LncRNA 4930581F22Rik promotes myogenic differentiation by regulating the ERK/MAPK signaling pathway.

The skeletal muscle is the largest organ in mammals and is the primary motor function organ of the body. Our previous research has shown that long non-coding RNAs (lncRNAs) are significant in the epigenetic control of skeletal muscle development. Here, we observed progressive upregulation of lncRNA 4930581F22Rik expression during skeletal muscle differentiation. Knockdown of lncRNA 4930581F22Rik hindered skeletal muscle differentiation and resulted in the inhibition of the myogenic markers MyHC and MEF2C. Furthermore, we found that lncRNA 4930581F22Rik regulates myogenesis via the ERK/MAPK signaling pathway, and this effect could be attenuated by the ERK-specific inhibitor PD0325901. Additionally, in vivo mice injury model results revealed that lncRNA 4930581F22Rik is involved in skeletal muscle regeneration. These results establish a theoretical basis for understanding the contribution of lncRNAs in skeletal muscle development and regeneration.

Characterization of a novel γ-type carbonic anhydrase, Sjγ-CA2, in Saccharina japonica: Insights into carbon concentration mechanism in macroalgae.

Carbonic anhydrase (CA) is a crucial component of CO2-concentrating mechanism (CCM) in macroalgae. In Saccharina japonica, an important brown seaweed, 11 CAs, including 5 α-, 3 ß-, and 3 γ-CAs, have been documented. Among them, one α-CA and one ß-CA were localized in the periplasmic space, one α-CA was found in the chloroplast, and one γ-CA was situated in mitochondria. Notably, the known γ-CAs have predominantly been identified in mitochondria. In this study, we identified a chloroplastic γ-type CA, Sjγ-CA2, in S. japonica. Based on the reported amino acid sequence of Sjγ-CA2, the epitope peptide for monoclonal antibody production was selected as 165 Pro-305. After purification and specificity identification, anti-SjγCA2 monoclonal antibody was employed in immunogold electron microscopy. The results illustrated that Sjγ-CA2 was localized in the chloroplasts of both gametophytes and sporophytes of S. japonica. Subsequently, immunoprecipitation coupled with LC-MS/MS analysis revealed that Sjγ-CA2 mainly interacted with photosynthesis-related proteins. Moreover, the first 65 amino acids at N-terminal of Sjγ-CA2 was identified as the chloroplast transit peptide by the transient expression of GFP-SjγCA2 fused protein in tabacco. Real-time PCR results demonstrated an up-regulation of the transcription of Sjγ-CA2 gene in response to high CO2 concentration. These findings implied that Sjγ-CA2 might contribute to minimizing the leakage of CO2 from chloroplasts and help maintaining a high concentration of CO2 around Rubisco.

Optimal choice of stapler and digestive tract reconstruction method after distal gastrectomy for gastric cancer: A prospective case-control study.

BACKGROUND: Gastric cancer is the most common cause of cancer-related deaths, and is classified according to its location in the proximal, middle, or distal stomach. Surgical resection is the primary approach for treating gastric cancer. This prospective study aimed to determine the best reconstruction method after distal gastrectomy for gastric cancer. AIM: To explore the efficacy of different staplers and digestive tract reconstruction (DTR) methods after radical gastrectomy and their influence on prognosis. METHODS: Eighty-seven patients who underwent radical gastrectomy for distal gastric cancer at our institution between April 2017 and April 2020 were included in this study, with a follow-up period of 12-26 mo. The patients were assigned to four groups based on the stapler and DTR plan as follows: Billroth Ⅰ (B-I) reconstruction + linear stapler group (group A, 22 cases), B-I reconstruction + circular stapler group (group B, 22 cases), Billroth II (B-II) reconstruction + linear stapler group (group C, 22 cases), and B-II reconstruction + circular stapler group (group D, 21 cases). The pathological parameters, postoperative gastrointestinal function recovery, postoperative complications, and quality of life (QOL) were compared among the four groups. RESULTS: No significant differences in the maximum diameter of the gastric tumors, total number of lymph nodes dissected, drainage tube removal time, QLQ (QOL questionnaire)-C30 and QLQ-STO22 scores at 1 year postoperatively, and incidence of complications were observed among the four groups (P > 0.05). However, groups A and C (linear stapler) had significantly lower intraoperative blood loss and significantly shorter anastomosis time, operation time, first fluid diet intake time, first exhaust time, and length of postoperative hospital stay (P < 0.05) than groups B and D (circular stapler). CONCLUSION: Linear staplers offer several advantages for postoperative recovery. B-I and B-II reconstruction methods had similar effects on QOL. The optimal solution can be selected according to individual conditions and postoperative convenience.

