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MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate the expression of targets genes by binding to the 3′-untranslated regions. They play vital roles in diverse biological processes, including the development of hepatic fibrosis (HF). HF is characterized by the accumulation of extracellular matrix (ECM) and hepatic stellate cells (HSCs) are considered a major cell type for producing ECM. Alteration of the HSC phenotype plays a crucial role in the HF pathological process. MiRNAs involved in various biological process, such as differentiation, apoptosis, migration, and their relevant signaling pathways, are expressed in HSCs; however, emerging evidence indicates that numerous miRNAs are abnormally expressed in activated HSCs. In this review, we summarize the categorization of miRNAs in HF and describe the relationships among them. We also discuss miRNAs recently discovered to be related to HF, and attempt to find potential miRNAs that may serve as novel biomarkers for use in HF treatment.

Manmade high-performance polymers are typically non-biodegradable and derived from petroleum feedstock through energy intensive processes involving toxic solvents and byproducts. While engineered microbes have been used for renewable production of many small molecules, direct microbial synthesis of high-performance polymeric materials remains a major challenge. Here we engineer microbial production of megadalton muscle titin polymers yielding high-performance fibers that not only recapture highly desirable properties of natural titin (i.e., high damping capacity and mechanical recovery) but also exhibit high strength, toughness, and damping energy — outperforming many synthetic and natural polymers. Structural analyses and molecular modeling suggest these properties derive from unique inter-chain crystallization of folded immunoglobulin-like domains that resists inter-chain slippage while permitting intra-chain unfolding. These fibers have potential applications in areas from biomedicine to textiles, and the developed approach, coupled with the structure-function insights, promises to accelerate further innovation in microbial production of high-performance materials., Here, the authors engineer microbial production of muscle titin fibers with highly desirable mechanical properties and provide structural analyses that explain the molecular mechanisms underlying high performance of this polymer with potential uses in biomedicine and textile industries, among others.

Hawthorn (Crataegus pinnatifida Bunge. var. major) is an edible and medicinal fruit that is very common in food and traditional Chinese medicine. Corosolic acid (CA), a pentacyclic triterpenoid, which is an active component of hawthorn (Crataegus pinnatifida Bunge. var. major), has been exhibiting various pharmacological activities such as antidiabetic, antibacterial, anticancer, antiinflammatory, and antioxidant effects. The study aimed to evaluate the effect of CA on non-alcoholic steatohepatitis (NASH) in mice induced by 60 kcal% high-fat diet (HFD) and carbon tetrachloride (CCl4 ). CA lowered liver index and serum AST, ALT, TG, and TC levels compared to those in the model group. Histological analyses of the liver tissues of mice treated with CA revealed significantly decreased number of lipid droplets and alleviated inflammation and fibrosis. CA inhibited the transcripts of pro-fibrogenic markers (including α-SMA, collagen I, and TIMP-1) and the levels of pro-inflammatory cytokines (including TNF-α, IL-1β, caspase-1, and IL-6) associated with hepatic fibrosis, and NF-κB translocation and TGF-β1/Smad2 and AMPK pathways. In addition, CA reduced lipid accumulation via the regulation of AMPK and NF-κB activation in FFA-induced steatotic HepG2 cells. CA also decreased α-SMA, collagen I expressions, and Smad2 phosphorylation, which were reduced by TGF-β1 treatment in LX2 cells. Our results suggested that CA ameliorated NASH through regulating TGF-β1/Smad2, NF-κB, and AMPK signaling pathways, and CA could be developed as a potential health functional food or therapeutic agent for NASH patients.

The gut microbiota is closely associated with colorectal neoplasia. While most metagenomics studies utilized fecal samples, circulating bacterial DNA in colorectal neoplasia patients remained unexplored. This proof-of-concept study aims to characterize alterations of circulating bacterial DNA in colorectal neoplasia patients. We performed WGS of plasma samples from 25 colorectal cancer (CRC) patients, 10 colorectal adenoma (CRA) patients and 22 healthy controls (HC). Bacterial relative abundance was measured by removing the host genome and mapping reads into bacterial genomes. By diversity analysis, we found plasma samples required less sample size to approach saturation than fecal samples, and species diversity in HC was slightly higher compared with CRC/CRA patients. The majority of circulating bacterial DNA came from bacterial genera which commonly associated with gastrointestine and oral tract. By differential analysis, a total of 127 significant species between CRC patients and HC were identified, on which basis 28 species with top predictive ability were selected and showed promise in preliminary discrimination between CRC/CRA and HC. In CRA patients, relative abundance of the selected 28 species more closely resembled those in CRC patients than HC. By comparing with fecal metagenomics studies, we found there was moderate positive correlation between fold changes of the overlapped fecal and circulating bacterial DNA. Finally, species correlation analysis revealed that CRC and HC displayed distinct patterns of species association. In conclusion, this study demonstrated alterations of circulating bacterial DNA in colorectal neoplasia patients, which had the potential to become non-invasive biomarkers for colorectal neoplasia screening and early diagnosis.

The essential human O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) is the sole enzyme responsible for modifying thousands of intracellular proteins with the monosaccharide O-GlcNAc. This unique modification plays crucial roles in human health and disease, but the substrate recognition of OGT remains poorly understood. Intriguingly, the only human enzyme reported to remove this modification, O-GlcNAcase (OGA), is O-GlcNAc modified. Here, we exploited a GlcNAc electrophilic probe (GEP1A) to rapidly screen OGT mutants in a fluorescence assay that can discriminate between altered OGT-sugar and -protein substrate binding to help elucidate the binding mode of OGT toward OGA protein substrate. Since OGT tetratricopeptide repeat (TPR) domain plays a key role in OGT-OGA binding, we screened 30 OGT TPR mutants, which revealed 15 "ladder like" asparagine or aspartate residues spanning TPRs 3-7 and 10-13.5 that affect OGA O-GlcNAcylation. By applying a truncated OGA construct, we found that OGA's N-terminal region or pseudo histone acetyltransferase domain is not required for its O-GlcNAcylation, suggesting OGT functionally interacts with OGA through its catalytic and/or stalk domains. This work represents the first effort to systemically investigate each OGT TPR and our findings will facilitate the development of new strategies to investigate the role of substrate-specific O-GlcNAcylation.

ALKBH5 is the main enzyme for m6A-based demethylation of RNAs and it has been implicated in many biological and pathophysiological processes. Here, we aimed to explore the potential involvement of ALKBH5 in osteosarcoma and decipher the underlying cellular/molecular mechanisms. We discovered downregulated levels of demethylase ALKBH5 were correlated with increased m6A methylation in osteosarcoma cells/tissues compared with normal osteoblasts cells/tissues. ALKBH5 overexpression significantly suppressed osteosarcoma cell growth, migration, invasion, and trigged cell apoptosis. In contrast, inhibition of ALKBH5 produced the opposite effects. Whereas ALKBH5 silence enhanced m6A methylations of pre-miR-181b-1 and YAP-mRNA exerting oncogenic functions in osteosarcoma. Moreover, upregulation of YAP or downregulation of mature miR-181b-5p displayed a remarkable attenuation of anti-tumor activities caused by ALKBH5. Further results revealed that m6A methylated pre-miR-181b-1 was subsequently recognized by m6A-binding protein YTHDF2 to mediate RNA degradation. However, methylated YAP transcripts were recognized by YTHDF1 to promote its translation. Therefore, ALKBH5-based m6A demethylation suppressed osteosarcoma cancer progression through m6A-based direct/indirect regulation of YAP. Thus, ALKBH5 overexpression might be considered a new approach of replacement therapy for osteosarcoma treatment.

Myotonic dystrophy type 1 (DM1) is the most common type of adult muscular dystrophy caused by the expanded triple-nucleotides (CUG) repeats. Myoblast in DM1 displayed many defects, including defective myoblast differentiation, ribonuclear foci, and aberrant alternative splicing. Despite many were revealed to function in DM1, microRNAs that regulated DM1 via directly targeting the expanded CUG repeats were rarely reported. Here we discovered that miR-322/-503 rescued myoblast defects in DM1 cell model by targeting the expanded CUG repeats. First, we studied the function of miR-322/-503 in normal C2C12 myoblast cells. Downregulation of miR-322/-503 significantly hindered the myoblast differentiation, while miR-322/-503 overexpression promoted the process. Next, we examined the role of miR-322/-503 in the DM1 C2C12 cell model. miR-322/-503 was downregulated in the differentiation of DM1 C2C12 cells. When we introduced ectopic miR-322/-503 expression into DM1 C2C12 cells, myoblast defects were almost fully rescued, marked by significant improvements of myoblast differentiation and repressions of ribonuclear foci formation and aberrant alternative splicing. Then we investigated the downstream mechanism of miR-322/-503 in DM1. Agreeing with our previous work, Celf1 was proven to be miR-322/-503′s target. Celf1 knockdown partially reproduced miR-322/-503′s function in rescuing DM1 C2C12 differentiation but was unable to repress ribonuclear foci, suggesting other targets of miR-322/-503 existed in the DM1 C2C12 cells. As the seed regions of miR-322 and miR-503 were complementary to the CUG repeats, we hypothesized that the CUG repeats were the target of miR-322/-503. Through expression tests, reporter assays, and colocalization staining, miR-322/-503 was proved to directly and specifically target the expanded CUG repeats in the DM1 cell model rather than the shorter ones in normal cells. Those results implied a potential therapeutic function of miR-322/-503 on DM1, which needed further investigations in the future.

