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Urocanic acid (UCA) is an endogenous small molecule that is elevated in skin, blood and brain after sunlight exposure, mainly playing roles in the periphery systems. Few studies have investigated the role of UCA in the central nervous system. In particular, its role in memory consolidation and reconsolidation is still unclear. In the present study, we investigated the effect of intraperitoneal injection of UCA on memory consolidation and reconsolidation in a novel object recognition memory (ORM) task. In the consolidation version of the ORM task, the protocol involved three phases: habituation, sampling and test. UCA injection immediately after the sampling period enhanced ORM memory performance; UCA injection 6 h after sampling did not affect ORM memory performance. In the reconsolidation version of the ORM task, the protocol involved three phases: sampling, reactivation and test. UCA injection immediately after reactivation enhanced ORM memory performance; UCA injection 6 h after reactivation did not affect ORM memory performance; UCA injection 24 h after sampling without reactivation did not affect ORM memory performance. This UCA-enhanced memory performance was not due to its effects on nonspecific responses such as locomotor activity and exploratory behavior. The results suggest that UCA injection enhances consolidation and reconsolidation of an ORM task, which further extends previous research on UCA effects on learning and memory.

Summary Skeletal muscle satellite cells (SCs) are stem cells responsible for muscle development and regeneration. Although CRISPR/Cas9 has been widely used, its application in endogenous SCs remains elusive. Here, we generate mice expressing Cas9 in SCs and achieve robust editing in juvenile SCs at the postnatal stage through AAV9-mediated short guide RNA (sgRNA) delivery. Additionally, we reveal that quiescent SCs are resistant to CRISPR/Cas9-mediated editing. As a proof of concept, we demonstrate efficient editing of master transcription factor (TF) Myod1 locus using the CRISPR/Cas9/AAV9-sgRNA system in juvenile SCs. Application on two key TFs, MYC and BCL6, unveils distinct functions in SC activation and muscle regeneration. Particularly, we reveal that MYC orchestrates SC activation through regulating 3D genome architecture. Its depletion results in strengthening of the topologically associating domain boundaries thus may affect gene expression. Altogether, our study establishes a platform for editing endogenous SCs that can be harnessed to elucidate the functionality of key regulators governing SC activities., Highlights • CRISPR/Cas9/AAV9-sgRNA system yields robust editing in juvenile SCs • Quiescent SCs are resistant to CRISPR/Cas9-mediated in vivo editing • Key TFs play diversified functions during SC early activation • MYC promotes SC activation through impinging on 3D chromatin architecture, In this article, Wang and colleagues generate a CRISPR/Cas9/AAV9-sgRNA in vivo genome editing system to achieve robust editing in juvenile satellite cells (SCs). Application of the system reveals diversified functions of key transcription factors (TFs) during SC activation and uncovers that MYC promotes SC activation through regulating 3D genome architecture. Quiescent SCs, however, are resistant to the in vivo editing.

Parathyroid hormone (PTH) plays an important role in maintaining mineral homeostasis by regulating calcium and phosphate levels. Clinical trials have shown that peptides of PTH (1-34), PTH-related protein (PTHrP 1-36), and the new peptide modeled on PTHrP, abaloparatide, can have different anabolic effects on osteoporotic subjects, but the underlying mechanisms are still unclear. The prevalence of moderate and major gingival recession has been shown to be higher in postmenopausal women with osteoporosis. In addition, there is a significant association between osteoporosis and tooth loss.We investigated the actions of these peptides on the cementoblasts and teeth of mice. The murine cementoblast line, OCCM-30, known to express collagen I (Col1a1), was treated with intermittent PTH (1-34), PTHrP (1-36), or abaloparatide for 6 h/d for 3 days. Microcomputed tomography was performed on the teeth of mice receiving daily injections of phosphate-buffered saline, PTH (1-34), or abaloparatide. Statistical differences were analyzed by a 2-way or 1-way analysis of variance followed by a Tukey's post-hoc test. Results are expressed as mean ± standard deviation, and P 0.05 was considered significant.Gene expression showed regulation of Bsp, Col1a1, Opg, Rankl, and Mmp13 by the 3 peptides in these cells. Western blots revealed that after intermittent treatment for 3 days, PTH (1-34) caused an increase in COL1A1 protein immediately after treatment. In contrast, abaloparatide showed a latent effect in increasing COL1A1 protein 18 hours after treatment. PTHrP had no effect on COL1A1 expression. Immunofluorescence confirmed the same result as the Western blots. Microcomputed tomography of teeth showed PTH (1-34) injections increased molar root mineral density in mice, whereas abaloparatide increased density in roots of incisors and molars.This study reveals the differential anabolic effects of intermittent PTH (1-34), PTHrP (1-36), and abaloparatide on cementoblasts, as revealed by COL1A1 expression and root mineral density. Abaloparatide may be a potential therapeutic approach for achieving improved cementogenesis.