The positive effects of postbiotic (SWF concentration®) supplemented diet on skin mucus, liver, gut health, the structure and function of gut microbiota of common carp (Cyprinus carpio) fed with high-fat diet.

Postbiotics are an emerging research interest in recent years, which shows that metabolites, lysate extracts, cell wall components and even culture supernatants of probiotics can also exhibit significant prebiotic effects. In this study postbiotic stress worry free concentration® (SWFC) were prepared from the composition of culture supernatant of Cetobacterium somerae and Lactococcus lactis. The positive effects of SWFC supplemented diets on the growth performance, skin mucus, liver and gut health, and intestinal microbiota profile of Cyprinus carpio fed with high fat diets were investigated. 180 C. carpio with an average body weight of (3.01 ± 0.01) g were selected and randomly divided into three groups. They were fed with one of the three experimental diets supplemented with SWFC of 0 (control), 0.2 and 0.3 g/kg for 98 days, afterwards indexes were detected. The results revealed that, addition of SWFC had no significant effect on growth performance of C. carpio, while it can improve the health of the fish remarkably. In addition, SWFC improved mucosal C3, T-AOC, SOD activities, and decreased lipid peroxidation product MDA level, which were notably better than those in the control group (P < 0.05). In terms of the liver health systems, C. carpio fed on the diet supplemented with 0.2 g/kg of SWFC, showed significant improvement of the liver injured by HFD and reduce the contents of serum ALT and AST, and liver TAG (P < 0.05; P < 0.01). The expression of inflammation-related and lipid synthesis genes revealed that SWFC0.2 group could noteworthy enhance antioxidant capacity, reduced the expression of pro-inflammatory factors (TNF-α, IL-1ß) and lipid synthesis genes (ACC, FAS, PPAR-ß, PPAR-γ), and up-regulated the expression of anti-inflammatory factors (TGF-ß). Additionally, intestinal morphology arose inflammatory cell infiltration, while intestinal integrity was better in SWFC groups compared with the control. Furthermore, the contents of serum LPS and LBP were remarkably lower in the SWFC0.2 group compared with the control (P < 0.01). The mRNA expression of genes related to gut health indicated that SWFC supplementation noteworthy up-regulated the expression of antioxidant (Nrf2, CAT, GPX), immune (Hepcidin, IL-10) and tight junction protein-related (ZO-1, Occludin). Simultaneously, the results of GF-zebrafish showed that the relative expression of anti-inflammatory genes (IL-1ß, TGF-ß) and antioxidant related genes (Nrf2, HO-1) were significantly up-regulated in SWFC groups. Data on intestinal microbiota profile verified that, at the phylum level, the abundance of Fusobacteria was remarkably elevated in the SWFC groups (P < 0.05), whereas the abundance of Firmicutes was declined noteworthy in SWFC0.2 and SWFC0.3 compared to the control group (P < 0.05; P < 0.01) respectively. At the genus level, the abundance of Cetobacterium in the SWFC groups were notably higher than those in the control group (P < 0.05), while the Vibrio content in the SWFC groups was significantly decreased (P < 0.05). PCoA result indicated that the intestinal microflora of SWFC0.2 group was abundant and diverse. Our results elucidate that dietary supplementation of SWFC protects C. carpio from HFD induced inflammatory response and oxidative stress, ameliorate skin mucus, liver and gut health, and improve the gut microbiota balance. Therefore, SWFC could be considered as an improving-fish-health additive, when supplemented to aquatic animal feed. With regards to how SWFC regulates the immunity and inflammatory responses and which signal transductions are involved remains unclear and more scientific evidences are needed to address these issues.

Nuclear-encoded CbbX located in chloroplast is essential for the activity of red-type Rubisco in Saccharina japonica.

