Identification and immunological evaluation of novel TLR2 agonists through structure optimization of Pam3CSK4

Toll-like receptor 2 (TLR2) is a bridge between innate immunity and adaptive immunity. TLR2 agonists have been exploited as potential vaccine adjuvants and antitumor agents. However, no TLR2 agonists have been approved by FDA up to now. To discover drug-like TLR2 selective agonists, a novel series of Pam₃CSK₄ derivatives were designed based on the crystal structure of hTLR2-hTLR1-Pam₃CSK₄ complex, synthesized and evaluated for their immune-stimulatory activities. Among them, 35c was identified as a murine-specific TLR2 agonist, while 35f was a human-specific TLR2 agonist. Besides, 35d (human and murine TLR2 agonist) showed TLR2 agonistic activity comparable to Pam₃CSK₄, which included: elevated IL-6 expression level (EC₅₀ = 83.08 ± 5.94 nM), up-regulated TNF-α and IL-6 mRNA expression and promoted maturation of DCs through activating the NF-κB signaling pathway. TLRs antibodies test showed that 35a and 35d were TLR2/1 agonists, while 35f was a TLR2/6 agonist.     

Identification of new dual FABP4/5 inhibitors based on a naphthalene-1-sulfonamide FABP4 inhibitor

Fatty acid binding protein 4 (FABP4) and fatty acid binding protein 5 (FABP5) are mainly expressed in adipocytes and/or macrophages and play essential roles in energy metabolism and inflammation. When FABP4 function is diminished, FABP5 expression is highly increased possibly as a functional compensation. Dual FABP4/5 inhibitors are expected to provide beneficial synergistic effect on treating diabetes, atherosclerosis, and inflammation-related diseases. Starting from our previously reported selective FABP4 inhibitor 8, structural biology information was used to modulate the selectivity profile and to design potent dual FABP4/5 inhibitors with good selectivity against FABP3. Two compounds A16 and B8 were identified to show inhibitory activities against both FABP4/5 and good selectivity over FABP3, which could also reduce the level of forskolin-stimulated lipolysis in mature 3T3-L1 adipocytes. Compared with compound 8, these two compounds exhibited better anti-inflammatory effects in lipopolysaccharide-stimulated RAW264.7 murine macrophages, with decreased levels of pro-inflammatory cytokines TNFα and MCP-1 and apparently inhibited IKK/NF-κB pathway.     

Highly sensitive biosensor based on aptamer and hybridization chain reaction for detection of cadmium ions

In this work, a highly sensitive biosensor for detecting cadmium ions (Cd²⁺) was developed based on a Cd²⁺-specific DNA aptamer and a hybridization chain reaction (HCR). The Cd²⁺ aptamer (named S0) was used to recognize Cd²⁺ and trigger the HCR. Without Cd²⁺, S0 initiated the HCR to form long nicked dsDNA structures to quench the fluorescence. Then, Cd²⁺ could bind with S0 to block HCR to recover fluorescence. This biosensor had high sensitivity with a detection limit of 0.36 nM and a linear range from 0 to 10 nM. Moreover, it showed a satisfactory selectivity and recovery rates.     

Ultra-Broadband Electromagnetically Induced Transparency in Metamaterial Based on Conductive Coupling

Plasmonics - This paper proposes a structure composed of a horizontal metal strip resonator (SR) and four C-shaped ring resonators (CRR) to obtain a broadband electromagnetic induction transparency...     

Pulsed electromagnetic fields regulate osteocyte apoptosis,RANKL/OPG expression,and its control of osteoclastogenesis depending on the presence of primary cilia

Growing evidence has shown that pulsed electromagnetic fields (PEMF) can modulate bone metabolism in vivo and regulate the activities of osteoblasts and osteoclasts in vitro. Osteocytes, accounting for 95% of bone cells, act as the major mechanosensors in bone for transducing external mechanical signals and producing cytokines to regulate osteoblastic and osteoclastic activities. Targeting osteocytic signaling pathways is becoming an emerging therapeutic strategy for bone diseases. We herein systematically investigated the changes of osteocyte behaviors, functions, and its regulation on osteoclastogenesis in response to PEMF. The osteocyte-like MLO-Y4 cells were exposed to 15 Hz PEMF stimulation with different intensities (0, 5, and 30 Gauss [G]) for 2 hr. We found that the cell apoptosis and cytoskeleton organization of osteocytes were regulated by PEMF with an intensity-dependent manner. Moreover, PEMF exposure with 5 G significantly inhibited apoptosis-related gene expression and also suppressed the gene and protein expression of the receptor activator of nuclear factor κB ligand/osteoprotegerin (RANKL/OPG) ratio in MLO-Y4 cells. The formation, maturation, and osteoclastic bone-resorption capability of in vitro osteoclasts were significantly suppressed after treated with the conditioned medium from PEMF-exposed (5 G) osteocytes. Our results also revealed that the inhibition of osteoclastic formation, maturation, and bone-resorption capability induced by the conditioned medium from 5 G PEMF-exposed osteocytes was significantly attenuated after abrogating primary cilia in osteocytes using the polaris siRNA transfection. Together, our findings highlight that PEMF with 5 G can inhibit cellular apoptosis, modulate cytoskeletal distribution, and decrease RANKL/OPG expression in osteocytes, and also inhibit osteocyte-mediated osteoclastogenesis, which requires the existence of primary cilia in osteocytes. This study enriches our basic knowledge for further understanding the biological behaviors of osteocytes and is also helpful for providing a more comprehensive mechanistic understanding of the effect of electromagnetic stimulation on bone and relevant skeletal diseases (e.g., bone fracture and osteoporosis).     