This study aimed to evaluate the feasibility of a wait-and-see strategy for non-small cell lung cancer (NSCLC) patients with special pleural dissemination lesions (r-pM1a and s-pM1a). Furthermore, the study characterized genomic alternations about disease progression. For this study, 131 NSCLC patients with a diagnosis of pM1a were retrospectively selected. Survival differences were evaluated among patients treated with three different initial postoperative treatments: chemotherapy, targeted therapy, and wait-and-see strategy. Whole-exome sequencing (WES) was performed on primary and metastatic tumors of 10 patients with dramatic progression and 13 patients with gradual progression. The wait-and-see group showed better progression-free survival (PFS) than the chemotherapy group (p

The incidence of melanoma is rising globally including China. Comparing to Caucasians, the incidence of non-cutaneous melanomas is significantly higher in Chinese. Herein, we performed genomic profiling of 89 Chinese surgically resected primary melanomas, including acral (n = 54), cutaneous (n = 22), and mucosal (n = 13), by hybrid capture-based next-generation sequencing. We show that mucosal melanomas tended to harbor more pathogenic mutations than other types of melanoma, though the biological significance of this finding remains uncertain. Chromosomal arm-level alterations including 6q, 9p, and 10p/q loss were highly recurrent in all subtypes, but mucosal melanoma was significantly associated with increased genomic instability. Importantly, 7p gain significantly correlated with unfavorable clinical outcomes in non-cutaneous melanomas, representing an intriguing prognostic biomarker of those subtypes. Furthermore, focal amplification of 4q12 (KIT, KDR, and PDGFRα) and RAD51 deletion were more abundant in mucosal melanoma, while NOTCH2 amplification was enriched in acral melanoma. Additionally, cutaneous melanomas had higher mutation load than acral melanomas, while mucosal melanomas did not differ from other subtypes in mutation burden. Together, our data revealed important features of acral and mucosal melanomas in Chinese including distinctive driver mutation pattern and increased genomic instability. These findings highlight the possibilities of combination therapies in the clinical management of melanoma.

Atopic dermatitis (AD) is a recurrent chronic inflammatory skin disease affecting up to 30% of the children population, and immuno-regulatory therapy that could modify the course of disease is urgently needed. Probiotics have demonstrated therapeutic effects on AD and could potentially regulate the disease process. However, the efficacy of probiotics for AD is inconsistent among different studies, which is mainly due to the elusive mechanism and different species and (or) strains used. In this study, we designed a mixture of five strains of probiotics (named IW5) and analyzed the effect and mechanism of IW5 on calcipotriol (MC903)-induced AD-like dermatitis. We found that IW5 significantly alleviated skin inflammation of the MC903-induced AD in mice. Administration with IW5 induced increased production of regulatory T cells and regulatory dendritic cells (DCregs) in the mesenteric lymph nodes. We also found that the diversity of the gut microbiota in the mice with MC903-induced dermatitis was increased after IW5 administration, and the level of butyrate in the gut was elevated. In cell culture, butyrate induced the production of DCregs. Our study revealed the therapeutic effects of a newly designed probiotics mixture and uncovered a possible mechanism, providing a foundation for future clinical studies.

Liver fibrosis is a reversible wound healing reaction characterized by abnormal accumulation of extracellular matrix (ECM) in response to liver injury. Recent studies have shown that it can be epigenetically regulated, especially by microRNAs (miRNAs). It has been acknowledged that activation of hepatic stellate cells (HSCs) is a pivotal step in the initiation and progression of liver fibrosis. Notably, our results showed that miR-195-3p was increased in HSCs isolated from CCl4-treated mice and that the increase was more pronounced as the degree of liver fibrosis increased. Moreover, treatment of LX-2 cells, a human immortalized hepatic stellate cell line, with TGF-β1 resulted remarkable upregulation of miR-195-3p. Gain-of-function and loss-of-function experiments have suggested that the increased levels of miR-195-3p inhibit the expression of phosphatase and tension homolog deleted on chromosome 10 (PTEN), a negative regulator of the PI3K/Akt/mTOR signaling pathway in liver fibrosis, thereby contributing to HSC activation and proliferation and promoting the expression of profibrotic genes, such as α-SMA and collagen I, in LX-2 cells, which accelerates the accumulation of fibrous extracellular matrix deposition in the liver, while knockdown of miR-195-3p induced the opposite effect. Taken together, these results provide evidence for the harmful role of miR-195-3p in CCl4-treated mouse liver fibrosis.

Summary N-Methyl-N-nitrosourea, an N-nitroso compound converted from dietary nitrite by Helicobacter pylori, causes somatic mutations in epithelial cells and induces gastric premalignancy. Here, we describe a detailed protocol for induction of gastric tumor and analysis of tumor phenotypes in mice. This model can be widely used for studying the initiation and growth of gastric cancer. For complete details on the use and execution of this protocol, please refer to Li et al. (2021)., Graphical abstract, Highlights • Preparation of N-methyl-N-nitrosourea (MNU) solution and Helicobacter pylori • Establishment of a murine gastric tumor model with MNU • Summary of development of gastric tumors and tumors in other organs, N-Methyl-N-nitrosourea, an N-nitroso compound converted from dietary nitrite by Helicobacter pylori, causes somatic mutations in epithelial cells and induces gastric premalignancy. Here, we describe a detailed protocol for induction of gastric tumor and analysis of tumor phenotypes in mice. This model can be widely used for studying the initiation and growth of gastric cancer.

Myotonic dystrophy type 1 (DM1) is an inherited neuromuscular disease caused by expanded CTG repeats in the 3′ untranslated region (3′UTR) of the DMPK gene. The myogenesis process is defective in DM1, which is closely associated with progressive muscle weakness and wasting. Despite many proposed explanations for the myogenesis defects in DM1, the underlying mechanism and the involvement of the extracellular microenvironment remained unknown. Here, we constructed a DM1 myoblast cell model and reproduced the myogenesis defects. By RNA sequencing (RNA-seq), we discovered that periostin (Postn) was the most significantly upregulated gene in DM1 myogenesis compared with normal controls. This difference in Postn was confirmed by real-time quantitative PCR (RT-qPCR) and western blotting. Moreover, Postn was found to be significantly upregulated in skeletal muscle and myoblasts of DM1 patients. Next, we knocked down Postn using a short hairpin RNA (shRNA) in DM1 myoblast cells and found that the myogenesis defects in the DM1 group were successfully rescued, as evidenced by increases in the myotube area, the fusion index, and the expression of myogenesis regulatory genes. Similarly, Postn knockdown in normal myoblast cells enhanced myogenesis. As POSTN is a secreted protein, we treated the DM1 myoblast cells with a POSTN-neutralizing antibody and found that DM1 myogenesis defects were successfully rescued by POSTN neutralization. We also tested the myogenic ability of myoblasts in the skeletal muscle injury mouse model and found that Postn knockdown improved the myogenic ability of DM1 myoblasts. The activity of the TGF-β/Smad3 pathway was upregulated during DM1 myogenesis but repressed when inhibiting Postn with a Postn shRNA or a POSTN-neutralizing antibody, which suggested that the TGF-β/Smad3 pathway might mediate the function of Postn in DM1 myogenesis. These results suggest that Postn is a potential therapeutical target for the treatment of myogenesis defects in DM1.

