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Ticks are hematophagous ectoparasites and cause a major public health threat worldwide. Development of anti-tick vaccines is regarded to be an optimal alternative for tick control. AV422, a unique protein in ticks, is secreted into hosts during blood-feeding, but its roles are not confirmed in Haemaphysalis flava ticks. We retrieved a gene fragment encoding AV422 from a transcriptome dataset of H. flava, and based on it, we reconstructed the full length of AV422 from H. flava (Hf-AV422) by rapid amplification of cDNA ends. Expression profiles of Hf-AV422 in whole ticks and organs of different engorgement levels were determined by qPCR. Then its opening reading frame (ORF) was expressed in Escherichia coli strain BL21 (DE3). The prothrombin time (PT), activated partial thromboplastin time (APTT) and thrombin time (TT) assays were conducted to test anticoagulant activities of the purified recombinant protein (rHf-AV422). The full length of AV422 was 1152 bp. Hf-AV422 showed to be conserved as indicated by multiple sequence alignment. Expression of Hf-AV422 was significantly higher in salivary glands and cuticles than in ovaries. Its expression in whole ticks decreased during engorgement with the highest levels in 1/4 engorged ticks. rHf-AV422 prolonged PT, APTT and TT when incubated with rabbit plasma. Our data demonstrated that Hf-AV422 is a conserved salivary protein with anticoagulant activity. Further studies are needed to test in detail its functional properties to ensure it an adequate antigen candidate for the development of broad-spectrum vaccines against ticks.

Laryngeal and hypopharyngeal carcinomas are common malignant tumors of the head and neck, and the incidence of both is increasing. Laryngopharyngeal reflux refers to the retrograde flow of gastric contents into the larynx, oropharynx, and/or nasopharynx. It remains controversial whether laryngopharyngeal reflux is a risk factor for laryngeal and hypopharyngeal cancers. The refluxing substances mainly include hydrochloric acid, pepsin, and occasionally bile acids and bile salts, as well as bacteria that colonize the gastrointestinal tract. Loss of epithelium in the mucous membrane of the larynx and hypopharynx is thought to be caused by pepsin. Here, we review the relationships between laryngopharyngeal reflux and both laryngeal and hypopharyngeal carcinomas, as well as the significance of pepsin, methods of clinical detection, and the mechanism of carcinogenesis.

In this study, the authors sequenced and characterized the complete mitochondrial (mt) genomes of two hard ticks of the genus Ixodes, I. nipponensis and Ixodes (Pholeoixodes) sp., which were 14 505 and 14 543 bp in length, respectively. Their mt genomes encoded 37 genes, including 13 protein-coding genes (PCGs), 22 transfer RNA genes and two ribosomal RNA genes, and have only one non-coding region. The gene order in their mt genomes was the same as that of other Ixodes spp. mt genomes. The average sequence identity, combined nucleotide diversity, non-synonymous/synonymous substitutions ratio analyses consistently demonstrated that cox1, rrnS, cox2, cox3 and cytb were the most conserved and atp8, nad6 and nad2 were the most variable genes across Ixodes mitogenomes. Phylogeny of the present Ixodes spp., and other selected hard tick species, based on concatenated amino acid sequences of PCGs, confirmed their position within the genus Ixodes and sub-family Ixodinae. The novel mt markers described herein will be useful for further studies of the population genetics, molecular epidemiology and systematics of hard ticks.

Molecular-based classifications of gastric cancer (GC) were recently proposed, but few of them robustly predict clinical outcomes. While mutation and expression signature of protein-coding genes were used in previous molecular subtyping methods, the noncoding genome in GC remains largely unexplored. Here, we developed the fast long-noncoding RNA analysis (FLORA) method to study RNA sequencing data of GC cases, and prioritized tumor-specific long-noncoding RNAs (lncRNAs) by integrating clinical and multi-omic data. We uncovered 1235 tumor-specific lncRNAs, based on which three subtypes were identified. The lncRNA-based subtype 3 (L3) represented a subgroup of intestinal GC with worse survival, characterized by prevalent TP53 mutations, chromatin instability, hypomethylation, and over-expression of oncogenic lncRNAs. In contrast, the lncRNA-based subtype 1 (L1) has the best survival outcome, while LINC01614 expression further segregated a subgroup of L1 cases with worse survival and increased chance of developing distal metastasis. We demonstrated that LINC01614 over-expression is an independent prognostic factor in L1 and network-based functional prediction implicated its relevance to cell migration. Over-expression and CRISPR-Cas9-guided knockout experiments further validated the functions of LINC01614 in promoting GC cell growth and migration. Altogether, we proposed a lncRNA-based molecular subtype of GC that robustly predicts patient survival and validated LINC01614 as an oncogenic lncRNA that promotes GC proliferation and migration.

