ENPP1 inhibits the transcription activity of the hepatitis B virus pregenomic promoter by upregulating the acetylation of LMNB1.

Current therapies for hepatitis B virus (HBV) infection can slow disease progression but cannot cure the infection, as it is difficult to eliminate or permanently silence HBV covalently closed circular DNA (cccDNA). The interaction between host factors and cccDNA is essential for their formation, stability, and transcriptional activity. Here, we focused on the regulatory role of the host factor ENPP1 and its interacting transcription factor LMNB1 in HBV replication and transcription to better understand the network of host factors that regulate HBV, which may facilitate the development of new antiviral drugs. Overexpression of ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) in Huh7 cells decreased HBV pregenomic RNA (pgRNA) and hepatitis B core antigen (HBcAg) expression levels, whereas knockdown of ENPP1 increased them. A series of HBV promoter and mutant plasmids were constructed, and a luciferase reporter assay showed that overexpression of ENPP1 caused inhibition of the HBV promoter and its mutants. A DNA pull-down assay showed that lamin B1 (LMNB1), but not ENPP1, interacts directly with the HBV enhancer II/ basic core promoter (EnhII/BCP). ZDOCK and PyMOL software were used to predict the interaction of ENPP1 with LMNB1. Overexpression of LMNB1 inhibited the activity of the HBV promoter and its mutant. The acetylation levels at the amino acids 111K, 261K, and 483K of LMNB1 were reduced compared to the control, and an LMNB1 acetylation mutant containing 111R, 261Q, 261R, 483Q, and 483R showed increased promoter activity. In summary, ENPP1 together with LMNB1 increased the acetylation level at 111K and 261K, and LMNB1 inhibited the activity of HBV promoter and downregulated the expression of pregenomic RNA and HBcAg. Our follow-up studies will investigate the expression, clinical significance, and relevance of ENPP1 and LMNB1 in HBV patient tissues, explore the effect of LMNB1 on post-transcriptional progression, and examine whether ENPP1 can reduce cccDNA levels in the nucleus.

Gut microbial community structure and function of Przewalski's horses varied across reintroduced sites in China.

Host-associated microbiota can significantly impact host fitness. Therefore, naturally occurring variations in microbiota may influence the health and persistence of their hosts. This finding is particularly important in reintroduced animals, as they typically experience habitat changes during translocations. However, little is known about how microbiomes are altered in response to conservation translocation. Here, we accessed the gut microbiome of Przewalski's horse (Equus przewalskii) populations in China from three nature reserves (i.e. Xinjiang Kalamaili Nature Reserve, KNR; Dunhuang Xihu National Nature Reserve, DXNNR; and Anxi Extreme-arid Desert Nature Reserve, AENR) using 16s rRNA gene and metagenome sequencing. The results showed that the microbial composition and function differed significantly across locations, while a subset of core taxa was consistently present in most of the samples. The abundance of genes encoding microbe-produced enzymes involved in the metabolism of carbohydrates, especially for glycoside hydrolases, was significantly higher in open-spaced KNR populations than in more confined AENR individuals. This study offers detailed and significant differential characters related to the microbial community and metabolic pathways in various reintroduced sites of Przewalski's horse, which might provide a basis for future microecological and conservation research on endangered reintroduced animals.

Comparative Analysis of Microbiome Metagenomics in Reintroduced Wild Horses and Resident Asiatic Wild Asses in the Gobi Desert Steppe.

The gut microbiome offers important ecological benefits to the host; however, our understanding of the functional microbiome in relation to wildlife adaptation, especially for translocated endangered species, is lagging. In this study, we adopted a comparative metagenomics approach to test whether the microbiome diverges for translocated and resident species with different adaptive potentials. The composition and function of the microbiome of sympatric Przewalski's horses and Asiatic wild asses in desert steppe were compared for the first time using the metagenomic shotgun sequencing approach. We identified a significant difference in microbiome composition regarding the microbes present and their relative abundances, while the diversity of microbe species was similar. Furthermore, the functional profile seemed to converge between the two hosts, with genes related to core metabolism function tending to be more abundant in wild asses. Our results indicate that sympatric wild equids differ in their microbial composition while harboring a stable microbial functional core, which may enable them to survive in challenging habitats. A higher abundance of beneficial taxa, such as Akkermansia, and genes related to metabolism pathways and enzymes, such as lignin degradation, may contribute to more diverse diet choices and larger home ranges of wild asses.

