Four-dimensional imaging with virtual reality to quantitatively explore jigsaw puzzle-like morphogenesis of Arabidopsis cotyledon pavement cells

In most dicotyledonous plants, leaf pavement cells exhibit complex jigsaw puzzle-like cell morphogenesis during leaf expansion. Although detailed molecular biological information and mathematical modeling of this jigsaw puzzle-like cell morphogenesis are now available, a full understanding of this process remains elusive. Recent reports have highlighted the importance of three-dimensional (3D) structures (i.e., anticlinal and periclinal cell wall) in understanding the mechanical models that describe this morphogenetic process. We believe that it is important to acquire 3D shapes of pavement cells over time, i.e., acquire and analyze four-dimensional (4D) information when studying the relationship between mechanical modeling and simulations and the actual cell shape. In this report, we have developed a framework to capture and analyze 4D morphological information of Arabidopsis thaliana cotyledon pavement cells by using both direct water immersion observations and computational image analyses, including segmentation, surface modeling, virtual reality and morphometry. The 4D cell models allowed us to perform time-lapse 3D morphometrical analysis, providing detailed quantitative information about changes in cell growth rate and shape, with cellular complexity observed to increase during cell growth. The framework should enable analysis of various phenotypes (e.g., mutants) in greater detail, especially in the 3D deformation of the cotyledon surface, and evaluation of theoretical models that describe pavement cell morphogenesis using computational simulations. Additionally, our accurate and high-throughput acquisition of growing cell structures should be suitable for use in generating in silico model cell structures.     

Molecular cloning of putative chloroplastic cysteine synthase in Leucaena leucocephala

Journal of Plant Research - Cysteine biosynthesis is directed by the successive commitments of serine acetyltransferase, and O-acetylserine (thiol) lyase (OASTL) compounds, which subsequently frame...     

A Highlights from MBoC Selection: Serine 129 phosphorylation of membrane-associated α-synuclein modulates dopamine transporter function in a G protein–coupled receptor kinase–dependent manner

Most α-synuclein (α-syn) deposited in Lewy bodies, the pathological hallmark of Parkinson disease (PD), is phosphorylated at Ser-129. However, the physiological and pathological roles of this modification are unclear. Here we investigate the effects of Ser-129 phosphorylation on dopamine (DA) uptake in dopaminergic SH-SY5Y cells expressing α-syn. Subcellular fractionation of small interfering RNA (siRNA)–treated cells shows that G protein–coupled receptor kinase 3 (GRK3), GRK5, GRK6, and casein kinase 2 (CK2) contribute to Ser-129 phosphorylation of membrane-associated α-syn, whereas cytosolic α-syn is phosphorylated exclusively by CK2. Expression of wild-type α-syn increases DA uptake, and this effect is diminished by introducing the S129A mutation into α-syn. However, wild-type and S129A α-syn equally increase the cell surface expression of dopamine transporter (DAT) in SH-SY5Y cells and nonneuronal HEK293 cells. In addition, siRNA-mediated knockdown of GRK5 or GRK6 significantly attenuates DA uptake without altering DAT cell surface expression, whereas knockdown of CK2 has no effect on uptake. Taken together, our results demonstrate that membrane-associated α-syn enhances DA uptake capacity of DAT by GRKs-mediated Ser-129 phosphorylation, suggesting that α-syn modulates intracellular DA levels with no functional redundancy in Ser-129 phosphorylation between GRKs and CK2.     

Hydroxyl Radical Generation Caused by the Reaction of Singlet Oxygen with a Spin Trap,DMPO, Increases Significantly in the Presence of Biological Reductants

Photosensitizers newly developed for photodynamic therapy of cancer need to be assessed using accurate methods of measuring reactive oxygen species (ROS). Little is known about the characteristics of the reaction of singlet oxygen (¹O²) with spin traps, although this knowledge is necessary in electron spin resonance (ESR)/spin trapping. In the present study, we examined the effect of various reductants usually present in biological samples on the reaction of ¹O² with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). The ESR signal of the hydroxyl radical (?OH) adduct of DMPO (DMPO-OH) resulting from ¹O²-dependent generation of ?OH strengthened remarkably in the presence of reduced glutathione (GSH), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), ascorbic acid, NADPH, etc. A similar increase was observed in the photosensitization of uroporphyrin (UP), rose bengal (RB) or methylene blue (MB). Use of 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO) as a spin trap significantly lessened the production of its ?OH adduct (DEPMPO-OH) in the presence of the reductants. The addition of DMPO to the DEPMPO-spin trapping system remarkably increased the signal intensity of DEPMPO-OH. DMPO-mediated generation of ?OH was also confirmed utilizing the hydroxylation of salicylic acid (SA). These results suggest that biological reductants enhance the ESR signal of DMPO-OH produced by DMPO-mediated generation of ?OH from ¹O², and that spin trap-mediated ?OH generation hardly occurs with DEPMPO.     

Periodontal tissue regeneration by combined implantation of adipose tissue-derived stem cells and platelet-rich plasma in a canine model

Background aimsOne goal of periodontal therapy is to regenerate periodontal tissues. Stem cells, growth factors and scaffolds and biomaterials are vital for the restoration of the architecture and function of complex tissues. Adipose tissue-derived stem cells (ASCs) are an ideal population of stem cells for practical regenerative medicine. In addition, platelet-rich plasma (PRP) can be useful for its ability to stimulate tissue regeneration. PRP contains various growth factors and may be useful as a cell carrier in stem cell therapies. The purpose of this study was to determine whether a mixture of ASCs and PRP promoted periodontal tissue regeneration in a canine model.MethodsAutologous ASCs and PRP were implanted into areas with periodontal tissue defects. Periodontal tissue defects that received PRP alone or non-implantation were also examined. Histologic, immunohistologic and x-ray studies were performed 1 or 2 months after implantation. The amount of newly formed bone and the scale of newly formed cementum in the region of the periodontal tissue defect were analyzed on tissue sections.ResultsThe areas of newly formed bone and cementum were greater 2 months after implantation of ASCs and PRP than at 1 month after implantation, and the radiopacity in the region of the periodontal tissue defect increased markedly by 2 months after implantation. The ASCs and PRP group exhibited periodontal tissue with the correct architecture, including alveolar bone, cementum-like structures and periodontal ligament-like structures, by 2 months after implantation.ConclusionsThese findings suggest that a combination of autologous ASCs and PRP promotes periodontal tissue regeneration that develops the appropriate architecture for this complex tissue.