The present research was to study the toxicology of artificially added Zn, Se and Sr in water. Specifically, we investigated the mortality and liver toxicity in zebrafish (Danio rerio), caused by different water concentrations of zinc sulfate (ZnSO4), sodium selenite (Na2SeO3), and strontium chloride hexahydrate (6H2O·SrCl2). Adult and embryo-larval zebrafish were used in the experiment. Analysis was performed of mortality, liver area and impermeability, delayed absorption area of the yolk sac, and liver tissue structure. The concentration change of sodium selenite exerted the most significant effect on the mortality of adult zebrafish, followed by that of strontium chloride hexahydrate, and zinc sulfate. Elevated strontium chloride hexahydrate concentration was associated with liver toxicity in zebrafish in the preliminary experiment. However, embryo-larval zebrafish were observed to die when the concentration of Zn2+ or Se4+ increased to a certain extent, without obvious liver toxicity. Our results indicated strontium chloride hexahydrate was hepatotoxic to embryo-larval zebrafish, which was manifested mainly as hepatomegaly and delayed absorption of the yolk sac. In addition, the artificially added strontium chloride hexahydrate destroyed liver tissue structure, resulting in hepatocyte enlargement, cell nucleus enlargement, blurred cytoplasmic boundaries, and formation of a vacuolar liver. These findings suggest the amount of strontium chloride hexahydrate added in soft drinks should be limited to certain levels.

Populations vary in their degree of ecological specialization. An intuitive, but often untested, hypothesis is that populations evolving under greater environmental heterogeneity will evolve to be less specialized. How important is environmental heterogeneity in explaining among-population variation in specialization? We assessed juvenile viability of 20 Drosophila melanogaster populations evolving under one of four regimes: (1) a salt-enriched environment, (2) a cadmium-enriched environment, (3) a temporally varying environment, and (4) a spatially varying environment. Juvenile viability was tested in both the original selective environments and a set of novel environments. In both the original and novel environments, populations from the constant cadmium regime had the lowest average viability and the highest variance in viability across environments but populations from the other three regimes were similar. Our results suggest that variation in specialization among these populations is most simply explained as a pleiotropic by-product of adaptation to specific environments rather than resulting from a history of exposure to environmental heterogeneity.

Background Fava beans (FBs) have long been used as food, and their principal disadvantage is derived from their haemotoxicity. We hypothesized that FB ingestion alters the intestinal gene expression pattern, thereby inducing an immune response. Results In-depth sequence analysis identified 769 differentially expressed genes (DEGs) associated with the intestine in FB-treated DBA/1 mouse intestines. The identified genes were shown to be associated with biological processes (such as response to stimulus and immune system processes), human disease pathways (such as infectious diseases, endocrine and metabolic diseases, and immune diseases), and organismal system pathways (such as the digestive system, endocrine system, environmental adaptation, and immune system). Moreover, plasma total immunoglobulin E (IgE), histamine, interleukin (IL)-4 and IL-13 levels were significantly increased when the mice were treated with FBs. Conclusions These results demonstrated that FBs affect the intestinal immune response and IgE and cytokine secretion in DBA/1 mice. Electronic supplementary material The online version of this article (10.1186/s40659-019-0216-9) contains supplementary material, which is available to authorized users.

Objective Neuroendocrine neoplasms (NENs) are relatively rare and heterogeneous malignancies with two major subtypes: low-grade neuroendocrine tumor (NET) and high-grade neuroendocrine carcinoma (NEC). Comprehensive molecular characterization of NENs is needed to refine our understanding of the biological underpinnings of different NEN subtypes and to predict disease progression more accurately. Methods We performed whole-exome sequencing (WES) of NEN samples from 49 patients (25 NETs and 24 NECs) arising from the stomach, intestines or lung. Clinicopathologic features were assessed and associated with molecular events. Results NENs generally harbor a low mutation burden, with TP53 being the top mutated gene found in 31% of patients. Consistent with other studies, p53 signaling pathway dysfunction is significantly enriched in NECs compared to NETs (P

O-GlcNAc transferase (OGT) glycosylates numerous proteins and is implicated in many diseases. To date, most OGT inhibitors lack either sufficient potency or characterized specificity in cells. We report the first targeted covalent inhibitor that predominantly reacts with OGT but does not affect other functionally similar enzymes. This study provides a new strategy to interrogate cellular OGT functions and to investigate other glycosyltransferases.

Chlorella pyrenoidosa was exposed to nonylphenol (NP) to investigate the tolerance, antioxidant response, removal efficiency, and biodegradation mechanism. We conducted studies on algal biomass, chlorophyll a content, and photosynthetic activity, and found that C. pyrenoidosa exhibited a high tolerance even at 8 mg L−1 of NP. Changes in peroxidase (POD) and superoxide dismutase (SOD) activities indicated that the NP-induced oxidative stress caused oxidant damage, which increased the malondialdehyde (MDA) content. After culturing for 120 h, the NP removal efficiency of C. pyrenoidosa was 89%, 59%, 49%, and 48% in the 2, 4, 6, and 8 mg L−1 treatment groups, respectively. Degradation intermediates determined by GC–MS suggested that the biodegradation of NP in C. pyrenoidosa originated from the long alkyl chain. In addition, transcriptome analysis indicated that NP affected photosynthesis, antioxidase, and oxidoreductase activity-related genes. In summary, our results indicated that C. pyrenoidosa is a species that exhibits high tolerance and biodegradation capacity toward NP.

Summary: mTOR, the sensor of nutrients and growth factors, has important roles in tissue homeostasis and tumorigenesis. However, how mTOR controls gastric epithelial cell turnover and gastric cancer development, a leading malignancy, remains poorly understood. Here, we provide genetic evidence that mTOR activation promotes proliferation and inhibits differentiation of Lgr5+ gastric epithelial progenitors (GEPs) in gastric homeostasis and tumorigenesis. mTOR signaling increases MEK1 and Smad1 expression and enhances activation of MEK1-ERKs and BMP-Smad1 pathways, respectively, in GEPs and gastric tumors. Mek1 deletion or inhibition rescues hyperproliferation, whereas Bmpr1a ablation or inhibition rescues differentiation defects of Tsc1−/− GEPs. Tsc1 deficiency in Lgr5+ GEPs accelerates gastric tumor initiation and development, which require MEK1-ERKs for hyperplasia and BMP-Smad1 for differentiation suppression. These findings reveal how mTOR signaling controls Lgr5+ GEP homeostasis and cancerization and suggest that ERKs and Smad1 signaling can be safely targeted to substitute mTOR inhibitors in gastric cancer therapy.

BackgroundAnti-programmed death (PD)-1 therapy has recently been used in recurrent or metastatic (R/M) nasopharyngeal carcinoma (NPC). The long-term survival and its biomarkers responding to anti-PD-1 treatment in patients with R/M NPC remain unclear.MethodsPatients with R/M NPC were enrolled between March 2016 and January 2018 from two phase I clinical trials. The median follow-up period was 24.7 months. Eligible patients progressed on standard chemotherapy had measurable disease by Response Evaluation Criteria in Solid Tumor V.1.1. Non-obligatory contemporaneous tumor samples were collected for whole-exome sequencing. The primary outcome was objective response rate (ORR). Duration of response (DOR), progression-free survival (PFS), and overall survival (OS) were secondary outcomes assessed in all patients.ResultsAmong 124 evaluable patients, anti-PD-1 therapy achieved an ORR of 29.8% and a durable clinical benefit rate of 60.5%. The median OS (mOS) was 17.1 months (95% CI 14.2 to 24.7), median PFS (mPFS) was 3.8 months (95% CI 3.4 to 6.0), and median DOR was 9.5 months. Significant OS benefit from treatment was observed in patients without liver metastasis (23.8 vs 13.3 months, p=0.006). Copy number deletion in genes encoding granzyme B or granzyme H (GZMB/H) was associated with poor treatment outcome (mPFS altered vs wildtype: 1.7 vs 3.6 months, p=0.03; mOS altered vs wildtype: 10.1 vs 18 months, p=0.012).ConclusionsAnti-PD-1 treatment provided promising clinical benefit in pretreated patients with R/M NPC. Copy number loss in either GZMB or GZMH genes was associated with reduced survival.

In the most recent classification of Rutaceae, Amyridoideae is the largest and most diverse subfamily. In Amyridoideae, the genera Phellodendron, Tetradium, Toddalia and Zanthoxylum were proposed as “proto-Rutaceae”due to substantial phytochemical similarities. In this study, we investigated the plastome varia-tions in eight species representing these four genera and Melicope. All plastomes exhibited a typical quadripartite structure with four regions (LSC, SSC, IRa and IRb). The whole chloroplast genome size ranged from 158,383 bp to 159,014 bp and the gene number ranged from 115 to 116. By comparative analyses, we found that there were structural variations at the LSC/IR and SSC/IR borders of the plastomes in the five genera, especially in Melicope. Three most divergent regions (trnH-psbA, trnE-trnT and psaB) were found from the LSC region, which had great potential for developing effective genetic markers. In addition, we conducted a phylogenomic analyses of the “proto-Rutaceae”and related taxa with plastomes data from 36 species. Our results showed that (1) Phellodendron, Tetradium, Toddalia and Zanthoxylum were confirmed as close relatives and grouped together as the ‘proto-Rutaceae’, (2) Phellodendron was sister to Tetradium, and Toddalia was deeply nested within Zanthoxylum, and (3) Toddalia asiatica was closely related to the Southwest Pacific and East Asian species of Zanthoxylum, and Melicope pteleifolia was more closely related to Acronychia than it is to Tetradium. This study provided new insights into the plastome structural varia-tions in subfamily Amyridoideae, and demonstrated that the plastomes data were sufficiently robust to explore implications of the phylogeny for the previous phylogenetic hypotheses.

