Combined use of dbcAMP and IBMX minimizes the damage induced by a long‐term artificial meiotic arrest in mouse germinal vesicle oocytes

Phosphodiesterase (PDE)‐mediated reduction of cyclic adenosine monophosphate (cAMP) activity can initiate germinal vesicle (GV) breakdown in mammalian oocytes. It is crucial to maintain oocytes at the GV stage for a long period to analyze meiotic resumption in vitro. Meiotic resumption can be reversibly inhibited in isolated oocytes by cAMP modulator forskolin, cAMP analog dibutyryl cAMP (dbcAMP), or PDE inhibitors, milrinone (Mil), Cilostazol (CLZ), and 3‐isobutyl‐1‐methylxanthine (IBMX). However, these chemicals negatively affect oocyte development and maturation when used independently. Here, we used ICR mice to develop a model that could maintain GV‐stage arrest with minimal toxic effects on subsequent oocyte and embryonic development. We identified optimal concentrations of forskolin, dbcAMP, Mil, CLZ, IBMX, and their combinations for inhibiting oocyte meiotic resumption. Adverse effects were assessed according to subsequent development potential, including meiotic resumption after washout, first polar body extrusion, early apoptosis, double‐strand DNA breaks, mitochondrial distribution, adenosine triphosphate levels, and embryonic development. Incubation with a combination of 50.0 μM dbcAMP and 10.0 μM IBMX efficiently inhibited meiotic resumption in GV‐stage oocytes, with low toxicity on subsequent oocyte maturation and embryonic development. This work proposes a novel method with reduced toxicity to effectively arrest and maintain mouse oocytes at the GV stage.     

The Arabidopsis AtAAE13.1 Gene Enhances Salt Stress Tolerance in Angiosperms and Gymnosperm Plant Cells

In Vitro Cellular & Developmental Biology - Plant - The Arabidopsis malonyl-CoA synthetase gene AAE13.1 (AtAAE13.1) plays important roles in cell metabolism, plant growth and development, and...     

Experimental models of maternal–fetal interface and their potential use for nanotechnology applications

During pregnancy, the placenta regulates the transfer of oxygen, nutrients, and residual products between the maternal and fetal bloodstreams and is a key determinant of fetal exposure to xenobiotics from the mother. To study the disposition of substances through the placenta, various experimental models are used, especially the perfused placenta, placental villi explants, and cell lineage models. In this context, nanotechnology, an area of study that is on the rise, enables the creation of particles on nanometric scales capable of releasing drugs aimed at specific tissues. An important reason for furthering the studies on transplacental transfer is to explore the potential of nanoparticles (NPs), in new delivery strategies for drugs that are specifically aimed at the mother, the placenta, or the fetus and that involve less toxicity. Due to the fact that the placental barrier is essential for the interaction between the maternal and fetal organisms as well as the possibility of NPs being used in the treatment of various pathologies, the aim of this review is to present the main experimental models used in studying the maternal–fetal interaction and the action of NPs in the placental environment.     

Tripartite motif protein 52 (TRIM52) promoted fibrosis in LX‐2 cells through PPM1A‐mediated Smad2/3 pathway

To investigate the roles of tripartite motif containing 52 (TRIM52) in human hepatic fibrosis in vitro, human hepatic stellate cell line LX‐2 cells were transfected with hepatitis B virus (HBV) replicon to establish HBV‐induced fibrosis in LX‐2 cells, and then treated with small interfering RNA‐mediated knockdown of TRIM52 (siTRIM52). LX‐2 cells without HBV replicon transfection were treated with lentiviruses‐mediated overexpression of TRIM52 and phosphatase magnesium dependent 1A (PPM1A). Fibrosis response of LX‐2 cells were assessed by the production of hydroxyproline (Hyp) and collagen I/III, as well as protein levels of α‐smooth muscle actin (α‐SMA). PPM1A and phosphorylated (p)‐Smad2/3 were measured to assess the mechanism. The correlation between TRIM52 and PPM1A was determined using co‐immunoprecipitation, and whether and how TRIM52 regulated the degradation of PPM1A were determined by ubiquitination assay. Our data confirmed HBV‐induced fibrogenesis of LX‐2 cells, as evidenced by significant increase in Hyp and collagen I/III and α‐SMA, which was associated with reduction of PPM1A and elevation of transforming growth factor‐β (TGF‐β), p‐Smad2/3, and p‐Smad3L. However, those changes induced by HBV were significantly attenuated with additional siTRIM52 treatment. Similar to HBV, overexpression of TRIM52 exerted promoted effect in the fibrosis of LX‐2 cells. Interestingly, TRIM52 induced the fibrogenesis of LX‐2 cells and the activation of TGF‐β/Smad pathway were significantly reversed by PPM1A overexpression. Furthermore, our data confirmed TRIM52 as a deubiquitinase that influenced the accumulation of PPM1A protein, and subsequently regulated the fibrogenesis of LX‐2 cells. TRIM52 was a fibrosis promoter in hepatic fibrosis in vitro, likely through PPM1A‐mediated TGF‐β/Smad pathway.     