Novel protein kinase C phosphorylated kinase inhibitor-matrine suppresses replication of hepatitis B virus via modulating the mitogen-activated protein kinase signal.

HBV (hepatitis B virus) infection still threatens human health. Therefore, it is essential to find new effective anti-HBV compounds. Here, we identified matrine as a novel inhibitor of PKC (protein kinase C) phosphorylated kinase by screening a natural compound library. After HepG2.215 cells were treated with matrine, we carried out a phosphorylated proteomics sequence study and analyzed the prediction of related kinase expression level. In the case of HBV infection, it was found that PKC kinase mediates the activation of mitogen-activated protein kinase (MAPK) signaling pathway known as son of sevenless (SOS) activation. It was also found that PKC kinase inhibits the expression of C-X-C Motif Chemokine Ligand 8 (CXCL8) by inhibiting the activity of activating transcription factor 2/ cAMP response element binding protein (ATF2/CREB), and this effect is independent of its activated MAPK signaling pathway. Finally, Western blot was used to detect the expression of MAPK, ATF2, CREB3 phosphorylation and nonphosphorylation in matrine-treated cells and PKC-treated cells. PKC phosphorylated kinase inhibitor-matrine suppresses the replication of HBV via modulating the MAPK/ATF2 signal. Matrine is a good clinical drug to enhance the autoimmunity in the adjuvant treatment of chronic HBV infection.

Clinical Diagnostic Value of Quantitative Hepatitis B Virus Core Antibody Test in Chronic Viral Hepatitis B.

The level of CHB virus (HBV) core antibody (HBcAb) is different in four stages of chronic HBV infection and may be used for differential diagnosis of the natural history of chronic HBV infection. To address this question, we examined multiple blood biomarkers and assessed the efficacy to diagnose different stages of chronic HBV infection. The quantitative detection of HBcAb, hepatitis B surface antigen (HBsAg), HBV DNA, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and platelet count (PLT) were determined in the serum of 73 cases of low-replicative phase (LR), 46 cases of immune-tolerant phase (IT), 44 cases of immune clearance phase (IC), and 57 cases of HBeAg-negative hepatitis (ENH). Differentiating performance of these serum protein levels was analyzed by receiver operating characteristic (ROC) curve analysis. Our results showed that the levels of HBcAb, ALT, and AST levels were significantly higher in IC and ENH than those in LR and IT (both P ≤ 0.001). The levels of HBV DNA and HBsAg were higher in IC and IT than those in LR and ENH (both P ≤ 0.001). Logistic regression models showed that HBcAb, HBsAg, HBV DNA, ALT, and AST were the independent variables, respectively, and when combined, they provided high diagnostic accuracy for the staging of CHB. To sum up, HBcAb quantification is a new index, which can reflect whether the liver is in the immune activation state of HBV infection, and is related to the inflammatory state of the host liver. The combined detection of HBcAb quantification and other indicators has showed promising efficiency for staging of IC and ENH and can assist the diagnosis and treatment of CHB.

LncRNA MIR205HG regulates melanomagenesis via the miR-299-3p/VEGFA axis.

In this study, we investigated the role of lncRNA MIR205HG in melanomagenesis. Quantitative real-time PCR (qRT-PCR) analysis showed that MIR205HG levels were significantly upregulated in melanoma cell lines compared to normal human melanocytes. Similarly, MIR205HG levels were significantly higher melanoma tissues than adjacent normal skin tissues (n=30). CCK-8 and flow cytometry assays showed that MIR205HG knockdown significantly decreased the viability of melanoma cells. Dual luciferase reporter and RNA pull-down assays confirmed that MIR205HG directly binds to microRNA (miR)-299-3p. Targetscan analysis and dual luciferase reporter assays showed that miR-299-3p directly binds to the 3'UTR of VEGFA mRNA. Wound healing and transwell invasion assays showed that MIR205HG knockdown decreased in vitro migration and invasiveness of melanoma cells, and these effects were reversed by treatment with miR-299-3p inhibitor. MIR205HG-silenced melanoma cells showed increased miR-299-3p expression and lower levels of both VEGFA mRNA and protein. Tumor volumes were significantly smaller in nude mice xenografted with MIR205HG knockdown melanoma cells than the controls. These results demonstrate that MIR205HG supports melanoma growth via the miR-299-3p/VEGFA axis. This makes MIR205HG a potential therapeutic target for the treatment of melanoma.

Parameter Estimation for Two-Dimensional Incoherently Distributed Source with Double Cross Arrays.