Plasmid-specific bacteriophages specifically infect bacteria carrying conjugal plasmids. While wastewater has been used as isolation source for such phages, to date, only the distribution and ecology of RNA phages specific to the F plasmid have been described, because they serve as a water quality indicator. Yet, several other plasmid classes have higher clinical and ecological relevance, and the distribution, fate, and ecology of the phages that target them remain uncharacterized. We aimed to (i) provide an experimental platform to quantify the abundance of plasmid-specific phages applicable to several different conjugal plasmid classes, (ii) describe the distribution of such phages in wastewater systems, and (iii) relate their abundance to plasmid abundance and to municipal wastewater treatment processes. We introduced four model conjugal plasmids, belonging to incompatibility groups IncP-1, IncN, IncHI1, or IncF into an avirulent Salmonella enterica strain, for which somatic phages are at low abundance in wastewater. These strains were used in double layer agar assays with waters from contrasting sources. Plasmid-specific phages were common in wastewater but rare in river water. Hospital wastewater contained significantly more IncP-1-, but fewer IncF- and IncN- specific phages than domestic wastewater. This pattern did not match that of plasmid abundance estimated by Inc group targeting high-throughput quantitative PCR. The comparison between influent and effluent of wastewater treatment plants revealed a reduction in phage concentration by ca. 2 log, without significant contribution of primary settling. Overall, the ubiquity of these phages hints at their importance for plasmid ecology, and can provide opportunities in water quality monitoring and in ecological management of mobile resistance genes. Plasmid-specific bacteriophages specifically infect bacteria carrying conjugal plasmids. While wastewater has been used as isolation source for such phages, to date, only the distribution and ecology of RNA phages specific to the F plasmid have been described, because they serve as a water quality indicator. Yet, several other plasmid classes have higher clinical and ecological relevance, and the distribution, fate, and ecology of the phages that target them remain uncharacterized. We aimed to (i) provide an experimental platform to quantify the abundance of plasmid-specific phages applicable to several different conjugal plasmid classes, (ii) describe the distribution of such phages in wastewater systems, and (iii) relate their abundance to plasmid abundance and to municipal wastewater treatment processes. We introduced four model conjugal plasmids, belonging to incompatibility groups IncP-1, IncN, IncHI1, or IncF into an avirulent Salmonella enterica strain, for which somatic phages are at low abundance in wastewater. These strains were used in double layer agar assays with waters from contrasting sources. Plasmid-specific phages were common in wastewater but rare in river water. Hospital wastewater contained significantly more IncP-1-, but fewer IncF- and IncN- specific phages than domestic wastewater. This pattern did not match that of plasmid abundance estimated by Inc group targeting high-throughput quantitative PCR. The comparison between influent and effluent of wastewater treatment plants revealed a reduction in phage concentration by ca. 2 log, without significant contribution of primary settling. Overall, the ubiquity of these phages hints at their importance for plasmid ecology, and can provide opportunities in water quality monitoring and in ecological management of mobile resistance genes.

Aerosol and particulate matter are playing significant roles in the regional climate and environment in the tropical rain forest of Southeast Asia. Both satellite and ground observations showed significant seasonal variations in the PM2.5 concentration and the aerosol optical properties during 2012–2014 in the Xishuangbanna tropical rain forest. The annual mean values of the PM2.5, aerosol optical depth (AOD), and the Angstrom exponent (α) were 34.3 ± 19.7 μg·m−3, 0.54 ± 0.37, and 1.36 ± 0.20 in the dry season and 16.90 ± 5.08 μg·m−3, 0.37 ± 0.10, and 1.07 ± 0.25 in the wet season, respectively. The results showed that 46.9% and 56.5% of the moderate-resolution imaging spectroradiometer (MODIS) C6 AOD data met the NASA accuracy requirements in the dry and wet season respectively and 17.1% and 17.7% of the seasonal mean systematically underestimated the ground-based data. There was a high correlation between PM2.5 and AOD. The range of the correlation coefficient (R2) was 0.69–0.85 in the dry season and 0.33–0.39 in the wet season. Linear regression functions of PM2.5 and MODIS AOD were developed and used to retrieve the spatial and temporal distributions of the PM2.5 in the tropical rain forest over the last decade (2006–2015). The annual mean PM2.5 increased slightly in the region. The range of the PM2.5 was 20–40 μg·m−3 in the wet season and 25–80 μg·m−3 in the dry season. In northern Thailand, northern Vietnam and the central district of Laos, PM2.5 was up to the range of 50–80 μg·m−3, which was mainly attributed to biomass burning in these areas.