Histone deacetylases (HDACs), especially HDAC2, play a role in alleviating liver fibrosis; however, the specific upstream regulation mechanism is unknown. Herein, TargetScan was used to predict the potential upstream targets of HDAC2, and the role of miR-455-3p was explored. The dual luciferase reporter assay showed that miR-455-3p binds to the 3' UTR of HDAC2 mRNA. Additionally, miR-455-3p was downregulated in the liver tissues of patients with cirrhosis and mice with liver fibrosis, as well as in primary HSCs isolated from fibrotic mouse livers and TGF-β-treated LX-2 cells. In contrast, it is highly expressed in the reversal stage of hepatic fibrosis and MDI-cultured LX-2 cells. Our functional analyses showed that miR-455-3p overexpression facilitated apoptosis and reduced the expression of pro-fibrotic markers and the proliferation of activated LX-2 cells. On the contrary, miR-455-3p inhibition converted inactivated LX-2 cells into activated, proliferative, fibrogenic cells. Interestingly, restoration of HDAC2 expression partially blocked the function of miR-455-3p. Downregulated miR-455-3p expression can be restored by DNA methyltransferases in activated LX-2 cells. Methylation-specific PCR, bisulfite sequencing PCR, and chromatin immunoprecipitation assays indicated that the methylation level of miR-455-3p promoter CpG islands was elevated in TGF-β-treated LX-2 cells and that miR-455-3p was downregulated in activated LX-2 cells by DNA hypermethylation, which is mediated by DNMT3b and DNMT1. In conclusion, miR-455-3p acts as a liver fibrosis suppressor by targeting HDAC2, and its deficiency further aggravates the reversal phase of fibrosis. Thus, the epigenetics mediated miR-455-3p/HDAC2 axis may serve as a novel potential therapeutic target for clinical treatment of hepatic fibrosis.

Gastric cancer is one of the most common and deadly cancer types. Currently, four subtypes have been identified with unique molecular alterations: Epstein–Barr virus (EBV)‐positive, microsatellite instability (MSI), chromosomal instability (CIN), and genomic stable (GS) tumors. Notably, many gastric tumors are associated with the bacterium Helicobacter pylori but the genomic landscape of this subgroup of tumors remains largely unknown. Targeted sequencing covering 425 genes was performed retrospectively on 1703 gastric tumor tissues and matched normal blood samples. Nonsynonymous mutations, copy‐number variation (CNV), and MSI status were called from human DNA reads; nonhuman DNA reads were mapped to NCBI microbial reference genome using Kraken and significant species were identified. Overall, 37 (2.76%) from a total of 1703 samples were EBV‐positive, 200 (11.74%) samples were H. pylori‐positive, and 10 samples were positive for both. Among the rest, 59 (3.46%) samples were MSI, 380 (22.31%) were CIN, and 1017 (59.72%) were GS. Most of the 200 H. pylori‐positive samples tend to be genome stable (85.5%, p, Previous group have found 4 molecular subtypes for gastric adenocarcinoma. We uncovered an additional subtype of H.pylori associated gastric cancer with distinct genomic profile.

Houttuynia cordata Thunb. (HCT) is a common vegetable native to southwest China, and grown for consumption. The results suggested that THg contents in all parts and MeHg in underground parts of HCT in Hg mining areas were much higher than those in non-Hg mining areas. The highest THg and MeHg content of HCT were found in the roots, followed by the other tissues in the sequence: roots > leaves > rhizomes > aboveground stems (THg), and roots > rhizomes > aboveground stems > leaves (MeHg). The average THg bioaccumulation factor (BCF) of HCT root in the Hg mining area and in non-Hg mining areas could reach 1.02 ± 0.71 and 0.99 ± 0.71 respectively, indicating that HCT is a Hg accumulator. And the THg and MeHg contents in all tissues of HCT, including the leaves, were significantly correlated with THg and MeHg content in the soil. Additionally, preferred dietary habits of HCT consumption could directly affect the Hg exposure risk. Consuming the aboveground parts (CAP) of HCT potentially poses a high THg exposure risk and consuming the underground parts (CUP) may lead to a relatively high MeHg exposure risk. Only consuming the rhizomes (OCR) of the underground parts could significantly reduce the exposure risk of THg and to some extent of MeHg. In summary, HCT should not be cultivated near the Hg contaminated sites, such as Hg tailings, as it is associated with a greater risk of Hg exposure and high root Hg levels, and the roots should be removed before consumption to reduce the Hg risk.

Background: The development of osteoporosis is often accompanied by autophagy disturbance, which also caused new osteoblasts defects from bone marrow mesenchymal stem cells (BMSCs). However, the underlying molecular mechanisms are still not fully understood. Methyltransferase-like 14 (METTL14) is the main enzyme for N6-methyladenosine (m6A), the most prevalent internal modification in mammalian mRNAs, and it has been implicated in many bioprocesses. We aimed to explore the biological functions of autophagy mediated by METTL14 in osteoporosis and the possibility of METTL14 as an effective target for the treatment of osteoporosis. Methods: Calcein staining and microcomputed tomography (μ-CT) were used to analyze the bone formation. immunohistochemical (IHC) staining was used to detect the expression of METTL14 and beclin-1 in bone tissues. The quantitative real time PCR (qRT-PCR) and western blotting were used to analyze the mRNA and protein expressions. Alizarin red staining (ARS) and alkaline phosphatase (ALP) staining were used to explore the osteogenic differentiation of BMSCs. The production of autophagy was determined by transmission electron microscopy (TEM) and mRFP-GFP-LC3 adenoviral. Moreover, RNA binding protein immunoprecipitation (RIP)-qPCR was used to investigate the m6A modification of beclin-1 and the binding capability of beclin-1 and IGF2BPs. Findings: Herein, we demonstrate that the expression of METTL14 is downregulated during the development of osteoporosis both in human and mice. In vivo, METTL14+/- knockdown mice exhibits elevated bone loss and impaired autophagy similar to the ovariectomized (OVX) mice, while injecting METTL14 overexpressing adenovirus delays the process of osteoporosis.In vitro, forced expression of METTL14 significantly facilitates the osteogenic differentiation ability of BMSCs through regulating the activation of autophagy by enhancing the expression of beclin-1 dependent on the m6 A modification; the opposite is true under METTL14 silencing condition. Subsequently, m6A-binding proteins IGF2BP1/2/3 recognize m6A methylated beclin-1 mRNA and promotes its translation via mediating RNA stabilization. Interpretation: Our study reveals the METTL14-IGF2BPs-beclin-1 signaling axis in BMSCs osteogenic differentiation and highlights the critical roles of METTL14 mediated m 6 A modification in osteoporosis. Funding: This work was supported by grants from the National Natural Science Fund of China (81972117), Natural Science Foundation of Heilongjiang Province of China for excellent youth (JJ2020JQ0004), The First Affiliated Hospital of Harbin Medical University Excellent Young Talents Funding (HYD2020JQ0013), Special fund for clinical research of Wu Jieping Medical Foundation (320.6750.2020-04-50), and CAMS Innovation Fund for Medical Sciences (CIFMS, 2020-I2M-5-003). Declaration of Interest: The authors declare that they have no competing interests. Date Availability Ethical Approval: The experiments were performed under the guidance of the Experimental Animal Ethic Committee of the Harbin Medical University.