A direction of arrival (DOA) estimator for two-dimensional (2D) incoherently distributed (ID) sources is presented under proposed double cross arrays, satisfying both the small interval of parallel linear arrays and the aperture equalization in the elevation and azimuth dimensions. First, by virtue of a first-order Taylor expansion for array manifold vectors of parallel linear arrays, the received signal of arrays can be reconstructed by the products of generalized manifold matrices and extended signal vectors. Then, the rotating invariant relations concerning the nominal elevation and azimuth are derived. According to the rotating invariant relationships, the rotating operators are obtained through the subspace of the covariance matrix of the received vectors. Last, the angle matching approach and angular spreads are explored based on the Capon principle. The proposed method for estimating the DOA of 2D ID sources does not require a spectral search and prior knowledge of the angular power density function. The proposed DOA estimation has a significant advantage in terms of computational cost. Investigating the influence of experimental conditions and angular spreads on estimation, numerical simulations are carried out to validate the effectiveness of the proposed method. The experimental results show that the algorithm proposed in this paper has advantages in terms of estimation accuracy, with a similar number of sensors and the same experimental conditions when compared with existing methods, and that it shows a robustness in cases of model mismatch.

Determine what to measure and how to measure in clinical trials for the treatment of pressure injury: A protocol for the development of a core outcome set.

BACKGROUND: A core outcome set (COS) is an agreed minimum set of outcomes that should be reported in all clinical trials in specific areas of health care. A considerable amount of trials did not report essential outcomes or outcomes measurement methods, which makes it challenging to evaluate the efficacy and safety of treatment strategies for pressure injury (PI) and produced significant heterogeneity of reported outcomes. It is necessary to develop a COS, which can be used for clinical trials in PI treatment. METHODS/DESIGN: The development of this COS will be guided by an advisory group composed of clinicians, senior nurses, patients, and methodologists. We will search six databases and 2 registry platforms to identify currently reported PI treatment outcomes and outcome measurement instruments in randomized controlled trials, meta-analysis, and systematic reviews. We will also conduct a semi-structured interview with clinicians, nurses, and adult PI patients to collect their opinions on important outcomes. Each outcome of the initial list generated from systematic review and interviews will be scored and reach a consensus through two rounds of international Delphi survey with all key stakeholders. A face-to-face consensus meeting with key stakeholders will be conducted to finish a final COS and recommend measurement instruments for each outcome. RESULTS: We will develop a COS that should be reported in future clinical trials to evaluate the effectiveness of PI treatment. DISCUSSION: The COS will follow current guidance to develop a high-quality COS in the field of PI treatment to reduce heterogeneity in trial reporting, facilitate valid comparisons of new therapies, and improve the quality of clinical trials.

[Intervention with thumbtack needling on spinal low back pain in Air Force crew: a randomized controlled trial].

OBJECTIVE: To observe the clinical effect on spinal low back pain (SLBP) in Air Force crew treated with novel thumbtack needling therapy and to analyze the relevant factors of the therapeutic effect. METHODS: A total of 120 Air Force crew with SLBP were randomized into a thumbtack needling group (40 cases), an external treatment group (40 cases, 1 case dropped off ) and a combined treatment group (40 cases, 1 case dropped off ). In the thumbtack needling group, the thumbtack needling therapy was adopted. The novel thumbtack needles were inserted at the lower No.6 region of the wrist-ankle acupuncture, Yaotongdian (EX-UE 7), Yaoyangguan (GV 3), etc. Each point was pressed and kneaded for 1 min each time, 3 or 4 times a day. The treatment for 3 days was taken as one course. At the interval of 2 days, 3 courses were required totally. In the external treatment group, shangshi zhitong plaster was compressed on the center of the tender site in the lumbar region, once daily, consecutively for 6 days as one course. At an interval of 1 day, 2 courses were required totally. In the combined treatment group, the treatments in the thumbtack needling group and the external treatment group were used in combination. Before and after treatment, McGill score and the score of Oswestry dysfunction index (ODI) were compared in the patients among the three groups. The average EMG (AEMG) and the mean power frequency (MPF) were analyzed by using JE-TB0810 electromyography (EMG) acquisition system to evaluate the erector spinae tension in the patients before and after treatment. The clinical effect was observed in the patient of each group and the safety was evaluated. Logistic analysis was performed on the relevant factors of therapeutic effect in the patients. RESULTS: Compared with the values before treatment, McGill scores and ODI scores were reduced (P<0.05), and AEMG and MPF increased in the patients of each group after treatment (P<0.05). After treatment, McGill scores and ODI scores in the thumbtack needling group and the combined treatment group were lower than those in the external treatment group (P<0.05), and AEMG and MPF were higher than the external treatment group (P<0.05). The total effective rates were 87.5% (35/40) and 87.2% (34/39) in the thumbtack needling group and the combined treatment group respectively and were higher than 64.1% (25/39) in the external treatment group (P<0.05). The incidence of the adverse reaction in the combined treatment group was higher than the other two groups (P<0.05). The weekly exercise frequency was the independent factor of the therapeutic effect (OR =12.166, P<0.001). CONCLUSION: The thumbtack needling therapy is significantly effective on spinal low back pain in Air Force crew and is of the safety. Hence, this therapy is applicable to be promoted in the primary care army hospital.