Saccharina japonica (Laminariales, Phaeophyta) is a brown alga and the major component of algae beds on the northwest coast of the Pacific Ocean. Rubisco, the key enzyme of CO2 fixation in photosynthesis, is inhibited by nonproductive binding of its substrate RuBP and other sugar phosphates. The inhibited Rubisco in eukaryotic phytoplankton of the red plastid lineage was reactivated by CbbXs, the red-type Rubisco activases, through the process of ATP-hydrolysis-powered remodeling. As well documented, CbbXs had two types of subunits encoded by the plastid or nuclear genome respectively. In this study, both proteins of S. japonica (SjCbbX-n and SjCbbX-p) were localized in the chloroplast illustrated by immuno-electron microscopy technique. GST pull-down detection verified SjCbbX-n could interact with SjCbbX-p. Two-dimensional electrophoresis-based Western blot analysis illustrated that the endogenous SjCbbXs could form heterohexamer in the ratio of 1:1. Activase activity assays showed that although both the recombinant proteins of SjCbbXs were functional, SjCbbX-n illustrated the significantly higher activase activity than SjCbbX-p. Notably, when the two proteins were mixed, the highest specific efficiencies of Rubisco were obtained. These results implied SjCbbX-n may be essential for Rubisco activation. Molecular evolutionary analysis of cbbx genes revealed that cbbx-n originated from the duplication of cbbx-p and then evolved independently under the positive selection pressure. This is the first report about the functional relationship between the two types of CbbXs in macroalge with the red-type Rubisco and provides useful information for revealing the mechanism of high photosynthetic efficiency of this important kelp.

Progress in the Clinical Assessment and Treatment of Myocardial Depression in Critically Ill Patient with Sepsis.

Myocardial inhibition is the main cause of death in patients with sepsis.In recent years, methodological differences in the diagnosis, assessment, and treatment of septic myocardial depression have been observed, and how to objectively and accurately evaluate the degree of myocardial depression and the timing of treatment strategies have generally been the focus of this area of research. Based on the relevant research at home and abroad, the current review summarizes the clinical characteristics, methodological diagnosis, and symptomatic treatment of septic myocardial depression. The aim of doing so is to provide a reference for the early identification and treatment of patients with sepsis and myocardial depression.

Study about the combination strategy of Bacillus subtilis wt55 with AiiO-AIO6 to improve the resistance of zebrafish to Aeromonas veronii infection.

Disease problems will seriously restrict the sustainable development of aquaculture, and the environmental-friendly prevention strategies are urgently needed. Probiotics and quorum-quenching enzyme are innovative strategies to control bacterial diseases. Firstly, the bacteriostatic activity of Bacillus subtilis wt55 strain and quenching enzyme AiiO-AIO6 on the growth of Aeromonas veronii were tested in vitro, and the results showed wt55 inhibit the growth of A. veronii, but AiiO-AIO6 did not. Then, the synergistic effects of simple combination of B. subtilis wt55 and AiiO-AIO6 were evaluated next. The results showed this combination could improve the survival rate and significantly reduce the number of invasive A. veronii in gut after challenge compared to the other groups, corresponding to the lower intestinal alkaline phosphatase activity. One of its effect mechanisms is the combination could inhibit the growth of A. veronii in vitro; the other is direct immersion of germ-free zebrafish proved AiiO-AIO6 did not directly regulate the innate immune response of the host, but wt55 did it, and the simple combination group could significantly reduce the expression of nuclear factor kappa-B (NF-κB) and proinflammatory cytokine interleukin-1ß (IL-1ß), increase the expression of lysozyme gene; and the third is intestinal microbiota also plays a regulatory role: the gut microbiota from combination group could significantly inhibit the expression of IL-1ß and NF-κB, and increased the expression of transforming growth factor-ß (TGF-ß) and lysozyme. Given the effectiveness of this simple combination, a B. subtilis quorum-quenching recombinant expression strain in which AiiO-AIO6 was surface displayed on the spores and secreted by vegetative cells was built. The results showed that the survival rate after challenge was lower than that of the group treated with AiiO-AIO6 or wt55 alone, and the expression of proinflammatory cytokine IL-1ß and NF-κB were significantly higher. Our study demonstrated the effectiveness of B. subtilis and AiiO-AIO6 simple combination and established an efficient B. subtilis expression system.

Influences of dietary Eucommia ulmoides leaf extract on the hepatic lipid metabolism, inflammation response, intestinal antioxidant capacity, intestinal microbiota, and disease resistance of the channel catfish (Ictalurus punctatus).

