Interactions between extracellular signal‐regulated kinase 1/2 and P38 Map kinase pathways in the control of RUNX2 phosphorylation and transcriptional activity

RUNX2, a key transcription factor for osteoblast differentiation, is regulated by ERK1/2 and p38 MAP kinase‐mediated phosphorylation. However, the specific contribution of each kinase to RUNX2‐dependent transcription is not known. Here we investigate ERK and p38 regulation of RUNX2 using a unique P‐RUNX2‐specific antibody. Both MAP kinases stimulated RUNX2 Ser319 phosphorylation and transcriptional activity. However, a clear preference for ERK1 versus p38α/β was found when the ability of these MAPKs to phosphorylate and activate RUNX2 was compared. Similarly, ERK1 preferentially bound to a consensus MAPK binding site on RUNX2 that was essential for the activity of either kinase. To assess the relative contribution of ERK1/2 and p38 to osteoblast gene expression, MC3T3‐E1 preosteoblast cells were grown in control or ascorbic acid (AA)‐containing medium ± BMP2/7. AA‐induced gene expression, which requires collagen matrix synthesis, was associated with parallel increases in P‐ERK and RUNX2‐S319‐P in the absence of any changes in P‐p38. This response was blocked by ERK, but not p38, inhibition. Significantly, in the presence of AA, BMP2/7 synergistically stimulated RUNX2 S319 phosphorylation and transcriptional activity without affecting total RUNX2 and this response was totally dependent on ERK/MAPK activity. In contrast, although p38 inhibition partially blocked BMP‐dependent transcription, it did not affect RUNX2 S319 phosphorylation, suggesting the involvement of other phosphorylation sites and/or transcription factors in this response. Based on this work, we conclude that extracellular matrix and BMP regulation of RUNX2 phosphorylation and transcriptional activity in osteoblasts is predominantly mediated by ERK rather than p38 MAPKs. © 2012 American Society for Bone and Mineral Research.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90254/1/561_ftp.pd

Preliminary screening, identification and biological characteristic analysis of Bacillus probiotics isolated from Cynoglossus semilaevis

To screen local probiotic strains to promote antibiotic-free farming, two potential probiotic strains (S3, S5) were recognized among 89 cultivable bacterial strains isolated from the intestine of healthy Cynoglossus semilaevis. The two potential probiotic isolates were analyzed in terms of their morphology, physiology, biochemistry, the similarity of 16S rDNA sequences, growth characteristics, enzyme production capacity, bacterial antagonism, and safety in C. semilaevis. The results revealed that the bacterial morphology and physiological and biochemical characteristics of S3 and S5 were similar to those of Bacillus subtilis. The 16S rDNA sequences had 99.9 % similarity to that of Bacillus subtilis MH 145363.1. Therefore, S3 and S5 were identified as B. subtilis. In addition, we found that S3 and S5 had a strong ability to secrete amylase, protease, and lipase. During the safety tests of S3 and S5 in C. semilaevis with high concentrations, C. semilaevis in immersion, injection, and feeding groups remained in good condition without falling ill or dying. Moreover, we found that S3 and S5 exhibited superior growth at 25~50℃, salinities of 10 to 40, and pH values of 5 to 9. Furthermore, S3 and S5 had significant bacteriostatic activity against Vibrio anguillarum, Aeromonas salmonicida, and Shewanella algae, which are the main pathogenic bacteria of mariculture fish. In summary, S3 and S5 showed superb inhibition of the pathogenic bacteria of marine fish, rapid growth, eurythermal and euryhaline features, and suitability for the intestinal environment of C. semilaevis. Thus, strains S3 and S5 have excellent commercial development potential. These results provide a basis for ecological disease prevention strategies and are also valuable for developing and utilizing probiotics