Temporal acuity is the ability to differentiate between sounds based on fluctuations in the waveform envelope. The proximity of successive sounds and background noise diminishes the ability to track rapid changes between consecutive sounds. We determined whether a physiological correlate of temporal acuity is also affected by these factors. We recorded the auditory brainstem response (ABR) from human listeners using a harmonic complex (S1) followed by a brief tone burst (S2) with the latter serving as the evoking signal. The duration and depth of the silent gap between S1 and S2 were manipulated, and the peak latency and amplitude of wave V were measured. The latency of the responses decreased significantly as the duration or depth of the gap increased. The amplitude of the responses was not affected by the duration or depth of the gap. These findings suggest that changing the physical parameters of the gap affects the auditory system&rsquo, s ability to encode successive sounds.

Extraskeletal Ewing's Sarcoma (EES) may harbor more than one tumor-specific genetic abnormality, leading to diagnostic difficulties. Case report: We report a nine-year-old boy with recurrent mass o...

The gut microbiota has been implicated in cancer and shown to modulate anticancer drug efficacy. Altered gut microbiota is associated with resistance to chemo drugs or immune checkpoint inhibitors (ICIs), whereas supplementation of distinct bacterial species restores responses to the anticancer drugs. Accumulating evidence has revealed the potential of modulating the gut microbiota to enhance the efficacy of anticancer drugs. Regardless of the valuable findings by preclinical models and clinical data of patients with cancer, a more thorough understanding of the interactions of the microbiota with cancer therapy helps researchers identify novel strategy for cancer prevention, stratify patients for more effective treatment and reduce treatment complication. In this review, we discuss the scientific evidence on the role of gut microbiota in cancer treatment, and highlight the latest knowledge and technologies leveraged to target specific bacteria that contribute to tumourigenesis. First, we provide an overview of the role of the gut microbiota in cancer, establishing the links between bacteria, inflammation and cancer treatment. Second, we highlight the mechanisms used by distinct bacterial species to modulate cancer growth, immune responses, as well as the efficacy of chemotherapeutic drugs and ICIs. Third, we demonstrate various approaches to modulate the gut microbiota and their potential in translational research. Finally, we discuss the limitations of current microbiome research in the context of cancer treatment, ongoing efforts to overcome these challenges and future perspectives.

BackgroundThe present study aimed to assess the perturbation in circular RNA (circRNA)/mRNA expression profiles and a circRNA-miRNA-mRNA coexpression network involved in the potential protective effect of diosgenin (DIO) on alveolar bone loss in rats subjected to ovariectomy (OVX).MethodsThe Wistar rats (female) manipulated with sham operation were classified as the SHAM group and the grouping of OVX rats administered with DIO, estradiol valerate or vehicle for 12 weeks was DIO group, EV group and OVX group respectively. Following treatments, the plasmatic levels of osteocalcin and tumor necrosis factor-alpha and the microstructure of alveolar bone were assayed. Based on microarray analyses, we identified differentially expressed (DE) circRNAs and mRNAs in alveolar bone of rats in both OVX and DIO group. The DE circRNAs and DE mRNAs involved in the bone metabolism pathway validated by RT-qPCR were considered key circRNAs/mRNAs. On the basis of these key circRNAs/mRNAs, we predicted the overlapping relative miRNAs of key circRNAs/mRNAs, and a circRNA-miRNA-mRNA network was built.ResultsDIO showed an anti-osteopenic effect on the rat alveolar bone loss induced by OVX. In total, we found 10 DE circRNAs (6 downregulated and 4 upregulated) and 614 DE mRNAs (314 downregulated and 300 upregulated) in samples of the DIO group compared with those of the OVX group. However, only one circRNA (rno_circRNA_016717) and seven mRNAs (Sfrp1,Csf1,Il1rl1,Nfatc4,Tnfrsf1a,Pik3c2g, andWnt9b) were validated by qRT-PCR and therefore considered key circRNA/mRNAs. According to these key circRNA/mRNAs and overlapping predicted miRNAs, a coexpression network was constructed. After network analysis, one circRNA-miRNA-mRNA axis (circRNA_016717/miR-501-5p/Sfrp1) was identified.ConclusionThe mechanism of DIO inhibiting alveolar bone loss after OVX is possibly relevant to the simultaneous inhibition of osteogenesis and osteoclastogenesis by mediating the expression of important molecules in the Wnt, PI3K, RANK/RANKL or osteoclastogenic cytokine pathways. The circRNA_016717/miR-501-5p/Sfrp1axis may play important roles in these processes.