The purpose of the study was to investigate the effects of Eucommia ulmoides leaf extract (ELE) on the common occurrence of liver steatosis, chronic inflammation, oxidative stress, disturbance of gut microbiota, and disease susceptibility in high-fat diet-fed channel catfish. Channel catfish fed three diets, including a high-fat diet (11% crude fat) and ELE-supplemented diets containing 1‰ or 2‰ ELE for 4 weeks. The results showed the contents of liver triacylglycerol of 1‰ and 2‰ ELE groups were reduced, and ELE treatments decreased the expression of lipogenesis related genes (srebp-1c, pparγ, and acc-1), and increased the expression of lipolysis related genes (pparα). In addition, the supplementation of ELE improved the inflammatory response of the liver and intestine. ELE could improve the destruction of intestinal morphology structure and increase the expression level of hif-1a and tight junction proteins (Occludin, Claudin2, Claudin15). 2‰ ELE significantly enhanced the antioxidant capacity of intestine by increasing the activity of SOD enzyme. Moreover, the supplement of ELE significantly increased the abundance of Cetobacterium and Romboutsia (p < 0.05). Compared with the control group, the expression of immune factor nf-κb had a significant decrease, and il-1ß showed a tendency to decrease in the ELE supplement groups after pathogenic bacteria challenge. In conclusion, the ELE alleviated fatty liver disease and inflammation response, improved the oxidative capacity and physiological structure of intestine, and improved the structure of intestinal microbiota and disease resistance in HFD-fed channel catfish.

Characterization of Long Non-coding RNAs Modified by m6A RNA Methylation in Skeletal Myogenesis.

Proper development of mammalian skeletal muscle relies on precise gene expression regulation. Our previous studies revealed that muscle development is regulated by both mRNA and long non-coding RNAs (lncRNAs). Accumulating evidence has demonstrated that N6-methyladenosine (m6A) plays important roles in various biological processes, making it essential to profile m6A modification on a transcriptome-wide scale in developing muscle. Patterns of m6A methylation in lncRNAs in developing muscle have not been uncovered. Here, we reveal differentially expressed lncRNAs and report temporal m6A methylation patterns in lncRNAs expressed in mouse myoblasts and myotubes by RNA-seq and methylated RNA immunoprecipitation (MeRIP) sequencing. Many lncRNAs exhibit temporal differential expression, and m6A-lncRNAs harbor the consensus m6A motif "DRACH" along lncRNA transcripts. Interestingly, we found that m6A methylation levels of lncRNAs are positively correlated with the transcript abundance of lncRNAs. Overexpression or knockdown of m6A methyltransferase METTL3 alters the expression levels of these lncRNAs. Furthermore, we highlight that the function of m6A genic lncRNAs might correlate to their nearby mRNAs. Our work reveals a fundamental expression reference of m6A-mediated epitranscriptomic modifications in lncRNAs that are temporally expressed in developing muscle.

Surface-Displayed Amuc_1100 From Akkermansia muciniphila on Lactococcus lactis ZHY1 Improves Hepatic Steatosis and Intestinal Health in High-Fat-Fed Zebrafish.

Fatty liver and intestinal barrier damage were widespread in most farmed fish, which severely restrict the development of aquaculture. Therefore, there was an urgent need to develop green feed additives to maintain host liver and intestinal health. In this study, a probiotic pili-like protein, Amuc_1100 (AM protein), was anchored to the surface of Lactococcus lactis ZHY1, and the effects of the recombinant bacteria AM-ZHY1 on liver fat accumulation and intestinal health were evaluated. Zebrafish were fed a basal diet, high-fat diet, and high-fat diet with AM-ZHY1 (108 cfu/g) or control bacteria ZHY1 for 4 weeks. Treatment with AM-ZHY1 significantly reduced hepatic steatosis in zebrafish. Quantitative PCR (qPCR) detection showed that the expression of the lipogenesis [peroxisome-proliferator-activated receptors (PPARγ), sterol regulatory element-binding proteins-1c (SREBP-1c), fatty acid synthase (FAS), and acetyl-CoA carboxylase 1 (ACC1)] and lipid transport genes (CD36 and FABP6) in the liver were significantly downregulated (p < 0.05), indicating that AM-ZHY1 could reduce liver fat accumulation by inhibiting lipid synthesis and absorption. Moreover, supplementing AM-ZHY1 to a high-fat diet could significantly reduce serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, indicating that liver injury caused by high-fat diets was improved. The expression of tumor necrosis factor (TNF)-a and interleukin (IL)-6 in the liver decreased significantly (p < 0.05), while IL-1ß and IL-10 did not change significantly in the AM-ZHY1 group. Compared to the high-fat diet-fed group, the AM-ZHY1 group, but not the ZHY1 group, significantly increased the expression of intestinal tight junction (TJ) proteins (TJP1a, claudina, claudin7, claudin7b, claudin11a, claudin12, and claudin15a; p < 0.05). Compared to the high-fat diet group, the Proteobacteria and Fusobacteria were significantly reduced and increased in the AM-ZHY1 group, respectively. In conclusion, the recombinant bacteria AM-ZHY1 has the capacity to maintain intestinal health by protecting intestinal integrity and improving intestinal flora structure and improving fatty liver disease by inhibiting lipid synthesis and absorption. This study will lay a foundation for the application of AM protein in improving abnormal fat deposition and restoring the intestinal barrier in fish.

