γ-secretase inhibitor disturbs the morphological development of differentiating neurons through affecting Notch/miR-342-5p.

During neurodevelopment, differentiation of neural stem/progenitor cells (NSPCs) into neurons are regulated by many factors including Notch signaling pathway. Herein, we report the effect of a Notch signaling blocker, i.e. γ -secretase inhibitor (GSI), on this differentiating process, especially on the morphological development. NSPCs were cultured and induced to differentiate with or without GSI. The neurite outgrowth was impeded by GSI application and the expression of a Notch signaling downstream effector miR-342-5p increased with the downregulated expression of Notch effectors Hes1 and Hes5. Upregulated expression of miR-342-5p in differentiating NSPCs could shorten the neurite length of progeny neurons, which was similar to the effect of GSI. To avoid the possible influence from astrocytes into neurons, we directly applied cultured neurons, on which GSI could shorten the processes and RBP-J knockdown could also reduce the neurite length. Similarly, transfection of miR-342-5p mimics or inhibitors into PC12 cells led to shorter or longer processes of cells compared with control ones. Furthermore, in differentiating NSPCs, GSI-induced shorter neurites could be partially rescued by miR-342-5p inhibitors, and STAT3 was one of the possible targets of miR-342-5p during this differentiating process as indicated by results of Western Blot test, luciferase reporter assay and GFP reporter assay. To further demonstrate the role of STAT3, it was introduced into GSI-treated neurons and the GSI-affected neurites could also be partially rescued. In conclusion, GSI could influence the morphological development of neurons and the possible mechanism involved Notch/miR-342-5p and STAT3. These results would be informative for future therapeutic research.

Evaluation of protein separation mechanism and pore size distribution in colloidal self-assembled nanoparticle sieves for on-chip protein sizing.

The separation behavior of 6.5-66 kDa proteins in silica particle array-based sieves formed by colloidal self-assembly in microchips is reported across a pore size range of 22-103 nm. The protein separation and resolution improves markedly with decreasing pore size. The variation of electrophoretic mobility with molecular weight of SDS-protein complexes and with particle size was evaluated using the Ogston sieving equation for a random pore gel structure, and using the modified Giddings equation developed by Wirth for uniform pore structures. The Wirth/Giddings equation provides the best fit for estimation of molecular weight of proteins, and demonstrates that even though experimental evidence shows there is some dispersion in measured pore sizes, these structures can best be described as having a uniform pore size.

Differential response of pig masseter to botulinum neurotoxin serotypes a and b.

INTRODUCTION: Pigs respond to direct administration of botulinum neurotoxins (BoNTs), although they are resistant to botulism. The human masseter is frequently targeted for BoNT therapy. We aimed to understand how BoNT affects chewing by injecting porcine masseters. METHODS: One masseter of minipigs was injected with BoNT serotype A or B at doses comparable to those used in humans. Masticatory function was evaluated electromyographically. Muscle force was measured during tetany. Four weeks after injection, strain gauges affixed to the mandible assessed bone strain during chewing. Masseter mass and fiber diameter were measured after euthanasia. RESULTS: BoNT-A had no measurable effect. In contrast, BoNT-B reduced electrical activity and muscle force, producing substantial asymmetry between injected and uninjected muscles. CONCLUSIONS: The pig masseter is highly resistant to direct injection of BoNT-A, but it is affected by BoNT-B.

Botulinum neurotoxin type A in the masseter muscle: effects on incisor eruption in rabbits.

INTRODUCTION: Botulinum neurotoxins are responsible for the paralytic food poisoning, botulism. Commercial formulations such as botulinum neurotoxin type A are increasingly used for various conditions, including cosmetic recontouring of the lower face by injection of the large masseter muscles. The paralysis of a major muscle of mastication lowers occlusal force and thus might affect tooth eruption. The purpose of this study was to investigate the effects of unilateral masseter muscle injection of botulinum neurotoxin type A on the rate of eruption of incisors in a rabbit model. We hypothesized that the teeth would overerupt in an underloaded environment. METHODS: Forty rabbits were injected with either botulinum neurotoxin type A or saline solution in 1 masseter muscle. Mastication and muscle force production were monitored, and incisor eruption rate was assessed by caliper measurement of grooved teeth. RESULTS: The injection of saline solution had no effect. The masseter muscle injected with botulinum neurotoxin type A showed a dramatic loss of force 3 weeks after injection despite apparently normal mastication. Incisor eruption rate was significantly decreased for the botulinum neurotoxin type A group, an effect attributed to decreased attrition. CONCLUSIONS: This study has implications for orthodontics. Although findings from ever-growing rabbit incisors cannot be extrapolated to human teeth, it is clear that botulinum neurotoxin type A caused a decrease in bite force that could influence dental eruption.

Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor-dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions.