Estrogen-related receptor γ (ERRγ) and 17β-hydroxysteroid dehydrogenase type 1 (HSD17B1) have important roles in cell invasion and in the proliferation of many types of cancer cells. However, it remains unknown whether ERRγ and HSD17B1 contribute to abnormal placental structure and dysfunction which characterize fetal growth restriction (FGR). Therefore, the aim of this study was to investigate the expression profiles of ERRγ and HSD17B1 in placenta tissues affected by FGR and to examine a possible molecular mechanism by which ERRγ is able to regulate HSD17B1 during development of FGR.Placenta tissues were collected from women affected by FGR (n = 28) and from women with appropriately gestational age (AGA) (n = 30). Relative mRNA and protein levels of ERRγ and HSD17B1 in both groups were assessed by quantitative real-time PCR, immunohistochemistry, and Western blot analyses. The effect of ERRγ on trophoblast function and its associated mechanistic details were studied in the trophoblast cell line, HTR-8/SVneo, which was transfected with small interfering RNA (siRNA) targeting ERRγ.Both mRNA and protein levels of ERRγ and HSD17B1 were significantly lower in FGR placentae (P 0.05). When ERRγ expression was knocked down in HTR-8/SVneo cells with siRNA, invasion and proliferation were inhibited. In addition, HSD17B1 expression was significantly decreased. In dual luciferase reporter assays, ERRγ stimulated transcription of HSD17B1 by targeting the ERRγ response element within its 5'-flanking promoter region.Aberrant ERRγ expression may contribute to the pathogenesis of FGR by regulating the transcriptional activity of HSD17B1.

This study proposes probabilistic seismic demand and capacity models for underground structures considering the spatial distribution characteristics of soil properties. The proposed models are then applied in a reliability analysis to estimate the seismic fragility of underground structures. First, the paper uses physics-based random fields to model different spatially varying soil properties. The random fields of soil properties are used to establish numerical models of the soil-underground structure system and simulate the structural response under pushover and seismic loads. Next, the paper develops probabilistic demand and capacity models based on the structural responses obtained from the numerical analyses. Utilizing the demand and capacity models, the paper estimates the seismic fragility of underground structures, and provides confidence intervals to capture the epistemic uncertainties in the estimates. As an illustration, the paper applies the proposed models to analyze the fragility of a typical rectangular underground structure and compares the fragility estimates under four selected seismic intensity levels. The results indicate that the proposed seismic demand model captures well the dependency between the maximum inter-story drift and peak ground velocity. The obtained fragility estimates can approximately assess the seismic fragility of similar underground structures located in similar sites.

Underground structures can be vulnerable due to seismic-induced deformation. A resilient underground structure is preferable for seismic response mitigation and quick recovery. In this study, a resilient central column with angular friction damper (RC-AFD) was constructed for the seismic performance upgrading of underground structures, and a practical design procedure was developed for the RC-AFD to enable the two-state control for underground structures subject to seismic excitations with multi-intensity. The RC-AFD consists of a self-centering column characterized by relaxed constraints at the top and bottom column that is restricted by unbounded prestressed tendons, whereas a pair of angular friction dampers are placed at the top of the central column. The mechanical model and its physical realization are detailed, whereas the two-state control mechanism is proposed by following the enhanced demand of seismic performance upgrading. The effectiveness and robustness of the proposed RC-AFD for structural seismic response mitigation of typical underground structures were investigated in terms of structural displacement, shear force, bending moments, and energy-based responses. The obtained results show that the two flexible connection joints concentrated at the top and bottom of the RC-AFD produce a significant isolation effect, through which the multiple responses of the central column can be reduced significantly. Benefiting from the two-state design and the employed angular friction damper with a large radius plate, the RC-AFD exhibits an enhanced energy dissipation capacity, which robustly dissipates the intense plastic energy for multi-intensity excitations. As a consequence, the plastic energy dissipation burden of the primary structures can be significantly relieved and residual deformation can be avoided due to the excellent self-centering ability.