Genome-Wide Identification and Comparative Analysis of the Teosinte Branched 1/Cycloidea/Proliferating Cell Factors 1/2 Transcription Factors Related to Anti-cancer Drug Camptothecin Biosynthesis in Ophiorrhiza pumila.

Ophiorrhiza pumila (O. pumila; Op) is a medicinal herbaceous plant, which can accumulate camptothecin (CPT). CPT and its derivatives are widely used as chemotherapeutic drugs for treating malignant tumors. Its biosynthesis pathway has been attracted significant attention. Teosinte branched 1/cycloidea/proliferating cell factors 1/2 (TCP) transcription factors (TFs) regulate a variety of physiological processes, while TCP TFs are involved in the regulation of CPT biosynthesis remain unclear. In this study, a systematic analysis of the TCP TFs family in O. pumila was performed. A total of 16 O. pumila TCP (OpTCP) genes were identified and categorized into two subgroups based on their phylogenetic relationships with those in Arabidopsis thaliana. Tissue-specific expression patterns revealed that nine OpTCP genes showed the highest expression levels in leaves, while the other seven OpTCPs showed a higher expression level in the stems. Co-expression, phylogeny analysis, and dual-luciferase (Dual-LUC) assay revealed that OpTCP15 potentially plays important role in CPT and its precursor biosynthesis. In addition, the subcellular localization experiment of candidate OpTCP genes showed that they are all localized in the nucleus. Our study lays a foundation for further functional characterization of the candidate OpTCP genes involved in CPT biosynthesis regulation and provides new strategies for increasing CPT production.

Identification and Characterization of a Periplasmic α-Carbonic Anhydrase (CA) in the Gametophytes of Saccharina japonica (Phaeophyceae).

Periplasmic or external carbonic anhydrases (CAs) have been well accepted as playing a crucial role in the acquisition of dissolved inorganic carbon; however, no cytological evidence or molecular information on these enzymes has been reported in seaweeds to date. In this study, the full-length cDNA sequence coding for a putative periplasmic Sjα-CA2 was cloned from the gametophytes of Saccharina japonica, an industrial brown seaweed. It was 1,728 bp in length and included a 263-bp 5'-untranslated region (UTR), a 577-bp 3'-UTR, and an 888-bp open reading frame encoding a protein precursor consisting of 295 amino acids. The mature protein, after removal of a predicted 28-residue signal peptide, was composed of 267 amino acids with a relative molecular weight of 29.27 kDa. Multisequence alignment and phylogenetic analysis indicated that it was a member of the α-CA family. Enzyme activity assays showed that the recombinant Sjα-CA2 in Escherichia coli possessed CO2 hydration and esterase activities, thus identifying this gene Sjα-CA2 in function. Immunogold electron microscopic observations with the prepared anti-Sjα-CA2 polyclonal antibody illustrated that Sjα-CA2 was located in periplasmic space of the kelp gametophyte cells. Quantitative real-time PCR results revealed that the transcription of Sjα-CA2 was induced by elevated HCO3- levels, but it was little changed while the kelp gametophytes were subjected to elevated CO2 concentrations. This study suggests that the periplasmic Sjα-CA2 might play a role in adapting to elevated environmental levels of HCO3- by dehydration of HCO3- to generate CO2 , which could be readily taken up by S. japonica gametophytes.

Isolation and characterization of a γ-carbonic anhydrase localized in the mitochondria of Saccharina japonica.

