Evaluation of Mechanical and Chemical Stimulations on Osteocalcin and Runx2 Expression in Mesenchymal Stem Cells

The osseous tissue repair and regeneration have great importance in orthopedic and maxillofacial surgery. Tissue engineering makes it possible to cure different tissue abnormalities using autologous grafts. It is now obvious that mechanical loading has essential role in directing cells to differentiation. In this study, the influence of cyclic uniaxial loading and its combination with chemical factors on expression of osteogenic markers was investigated. Rat bone marrow-derived stem cells were isolated and cultured. In one group cells were maintained in chemical induction medium. In another group cells were subjected to cyclic uniaxial strain with 3% amplitude and 0.3 Hz frequency for 24 hours and in the last group cells were affected by induction medium and physical stimulation. TaqMan Real time PCR and immunocytochemistry were done to evaluate gene expression variations. Moreover, a small incision was made to access the bone of the cranium and induced cells were seeded on collagen based scaffolds and finally the cell seeded scaffolds were implanted. Results indicated that mechanical loading alone caused a phenomenal increase in Runx2 and osteocalcin expression. Remarkable increment in gene expression was gained when induction medium were added to mechanical stimulation. The order of chemical and mechanical stimulation caused different effects and results were much better when the cells were affected by mechanical strain at first. Histological analysis showed mechanical stimulation could promote bone ingrowth in vivo. These evidences demonstrated that combination of chemical factors with mechanical strain was much more effective for directing osteogenesis since these elements have synergistic effects.     

Pancreatic β-cell regeneration: From molecular mechanisms to therapy

Pancreatic β cells are a type of cells that are present in the islets of Langerhans. These cells are highly specialized for the secretion of insulin in response to low increasing of blood glucose levels. Hence, pancreatic β cells could contribute to maintaining systemic glucose homeostasis. Increasing evidence has revealed that a variety of internal (ie, genetic and epigenetic factors) and external factors (ie, radical-oxidative stress) are involved in the protection and/or regeneration of pancreatic β cells. The pathways regulating β-cell replication have been intensely investigated. Glucose has an important role in cell cycle entry of quiescent β cells, which exerts its effect via glucose metabolism and unfolded proteins. A variety of growth factors, hormones, and signaling pathways (ie, calcium-calcineurin nuclear factor of activated T cells) are others factors that could affect β-cell replication under different conditions. Therefore, a greater understanding of the underlying pathways involved in the regeneration and protection of pancreatic β cells could lead to finding and developing new therapeutic approaches. Utilization of stem cells and various phytochemical agents have provided new aspects for preventing β-cell degeneration and stimulating the endogenous regeneration of islets. Thus, these therapeutic platforms could be used as potential therapies in the treatment of insulin-dependent diabetes mellitus. Here, we summarized the various mechanisms involved in pancreatic β-cell regeneration. Moreover, we highlighted different therapeutic approaches which could be used for the regeneration of pancreatic β cells.     

Signaling roadmap to epithelial–mesenchymal transition in pterygium,TWIST1 centralized

Pterygium as a complex disease shares common features with other malignant cells in its onset recurrence and especially epithelial–mesenchymal transition (EMT) transition. Although using different approaches including conjunctival autografts, amniotic membrane, radiotherapy, mitomycin C (MMC) has shown promising insights in the inhibition of pterygium recurrence, it needs to be investigated in more details in molecular pathways to present adjuvant target therapy. In this study, we aimed to evaluate the expression of and then illustrate the role of signaling pathways on EMT in pterygium. Using real-time polymerase chain reaction, the twist-related protein 1 (TWIST1) expression was compared in primary pterygium and normal conjunctiva. This study assessed the mRNA expression, as well as the association between the clinicopathological indices and the gene expression level. The expression level of TWIST1 was overexpressed in 36% of our cohort ( n = 76). There was a significant positive correlation between recurrence with grade T, grade V and a significant negative correlation with growth activity. Our vast literature review on different signaling pathways in pterygium showed that EMT has centralization role in recurrence of this disease. Our data confirmed that EMT is important in the recurrence of pterygium samples and different signaling pathways end up activating the EMT markers. It is suggested to evaluate the environmental factors and their correlation with molecular markers to select favorable treatment for this kind of diseases.     

The role of miRNA-377 as a tumor suppressor in lung cancer by negative regulation of genes belonging to ErbB signaling pathway

Molecular Biology Reports - The ErbB signaling pathway plays important role in the pathogenesis of lung cancer. We explored the role of miRNA-377 as a tumor suppressor in NSCLC through silencing of...     