Background Altered microbiome composition and aberrant promoter hypermethylation of tumor suppressor genes (TSGs) are two important hallmarks of colorectal cancer (CRC). Here we performed concurrent 16S rRNA gene sequencing and methyl-CpG binding domain-based capture sequencing in 33 tissue biopsies (5 normal colonic mucosa tissues, 4 pairs of adenoma and adenoma-adjacent tissues, and 10 pairs of CRC and CRC-adjacent tissues) to identify significant associations between TSG promoter hypermethylation and CRC-associated bacteria, followed by functional validation of the methylation-associated bacteria. Results Fusobacterium nucleatum and Hungatella hathewayi were identified as the top two methylation-regulating bacteria. Targeted analysis on bona fide TSGs revealed that H. hathewayi and Streptococcus spp. significantly correlated with CDX2 and MLH1 promoter hypermethylation, respectively. Mechanistic validation with cell-line and animal models revealed that F. nucleatum and H. hathewayi upregulated DNA methyltransferase. H. hathewayi inoculation also promoted colonic epithelial cell proliferation in germ-free and conventional mice. Conclusion Our integrative analysis revealed previously unknown epigenetic regulation of TSGs in host cells through inducing DNA methyltransferase by F. nucleatum and H. hathewayi, and established the latter as CRC-promoting bacteria.

Background Invasion of pituitary growth hormone-secreting adenoma into surrounding tissues poses a challenge for complete resection in surgery, which is the main reason for recurrence of this type of cancer. Studies have shown that abnormal methylation of RASSF10 can promote the expression of MDM2 and regulate the tumor microenvironment by affecting the secretion of exosomes. In the present study, we aim to uncover the specific underlying mechanism of this effect. Method Transwell co-culture assays was performed using GT1.1 cells or exosomes and RAW264.7 cells. RAW264.7 cells were collected for invasion, proliferation and apoptosis assays, RT-qPCR and western blotting. RNA-seq was performed and used to assess the potential molecular pathways of the effect of GT1.1 cell-exosomes on RAW264.7 cells. Results GT1.1 cells with reduced RASSF10 expression could promote the proliferation and migration of RAW264.7 cells, and promote their expression of osteoclast markers TRAP and CK. The effect of GT1.1 cell exosomes on the RAW264.7-cell phenotype was shown to be achieved through the RASSF10-MDM2 pathway. RNA-seq allowed the identification of PI3K-AKT, MAPK, and calcium signaling as important in this regulation system of RASSF10-MDM2. Conclusion Our results indicate that GT1.1 cells activate PI3K-AKT, MAPK and calcium signaling via the RASSF10-MDM2 pathway, and promote the differentiation of RAW264.7 cells into osteoclasts through exosomes. This study may provide new ideas to aid in early diagnosis, prognostic assessment and treatment of aggressive pituitary adenomas.

TelN and tos are a unique DNA linearization unit isolated from bacteriophage N15. While being transferable, the TelN cleaving-rejoining activities remained stable to function on tos in both bacterial and mammalian environments. However, TelN contribution in linear plasmid replication in mammalian cells remains unknown. Herein, we investigated the association of TelN in linear tos-containing DNA (tos-DNA) replication in mammalian cells. Additionally, the mammalian origin of replication (ori) that is well-known to initiate the replication event of plasmid vectors was also studied. In doing so, we identified that both TelN and mammalian initiation sites were essential for the replication of linear tos-DNA, determined by using methylation sensitive DpnI/MboI digestion and polymerase chain reaction (PCR) amplification approaches. Furthermore, we engineered the linear tos-DNA to be able to retain in mammalian cells using S/MAR technology. The resulting S/MAR containing tos-DNA was robust for at least 15 days, with (1) continuous tos-DNA replication, (2) correct splicing of gene transcripts, and (3) stable exogenous gene expression that was statistically comparable to the endogenous gene expression level. Understanding the activities of TelN and tos in mammalian cells can potentially provide insights for adapting this simple DNA linearization unit in developing novel genetic engineering tools, especially to the eukaryotic telomere/telomerase study.