Restoration and management for cliff‐nesting birds in Mediterranean mining sites: the Sand Martin case study

Habitat enhancement for birds is frequently implemented during mine site restoration. Cliff‐nesting birds often colonize anthropogenic environments such as mining areas (aggregate sites and quarries for aggregate and cement production). Mining activity can compromise breeding success, causing cliff‐nesting birds to depend on the management and restoration of mining areas. The objective of our study is to assess the importance of mine site habitats for Sand Martin conservation and reconcile mining activity with breeding success in Mediterranean environments. We studied Sand Martin breeding habitat preferences in mining areas at three spatial scales. At the mining site scale, we compared 10 mining sites with Sand Martin burrows with 19 mining sites without burrows. At the colony scale (vertical structures with colonies), we evaluated the relationships between the number of breeding pairs, number of burrows, and colony characteristics within 30 distinct Sand Martin colonies. At the burrow scale, we compared the characteristics of the available vertical structure with the areas used by Sand Martins. At the mining site scale, Sand Martins preferred more surface of water bodies, shorter distances to flowing water, older sites, and mining sites which produce aggregates instead of cement. At the colony scale, Sand Martins preferred southwest orientations and stockpiles to vertical extraction faces. At the burrow scale, birds preferred the most vertical areas of the face. Our results support the need for effective habitat restoration and improved management for more effective Sand Martin conservation within mining areas. Simple interventions can enhance habitat quality and conservation of cliff‐nesting birds.     

Potential of restored gravel pits to provide suitable habitats for Eurasian otters in anthropogenic landscapes

In anthropogenic landscapes, which are usually characterized by the existence of highly heterogeneous patchworks of habitats with different conservation status, restored gravel pits have the potential to play a conservation role for semiaquatic species such as Eurasian otters (Lutra lutra). Here, we report an otter habitat suitability analysis on a complex fluvial system in an anthropogenic landscape to understand the role of different artificial water bodies (i.e. irrigation channels and ditches) and natural water bodies (i.e. rivers) related to a restored gravel pit lagoon system as providers of suitable habitats for otters. We implemented seven sampling campaigns during 2016 and 2017 consisting of 19 transects across all existing types of water bodies. We integrated 34 environmental variables with otter habitat use, measured by three spraint marking intensity indicators. We found that otter use of water bodies was not related to the natural or artificial origin of the water. Three key factors influenced habitat suitability: riparian vegetation tree cover, forestland use (either natural or planted) within 100 m from the banks, and the level of human disturbance. Our results suggest that otters' tolerance of human activities might be lower in key areas of their territory compared to areas they pass through or explore. We argue that restored gravel pit lagoon systems can potentially play a role in providing suitable habitats for otters in anthropogenic landscapes.     

Phosphorylation of Dab2 is involved in inhibited VEGF‐VEGFR‐2 signaling induced by downregulation of syndecan‐1 in glomerular endothelial cell

Disabled‐2 (Dab2) and PAR‐3 (partitioning defective 3) are reported to play critical roles in maintaining retinal microvascular endothelial cells biology by regulating VEGF‐VEGFR‐2 signaling. The role of Dab2 and PAR‐3 in glomerular endothelial cell (GEnC) is unclear. In this study, we found that, no matter whether with vascular endothelial growth factor (VEGF) treatment or not, decreased expression of Dab2 could lead to cell apoptosis by preventing activation of VEGF‐VEGFR‐2 signaling in GEnC, accompanied by reduced membrane VEGFR‐2 expression. And silencing of PAR‐3 gene expression caused increased apoptosis of GEnC by inhibiting activation of VEGF‐VEGFR‐2 signaling and membrane VEGFR‐2 expression. In our previous research, we found that the silencing of syndecan‐1 gene expression inhibited VEGF‐VEGFR‐2 signaling by modulating internalization of VEGFR‐2. And our further research demonstrated that downregulation of syndecan‐1 lead to no significant change in the expression of Dab2 and PAR‐3 both at messenger RNA and protein levels in GEnC, while phosphorylation of Dab2 was significantly increased in GEnC transfected with Dab2 small interfering RNA (siRNA) compared with control siRNA. Atypical protein kinase C (aPKC) could induce phosphorylation of Dab2, thus negatively regulating VEGF‐VEGFR‐2 signaling. And we found that decreased expression of syndecan‐1 lead to activation of aPKC, and aPKC inhibitor treatment could block phosphorylation of Dab2 in GEnC. Besides, aPKC inhibitor treatment could activate VEGF‐VGEFR‐2 signaling in GEnC transfected with syndecan‐1 siRNA in a dose‐dependent manner. In conclusion, we speculated that phosphorylation of Dab2 is involved in preventing activation of VEGF‐VEGFR‐2 signaling in GEnC transfected with syndecan‐1 siRNA. This provides a new target for the therapy of GEnC injury and kidney disease.     

Protosappanin‐A and oleanolic acid protect injured podocytes from apoptosis through inhibition of AKT‐mTOR signaling

Protosappanin‐A (PrA) and oleanolic acid (OA), which are important effective ingredients isolated from Caesalpinia sappan L., exhibit therapeutic potential in multiple diseases. This study focused on exploring the mechanisms of PrA and OA function in podocyte injury. An in vitro model of podocyte injury was induced by the sC5b‐9 complex and assays such as cell viability, apoptosis, immunofluorescence, quantitative real‐time polymerase chain reaction, and western blot were performed to further investigate the effects and mechanisms of PrA and OA in podocyte injury. The models of podocyte injury were verified to be successful as seen through significantly decreased levels of nephrin, podocin, and CD2AP and increased level of desmin. The sC5b‐9‐induced podocyte apoptosis was inhibited in injured podocytes treated with PrA and OA, accompanied by increased protein levels of nephrin, podocin, CD2AP, and Bcl2 and decreased levels of desmin and Bax. The p‐AKT/p‐mTOR levels were also reduced by treatment of PrA and OA while AKT/mTOR was unaltered. Further, the effects of PrA and OA on injured podocytes were similar to that of LY294002 (a PI3K‐AKT inhibitor). PrA and OA were also seen to inhibit podocyte apoptosis and p‐AKT/p‐mTOR levels induced by IGF‐1 (a PI3K‐AKT activator). Our data demonstrate that PrA and OA can protect podocytes from injury or apoptosis, which may occur through inhibition of the abnormal activation of AKT‐mTOR signaling.