Recently, a new subgroup of T cells, named peripheral helper T (Tph) cells, has been implicated in autoimmune pathogenesis. An imbalance of Tph cell subsets influences the severity of immune-related diseases. However, the characteristics and roles of Tph cell subsets in psoriasis remain unknown. Programmed cell death 1-positive, chemokine C-X-C receptor (CXCR) 5-negative Tph cells can be divided into 3 subgroups based on differential expression of chemokine CXCR3 and chemokine C-C receptor (CCR) 6. CXCR3+CCR6- Tph cells are classified as Tph1, CXCR3-CCR6- Tph cells are classified as Tph2, and CXCR3-CCR6+ Tph cells are classified as Tph17. In this study, conditions of circulating Tph cell subsets and CD4+CXCR5+ follicular helper T (Tfh) cells in 27 patients with psoriasis and 13 healthy individuals were detected by flow cytometry. The level of plasma chemokine C-X-C ligand (CXCL) 13 was measured by enzyme-linked immunosorbent assay. The correlations between the above indexes and disease severity were explored. In the peripheral blood of patients with psoriasis, Tph17 cells had an activated, proliferative phenotype; the quantity of the cells correlated with disease severity. Plasma CXCL13 levels were elevated in psoriasis and associated with disease severity and the frequency of Tph17 cells. CD4+CXCR5+ Tfh cells were increased in patients and positively correlated with disease severity, the frequency of Tph17 cells, and plasma CXCL13 levels. Our results suggest that Tph17 cells and the CXCL13/CXCR5 axis may be involved in the pathogenesis of psoriasis and represent new immunotherapeutic targets for treating psoriasis.

The carbon and oxygen isotopic composition of water and assimilates in plants reveals valuable information on plant responses to climatic conditions. Yet, the carbon and oxygen uptake, incorporation and allocation processes determining isotopic compositions are not fully understood. We carried out a dual-isotope labeling experiment at high humidity with 18O-enriched water (H218O) and 13C-enriched CO2 (13CO2) with attached Scots pine (Pinus sylvestris L.) branches and detached twigs of hemiparasitic mistletoes (Viscum album ssp. austriacum) in a naturally dry coniferous forest, where also a long-term irrigation takes place. After 4 h of label exposure, we sampled previous- and recent-year leaves, twig phloem and twig xylem over 192 h for the analysis of isotope ratios in water and assimilates. For both species, the uptake into leaf water and the incorporation of the 18O-label into leaf assimilates was not influenced by soil moisture, while the 13C-label incorporation into assimilates was significantly higher under irrigation compared with control dry conditions. Species-specific differences in leaf morphology or needle age did not affect 18O-label uptake into leaf water, but the incorporation of both tracers into assimilates was two times lower in mistletoe than in pine. The 18O-label allocation in water from pine needles to twig tissues was two times higher for phloem than for xylem under both soil moisture conditions. In contrast, the allocation of both tracers in pine assimilates were similar and not affected by soil moisture, twig tissue or needle age. Soil moisture effects on 13C-label but not on 18O-label incorporation into assimilates can be explained by the stomatal responses at high humidity, non-stomatal pathways for water and isotope exchange reactions. Our results suggest that non-photosynthetic 18O-incorporation processes may have masked prevalent photosynthetic processes. Thus, isotopic variation in leaf water could also be imprinted on assimilates when photosynthetic assimilation rates are low.

Pine Mistletoe (Viscum album ssp. austriacum) is a hemi-parasite shrub species, whose survival and development rely on water and mineral resources obtained through the xylem sap from the host tree. Mistletoe can produce photosynthates in its green organs on its own. On the other side, as they are connected to the phloem of the host tree, they may also be able to retrieve carbon assimilates from their host and/or vice versa. However, the dynamics and the steering factors of this relationship remains unclear. We conducted 13C- labelling experiments with mature Scots pine (Pinus sylvestris) infected by mistletoes in a long-term (15 years) irrigation experiment in Switzerland (Pfynwald, Valais) to investigate the transport of carbon assimilates and nutrients between the host and the parasite under different soil moisture (600 mm vs 1200 mm of precipitation per year). Three irrigated and three control (natural xeric) pine trees infected by mistletoes were 13C labelled for 4 hours. During the 13C labelling of the trees, the following two experiments were carried out. 1) Wrapping experiment: 3-4 clusters of mistletoes on the labelled trees were wrapped with aluminium foil and enclosed in plastic bags to prevent mistletoe photosynthesis using 13C-enriched CO2 and to investigate a potential host-mistletoe carbon transfer. 2) Girdling and removal experiment: The phloem of 12 host tree branches (6 control & 6 irrigated) infected by mistletoes was manually removed (ca. 2 cm in width, basipetally of the mistletoes) to stop the downward transport of photosynthates from the girdled branches. Additionally, host needles or mistletoes were removed from the girdled branches to investigate the respective contribution of photosynthesis by the host needles vs. mistletoes to the host branch carbon balance. In the wrapping experiment, wrapped and unwrapped mistletoe leaves and stems were harvested at 10 times points over 6 months. In the girdling and removal experiment, needles, twigs (i.e. xylem and phloem) and mistletoe leaves were harvested at 7 time points over 14 days. In both experiment, bulk organic matter of each tissue was analysed to trace the 13C signal. We found that there was no 13C signal in the wrapped mistletoe leaves, indicating that there was no C-transfer from the host to the mistletoes via the phloem sap. Mistletoe removal from girdled branches decreased 13C-labelled carbon assimilates in host xylem and phloem. Meanwhile, when needles were completely removed from the girdled branches, host xylem and phloem were still able to acquire 13C from the mistletoes. These results suggest that mistletoes can support the host with carbon resources, which might be especially important when the host is C resource limited.Our study provides new insights into the relationship between the hemi-parasite and its host tree. Mistletoes may play a role as C source providers to support the host and maintain a symbiotic relationship to survive.

Osteosarcoma (OS) is a primary malignant bone tumor in early adolescence with high metastasis and death rates. Although the combination of polychemotherapy and surgical excision increased the survival rates up to 60%, the prognosis remains poor for most patients with metastatic or recurrent osteosarcoma. However, the exact pathogenic mechanism and pivotal elements regulating tumor invasion and metastasis are largely unknown. Circular RNAs (circRNAs) are novel endogenous non-coding RNA (ncRNA) molecules that generate the cyclic structure from back splicing. An increasing number of studies show that circRNAs can regulate transcriptional or posttranscriptional gene expression by acting as microRNA (miRNA) sponges and are involved in regulation of many important biological processes. The deregulation of some circRNAs was demonstrated in osteosarcoma. Furthermore, some circRNAs were identified to play essential roles in osteosarcoma occurrence, invasion, and metastasis. This review summarizes the regulatory effect of circRNAs in the occurrence and development of osteosarcoma, concentrating on deregulation, regulatory mechanisms, and functions of circRNAs and their potential value as biomarkers and therapy.

Long noncoding RNAs (lncRNAs) are critical gene expression regulators and are involved in several bone diseases. To explore the potential roles of lncRNAs in osteonecrosis of the femoral head (ONFH), we investigated for the first time the lncRNA expression profile of bone marrow mesenchymal stem cells (BMSCs) from patients with steroid-induced ONFH (SONFH) with microarray and bioinformatics analysis. A total of 1878 lncRNAs and 838 mRNAs were significantly up-regulated while 1842 lncRNAs and 1937 mRNAs were statistically down-regulated in the SONFH group compared with control group. The results validated by qRT-PCR were consistent with the microarray profiling data, especially involved in upregulation and downregulation of critical lncRNAs as well as mRNAs expression related to adipogenic and osteogenic differentiation. Pathway analyses revealed 40 signaling pathways with significant differences, especially the signaling pathways to regulate stem cell pluripotency. The CNC and ceRNA network indicated that lncRNA RP1-193H18.2, MALAT1 and HOTAIR were associated with abnormal osteogenic and adipogenic differentiation of BMSCs in the patients with SONFH. Our results suggest the lncRNA expression profiles were closely associated with the abnormal adipogenic and osteogenic transdifferentiation of BMSCs during the development of SONFH and explore a new insight into the molecular mechanisms of SONFH.

BACKGROUND The oxidative stress environment of pathological tissue has an adverse effect on the survival of bone marrow mesenchymal stem cells (BMSCs) transplantation. Ginkgo biloba L. extract (EGB) has a potent antioxidant effect. In this research, we assessed the protective effects of EGB and EGB-Containing Serum (EGB CS) on BMSCs against injury induced by hydrogen peroxide (H2O2). MATERIAL AND METHODS BMSCs were pretreated with EGB or EGB CS and treated with H2O2. The cell counting kit-8 (CCK-8) method was utilized to detect cell viability. The DCFH-DA Fluorescent Kit method was used to detect intracellular ROS level. Malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and (CAT) were determined. The Hoechst staining assay and qRT-PCR assay were utilized to evaluate the effect of EGB on cell apoptosis. Mitogen-activated protein kinases (MAPKs) signaling pathway were detected by western blot analysis. RESULTS Compared to the H2O2 group, the number of apoptotic cells in the EGB and EGB CS pretreated groups significantly decreased. The mRNA expression ratio of Bax/Bcl-2 was also decreased. EGB and EGB CS can reduce the production of ROS in BMSCs exposed to H2O2. SOD, GSH-Px and CAT activities were significantly higher compared with those with H2O2 group. Furthermore, EGB or EGB CS pretreatment decreased the protein levels of p-p38MAPK and p-JNK in BMSCs compared to the H2O2 group. CONCLUSIONS Our findings suggested that EGB and EGB CS have protective effect on BMSCs against oxidative stress injury and increase the survival rate of BMSCs transplantation by regulating p38MAPK and JNK signaling.