Hepatic deficiency of selenoprotein S exacerbates hepatic steatosis and insulin resistance

Nonalcoholic fatty liver disease (NAFLD) is closely associated with insulin resistance (IR) and type 2 diabetes mellitus (T2DM), which are all complex metabolic disorders. Selenoprotein S (SelS) is an endoplasmic reticulum (ER) resident selenoprotein involved in regulating ER stress and has been found to participate in the occurrence and development of IR and T2DM. However, the potential role and mechanism of SelS in NAFLD remains unclear. Here, we analyzed SelS expression in the liver of high-fat diet (HFD)-fed mice and obese T2DM model (db/db) mice and generated hepatocyte-specific SelS knockout (SelS^H-KO) mice using the Cre-loxP system. We showed that hepatic SelS expression levels were significantly downregulated in HFD-fed mice and db/db mice. Hepatic SelS deficiency markedly increased ER stress markers in the liver and caused hepatic steatosis via increased fatty acid uptake and reduced fatty acid oxidation. Impaired insulin signaling was detected in the liver of SelS^H-KO mice with decreased phosphorylation levels of insulin receptor substrate 1 (IRS1) and protein kinase B (PKB/Akt), which ultimately led to disturbed glucose homeostasis. Meanwhile, our results showed hepatic protein kinase Cɛ (PKCɛ) activation participated in the negative regulation of insulin signaling in SelS^H-KO mice. Moreover, the inhibitory effect of SelS on hepatic steatosis and IR was confirmed by SelS overexpression in primary hepatocytes in vitro. Thus, we conclude that hepatic SelS plays a key role in regulating hepatic lipid accumulation and insulin action, suggesting that SelS may be a potential intervention target for the prevention and treatment of NAFLD and T2DM.Subject terms: Metabolic syndrome, Obesity     

Yersinia spp. Identification Using Copy Diversity in the Chromosomal 16S rRNA Gene Sequence

API 20E strip test, the standard for Enterobacteriaceae identification, is not sufficient to discriminate some Yersinia species for some unstable biochemical reactions and the same biochemical profile presented in some species, e.g. Yersinia ferderiksenii and Yersinia intermedia, which need a variety of molecular biology methods as auxiliaries for identification. The 16S rRNA gene is considered a valuable tool for assigning bacterial strains to species. However, the resolution of the 16S rRNA gene may be insufficient for discrimination because of the high similarity of sequences between some species and heterogeneity within copies at the intra-genomic level. In this study, for each strain we randomly selected five 16S rRNA gene clones from 768 Yersinia strains, and collected 3,840 sequences of the 16S rRNA gene from 10 species, which were divided into 439 patterns. The similarity among the five clones of 16S rRNA gene is over 99% for most strains. Identical sequences were found in strains of different species. A phylogenetic tree was constructed using the five 16S rRNA gene sequences for each strain where the phylogenetic classifications are consistent with biochemical tests; and species that are difficult to identify by biochemical phenotype can be differentiated. Most Yersinia strains form distinct groups within each species. However Yersinia kristensenii, a heterogeneous species, clusters with some Yersinia enterocolitica and Yersinia ferderiksenii/intermedia strains, while not affecting the overall efficiency of this species classification. In conclusion, through analysis derived from integrated information from multiple 16S rRNA gene sequences, the discrimination ability of Yersinia species is improved using our method.     

Complete genome sequence and pathogenic genes analysis of <Emphasis Type="Italic">Pectobacterium atroseptica</Emphasis> JG10-08

Pectobacterium atroseptica is known as a rod-shaped gram-negative bacterial pathogen associated with the blackleg of potato. P. atroseptica has been widely identified as the predominant agent causing tuber rot in temperate regions, a disease that leads to severe economic losses to potato industry. In this study, we provide the complete genome sequence of P. atroseptica JG10-08, which revealed that P. atroseptica strain JG10-08 carries a single 5,004,926 bp chromosome with 51.15% G+C content and harbors 4252 predicted coding genes. Phylogenetic analysis based on the genome sequences showed a close evolutionary relationship between P. atroseptica and Pectobacterium wasabiae. We discovered total 168 genes were potentially related to pathogenesis including 9 strain-specific genes encoding toxins on the genome of JG10-08. Further comparison with other species in Pectobacterium revealed a better understanding of pathogenic factors, especially secretion systems in P. atroseptica JG10-08. Collectively, the results of this research provide a solid foundation for discovering the underlying pathogenic mechanisms of P. atroseptica and offer the information to develop more effective strategies against blackleg of potatoes.     

外源木聚糖酶在枯草芽胞杆菌中信号肽筛选

[目的]本试验旨在筛选引导表达外源木聚糖酶基因高效分泌的信号肽,为枯草芽胞杆菌木聚糖酶高效分泌表达系统提供元件.[方法]构建信号肽筛选载体,载体是以含壮观霉素抗性基因的大肠-枯草穿梭载体为基本骨架,目标蛋白为耐碱性木聚糖酶,可在麦芽糖启动子Pglv诱导下表达.从枯草芽胞杆菌A1747基因组中扩增获得24个Sec途径信号肽,并将其全部链接到至筛选载体上,并在枯草芽胞杆菌WB700中实现表达分泌.重组菌在3%麦芽糖诱导下培养24h后用DNS法测定上清酶活.[结果]成功构建信号肽筛选载体pGPSX及24个表达载体,实现木聚糖酶表达分泌.且不同信号肽对于引导外源木聚糖酶分泌能力不同,其中YnfF信号肽引导分泌目标蛋白效率最高,上清酶活为37.2IU/mL.[结论]试验证明在枯草杆菌中对外源蛋白进行信号肽筛选是提高其分泌的有效途径,并获得了针对木聚糖酶高效分泌信号肽YnfF.