Discoidin Receptor 2 Controls Bone Formation and Marrow Adipogenesis

Cell–extracellular matrix (ECM) interactions play major roles in controlling progenitor cell fate and differentiation. The receptor tyrosine kinase, discoidin domain receptor 2 (DDR2), is an important mediator of interactions between cells and fibrillar collagens. DDR2 signals through both ERK1/2 and p38 MAP kinase, which stimulate osteoblast differentiation and bone formation. Here we show that DDR2 is critical for skeletal development and differentiation of marrow progenitor cells to osteoblasts while suppressing marrow adipogenesis. Smallie mice (Ddr2slie/slie), which contain a nonfunctional Ddr2 allele, have multiple skeletal defects. A progressive decrease in tibial trabecular bone volume/total volume (BV/TV) was observed when wild‐type (WT), Ddr2wt/slie, and Ddr2slie/slie mice were compared. These changes were associated with reduced trabecular number (Tb.N) and trabecular thickness (Tb.Th) and increased trabecular spacing (Tb.Sp) in both males and females, but reduced cortical thickness only in Ddr2slie/slie females. Bone changes were attributed to decreased bone formation rather than increased osteoclast activity. Significantly, marrow fat and adipocyte‐specific mRNA expression were significantly elevated in Ddr2slie/slie animals. Additional skeletal defects include widened calvarial sutures and reduced vertebral trabecular bone. To examine the role of DDR2 signaling in cell differentiation, bone marrow stromal cells (BMSCs) were grown under osteogenic and adipogenic conditions. Ddr2slie/slie cells exhibited defective osteoblast differentiation and accelerated adipogenesis. Changes in differentiation were related to activity of runt‐related transcription factor 2 (RUNX2) and PPARγ, transcription factors that are both controlled by MAPK‐dependent phosphorylation. Specifically, the defective osteoblast differentiation in calvarial cells from Ddr2slie/slie mice was associated with reduced ERK/MAP kinase and RUNX2‐S319 phosphorylation and could be rescued with a constitutively active phosphomimetic RUNX2 mutant. Also, DDR2 was shown to increase RUNX2‐S319 phosphorylation and transcriptional activity while also increasing PPARγ‐S112 phosphorylation, but reducing its activity. DDR2 is, therefore, important for maintenance of osteoblast activity and suppression of marrow adipogenesis in vivo and these actions are related to changes in MAPK‐dependent RUNX2 and PPARγ phosphorylation. © 2016 American Society for Bone and Mineral Research.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/135235/1/jbmr2893_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/135235/2/jbmr2893.pd

Tracking circadian rhythms of bone mineral deposition in murine calvarial organ cultures

Osteoblasts, which orchestrate the deposition of small apatite crystals through the expression of nucleating proteins, have been shown to also express clock genes associated with the circadian signaling pathway. We hypothesized that protein‐mediated bone mineralization may be linked to circadian oscillator mechanisms functioning in peripheral bone tissue. In this study, Per1 expression in ex vivo neonatal murine calvaria organ cultures was monitored for 6 days using a Per1 ‐ luciferase transgene as a bioluminescent indicator of clock function. Fluctuations in Per1 expression had a period of 25 ± 4 hours ( n  = 14) with early expression at CT09:59 ± 03:37 (CT = circadian time). We also established the kinetics of mineral deposition in developing bone by using noninvasive Raman microscopy to track mineral accumulation in calvarial tissue. The content and quality of newly deposited mineral was continually examined at the interparietal bone/fontanel boundary for a period of 6 days with 1‐hour temporal resolution. Using this approach, mineralization over time exhibited bursts of mineral deposition followed by little or no deposition, which was recurrent with a periodicity of 26.8 ± 9.6 hours. As many as six near‐daily mineralization events were observed in the calvaria before deposition ceased. Earliest mineralization events occurred at CT16:51 ± 03:45, which is 6 hours behind Per1 expression. These findings are consistent with the hypothesis that mineralization in developing bone tissue is regulated by a local circadian oscillator mechanism.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/99084/1/jbmr1924.pd

The mechanism of pore segregation in the sintered nano Ag for high temperature power electronics applications