BACKGROUND Steroid-induced osteonecrosis of the femoral head (SONFH) is a common orthopedic disease associated with the application of glucocorticoid (GC). In this study, we detected the microRNAs (miRNAs) differentially expressed in bone marrow mesenchymal stem cells (BMSCs) from SONFH patients, and target gene predictions were performed, and the functions of the target genes was verified. MATERIAL AND METHODS BMSCs collected from patients with SONFH and femoral neck fracture (FNF) constituted the SONFH group (n=3) and FNF (control) group (n=3), respectively. MiRNA microarray analysis was utilized to detect the differentially expressed miRNAs, and quantitative real-time polymerase chain reaction (qRT-PCR) was used to verify the microarray results. The target genes and functions of the differentially expressed miRNAs were analyzed using a bioinformatics database. RESULTS The microarray results revealed that compared with the control group, 22 miRNAs were identified differentially expressed in the SONFH group, with 17 upregulated and 5 downregulated. Further qRT-PCR validation of differentially expressed miRNAs confirmed that hsa-miR-601, hsa-miR-452-3p, hsa-miR-647, and hsa-miR-516b-5p were significantly increased, whereas hsa-miR-122-3p was significantly decreased. During osteogenic differentiation, hsa-miR-601, hsa-miR-452-3p, hsa-miR-647, hsa-miR-516b-5p, and hsa-miR-127-5p were significantly downregulated, whereas hsa-miR-122-3p was significantly upregulated, and miRNAs showed a converse tendency during adipogenic differentiation. CONCLUSIONS Six miRNAs associated with osteogenic and adipogenic differentiation were identified differentially expressed in the BMSCs of SONFH patients; these miRNAs may serve as novel biomarkers or therapeutic targets for SONFH.

Defoliation by herbivores commonly imposes negative effects on plants, and physiological integration (resource sharing) can enhance the ability of guerilla clonal plants to tolerate stresses. Here we examined whether physiological integration can increase the ability of phalanx clonal plants to withstand defoliation. On a high mountain grassland in southwestern China, we subjected the phalanx clonal plant Iris delavayi within 10 cm × 10 cm plots to three levels of defoliation intensity, i.e., control (no defoliation), moderate (50% shoot removal to simulate moderate herbivory) and heavy defoliation (100% shoot removal to simulate heavy herbivory), and kept rhizomes at the plot edges connected (allowing physiological integration) or disconnected (preventing integration) with intact ramets outside the plots. Defoliation significantly reduced leaf biomass, root biomass and ramet number of I. delavayi . Clonal integration did not affect the growth of I. delavayi under control, but significantly increased total biomass, rhizome and root biomass under heavy defoliation, and leaf biomass and ramet number under moderate defoliation. We conclude that clonal integration associated with resource reallocation plays an important role in maintaining the productivity of the alpine and subalpine grassland ecosystems in SW China where clonal plants are a dominant component of the grasslands and are commonly extensively managed with moderate grazing intensity. Our results also help to better understand the adaption and tolerance of phalanx clonal plants subjected to long-term grazing in the high mountain environment.

The paper studied the absorption of Hg by five common herb species selected, Opuntia stricta , Aloe vera , Setcreasea purpurea , Chlorophytum comosum and Oxalis corniculata , in solution with different levels of mercury. We compared the accumulation of roots and shoots of the plants selected in medium containing different concentrations of mercury. The study was to explore what extent of mercury the five plant species were suitable for absorbing and transferring. The mercury amount uptake by five herb species was tested by CVAAS. The results demonstrated that the effect of different concentrations of mercury on the accumulation condition of roots was greater than that of shoots. There was an ideal Hg concentration for transfer by each plant species. Oxalis corniculata was the most suitable for transferring Hg and was more suitable for repairing soils with Hg at concentrations of less than 500 μg/L.

Polyacetylene glycosides, (6Z, 12E)-tetradecadiene-8,10-diyne-1-ol-3(R)-O-β-D-glucopyranoside (trivially named coreoside E) and (6Z, 12E)-tetradecadiene-8,10-diyne-1-ol-3(R)-O-β-L-arabinopyranosyl-(1 → 2)-β-D-glucopyranoside (trivially named coreoside F), were isolated from buds of Coreopsis tinctoria Nutt., together with one known compound, coreoside B. Their chemical structures were elucidated by extensive spectroscopic analysis and on the basis of their chemical reactivities. Coreoside E exhibited high levels of antimicrobial activity against Staphylococcus aureus and Bacillus anthracis with minimum inhibitory concentrations of 27 ± 0.27 and 18 ± 0.40 μM, respectively, whereas coreoside F and coreoside B showed weak antimicrobial activity against S. aureus and B. anthracis.

Positively charged drugs usually have enhanced water solubility, cellular uptake efficiency and anticancer activity. However, the common quaternized and protonated cationic photosensitizers both have some drawbacks such as needing potentially dangerous agents for preparation and easily being deprotonated in alkaline circumstance. Arginine is unique among the amino acids as its guanidine group has exceptionally high basicity in aqueous solution, which may make it positively charged in a wide range of pH. In this paper, two arginine substituted zinc phthalocyanines (ArgEZnPc and ArgZnPc) were reported. They can be positively charged in the range of pH 5–9. Moreover, the photobiological, photochemical properties, subcellular localization, and in vitro anticancer activities of the them were also carried out. The results show that ArgZnPc may be not a good photosensitizer because of its poor photobiological activities though it is positively charged in a wide range of pHs. This may be attributed to the formation of inner salts between guanidine and carboxyl groups of ArgZnPc, which weakens its photobiological and in vitro anticancer activity. While in contrast, ArgEZnPc shows preferential localization in the lysosomes of HeLa cells, exhibits high water solubility, excellent 1O2 and intracellular reactive oxygen species generation efficiency as well as high in vitro anticancer activity, making it a promising photosensitizer for photodynamic therapy.

The critical roles of IGFBP3 in regulating the osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells strongly indicate its potential effects on the pathogenesis of osteonecrosis of the femoral head (ONFH). In this study, we investigated the association of IGFBP3 gene polymorphism and its protein expression with the development of ONFH to further explore its molecular pathogenesis. Ligase detection reactions and enzyme-linked immunosorbent assay were used to detect the polymorphisms of rs2453839[C/T] and rs3110697[A/G] and the serum protein expression of IGFBP3 gene in 182 cases and 179 controls, respectively. The serum lipids level was also measured by automatic biochemistry analyzer. The results revealed that the recessive model of rs3110697 and the dominant model of rs2453839 were significantly associated with the increased risk of ONFH (p = 0.048, p = 0.047, respectively). The genotypes of rs2453839 were also significantly related to the clinical stages of ONFH (p = 0.017). More importantly, the serum protein expression of IGFBP3 and insulin-like growth factor 1 (IGF1) in the ONFH group were statistically increased compared with the control group (p = 0.044, p = 0.007). The serum triglyceride and low-density lipoprotein cholesterol level in the ONFH group were significantly higher than the control group (p = 0.01, p = 0.005, respectively), but the serum high-density lipoprotein cholesterol level of the ONFH group was dramatically lower than the control group (p = 0.0001). Our results showed that both the gene polymorphisms of IGFBP3 and the abnormal protein expression of serum IGFBP3 and IGF1 closely associated with the development of ONFH.

In most defoliation experiments with evergreen trees, leaves are removed proportionally across all leaf age classes or only the younger ones are removed. However, it remains unclear how the ecophysiological functions of older foliage potentially differ from those of younger leaves. Hence, we conducted a field experiment with five intensities of artificial defoliation (0%, 11% needle biomass loss = all ≥ 5-year-old needles removed, 27% loss = all ≥ 4-year-old needles removed, 46% loss = all ≥ 3-year-old needles removed, and 71% loss = all ≥ 2-year-old needles removed) of Abies alba trees grown under low-light (LL, 28% of full sunlight) and high-light conditions (HL, 70%). Photosynthesis of the remaining one-year-old needles; concentrations of non-structural carbohydrates (NSCs = soluble sugars + starch), total nitrogen (N), and soluble protein (SP) in stem sapwood and one-year-old needles; and tree height and stem radial growth rates were measured. We found that growth light conditions, rather than old needle defoliation up to a needle biomass loss of 70%, significantly affected growth, photosynthetic capacity and concentrations of tissue N, SP, and NSCs, with levels of all parameters greater in HL trees than in LL tress. Defoliation-induced compensatory growth was found in HL trees only, whereas tree height growth rates tended to decrease with increasing defoliation intensity in LL trees. Our results indicate that light availability, rather than damage to older foliage, plays an important role in determining plant physiology and growth. These results from A. alba trees suggest that the resource uptake by older foliage almost balances their resource consumption, leading to the conclusion that the older needles from evergreen trees physiologically contribute less to the whole-tree carbon and nitrogen balance. Our findings also suggest that the effects of older foliage defoliation on forest survival and productivity may have been overestimated in past forest ecosystem management practices, while the physiological functions of the youngest foliage may have been underestimated.