Tick blood meals are stored and digested in their midguts. Blood digestion is complex, and many proteins are involved. Study of the tick-derived proteins in the midgut content may aid in the discovery of active molecules that would be useful for anti-tick vaccines. We analyzed the midgut content proteomes of partially engorged female Haemaphysalis flava, fully engorged female H. flava, and hedgehog serum using liquid chromatography tandem-mass spectrometry and label-free quantitation. In this study, high-confidence protein profiling of tick midgut content was determined. Based on the search against our in-house transcriptome database, the 28 high-confidence proteins were identified. Of these, 17 were identified as tick-derived, and the rest were of unspecified origin (proteins that could not be differentiated as host-derived or tick-derived proteins). The function of these midgut content proteins identified here may involve nutrient transportation, anti-coagulation, erythrocyte lysis, detoxification, lipid metabolism, and immunization. The presence of hemoglobin suggested that the red blood cells were lysed in the gut lumen. The midgut contents contain a large amount of fibrinogen and it has the ability to clot immediately. The midgut contained mostly host-derived proteins, and these host proteins provide rich nutrients for tick development and reproduction. However, some intracellular proteins were also identified, suggesting the possibility of shedding of the midgut epithelium and ingestion of saliva during feeding. This finding advances our understanding of the digestive mechanism and will be useful in the screening of vaccine antigens.

Diabetic nephropathy (DN) is a major and severe complication of diabetes mellitus. Ferulic acid (FA), a phenolic compound widespread in fruits and plants, displays a variety of pharmacological activities including regulating blood glucose and lipids, anti-oxidation, anti-inflammation and anti-fibrosis. The study was aimed to investigate the renal protective effects of FA on diabetic rats and elucidate the underlying mechanisms. FA (100 mg kg-1, i.g., once a day) was administered to DN rats for 8 weeks. The organ coefficient of kidneys was calculated. Levels of UP, BUN, Cr, FBG, TC and TG in serum were measured. Activities of SOD, CAT and GPx and the content of MDA in renal tissues were assayed. Pathological changes in renal tissues were observed by HE staining, PAS staining, PASM staining, Masson staining and transmission electron microscopy. p-NF-κB p65, TNF-α, TGF-β1, collagen IV, nephrin and podocin protein expressed in renal tissues were determined by immunohistochemistry and western blotting. Results showed that FA significantly improved the kidney organ coefficient, decreased the UP, BUN, Cr, FBG, TC and TG levels in serum, increased SOD, CAT and GPx activities, reduced MDA content in renal tissues and alleviated pathological injury of the renal tissues. What's more, long-term treatment with FA considerably down-regulated the expressions of p-NF-κB p65, TNF-α, TGF-β1 and collagen IV proteins, and up-regulated the expressions of nephrin and podocin proteins in renal tissues. FA could be a renoprotective agent by attenuating oxidative stress, inflammation, and fibrosis, as well as improving podocyte injury in STZ-induced DN rats.

Adipose‐derived mesenchymal stem cells (ASCs) exhibit great potential in regenerative medicine, and in vitro expansion is frequently necessary to obtain a sufficient number of ASCs for clinical use. Fibroblast growth factor 2 (FGF2) is a common supplement in the ASC culture medium to enhance cell proliferation. To achieve clinical applicability of ASC‐based products, prolonged culture of ASCs is sometimes required to obtain sufficient quantity of ASCs. However, the effect of FGF2 on ASCs during prolonged culture has not been previously determined. In this study, ASCs were subjected to prolonged in vitro culture with or without FGF2. FGF2 maintained the small cell morphology and expedited proliferation kinetics in early ASC passages. After prolonged in vitro expansion, FGF2‐treated ASCs exhibited increased cell size, arrested cell proliferation, and increased cellular senescence relative to the control ASCs. We observed an upregulation of FGFR1c and enhanced expression of downstream STAT3 in the initial passages of FGF2‐treated ASCs. The application of an FGFR1 or STAT3 inhibitor effectively blocked the enhanced proliferation of ASCs induced by FGF2 treatment. FGFR1c upregulation and enhanced STAT3 expression were lost in the later passages of FGF2‐treated ASCs, suggesting that the continuous stimulation of FGF2 becomes ineffective because of the refractory downstream FGFR1 and the STAT3 signaling pathway. In addition, no evidence of tumorigenicity was noted in vitro and in vivo after prolonged expansion of FGF2‐cultured ASCs. Our data indicate that ASCs have evolved a STAT3‐dependent response to continuous FGF2 stimulation which promotes the initial expansion but limits their long‐term proliferation., Fibroblast growth factor 2 (FGF2) exerts positive effects in the early passages of adipose‐derived mesenchymal stem cells (ASCs), including maintenance of cellular morphology, less senescent cells, and expedition of proliferative kinetics. However, after prolonged in vitro expansion with FGF2, decreased proliferation with more senescent cells was noted, which was probably mediated through FGFR1 and the downstream STAT3 signaling pathway.