Saccharina japonica is an ecologically and economically important seaweed that is dominant in the rocky shores of cold-temperate regions, forms the major component of productive beds, and affects marine environments. S. japonica exhibits a high photosynthetic efficiency in natural seawater with low dissolved CO2 concentration, thus suggesting the presence of its carbon-concentrating mechanism (CCM). However, the genes, proteins, and pathways involved in the CCM of S. japonica have not been fully identified and characterized. Carbonic anhydrase (CA) is a crucial component of CCM in macroalgae. In this study, the cloning, characterization, and subcellular localization of a specific CA were described. Multisequence alignment and phylogenetic analysis indicated that this CA belonged to the gamma (Sjγ-CA) class. This enzyme has a full-length cDAN of 1370 bp, encodes a protein with 246 amino acids (aa; ca. 25.7 kDa), and contains the mitochondrial transit peptide of 16 aa and LbH_gama_CA_like domain of 159 aa that defined the γ-CA region. The Sjγ-CA was successfully expressed in E. coli BL21 and purified as an active recombinant CA. Immunogold electron microscopy and fluorescence localization illustrated that this enzyme is localized in the mitochondria, and its transcription level is up-regulated by low CO2 concentration. These findings showed that Sjγ-CA is a possible component of the CCM in S. japonica. This work is the first to report about the mtCA of macroalgae and provides a basis for further analysis on seaweed CCM.

Sustainable lipid and lutein production from Chlorella mixotrophic fermentation by food waste hydrolysate.

Bioconversion of food waste into value-added products is a promising way to tackle the global food waste management problem. In this study, a novel valorisation strategy for bioenergy and lutein production via microalgal fermentation was investigated. Significant amount of glucose was recovered from enzymatic hydrolysis of food waste. The resultant hydrolysate was then utilised as culture medium in mixotrophic cultivation of Chlorella sp. to obtain high levels of lipid and lutein, whose accumulation patterns were consistent with molecular analyses. The resultant algal lipid derived from microalgal biomass using food hydrolysate was at high quality in terms of biodiesel properties. Further, in semi-continuous fermentation, the average algal biomass was 6.1 g L-1 with 2.5 g L-1 lipid and 38.5 mg L-1 lutein using hydrolysate with an initial glucose concentration of 10 g L-1. Meanwhile, the resultant algal biomass was 6.9 g L-1 with 1.8 g L-1 lipid and 63.0 mg L-1 lutein using hydrolysate with an initial glucose concentration of 20 g L-1, which suggests food waste hydrolysate could trigger algal products preferences. The experimental results of this study suggested the potential of microalgae as a platform for bioconversion of food waste into high-value products, especially sustainable bioenergy.

Expression of HIF-1α is a predictive marker of the efficacy of neoadjuvant chemotherapy for locally advanced cervical cancer.

Platinum-based, arterial infusion chemotherapy as a neoadjuvant chemotherapy (NACT) followed by hysterectomy may be efficient for the treatment of locally advanced cervical cancer and improve prognosis. It is important to predict whether the NACT would be effective before it is launched. Hypoxia inducible factor-1α (HIF-1α) is the master transcriptional regulator of the cellular response to altered oxygen concentration. HIF-1α protein expression is elevated in numerous human malignancies, contributes to poor disease outcome, and has been reported to induce tumorigenesis and chemoresistance. In the present study, patients with International Federation of Gynecology and Obstetrics stage IIB-IIIB cervical cancer (n=59) between 2008 and 2014 were assessed for HIF-1α expression by immunohistochemistry. Tumor samples were obtained by biopsy before any treatment. A double-path chemotherapy regimen, paclitaxel (intravenous) plus cisplatin (intra-arterial injection into the uterine region), was used as NACT. The patients were then separated into two groups according to NACT response: One group comprised patients with NACT, for whom the response to treatment was efficient resulting in complete/partial remission of the tumor (CR + PR group; n=52), the other group contained patients with NACT, for whom the result of the treatment was a stable/progressive disease (SD + PD group; n=7). HIF-1α expression was tested in paraffin-embedded sections using immunohistochemistry. HIF-1α expression was significantly higher in the SD + PD group compared with the CR + PR group (P=0.029). The overall survival time was significantly longer in the CR + PR group compared with the SD + PD group (P<0.001). When the patients were divided into two groups based on HIF-1α expression levels. Low (weighted score ≤4, n=39) and high (weighted score ≥6, n=20) expression level groups; the low HIF-1α expression group was significantly more susceptible to NACT treatment (P=0.025). Cox hazard analysis revealed that a high level of HIF-1α expression and lymph node metastases were significant independent predictors of poor overall survival (P=0.025, HR=6.354; P=0.020, HR=6.909, respectively). These results indicated that the expression of HIF-1α may be able to predict the efficiency of NACT and may be considered an independent prognostic factor for stage IIB-IIIB cervical cancer.