The ability of plants to tolerate freezing limits their geographical distribution. Therefore, winter warming may shift a species’ occurrence northwards and/or to higher altitudes. In Europe, the hemiparasitic vascular plant Viscum album (mistletoe) has two common and widespread subspecies: V. a. ssp. album and V. a. ssp. austriacum. The former has a more northern geographic distribution than the latter. Therefore we hypothesised that seeds of V. a. ssp. album are more tolerant to freezing than those of V. a. ssp. austriacum. From these two mistletoe subspecies V. a. ssp. austriacum is, in some managed forest areas, considered a novel threat to tree growth and forest health. Berries of V. a. ssp. album were collected from Sweden, Poland and Switzerland and berries of V. a. ssp. austriacum were collected from Poland, Switzerland and Spain. After storage at −3 °C, seeds were extracted from the pulp of berries and exposed to eight different temperatures between −8 °C and −30 °C, with the storage temperature serving as the control. After freezing treatments, germination of seeds was monitored. In addition, differential thermal analysis was used to measure freeze tolerance of seeds. The seeds of V. a. ssp. album tolerated lower temperatures than seeds of V. a. ssp. austriacum. The temperature at which 50% of seeds lost their ability to germinate (LT50) was −15 °C in V. a. ssp. austriacum and between −15 °C and −19 °C in V. a. ssp. album. The results of differential thermal analysis to determine the freezing point of seeds supported these findings. The freezing tolerance of mistletoe seeds was relatively well coupled with the winter climate at the edge of their current geographic distribution. Based on our results, the warming of winters may eliminate the abiotic barrier that has thus far limited mistletoes’ expansion, opening a window of opportunity for these parasites to increase their abundance and shift their distribution range towards higher latitudes and altitudes. Although mistletoes play important ecological roles in forest ecosystems, their recent increase has raised concern among forest managers, because they may cause a substantial reduction in tree growth in single species dominated stands. Increasing tree species diversity might be an effective method for limiting future mass infestations in homogeneous managed forests.

A novel esterase gene (estW) from soil bacterium Streptomyces lividans TK64 was successfully cloned using a pair of homologous primers. The estW gene encoded a protein (EstW) of 289 amino acid residues with a predicted molecular weight of 31.43 kDa. Sequence alignment revealed that EstW show relatively high levels of homology to other lipolytic enzymes characterized from Streptomyces and phylogenetic analysis suggested EstW belongs to the bacterial lipase/esterase family I. The estW gene was expressed at a high level in Escherichia coli and the recombinant enzyme was purified to homogeneity. The purified EstW was characterized via hydrolysis of various p-nitrophenyl esters and the best substrate was found to be p-nitrophenyl acetate (pNPA). Maximal activity of the recombinant protein was observed at pH 8.0 and 50 °C with pNPA as the substrate. The calculated activation energy (Ea ) of the esterase reaction was 9.12 kcal/mol. Half-life of EstW at 95 °C was approximately 12.5 H, making it the most thermostable esterase among all of the known lipolytic enzymes from Streptomyces, and the thermostability of EstW was similar to those of some enzymes characterized from the thermophilic bacteria. EstW exhibited relatively high tolerance to several detergents and required no cations for its maximal activity. The unique properties of EstW, namely its high thermostability and stability in the presence of organic solvents, may render it a potential candidate for industrial applications.

A series of novel benzimidazole quinolones as potential antimicrobial agents were designed and synthesized. Most of the prepared compounds exhibited good or even stronger antimicrobial activities in comparison with reference drugs. The most potent compound 15m was membrane active and did not trigger the development of resistance in bacteria. It not only inhibited the formation of biofilms but also disrupted the established Staphylococcus aureus and Escherichia coli biofilms. It was able to inhibit the relaxation activity of E. coli topoisomerase IV at 10 μM concentration. Moreover, this compound also showed low toxicity against mammalian cells. Molecular modeling and experimental investigation of compound 15m with DNA suggested that this compound could effectively bind with DNA to form a steady 15m-DNA complex which might further block DNA replication to exert the powerful bioactivities.

A series of novel N,N-bis(3-aminopropyl)methylamine-bridged bis-naphthalimide derivatives NI1–6 were designed and synthesized. Their cytotoxic activities against Hela, MCF-7, SGC-7901 and A549 cells were investigated. Compounds NI1–6 exhibited selectively cytotoxic activities in the tested cell lines and lower cytotoxic activities against MCF-7 cells were found except for compound NI1. NI1, the N-(2-hydroxyethyl)piperazine-modified naphthalimide, showed potent cytotoxic activity in the tested cell lines with IC50 values of 2.31, 2.94, 0.88 and 1.21 μM, respectively, better than the control drug (amonafide). Furthermore, its DNA binding properties, fluorescence imaging and collective apoptosis were investigated. Interestingly, NI1 as a DNA intercalator showed fluorescence enhancement upon binding with Ct-DNA and exhibited different impacts on the cell cycle compared with amonafide. Moreover, compound NI1 showed no significant hematoxicity and cardiotoxicity.

Background: Gastric cancer is one of the most common and deadly cancer types. Currently 4 subtypes have been identified with unique molecular alterations: Epstein-Barr virus (EBV)-positive, microsatellite instability (MSI), chromosomal instability (CIN), and genomic stable (GS) tumors. However, many gastric tumors are associated with the bacterium Helicobacter pylori but the genomic landscape of this subgroup of tumors remains largely unknown. Methods: Targeted-sequencing covering 425 genes were performed retrospectively on 1759 gastric tumor tissues and matched normal blood samples. The mean sequencing depth of tumors was 1159X. Nonsynonymous mutations, copy-number variation (CNV) and MSI status were called from human DNA reads; non-human DNA reads were mapped to NCBI microbial reference genome using Kraken and significant species were identified. Results: Overall, 37 (2.67%) samples were EBV-positive, 200 (11.37%) samples were H. pylori-positive, and 10 (0.57%) samples were positive for both. Other notable species identified in tumor samples were E. coli and C. acnes. No bacteria or viral reads were found in normal blood. Among the rest, 59 (3.35%) samples were MSI, 380 (21.6%) were CIN, and 1017 (57.82%) were GS. H. pylori-positive samples tend to be GS (85.5%, p < 0.001) and microsatellite stable (95%, p = 0.04). The most common mutations identified in H. pylori-positive tumors were TP53 (47%), CDH1 (12%) and ARID1A (8.5%). The top CNV events were amplification in MCL1 (14.5%), TERC (9%), CCNE1 (8.5%), PTPRD (8.5%), and MYC (7.5%). Compared to other GS tumors, mutation in AKT3 (p = 0.002) and amplification of TERC (p = 0.008) as well as MCL1 (p = 0.0.029) were significantly enriched in H. pylori-positive tumors. There was no difference in patients' sex or age among subtypes (p >= 0.05). Conclusion: This study characterized the genomic landscape of H. pylori positive gastric tumors using targeted-sequencing, which allowed direct comparison with other subtypes of gastric cancer. Identification of DNA reads from infectious agents in the tumor samples indicates that deep sequencing platform has promising utility in uncovering the characteristics of microbial environment of tumor. Co-infection between H. pylori and EBV occurred at a low frequency. H. pylori-positive tumors were GS and microsatellite stable with unique molecular features. Future study examining difference in treatment response among these subtypes of gastric cancer are warranted. Citation Format: Ao Wang, Hua Bao, Pengfei Hu, Yong Wu, Jinfeng Zhang, Xue Wu, Xiaonan Wang, Yang Shao. Distinct genomic profile in H. pylori associated gastric cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5439.