Testicular damage is the anomaly that will often accompany diabetes mellitus. Thus, this study aimed to investigate the role that total flavonoids of Epimedium (TFE) played against streptozotocin (STZ)-induced diabetic testicular dysfunction and to elucidate the mechanism of action. The diabetic rat model was induced by vein injection of STZ in healthy rats. Thirty male healthy Spraque-Dawley rats were randomly divided into following groups: the control group, the diabetic group, and the diabetic + TFE group. Gastrointestinal administration begins at fifth week of TFE for 6 weeks. After TFE administration, all animals were euthanized. Testicular tissue samples and blood samples of rats were collected for histopathological examination and for determination of levels of various biomarkers including blood glucose, testosterone, testicular enzymes, and oxidative stress indicators. All testes were weighted to calculate the testicular organ index. Hematoxylin-eosin staining was used for observing the testis and epididymis pathological changes. Protein expression (monocyte chemoattractant protein-1, transforming growth factor-beta-1, interleukin-6, and 3-beta-hydroxysteroid dehydrogenase) was detected by immunohistochemistry and western blot techniques. There was a significant difference in the changes between the diabetes group and the control group. As a result of treat with TFE, the blood glucose decreased but there was no significant difference, and other indicators showed significant improvement. TFE may protect against STZ-induced testicular injury by suppressing inflammation and oxidative stress.

Background Glioblastoma (GBM) is a malignant human brain tumor that has an extremely poor prognosis. Classic mutations such as IDH (isocitrate dehydrogenase) mutations, EGFR (epidermal growth factor receptor) alternations, and MGMT (O6-methylguanine-methyltransferase) promoter hypermethylation have been used to stratify patients and provide prognostic significance. Epigenetic perturbations have been demonstrated in glioblastoma tumorigenesis. Despite the genetic markers used in the management of glioblastoma patients, new biomarkers that could predict patient survival independent of known biomarkers remain to be identified. Methods ATAC-seq (assay for transposase accessible chromatin followed by sequencing) and RNA-seq have been used to profile chromatin accessible regions using glioblastoma patient samples with short-survival versus long-survival. Cell viability, cell cycle, and Western blot analysis were used to characterize the cellular phenotypes and identify signaling pathways. Results Analysis of chromatin accessibility by ATAC-seq coupled with RNA-seq methods identified the GSTM1 (glutathione S-transferase mu-1) gene, which featured higher chromatin accessibility in GBM tumors with short survival. GSTM1 was confirmed to be a significant prognostic marker to predict survival using a different GBM patient cohort. Knockdown of GSTM1 decreased cell viability, caused cell cycle arrest, and decreased the phosphorylation levels of the NF-kB (nuclear factor kappa B) p65 subunit and STAT3 (signal transducer and activator of transcription 3) (pSer727). Conclusions This report demonstrates the use of ATAC-seq coupled with RNA-seq to identify GSTM1 as a prognostic marker of GBM patient survival. Activation of phosphorylation levels of NF-kB p65 and STAT3 (pSer727) by GSTM1 is shown. Analysis of chromatin accessibility in patient samples could generate an independent biomarker that can be used to predict patient survival.