The final publication is available at Elsevier via http://dx.doi.org/10.1016/j.matlet.2018.06.007 © 2018. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/It is widely accepted that nano pores of sintered nanoparticles can coalesce into micro pores during high temperature service. When applying sintered nanoparticles in power electronics, the pore segregation and delamination seriously degrade the bond strength and thermal conductivity, but the reason is still not well understood. In this paper, both finite element analysis and experimental results confirmed that thermal stress is the main driving force for this phenomenon, which was not considered in the previous study. The effect of pore segregation on performance and reliability of power devices is also discussed.National Natural Science Foundation of China (51520105007, 51775299) National Key Research and Development Program of China (2017YFB1104900

The prognostic value of multiparametric cardiac magnetic resonance in patients with systemic light chain amyloidosis

BackgroundLate gadolinium enhancement (LGE) is a classic imaging modality derived from cardiac magnetic resonance (CMR), which is commonly used to describe cardiac tissue characterization. T1 mapping with extracellular volume (ECV) and native T1 are novel quantitative parameters. The prognostic value of multiparametric CMR in patients with light chain (AL) amyloidosis remains to be thoroughly investigated.MethodsA total of 89 subjects with AL amyloidosis were enrolled from April 2016 to January 2021, and all of them underwent CMR on a 3.0 T scanner. The clinical outcome and therapeutic effect were observed. Cox regression was used to investigate the effect of multiple CMR parameters on outcomes in this population.ResultsLGE extent, native T1 and ECV correlated well with cardiac biomarkers. During a median follow-up of 40 months, 21 patients died. ECV (hazard ratio [HR]: 2.087 for per 10% increase, 95% confidence interval [CI]: 1.379-3.157, P < 0.001) and native T1 (HR: 2.443 for per 100 ms increase, 95% CI: 1.381-4.321, P=0.002) were independently predictive of mortality. A novel prognostic staging system based on median native T1 (1344 ms) and ECV (40%) was similar to Mayo 2004 Stage, and the 5-year estimated overall survival rates in Stage I, II, and III were 95%, 80%, and 53%, respectively. In patients with ECV > 40%, receiving autologous stem cell transplantation had higher cardiac and renal response rates than conventional chemotherapy.ConclusionBoth native T1 and ECV independently predict mortality in patients with AL amyloidosis. Receiving autologous stem cell transplantation is effective and significantly improves the clinical outcomes in patients with ECV > 40%

Reversal of cancer gene expression identifies repurposed drugs for diffuse intrinsic pontine glioma

Diffuse intrinsic pontine glioma (DIPG) is an aggressive incurable brainstem tumor that targets young children. Complete resection is not possible, and chemotherapy and radiotherapy are currently only palliative. This study aimed to identify potential therapeutic agents using a computational pipeline to perform an in silico screen for novel drugs. We then tested the identified drugs against a panel of patient-derived DIPG cell lines. Using a systematic computational approach with publicly available databases of gene signature in DIPG patients and cancer cell lines treated with a library of clinically available drugs, we identified drug hits with the ability to reverse a DIPG gene signature to one that matches normal tissue background. The biological and molecular effects of drug treatment was analyzed by cell viability assay and RNA sequence. In vivo DIPG mouse model survival studies were also conducted. As a result, two of three identified drugs showed potency against the DIPG cell lines Triptolide and mycophenolate mofetil (MMF) demonstrated significant inhibition of cell viability in DIPG cell lines. Guanosine rescued reduced cell viability induced by MMF. In vivo, MMF treatment significantly inhibited tumor growth in subcutaneous xenograft mice models. In conclusion, we identified clinically available drugs with the ability to reverse DIPG gene signatures and anti-DIPG activity in vitro and in vivo. This novel approach can repurpose drugs and significantly decrease the cost and time normally required in drug discovery

Deviant Dynamics of Resting State Electroencephalogram Microstate in Patients With Subjective Tinnitus