Comparative proteomic profiles of Pinus monticola needles during early compatible and incompatible interactions with Cronartium ribicola

The proteomic profiles of primary needles from Cr2-resistant and cr2-susceptible Pinus monticola seedlings were analysed post Cronartium ribicola inoculation by 2-DE. One hundred-and-five protein spots exhibiting significant differential expression were identified using LC–MS/MS. Functional classification showed that the most numerous proteins are involved in defence signalling, oxidative burst, metabolic pathways, and other physiological processes. Our results revealed that differential expression of proteins in response to C. ribicola inoculation was genotype- and infection-stage dependent. Responsive proteins in resistant seedlings with incompatible white pine blister rust (WPBR) interaction included such well-characterized proteins as heat shock proteins (HSPs), reactive oxygen species (ROS) scavenging enzymes, and intermediate factors functioning in the signal transduction pathways triggered by well-known plant R genes, as well as new candidates in plant defence like sugar epimerase, GTP-binding proteins, and chloroplastic ribonucleoproteins. Fewer proteins were regulated in susceptible seedlings; most of them were in common with resistant seedlings and related to photosynthesis among others. Quantitative RT-PCR analysis confirmed HSP- and ROS-related genes played an important role in host defence in response to C. ribicola infection. To the best of our knowledge, this is the first comparative proteomics study on WPBR interactions at the early stages of host defence, which provides a reference proteomic profile for other five-needle pines as well as resistance candidates for further understanding of host resistance in the WPBR pathosystem.     

Intervessel pit membrane thickness best explains variation in embolism resistance amongst stems of Arabidopsis thaliana accessions

Background and AimsThe ability to avoid drought-induced embolisms in the xylem is one of the essential traits for plants to survive periods of water shortage. Over the past three decades, hydraulic studies have been focusing on trees, which limits our ability to understand how herbs tolerate drought. Here we investigate the embolism resistance in inflorescence stems of four Arabidopsis thaliana accessions that differ in growth form and drought response. We assess functional traits underlying the variation in embolism resistance amongst the accessions studied using detailed anatomical observations.MethodsVulnerability to xylem embolism was evaluated via vulnerability curves using the centrifuge technique and linked with detailed anatomical observations in stems using light microscopy and transmission electron microscopy.Key ResultsThe data show significant differences in stem P_50, varying 2-fold from −1.58 MPa in the Cape Verde Island accession to −3.07 MPa in the woody soc1 ful double mutant. Out of all the anatomical traits measured, intervessel pit membrane thickness (T_PM) best explains the differences in P₅₀, as well as P₁₂ and P₈₈. The association between embolism resistance and T_PM can be functionally explained by the air-seeding hypothesis. There is no evidence that the correlation between increased woodiness and increased embolism resistance is directly related to functional aspects. However, we found that increased woodiness is strongly linked to other lignification characters, explaining why mechanical stem reinforcement is indirectly related to increased embolism resistance.ConclusionsThe woodier or more lignified accessions are more resistant to embolism than the herbaceous accessions, confirming the link between increased stem lignification and increased embolism resistance, as also observed in other lineages. Intervessel pit membrane thickness and, to a lesser extent, theoretical vessel implosion resistance and vessel wall thickness are the missing functional links between stem lignification and embolism resistance.     

Colocalization of the (Pro)renin Receptor/Atp6ap2 with H+-ATPases in Mouse Kidney but Prorenin Does Not Acutely Regulate Intercalated Cell H+-ATPase Activity

