaf vacuoles, while MC enhanced temporarily the same ability, but in roots. Farold®40, instead, exhibited increases in SOS1 and HKT1 expression mainly at leaf level in the attempt to retrieve Na+ from xylem, while Abbé Fétel differently altered the expression of these genes in roots. Finally, each genotype showed a peculiar response to salt stress that was the sum of its ability in Na+ exclusion, osmotic tolerance and tissue tolerance. In the long-term experiment, potted trees of the pear variety Abbé Fétel grafted on different rootstocks (MC, BA29 and Farold®40), or own rooted and also rootstocks only were subjected to a salt stress through saline water irrigation with an electrical conductivity of 5 dS/m for two years. The purposes of this study were to evaluate salinity effects on physiological (shoot length, number of buds, photosynthesis, etc.) and yield parameters of cultivar Abbé Fétel in the different combinations and to determine the salt amount that pear is able to tolerate over the years. With this work, we confirmed the previous hypothesis that pear, despite being classified as a salt-sensitive fruit tree, can be cultivated for two years under saline water irrigation, without showing any salt toxicity symptoms or severe drawbacks on plant development and production. Among different combinations, Abbé Fétel grafted on MC resulted interesting for its peculiar behaviors under salt stress conditions. In the near future, further investigations on physiological and molecular aspects will be necessary to enrich and broaden the knowledge of salt stress responses in pear.

Utilizzo di dispositivi magnetici nel trattamento della patologia artrosica del ginocchio

The use of scaffolds for Tissue Engineering (TE) is increasing due to their efficacy in helping the body rebuild damaged or diseased tissue. Hydroxyapatite (HA) is the most suitable bioactive ceramic to be used in orthopaedic reconstruction since it replicates the mineral component of the hard tissues, and it has therefore excellent biocompatibility properties. The temporal and spatial control of the tissue regeneration process is the limit to be overcome in order to treat large bone and osteochondral defects. In this thesis we describe the realization of a magnetic scaffolds able to attract and take up growth factors or other bio-agents in vivo via a driving magnetic force. This concept involves the use of magnetic nanoparticles (MNP) functionalized with selected growth factors or stem cells. These functionalized MNP act as shuttles transporting the bio-agents towards and inside the scaffold under the effect of the magnetic field, enhancing the control of tissue regeneration processes. This scaffold can be imagined as a fixed “station” that provides a unique possibility to adjust the scaffold activity to the specific needs of the healing tissue. Synthetic bone graft substitutes, made of collagen or biomineralized collagen (i.e. biomimetic Hydroxyapatite/collagen composites) were used as starting materials for the fabrication of magnetic scaffolds. These materials are routinely used clinically to replace damaged or diseased cartilaginous or bone tissue. Our magnetization technique is based on a dip-coating process consisting in the infilling of biologically inspired porous scaffolds with aqueous biocompatible ferrofluids’ suspensions. In this technique, the specific interconnected porosity of the scaffolds allows the ferrofluids to be drawn inside the structure by capillarity. A subsequent freeze-drying process allows the solvent elimination while keeping very nearly the original shape and porosity of the scaffolds. The remaining magnetic nanoparticles, which are trapped in the structure, lead to the magnetization of the HA/Collagen scaffold. We demonstrate here the possibility to magnetize commercially available scaffolds up to magnetization values that are used in drug delivery processes. The preliminary biocompatibility test showed that the investigated scaffolds provide a suitable micro-environment for cells. The biocompatibility of scaffold facilitates the growth and proliferation of osteogenic cells.

Rilascio dei biomarkers di danno miocardico dopo iniezione diretta per via percutanea di cellule staminali e terapia genica

Aims: We aimed to quantify the release of bio-markers of myocardial damage in relation to direct intramyocardial injections of genes and stem cells in patients with severe coronary artery disease. Methods and Results: We studied 71 patients with “no-option” coronary artery disease. Patients had, via the percutaneous transluminal route, a total of 11±1 (mean ± SD) intramyocardial injections of vascular endothelial growth factor genes (n=56) or mesenchymal stromal cells (n=15). Injections were guided to an ischemic area by electromechanical mapping, using the NOGA™/Myostar™ catheter system. ECG was monitored continuously until discharge. Plasma CKMB (upper normal laboratory limit=5 μg/l) was 2 μg/l (2-3) at baseline; increased to 6 (5-9) after 8 hours (p < 0.0001) and normalized to 4 (3-5) after 24 hours. A total of 8 patients (17%), receiving a volume of 0.3 ml per injection, had CKMB rises exceeding 3 times the upper limit, whereas no patient in the group receiving 0.2 ml had a more than two fold CKMB increase. No patient developed new ECG changes. There were no clinically important ventricular arrhythmias and no death. Conclusion: Direct Intramyocardial injections of stem cells or genes lead to measurable release of cardiac bio-markers, which was related to the injected volume.