Coxsackievirus A6 (CV-A6) is an emerging pathogen associated with hand, foot, and mouth disease (HFMD). Its genetic characterization and pathogenic properties are largely unknown. Here, we report 39 circulating CV-A6 strains isolated in 2013 from HFMD patients in northeast China. Three major clusters of CV-A6 were identified and related to CV-A6, mostly from Shanghai, indicating that domestic CV-A6 strains were responsible for HFMD emerging in northeast China. Four full-length CV-A6 genomes representing each cluster were sequenced and analyzed further. Bootscanning tests indicated that all four CV-A6-Changchun strains were most likely recombinants between the CV-A6 prototype Gdula and prototype CV-A4 or CV-A4-related viruses, while the recombination pattern was related to, yet distinct from, the strains isolated from other regions of China. Furthermore, different CV-A6 strains showed different capabilities of viral replication, release, and pathogenesis in a mouse model. Further analyses indicated that viral protein 2C contributed to the diverse pathogenic abilities of CV-A6 by causing autophagy and inducing cell death. To our knowledge, this study is the first to report lethal and nonlethal strains of CV-A6 associated with HFMD. The 2C protein region may play a key role in the pathogenicity of CV-A6 strains. IMPORTANCE Hand, foot, and mouth disease (HFMD) is a major and persistent threat to infants and children. Besides the most common pathogens, such as enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16), other enteroviruses are increasingly contributing to HFMD. The present study focused on the recently emerged CV-A6 strain. We found that CV-A6 strains isolated in Changchun City in northeast China were associated with domestic origins. These Changchun viruses were novel recombinants of the CV-A6 prototype Gdula and CV-A4. Our results imply that measures to control CV-A6 transmission are urgently needed. Further analyses revealed differing pathogenicities in strains isolated in a neonatal mouse model. One of the possible causes has been narrowed down to the viral protein 2C, using phylogenetic studies, viral sequences, and direct tests on cultured human cells. Thus, the viral 2C protein is a promising target for antiviral drugs to prevent CV-A6-induced tissue damage.

Background Ovarian cancer is one of the most deadly gynecological malignancies and inclined to recurrence and drug resistance. Previous studies showed that the tumorigenesis of ovarian cancers and their major histotypes are associated with genomic instability caused by defined sets of pathogenic mutations. In contrast, the mechanism that influences the development of drug resistance and disease recurrence is not well elucidated. Solid tumors are prone to chromosomal instability (CIN) and micronuclei formation (MN). Although MN is traditionally regarded as the outcome of genomic instability, recent investigation on its origin and final consequences reveal that the abnormal DNA metabolism in MN is a driver force for some types of catastrophic genomic rearrangements, accelerating dramatic genetic variation of cancer cells. Methods We used Indirect Immunofluorescent staining to visualize micronuclei and activation of DNA repair factors in ovarian cancer cell lines and biopsies. Results We show that ovarian cancer cells are disposed to form micronuclei upon genotoxic insults. Double strand DNA breaks (DSBs)-triggered insurgence of micronuclei is associated with unrepaired chromosomes passing through mitosis. According to their morphology and DNA staining, micronuclei compartments are divided into early and late stages that can be further characterized by differential staining of γH2AX and 53BP1. We also show that MN compartments do not halt controlled DNA metabolism as sequestered nuclear repair factors are enriched at DNA breaks in MN compartments and efficiently process DNA ends to generate single-stranded DNA (ssDNA) structures. Interestingly, unknown factors are required for DNA end processing in MN in addition to the nuclear resection machinery. Finally, these hallmarks of micronuclei evolution depicted in cell culture were recapitulated in different stages of ovarian cancer biopsies. Conclusions In aggregate, our findings demonstrate that ovarian cancer cells are inclined to form micronuclei that undergo robust DNA metabolism and generate ssDNA structures, potentially destabilizing genomic structures and triggering genetic variation.

The present study investigated remediation of mercury-contaminated soils using Oxalis corniculata L. combined with various enhancers (sodium thiosulfate, ammonium thiosulfate, ethylenediaminetetraacetic acid and diethylenetriaminepentaacetic acid). The experiment was conducted using Oxalis corniculata seedlings planted in pots containing mercury loaded soils. Investigations included analysis of soil properties, plant growth conditions, ability of the plants to accumulate and extract mercury, and rhizosphere microorganism distribution. The maximal mercury content of the aerial parts and the mercury-translocation ratio of Oxalis corniculata treated with enhancers increased compared to Oxalis corniculata without enhancers. Compared with no enhancers, the theoretical reduction in phytoremediation time was about 50%, 25%, 20% and 21% when Oxalis corniculata was treated with sodium thiosulfate (Na2S2O3), ammonium thiosulfate ((NH4)2S2O3), ethylenediaminetetraacetic acid (EDTA) and diethylenetriaminepentaacetic acid (DTPA), respectively. The results indicated that the dominant species in rhizosphere soils varied with different enhancers. However, the evenness of background soils, rhizosphere soils of Oxalis corniculata, Oxalis corniculata treated with Na2S2O3, (NH4)2S2O3, EDTA and DTPA was not largely different at 0.62, 0.61, 0.57, 0.64, 0.61 and 0.63, respectively. These findings demonstrate that Oxalis corniculata treated with Na2S2O3 has the potential to recover and reclaim mercury-contaminated soils in pots.

The incidence of lung cancer and lung cancer-associated mortality have markedly increased worldwide, and gene-targeted therapy has emerged as a promising treatment strategy. The present study aimed to explore the targeted antitumor effect of the herpes simplex virus-thymidine kinase/human interleukin-12 (HSV-TK/hIL-12) fusion gene regulated by the human secretory leukocyte protease inhibitor (hSLPI) promoter of human non-small cell lung cancer (hNSCLC). There were four recombinant eukaryotic expression vectors: pcDNA3.1-CMV-TK, pcDNA3.1-CMV-TK/hIL-12, pcDNA3.1-phSLP-TK and pcDNA3.1-phSLP-TK/hIL-12. These were constructed and transfected into the A549, SPC-A1 and HepG2 cell lines in vitro. The expression of the HSV-TK/hIL-12 fusion gene was detected with reverse transcription-polymerase chain reaction (RT-PCR), and the content of hIL-12 was measured using an ELISA. The antitumor effect of the fusion gene on the A549, SPC-A1 and HepG2 cell lines was determined using an MTT assay. Analysis of the experimental data demonstrated that genes regulated by the cytomegalovirus promoter were expressed at the same level in three different tumor cell lines. Genes regulated by the hSLPI promoter were expressed in the A549 and SPC-A1 cell lines, but not in the HepG2 cell line. Coincidentally, the hIL-12 expression levels were similar to those observed in previous RT-PCR findings. In the Pcmv-TK/Pcmv-TK-hIL-12 group for all three cell lines, as well as in the PSLPI-TK/PSLPI-TK-hIL-12 group for the A549 and SPC-A1 cell lines, the cell survival rate declined significantly and the fusion gene transfection group indicated a lower cell survival rate, when compared with single gene transfection group. The present study indicated that the fusion gene regulated by the hSLPI promoter had a targeted antitumor effect on hNSCLC, and that the combined suicide gene and immune gene therapy had a stronger antitumor effect, compared with single gene therapy.

Remobilization is an important mechanism of resource conservation in plants. However, our understanding of whether the responses of resource remobilization to global warming differ between deciduous and evergreen trees remains unclear. We assessed resource remobilization from leaves to 1-year-old shoots in a deciduous (Betula ermanii) and an evergreen (Quercus aquifolioides) species along elevational gradients (i.e., temperature gradient) at the end of growing season. We aimed to test the hypotheses that the reallocation rate increased with increasing elevation and that more resources were reallocated from leaves to storage tissues in deciduous species than in evergreen species. We analyzed the concentrations of non-structural carbohydrates (NSC), nitrogen (N) and phosphorus (P), and compared the differences in remobilization efficiency of NSC, N, and P between leaves and shoots within each species and between the two species along the elevational gradients. Due to the different strategies of evergreen and deciduous species in nutrients use, the deciduous species had higher N and P remobilization rate, but lower remobilization rate of sugars, starch, and NSC than the evergreen species at the end of growing season. The remobilization rate of NSC, N, and P was significantly higher in trees at their upper limits compared to lower elevations. Our results suggest that trees reallocate resources from leaves to storage tissues before leaf senescence or at the end of growing season, to increase the resource use efficiency and to adapt to the harsh alpine environments. These results contribute to better understanding of the alpine treeline phenomenon in a changing world.

Elevation is a complex environmental factor altering temperature, light, moisture and soil nutrient availability, and thus may affect plant growth and physiology. Such effects of elevation may also depend on seasons. Along an elevational gradient of the Balang Mountain, southwestern China, we sampled soil and 2-year old leaves, 2-year old shoots, stem sapwood and fine roots (diameter