Dissemination of multidrug-resistant, particularly tigecycline-resistant, Acinetobacter baumannii is of critical importance, as tigecycline is considered a last-line antibiotic. Acquisition of tet(X), a tigecycline-inactivating enzyme mostly found in strains of animal origin, imparts tigecycline resistance to A. baumannii. Herein, we investigated the presence of tet(X) variants among 228 tigecycline-non-susceptible A. baumannii isolates from patients at a Taiwanese hospital via polymerase chain reaction using a newly designed universal primer pair. Seven strains (3%) carrying tet(X)-like genes were subjected to whole genome sequencing, revealing high DNA identity. Phylogenetic analysis based on the PFGE profile clustered the seven strains in a clade, which were thus considered outbreak strains. These strains, which were found to co-harbor the chromosome-encoded tet(X6) and the plasmid-encoded blaOXA-72 genes, showed a distinct genotype with an uncommon sequence type (Oxford ST793/Pasteur ST723) and a new capsular type (KL129). In conclusion, we identified an outbreak clone co-carrying tet(X6) and blaOXA-72 among a group of clinical A. baumannii isolates in Taiwan. To the best of our knowledge, this is the first description of tet(X6) in humans and the first report of a tet(X)-like gene in Taiwan. These findings identify the risk for the spread of tet(X6)-carrying tigecycline- and carbapenem-resistant A. baumannii in human healthcare settings.

Parkinson's disease (PD), the most common neurodegenerative motor disorder, is currently incurable. Although many studies have provided insights on the substantial influence of genetic factors on the occurrence and development of PD, the molecular mechanism underlying the disease is largely unclear. Previous studies have shown that point mutations in the phospholipase A2 group VI gene (PLA2G6) correlate with young-onset dystonia-parkinsonism type 14 (PARK14). However, limited information is available regarding the pathogenic role of this gene and the mechanism underlying its function. To study the role of PLA2G6 mutations, we first used zebrafish larvae to screen six PLA2G6 mutations and revealed that injection of D331Y, T572I, and R741Q mutation constructs induced phenotypes such as motility defects and reduction in dopaminergic neurons. The motility defects could be alleviated by treatment with L-3, 4-dihydroxyphenylalanine (L-dopa), indicating that these mutations are pathological for PARK14 symptoms. Furthermore, the injection of D331Y and T572I mutation constructs reduced phospholipase activity of PLA2G6 and its lipid metabolites, which confirmed that these two mutations are loss-of-function mutations. Metabolomic analysis revealed that D331Y or T572I mutation led to higher phospholipid and lower docosahexaenoic acid (DHA) levels, indicating that reduced DHA levels are pathological for defective motor functions. Further, a dietary DHA supplement relieved the motility defects in PLA2G6D331Y/D331Y knock-in mice. This result revealed that the D331Y mutation caused defective PLA2G6 phospholipase activity and consequently reduced the DHA level, which is the pathogenic factor responsible for PARK14. The results of this study will facilitate the development of therapeutic strategies for PARK14.

In higher-order conditioning paradigms, such as sensory preconditioning or second-order conditioning, discrete (e.g., phasic) or contextual (e.g., static) stimuli can gain the ability to elicit learned responses despite never being directly paired with reinforcement. The purpose of this mini-review is to examine the neuroanatomical basis of high-order conditioning, by selectively reviewing research that has examined the role of the retrosplenial cortex (RSC) in sensory preconditioning and second-order conditioning. For both forms of higher-order conditioning, we first discuss the types of associations that may occur and then review findings from RSC lesion/inactivation experiments. These experiments demonstrate a role for the RSC in sensory preconditioning, suggesting that this cortical region might contribute to higher-order conditioning via the encoding of neutral stimulus-stimulus associations. In addition, we address knowledge gaps, avenues for future research, and consider the contribution of the RSC to higher-order conditioning in relation to related brain structures.

Amblyomma javanense is an important ectoparasite of Manis javanica, although the population genetics, molecular biology and systematics of A. javanense remain poorly understood. In the present study, the mitochondrial genome of A. javanense was sequenced using the Illumina HiSeq sequencing platform (Illumina, San Diego, CA, U.S.A.) and compared with the genomes of two closely related species: Amblyomma fimbriatum and Amblyomma americanum. The intraspecies and interspecies relationships of A. javanense and another 21 selected species were investigated by constructing a maximum-likelihood tree and a neighbour-joining tree. The mitochondrial genome of A. javanense was 14 780 bp in length and contained 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes and two control regions. The results of the comparisons indicate that there is great similarity among these three species, and both trees indicate that A. javanense is a member of the Amblyomminae. The study of A. javanense of pangolins also indicates the premise and foundation of the relationship between the parasite and other species.