BACKGROUND: Tumor neovascularization (TNV) is a common pathologic basis for malignant growth and metastasis. However, the mechanism of TNV pathogenesis is not fully understood. Ras homolog gene family, member A (RhoA), a Rho guanosine triphosphatase (GTPase) family member, may be involved in a hypoxia-induced vascular endothelial growth factor (VEGF) pathway that regulates TNV angiogenesis through an unclear mechanism. METHODS: The regulation of RhoA on p53, the p53 binding protein homolog murine double minute 2 (MDM2), and VEGF was analyzed in hypoxic MCF-7 cells using Western blot analysis, real-time polymerase chain reaction (PCR) analysis, coimmunoprecipitation, and immunofluorescence staining assays. Changes in proliferation, invasion, migration, stress fiber formation, and tube formation were detected in an MCF-7 human umbilical vein endothelial cell (HUVEC) coculture system. Correlations of RhoA expression with MDM2, wild-type p53 (wt-p53), and VEGF expression in breast cancer tissues and relations between RhoA and breast cancer clinical features were analyzed by immunohistochemistry. RESULTS: Activated RhoA down-regulated p53 protein, which increased VEGF expression in hypoxic MCF-7 cells; whereas p53 messenger RNA levels were not altered. In addition, the ubiquitin-mediated degradation of p53 was enhanced by active RhoA. RhoA and MDM2 colocalized in the cytoplasm of hypoxic MCF-7 cells and interacted with each other physically. Furthermore, nutlin-3, a specific MDM2 inhibitor, was capable of reducing activated RhoA-induced p53 protein stability and attenuating VEGF accumulation. In an MCF-7-HUVEC coculture system, nutlin-3 effectively inhibited HUVEC proliferation, invasion, migration, stress fiber formation, and tube formation mediated by activated RhoA under hypoxic conditions. Data from 129 clinical breast cancer specimens with wt-p53 revealed that high RhoA expression was correlated with high MDM2 expression, low wt-p53 expression, and high VEGF expression. CONCLUSIONS: The current data suggested that activated RhoA promotes VEGF expression and hypoxia-induced angiogenesis through the up-regulation of MDM2 to decrease p53 stability.

Segmental curvilinear distraction osteogenesis.

Curvilinear distraction is currently under investigation to reconstruct curved maxillofacial bone defects. However, previous studies have revealed the discrepancy between the contour of the regenerated bone in the distraction gap and the curvilinear pathway of the transport disc. We hypothesize that the discrepancy is because of the conflict of the distraction vector and the strain vector during the consolidation. In curvilinear distraction osteogenesis, the distraction vector varies, while the strain vector during the consolidation phase is fixed-linear, from the beginning to the end of the distraction pathway. Here we bring forward a solution of segmental curvilinear distraction osteogenesis to divide the curvilinear distraction into several segments, with respective consolidation for each distraction gap. If this hypothesis is verified, the segmental distraction curvilinear distraction will benefit the reconstruction of complicated long-range maxillofacial bone defects.

Botulinum toxin in masticatory muscles: short- and long-term effects on muscle, bone, and craniofacial function in adult rabbits.

Paralysis of the masticatory muscles using botulinum toxin (BTX) is a common treatment for cosmetic reduction of the masseters as well as for conditions involving muscle spasm and pain. The effects of this treatment on mastication have not been evaluated, and claims that the treatment unloads the jaw joint and mandible have not been validated. If BTX treatment does decrease mandibular loading, osteopenia might ensue as an adverse result. Rabbits received a single dose of BTX or saline into one randomly chosen masseter muscle and were followed for 4 or 12 weeks. Masticatory muscle activity was assessed weekly, and incisor bite force elicited by stimulation of each masseter was measured periodically. At the endpoint, strain gages were installed on the neck of the mandibular condyle and on the molar area of the mandible for in vivo bone strain recording during mastication and muscle stimulation. After termination, muscles were weighed and mandibular segments were scanned with micro CT. BTX paralysis of one masseter did not alter chewing side or rate, in part because of compensation by the medial pterygoid muscle. Masseter-induced bite force was dramatically decreased. Analysis of bone strain data suggested that at 4 weeks, the mandibular condyle of the BTX-injected side was underloaded, as were both sides of the molar area. Bone quantity and quality were severely decreased specifically at these underloaded locations, especially the injection-side condylar head. At 12 weeks, most functional parameters were near their pre-injection levels, but the injected masseter still exhibited atrophy and percent bone area was still low in the condylar head. In conclusion, although the performance of mastication was only minimally harmed by BTX paralysis of the masseter, the resulting underloading was sufficient to cause notable and persistent bone loss, particularly at the temporomandibular joint.

A systematic evaluation of the role of crystalline order in nanoporous materials on DNA separation.

The role of order within a porous separation matrix on the separation efficiency of DNA was studied systematically. DNA separation was based on a ratchet mechanism. Monodisperse colloidal suspensions of nanoparticles were used to fabricate highly ordered separation media with a hexagonal close-packed structure. Doping with a second particle size yielded structures with different degrees of disorder, depending upon the volume fraction of each particle size. Radial distribution functions and orientational order parameters were calculated from electron micrographs to characterize the scale of disorder. The peak separation distance, band broadening, and separation resolution of DNA molecules was quantified for each structure. DNA separation parameters using pulsed fields and the ratchet effect showed a strong dependence on order within the porous nanoparticle array. Ordered structures gave large separation distances, smaller band broadening and better resolution than highly disordered, nearly random, porous structures. The effect dominated these three parameters when compared to the effect of pore size. However, the effect of order on separation performance was not monotonic. A small, but statistically significant improvement was seen in structures with short range order compared to those with long range order.