The (Pro)renin receptor (P)RR/Atp6ap2 is a cell surface protein capable of binding and non-proteolytically activate prorenin. Additionally, (P)RR is associated with H⁺-ATPases and alternative functions in H⁺-ATPase regulation as well as in Wnt signalling have been reported. Kidneys express very high levels of H⁺-ATPases which are involved in multiple functions such as endocytosis, membrane protein recycling as well as urinary acidification, bicarbonate reabsorption, and salt absorption. Here, we wanted to localize the (P)RR/Atp6ap2 along the murine nephron, exmaine whether the (P)RR/Atp6ap2 is coregulated with other H⁺-ATPase subunits, and whether acute stimulation of the (P)RR/Atp6ap2 with prorenin regulates H⁺-ATPase activity in intercalated cells in freshly isolated collecting ducts. We localized (P)PR/Atp6ap2 along the murine nephron by qPCR and immunohistochemistry. (P)RR/Atp6ap2 mRNA was detected in all nephron segments with highest levels in the collecting system coinciding with H⁺-ATPases. Further experiments demonstrated expression at the brush border membrane of proximal tubules and in all types of intercalated cells colocalizing with H⁺-ATPases. In mice treated with NH₄Cl, NaHCO₃, KHCO₃, NaCl, or the mineralocorticoid DOCA for 7 days, (P)RR/Atp6ap2 and H⁺-ATPase subunits were regulated but not co-regulated at protein and mRNA levels. Immunolocalization in kidneys from control, NH₄Cl or NaHCO₃ treated mice demonstrated always colocalization of PRR/Atp6ap2 with H⁺-ATPase subunits at the brush border membrane of proximal tubules, the apical pole of type A intercalated cells, and at basolateral and/or apical membranes of non-type A intercalated cells. Microperfusion of isolated cortical collecting ducts and luminal application of prorenin did not acutely stimulate H⁺-ATPase activity. However, incubation of isolated collecting ducts with prorenin non-significantly increased ERK1/2 phosphorylation. Our results suggest that the PRR/Atp6ap2 may form a complex with H⁺-ATPases in proximal tubule and intercalated cells but that prorenin has no acute effect on H⁺-ATPase activity in intercalated cells.     

Susceptibility of the predatory stinkbug <Emphasis Type="Italic">Picromerus bidens</Emphasis> to selected insecticides

The Palaearctic stinkbug Picromerus bidens L. (Heteroptera: Pentatomidae) has been considered as a potential biocontrol agent of several defoliator pests in various agricultural and forest ecosystems. It may therefore be a valuable alternative for the Nearctic pentatomid Podisus maculiventris (Say) (Heteroptera: Pentatomidae), the augmentative use of which has largely been abandoned in Europe given its possible environmental impact as an exotic polyphagous predator. However, no study has yet documented the impact of insecticides on P. bidens, which is essential to evaluate the possible combination of this predatory pentatomid with current insecticide applications in integrated pest management (IPM) programmes. This study reports on laboratory experiments investigating the susceptibility of P. bidens to five insecticides with different modes of action: the pyrethroid deltamethrin, the diacylhydrazine methoxyfenozide, the juvenile hormone mimic pyriproxyfen, the spinosyn spinosad, and the neonicotinoid imidacloprid, all of which are used to control defoliator pests. Fourth-instar nymphs and female adults of the predator were exposed to formulated materials of the insecticides by residual contact. Methoxyfenozide and spinosad did not cause significant mortality to 4th instars and female adults of P. bidens. In contrast, deltamethrin and imidacloprid were harmful to nymphs and adults of the predator, with LC₅₀ values ranging between 1.5 and 9.9 mg a.i./l. Pyriproxyfen was toxic to 4th instars with an LC₅₀ value of 13.9 mg a.i./l but did not affect female adults. Reproduction and longevity of P. bidens were not adversely affected when the predator was exposed to field concentrations of spinosad, methoxyfenozide, or pyriproxyfen, or sublethal concentrations (around LC₁₀) of deltamethrin and imidacloprid. The results from residual contact experiments suggest that methoxyfenozide, spinosad, and to a lesser extent, pyriproxyfen may be compatible with P. bidens in an IPM programme. Further experiments assessing food chain toxicity of these compounds should offer a more complete picture of their selectivity.     

Development of Metal-Enhanced Fluorescence-Based Aptasensor for Thrombin Detection Using Silver Dendritic Nanostructures

Metal-enhanced fluorescence (MEF) phenomenon has shown a promising potential in the field of fluorescence-based biological sensing. In this study, we optimized the electroless metal deposition method to fabricate silver dendritic nanostructures as effective MEF active substrates. Then, an aptasensor was developed for thrombin detection using the established surfaces. For this purpose, thiolated 29-mer thrombin-binding aptamers (TBA₂₉ (12T) SH) as capturing aptamer were immobilized on the surface of silver dendritic nanostructures, then thrombin was sandwiched between the capturing aptamer and Cy5-labeled 15-mer thrombin aptamer (TBA_15-Cy5). Quantitative analysis was performed through fluorescence signal measurement. The established aptasensor presented satisfactory sensitivity and selectivity and exhibited a limit of detection (LOD) as low as 32 pM. This aptasensor was also able to detect thrombin in the human serum at picomolar levels. Furthermore, the ease and relatively low-cost of fabrication of this platform introduce it as a tool with great potential for the clinical diagnosis of diseases and also for improving sensitivity of a variety of technologies which exploit fluorescent dyes for analyte detection, at ultra-trace levels, in complex matrices.