Reciprocal Regulation Between Forkhead Box M1/NF-κB and Methionine Adenosyltransferase 1A Drives Liver Cancer.

BACKGROUND AND AIMS: Forkhead box M1 (FOXM1) and nuclear factor kappa B (NF-ĸB) are oncogenic drivers in liver cancer that positively regulate each other. We showed that methionine adenosyltransferase 1A (MAT1A) is a tumor suppressor in the liver and inhibits NF-ĸB activity. Here, we examined the interplay between FOXM1/NF-κB and MAT1A in liver cancer. APPROACH AND RESULTS: We examined gene and protein expression, effects on promoter activities and binding of proteins to promoter regions, as well as effects of FOXM1 inhibitors T0901317 (T0) and forkhead domain inhibitory-6 (FDI-6) in vitro and in xenograft and syngeneic models of liver cancer. We found, in both hepatocellular carcinoma and cholangiocarcinoma, that an induction in FOXM1 and NF-κB expression is accompanied by a fall in MATα1 (protein encoded by MAT1A). The Cancer Genome Atlas data set confirmed the inverse correlation between FOXM1 and MAT1A. Interestingly, FOXM1 directly interacts with MATα1 and they negatively regulate each other. In contrast, FOXM1 positively regulates p50 and p65 expression through MATα1, given that the effect is lost in its absence. FOXM1, MATα1, and NF-κB all bind to the FOX binding sites in the FOXM1 and MAT1A promoters. However, binding of FOXM1 and NF-κB repressed MAT1A promoter activity, but activated the FOXM1 promoter. In contrast, binding of MATα1 repressed the FOXM1 promoter. MATα1 also binds and represses the NF-κB element in the presence of p65 or p50. Inhibiting FOXM1 with either T0 or FDI-6 inhibited liver cancer cell growth in vitro and in vivo. However, inhibiting FOXM1 had minimal effects in liver cancer cells that do not express MAT1A. CONCLUSIONS: We have found a crosstalk between FOXM1/NF-κB and MAT1A. Up-regulation in FOXM1 lowers MAT1A, but raises NF-κB, expression, and this is a feed-forward loop that enhances tumorigenesis.

Increased attention to snake images in cynomolgus monkeys: an eye-tracking study.

Previous studies have revealed faster detection of snake images in humans and non-human primates (NHPs), suggesting automatic detection of evolutionary fear-relevant stimuli. Furthermore, human studies have indicated that general fear-relevance rather than evolutionary relevance is more effective at capturing attention. However, the issue remains unclarified in NHPs. Thus, in the present study, we explored the attentional features of laboratory-reared monkeys to evolutionary and general fear-relevant stimuli (e.g., images of snakes, capturing gloves). Eye-tracking technology was utilized to assess attentional features as it can provide more accurate latency and variables of viewing duration and frequency compared with visual search task (VST) and response latency adopted in previous studies. In addition, those with autism spectrum disorder (ASD) show abnormal attention to threatening stimuli, including snake images. Rett syndrome (RTT) is considered a subcategory of ASD due to the display of autistic features. However, the attentional features of RTT patients or animal models to such stimuli remain unclear. Therefore, we also investigated the issue in MECP2 gene-edited RTT monkeys. The influence of different cognitive loads on attention was further explored by presenting one, two, or four images to increase stimulus complexity. The eye-tracking results revealed no significant differences between RTT and control monkeys, who all presented increased viewing (duration and frequency) of snake images but not of aversive stimuli compared with control images, thus suggesting attentional preference for evolutionary rather than general fear-relevant visual stimuli. Moreover, the preference was only revealed in visual tasks composed of two or four images, suggesting its cognitive-load dependency.

AidB, a Novel Thermostable N-Acylhomoserine Lactonase from the Bacterium Bosea sp.