Central-column failure can cause large underground structures to collapse during strong earthquakes. The use of lead-filled steel-tube dampers (LFSTDs) in the seismic design of such structures is proposed to mitigate or prevent such an eventuality. Three-dimensional nonlinear dynamic analyses of the Daikai subway station with and without LFSTDs under the action of the Hyogoken–Nambu earthquake has been performed, considering soil-structure interaction. The feasibility of the proposed method was verified from four aspects—force, deformation, damage, and energy dissipation. The effects of variability in seismic inputs, strength ratio, and arrangement mode of dampers on the structural dynamic response characteristics have been discussed. Results reveal that the application of LFSTDs to large underground structures remarkably reduces the force, deformation, and damage associated with the central column under seismic loading. Additionally, a strength ratio of 0.4 is recommended when using LFSTDs, whereas in cases without LFTSTDs, the strength of the central column must be appropriately enhanced.

Runx2 (Runt-related transcription factor 2) is a key transcription factor which is associated with osteoblast differentiation and expressed in ER+ (estrogen receptor positive) human breast cancer cell lines. Runx2 also participates in mammary gland development. Deregulation of RNA Pol III genes (polymerase III-dependent genes) is tightly linked to tumor development, while Brf1 (TFIIB-related factor 1) specifically regulates these gene transcription. However, nothing is known about the effect of Runx2 on Brf1 expression and Pol III gene transcription. Expression of Runx2, Brf1 and Pol III genes from the samples of human breast cancer and cell culture model were determined by the assays of RT-qPCR, immunoblot, luciferase reporter activity, immunohistochemistry, chromatin immunoprecipitation and Immunofluorescence. High expression of Runx2 is observed in the cases of breast cancer. The patients of high Runx2 expression at early stages display longer survival period, whereas the cases of high Runx2 at advanced stages reveal faster recurrence. The identification of signaling pathway indicates that JNK1 and c-Jun mediate Runx2 transcription. Repression of Runx2 reduces Brf1 expression and Pol III gene transcription. Further analysis indicates that Runx2 is colocalized with Brf1 in nucleus of breast cancer tissue. Both Runx2 and Brf1 synergistically modulate Pol III gene transcription. These studies indicate that Brf1 overexpression is able to be used as an early diagnosis biomarker of breast cancer, while high Runx2 expression indicates long survival period and faster recurrence. Runx2 mediates the deregulation of Brf1 and Pol III genes and its abnormal expression predicts the worse prognosis of breast cancer.

Histone deacetylases (HDACs) are crucial regulators of gene expression in transcriptional co-repressor complexes. Previously, we reported that HDAC4 was a basal repressor of matrix metalloproteinase-13 (MMP-13) transcription and parathyroid hormone (PTH) regulates HDAC4 to control MMP-13 promoter activity through dissociation from Runx2. Here, we show that PTH induces the protein kinase A (PKA)-dependent phosphorylation of HDAC4 in the nucleus of the rat osteoblastic cell line, UMR 106-01. We demonstrate that PKA-dependent phosphorylated HDAC4 is released from Runx2 bound to the MMP-13 promoter in these cells. Point mutation of Ser-740 in rHDAC4 prevents the release of HDAC4 from Runx2 on the MMP-13 promoter and also prevents the PTH stimulation of MMP-13 transcription. Thus, PTH-induced phosphorylation of rHDAC4 at Ser-740 is crucial for regulating MMP-13 transcription in osteoblasts. PTH causes degradation of HDAC4, and this product appears in the cytoplasm. The cytoplasmic degradation of HDAC4 is blocked by PKA and lysosomal inhibitors, but is not affected by proteasome, caspase-3, or serine and aspartic protease inhibitors. In addition, the phosphatase inhibitor, okadaic acid, prevents degradation indicating that dephosphorylation is associated with degradation. These mechanisms regulating HDAC4 and their roles in such processes are crucial for bone and chondrocyte development. Our data support a link between PTH regulating HDAC4 phosphorylation by PKA, trafficking, partial degradation, and the control of MMP-13 transcription through association with Runx2.