Given the importance of central reorganization and tinnitus, we undertook the current study to investigate changes to electroencephalogram (EEG) microstates and their association with the clinical symptoms in tinnitus. High-density (128 channel) EEG was used to explore changes in microstate features in 15 subjects with subjective tinnitus and 17 age-matched controls. Correlations between microstate parameters and subjective tinnitus symptoms were also analyzed. An increased presence of class A microstate and decreased presence of class D microstate were found in coverage and lifespan of microstate parameters in the tinnitus patients. Syntax analysis also demonstrated an aberrant pattern of activity, with reduced transitions from class D to class B in tinnitus patients. Moreover, a significant positive correlation of tinnitus loudness with increased lifespan of microstate class C was found. Significant differences in temporal characteristics and syntax of the EEG microstate classes were found at rest between tinnitus patients and the healthy subjects. Our study indicates that EEG microstates may provide a possible valuable method to study large-scale brain networks, which may in turn be beneficial to investigation of the neurophysiological mechanisms behind tinnitus

EbMYBP1, a R2R3-MYB transcription factor, promotes flavonoid biosynthesis in Erigeron breviscapus

Erigeron breviscapus, a traditional Chinese medicinal plant, is enriched in flavonoids that are beneficial to human health. While we know that R2R3-MYB transcription factors (TFs) are crucial to flavonoid pathway, the transcriptional regulation of flavonoid biosynthesis in E. breviscapus has not been fully elucidated. Here, EbMYBP1, a R2R3-MYB transcription factor, was uncovered as a regulator involved in the regulation of flavonoid accumulation. Transcriptome and metabolome analysis revealed that a large group of genes related to flavonoid biosynthesis were significantly changed, accompanied by significantly increased concentrations of the flavonoid in EbMYBP1-OE transgenic tobacco compared with the wild-type (WT). In vitro and in vivo investigations showed that EbMYBP1 participated in flavonoid biosynthesis, acting as a nucleus-localized transcriptional activator and activating the transcription of flavonoid-associated genes like FLS, F3H, CHS, and CHI by directly binding to their promoters. Collectively, these new findings are advancing our understanding of the transcriptional regulation that modulates the flavonoid biosynthesis

Aberrant Functional and Causal Connectivity in Acute Tinnitus With Sensorineural Hearing Loss

Purpose: The neural bases in acute tinnitus remains largely undetected. The objective of this study was to identify the alteration of the brain network involved in patients with acute tinnitus and hearing loss. Methods: Acute tinnitus patients (n = 24) with hearing loss and age-, sex-, education-matched healthy controls (n = 21) participated in the current study and underwent resting-state functional magnetic resonance imaging (fMRI) scanning. Regional homogeneity and amplitude of low-frequency fluctuation were used to investigate the local spontaneous neural activity and functional connectivity (FC), and Granger causality analysis (GCA) was used to analyze the undirected and directed connectivity of brain regions. Results: Compared with healthy subjects, acute tinnitus patients had a general reduction in FC between auditory and non-auditory brain regions. Based on FC analysis, the superior temporal gyrus (STG) revealed reduced undirected connectivity with non-auditory brain regions including the amygdala (AMYG), nucleus accumbens (NAc), the cerebellum, and postcentral gyrus (PoCG). Using the GCA algorithm, increased effective connectivity from the right AMYG to the right STG, and reduced connectivity from the right PoCG to the left NAc was observed in acute tinnitus patients with hearing loss. The pure-tone threshold was positively correlated with FC between the AMYG and STG, and negatively correlated with FC between the left NAc and the right PoCG. In addition, a negative association between the GCA value from the right PoCG to the left NAc and the THI scores was observed. Conclusion: Acute tinnitus patients have aberrant FC strength and causal connectivity in both the auditory and non-auditory cortex, especially in the STG, AMYG, and NAc. The current findings will provide a new perspective for understanding the neuropathophysiological mechanism in acute tinnitus