Many Gram-negative bacteria employ N-acylhomoserine lactones (AHLs) as quorum-sensing (QS) signal molecules to regulate virulence expression in a density-dependent manner. Quorum quenching (QQ) via enzymatic inactivation of AHLs is a promising strategy to reduce bacterial infections and drug resistance. Herein, a thermostable AHL lactonase (AidB), which could degrade different AHLs, with or without a substitution of carbonyl or hydroxyl at the C-3 position, was identified from the soil bacterium Bosea sp. strain F3-2. Ultrahigh-performance liquid chromatography analysis demonstrated that AidB is an AHL lactonase that hydrolyzes the ester bond of the homoserine lactone (HSL) ring. AidB was thermostable in the range 30 to 80°C and showed maximum activity after preincubation at 60°C for 30 min. The optimum temperature of AidB was 60°C, and the enzyme could be stably stored in double-distilled water (ddH2O) at 4°C or room temperature. AidB homologs were found only in Rhizobiales and Rhodospirillales of the Alphaproteobacteria AidB from Agrobacterium tumefaciens and AidB from Rhizobium multihospitium (with amino acid identities of 50.6% and 52.8% to AidB, respectively) also showed thermostable AHL degradation activity. When introduced into bacteria, plasmid-expressed AidB attenuated pyocyanin production by Pseudomonas aeruginosa PAO1 and the pathogenicity of Pectobacterium carotovorum subsp. carotovorum Z3-3, suggesting that AidB is a potential therapeutic agent by degrading AHLs.IMPORTANCE A quorum-sensing system using AHLs as the signal in many bacterial pathogens is a critical virulence regulator and an attractive target for anti-infective drugs. In this work, we identified a novel AHL lactonase, AidB, from a soil bacterial strain, Bosea sp. F3-2. The expression of aidB reduced the production of AHL signals and QS-dependent virulence factors in Pseudomonas aeruginosa and Pectobacterium carotovorum The homologs of AidB with AHL-degrading activities were found only in several genera belonging to the Alphaproteobacteria Remarkably, AidB is a thermostable enzyme that retained its catalytic activity after treatment at 80°C for 30 min and exhibits reliable storage stability at both 4°C and room temperature. These properties might make it more suitable for practical application.

Identification of Benzyloxy Carbonimidoyl Dicyanide Derivatives as Novel Type III Secretion System Inhibitors via High-Throughput Screening.

The type III secretion system (T3SS) in many Gram-negative bacterial pathogens is regarded as the most critical virulence determinant and an attractive target for novel anti-virulence drugs. In this study, we constructed a T3SS secretion reporter containing the ß-lactamase gene fused with a signal peptide sequence of the T3SS effector gene, and established a high-throughput screening system for T3SS inhibitors in the plant pathogenic bacterium Acidovorax citrulli. From a library of 12,000 chemical compounds, we identified a series of benzyloxy carbonimidoyl dicyanide (BCD) derivatives that effectively blocked T3SS-dependent ß-lactamase secretion. Substitution of halogens or nitro groups at the para-position on the benzene ring contributed to an increased inhibitory activity. One representative compound, BCD03 (3,4-dichloro-benzyloxy carbonimidoyl dicyanide), dramatically reduced pathogenicity of A. citrulli on melon seedlings, and attenuated hypersensitive responses in the non-host Nicotiana tabacum caused by pathogenic bacteria A. citrulli, Xanthomonas oryzae pv. oryzae and Pseudomonas syringae pv. tomato at sub-MIC concentrations. Western blotting assay further confirmed that BCD03 inhibited effector secretion from the above bacteria via T3SS in the liquid medium. Taken together, our data suggest that BCD derivatives act as novel inhibitors of T3SS in multiple plant bacterial pathogens.

Ability of prebiotic polysaccharides to activate a HIF1α-antimicrobial peptide axis determines liver injury risk in zebrafish.

Natural polysaccharides have received much attention for their ability to ameliorate hepatic steatosis induced by high-fat diet. However, the potential risks of their use have been less investigated. Here, we show that the exopolysaccharides (EPS) from Lactobacillus rhamnosus GG (LGG) and L. casei BL23 reduce hepatic steatosis in zebrafish fed a high-fat diet, while BL23 EPS, but not LGG EPS, induce liver inflammation and injury. This is due to the fact that BL23 EPS induces gut microbial dysbiosis, while LGG EPS promotes microbial homeostasis. We find that LGG EPS, but not BL23 EPS, can directly activate intestinal HIF1α, and increased HIF1α boosts local antimicrobial peptide expression to facilitate microbial homeostasis, explaining the distinct compositions of LGG EPS- and BL23 EPS-associated microbiota. Finally, we find that liver injury risk is not confined to Lactobacillus-derived EPS but extends to other types of commonly used natural polysaccharides, depending on their HIF1α activation efficiency.