First reports of nasal and traumatic myiasis infection in endangered Przewalski's horses (Equus ferus przewalskii).

Myiasis has great economic and medical importance. However, myiasis in wildlife that is caused by oestroid flies is relatively rarely recorded compared with that in humans and domestic animals. Recently, during our research on the conservation of Przewalski's horse (PH), we observed two new records of oestroid flies parasitizing wildlife in China. The first is the horse nasal bot fly, Rhinoestrus sp. (Diptera: Oestridae), found in a dead PH from Kalamaili Nature Reserve. One morphotype (R. purpureus-like) was identified. The second is the Wohlfahrt's wound myiasis fly, Wohlfahrtia magnifica (Schiner, 1862) (Diptera: Sarcophagidae), which was collected from an open wound of a PH in the Wild Horse Breeding Research Centre. These observations extend the records of known hosts of these two oestroid myiasis agents. To the knowledge of the authors, infestation by Rhinoestrus and Wohlfahrtia larvae causing myiasis in wildlife has not been reported in China previously.

Genetic diversity of common Gasterophilus spp. from distinct habitats in China.

BACKGROUND: Gasterophilus species are widely distributed around the world. The larvae of these flies parasitize the digestive tract of equids and cause damage, hindering horse breeding and protection of endangered species. However, study of the genetic structure of geographically distinct Gasterophilus populations is lacking. Here, we analyzed the genetic diversity of Gasterophilus pecorum, G. intestinalis, G. nasalis and G. nigricornis from three typical grasslands (meadow, desert and alpine steppes) in China as compared to published sequences from Italy, Poland and China (Daqing and Yili), based on the mitochondrial cytochrome c oxidase cox1 and cox2 gene sequences. RESULTS: Haplotype diversity and nucleotide diversity of mitochondrial genes was generally high in all Gasterophilus populations. Due to the unique natural climatic conditions of the alpine steppe, there were high levels of genetic differentiation among different geographical populations of G. pecorum and G. nasalis, indicating that environmental variations influenced population genetic structure. Frequent exchanges between meadow and desert steppe Gasterophilus species resulted in low genetic differentiation. The highest exchange rates were found among G. intestinalis populations. Genetic differentiation was only observed on a large geographical scale, which was confirmed by analyzing population genetic structure. Three species, G. pecorum, G. intestinalis and G. nasalis, from meadow steppe showed a high emigration rate, indicating that the direction of Gasterophilus dispersal in China was from east to west. CONCLUSIONS: Our results show that the four Gasterophilus species have a high level of genetic diversity and different degrees of genetic differentiation and gene flow among different populations of the same species, reflecting their potential to adapt to the environment and the environmental impact on genetic structure. Knowledge of the genetic structure, population history, and migration will help understand the occurrence and prevalence of gasterophilosis and provide a basis for controlling the local spread of Gasterophilus spp.

Combination of chemical fingerprint and bioactivity evaluation to explore the antibacterial components of Salvia miltiorrhizae.

The aim of this study was to explore the possible antibacterial components of Salvia miltiorrhizae on Pseudomonas aeruginosa using a combination of chemical fingerprint and bioactivity evaluation. The chemical fingerprints of 32 batches of S. miltiorrhizae samples from different sources were developed using high-performance liquid chromatography with diode array detection, and then were evaluated by similarity analysis and hierarchical clustering analysis. Anti-P. aeruginosa activity was determined by microcalorimetry. Some crucial thermokinetic parameters obtained from the heat-flow power-time curves of P. aeruginosa growth in the absence or presence of these S. miltiorrhizae samples were evaluated using principal component analysis. Thereafter, multiple linear regression analysis was used to analyze the fingerprint-activity relationship between the chemical fingerprints and anti-P. aeruginosa activity. This established the related equation between the inhibition ratio (I, %) of S. miltiorrhizae samples on P. aeruginosa and the peak areas of the common peaks. The results showed that the 32S. miltiorrhizae samples could be grouped into three clusters according to their chemical fingerprints and anti-P. aeruginosa activities. Protocatechualdehyde, salvianolic acid B, together with three unidentified compounds might be the major components that contributed largely to the antibacterial properties of S. miltiorrhizae and should be the focus of S. miltiorrhizae quality control. Thus, this study provided a preferred way for exploring the bioactive components of medicinal plants.