Background The sustainability of the rural health-care system is under pressure in Gongcheng county, Guangxi, China, mainly owing to disparities between rural and urban medical resources, and preference of local patients to seek health care in urban tertiary hospitals. Our aim is to reform the county's primary care model to improve rural health-care services and increase its use, a critical component of the current national health-care reform plan. In this study, we primarily test the feasibility of implementing artificial intelligence (AI) powered bidirectional referral between rural clinics and the county hospitals on a new internet-based platform. We focused on developing the new web platform and primary care model and implementing it throughout the whole county health system, which serves over 300 000 residents. Methods Our reform adopts the University of California, San Francisco primary care model augmented with AI on a cloud platform. The main county hospital of Gongcheng county, Guangxi, China, and its associated rural clinics used the AI-powered Internet+ Primary Care Model (AiPCM) platform to provide patient education, patient engagement, bidirectional referral, panel management, and health coaching. The aim was for the three-level county health-care system to operate seamlessly and cost-effectively. In conjunction, the second key innovation required to make AiPCM work in rural China was deployed; an online health-care team from Guilin Medical University provided support to rural doctors in the form of professional training, online support, and a chatbot to assist village doctors in making referrals and managing the health care of rural residents. Findings We developed a new web platform to smoothly integrate AiPCM into existing clinical workflows at county, town and village levels. The main county people's hospital, two town health centres, and 17 village clinics joined the web platform in the first phase of the pilot study. The internet-based hub and spokes model allows the main county hospital (hub), and town health centres and village clinics (spokes) to care collaboratively for patients through bidirectional referral mechanisms. County hospital teams focus on diagnosis and treatment of patients referred from town health centres. Town health centre teams provide panel management for patients at risk and refer patients to county hospitals when needed. Village clinic doctors engage villagers and refer patients to town health centres when needed. County hospital specialists provide feedback to the referring doctors for them to continue caring for the patient locally. As care data are collected, a chatbot can be trained to further assist village and town doctors to make referrals and provide initial primary care. The AiPCM cloud platform can allow Guilin Medical University teams to add more capabilities to the county health system, including doctor training, patient coaching, referral assistance, chatbot training, and patient education. Thus, it is feasible to use AiPCM on a web platform to effectively implement bidirectional referrals, which should encourage more patients to see local doctors. Interpretation With AiPCM and technical support from academic institutions, the new primary care model can be implemented in rural China. Next, we will evaluate the effectiveness of AiPCM, its effect on the number of rural residents receiving the health care that they require, and patient clinical outcomes. Funding None.

Osteoblast differentiation is tightly regulated by post transcriptional regulators such as microRNAs (miRNAs). Several bioactive materials including nano-bioglass ceramic particles (nBGC) influence differentiation of the osteoblasts, but the molecular mechanisms of nBGC-stimulation of osteoblast differentiation via miRNAs are not yet determined. In this study, we identified that nBGC-treatment stimulated miR-30c expression in human osteoblastic cells (MG63). The bioinformatics tools identified its regulatory network, molecular function, biological processes and its target genes involved in negative regulation of osteoblast differentiation. TGIF2 and HDAC4 were found to be its putative target genes and their expression was down regulated by nBGC-treatment in MG63 cells. Thus, this study advances our understanding of nBGC action on bone cells and supports utilization of nBGC in bone tissue engineering.

Bladder cancer is one of the few cancers that have been linked to carcinogens in the environment and tobacco smoke. Of the carcinogens tested in mouse chemical carcinogenesis models, N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) is one that reproducibly causes high-grade, invasive cancers in the urinary bladder, but not in any other tissues. However, the basis for such a high-level tissue-specificity has not been explored. Using mutagenesis in lacI (Big Blue™) mice, we show here that BBN is a potent mutagen and it causes high-level of mutagenesis specifically in the epithelial cells (urothelial) of the urinary bladder. After a 2-6-week treatment of 0.05% BBN in the drinking water, mutagenesis in urothelial cells of male and female mice was about two orders of magnitude greater than the spontaneous mutation background. In contrast, mutagenesis in smooth muscle cells of the urinary bladder was about five times lower than in urothelial tissue. No appreciable increase in mutagenesis was observed in kidney, ureter, liver or forestomach. In lacI (Big Blue™) rats, BBN mutagenesis was also elevated in urothelial cells, albeit not nearly as profoundly as in mice. This provides a potential explanation as to why rats are less prone than mice to the formation of aggressive form of bladder cancer induced by BBN. Our results suggest that the propensity to BBN-triggered mutagenesis of urothelial cells underlies its heightened susceptibility to this carcinogen and that mutagenesis induced by BBN represents a novel model for initiation of bladder carcinogenesis.