Trace analysis of multi-class pesticide residues in Chinese medicinal health wines using gas chromatography with electron capture detection.

A method is described for multi-residue, high-throughput determination of trace levels of 22 organochlorine pesticides (OCPs) and 5 pyrethroid pesticides (PYPs) in Chinese medicinal (CM) health wines using a QuEChERS (quick, easy, cheap, effective, rugged, and safe) based extraction method and gas chromatography-electron capture detection (GC-ECD). Several parameters were optimized to improve preparation and separation time while still maintaining high sensitivity. Validation tests of spiked samples showed good linearities for 27 pesticides (R = 0.9909-0.9996) over wide concentration ranges. Limits of detection (LODs) and quantification (LOQs) were measured at ng/L levels, 0.06-2 ng/L and 0.2-6 ng/L for OCPs and 0.02-3 ng/L and 0.06-7 ng/L for PYPs, respectively. Inter- and intra-day precision tests showed variations of 0.65-9.89% for OCPs and 0.98-13.99% for PYPs, respectively. Average recoveries were in the range of 47.74-120.31%, with relative standard deviations below 20%. The developed method was then applied to analyze 80 CM wine samples. Beta-BHC (Benzene hexachloride) was the most frequently detected pesticide at concentration levels of 5.67-31.55 mg/L, followed by delta-BHC, trans-chlordane, gamma-BHC, and alpha-BHC. The validated method is simple and economical, with adequate sensitivity for trace levels of multi-class pesticides. It could be adopted by laboratories for this and other types of complex matrices analysis.

Antibacterial evaluation of flavonoid compounds against E. coli by microcalorimetry and chemometrics.

Fighting against multidrug-resistant bacteria requires reliable methods to evaluate the effect of antibacterial agents. As a universal, non-destructive, and highly sensitive tool, microcalorimetry has been used in many biological investigations to provide continuous real-time monitoring of the metabolic activity. This method, based on heat-flow output, was used to evaluate the influence of two flavonoid compounds (liquiritigenin and liquiritin) on Escherichia coli. Some crucial information, such as the thermogenic power-time curve and thermokinetic parameters of E. coli growth affected by the two compounds, was obtained and further studied by chemometric techniques including similarity analysis, multivariate analysis of variance, and principal component analysis. By comparing the values of two main parameters, k 2 (growth rate constant of the second exponential phase) and Q 1 (heat output of the first exponential growth phase) of E. coli based on the box and whisker plot, liquiritigenin and liquiritin could be differentiated according to their antibacterial effects; liquiritin with IC50 (half-inhibitory concentration) of 198.6 µg mL(-1) expressed a stronger antibacterial effect than liquiritigenin with IC50 of 337.8 µg mL(-1). The glucoside group in liquiritin containing four additional free hydroxyls in the diphenylpropane skeleton was crucial for inducing the antibacterial effect. Liquiritin might be a promising candidate against E. coli. This study provides a valuable method for searching for novel antibacterial agents using microcalorimetry with chemometrics.

[Ancient needling method--essence of ZHANG Shi-jie: a famous acupuncturist].

ZHANG Shi-jie is one of the 500 famous TCM doctors designated by the State Administration of TCM and Beijing Municipal Health Bureau. ZHANG advocates ancient needling method and uses a unique treating method which includes comprehensive analysis of the four examinations and analogy; in his ancient treatment, he usually selects few acupoints and prefers Taixi (KI 3), he insists on stopping needling after the harmonious of qi and needling on alternative days; theoretically, ZHANG is versatile and full of learning, he follows the rule of yin and yang and adjusts his ways to cultivate the health; in his treatment, ZHANG considers the patients in diagnosis and treatment and combines the acupuncture with drugs; in teaching, he is strict and rigorous, on one hand, he is ruthless, but on the other hand, he is patient, demonstrating the sincere shining example of great doctors.