Rhipicephalus microplus salivary gland secretes a number of complex bioactive proteins during feeding. These components are important in feeding and affect anti-coagulation, anti-inflammation and also have anti-microbial effects. In this study, tick saliva was collected from partially engorged female (PEF) and fully engorged female (FEF) ticks. Liquid chromatography tandem-mass spectrometry (LC-MS/MS) and isobaric tags for relative and absolute quantification (iTRAQ) were used to identify and quantify R. microplus salivary proteins. A total of 322 unique peptides were detected and 151 proteins were characterized in both PEF and FEF. Of these, 41 proteins are considered as high-confidence proteins. Fifteen high-confidence proteins were upregulated and six high-confidence proteins were downregulated (p 0.05; PEF:FEF ratio ≥ 1.2 or PEF:FEF ratio ≤ 0.83); 17 high-confidence proteins are slightly changed (PEF:FEF ratio 0.83 and 1.2). These high-confidence proteins are involved in several physiological roles, including egg development, transportation of proteins, immunity and anti-microorganism, anti-coagulant, and adhesion. In comparison with PEF, the number of upregulated proteins exceeded the number of proteins downregulated. Salivary protein may be induced by the blood-meal and these proteins contribute to successful feeding.

Ticks and tick-borne diseases are major global health threats. During blood feeding, ticks insert their hypostomes into hosts and inject an array of anticoagulant molecules to maintain fluidity of the blood-meal. These anticoagulant molecules may provide insights into understanding the feeding biology of ticks and to develop vaccines against infestations. In Haemaphysalis flava, the heat shock cognate 70 (HSC70), a member of the heat shock protein (HSP) family, is differentially expressed in salivary glands at different levels of engorgement during blood feeding. However, its function in ticks is largely not known. The present study was designed to explore the possible effects of HSC70 on the plasma. The open reading frame (ORF) of HSC70 was expressed in a prokaryotic system, and recombinant HSC70 (rHSC70) was purified and characterized. The anticoagulation activity of rHSC70 was estimated by measuring prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT) and fibrinogen (FIB) with/without its inhibitor, VER155008. The results demonstrated that rHSC70 from H. flava extended TT (P 0.001) and FIB clotting times (300 s), but showed little effect on PT and APTT. Adding an inhibitor reversed anticlotting effects of rHSC70 on TT and FIB. These data indicate that rHSC70 is an anticoagulant agent, and the anticlotting activity likely attributes to the inhibition of thrombin and the transformation of fibrinogen into fibrin.

Background Many molecules and signaling pathways involved in neural development play a role in neurodegenerative diseases and brain tumor progression. Peroxisome proliferator-activated receptor (PPAR) proteins regulate the differentiation of tissues and the progression of many diseases. However, the role of these proteins in neural development is unclear. Results We examined the function of Pparα in the neural development of zebrafish. Two duplicate paralogs for mammalian PPARA/Ppara, namely pparaa and pparab, are present in the zebrafish genome. Both pparaa and pparab are expressed in the developing central nervous system in zebrafish embryos. Inhibiting the function of Pparα by using either the PPARα/Pparα antagonist GW6471 or pparaa or pparab truncated constructs produced identical phenotypes, which were sufficient to reduce the proliferation of neuronal and glial precursor cells without affecting the formation of neural progenitors. Conclusions We demonstrated that both Pparαa and Pparαb proteins are essential regulators of the proliferation of neuronal and glial precursors. This study provides a better understanding of the functions of PPARα/Pparα in neural development and further expands our knowledge of the potential role of PPARα/Pparα in neurological disorders and brain tumors. Developmental Dynamics 247:1264-1275, 2018. © 2018 Wiley Periodicals, Inc.

Haemaphysalis flava (Acari: Ixodidae) harbors pathogenic microorganisms and transfers these to hosts during blood feeding. Proteomic analysis in the midgut contents of H. flava detected glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and contig 1683 was retrieved as a GAPDH gene fragment by searching our previous transcriptomic library. In the study, the 5' and 3' ends of contig 1683 were cloned by rapid amplification of cDNA ends (RACE) and a full length, 1340 bp cDNA of Hf-GAPDH was obtained. The open-reading frame had 999 bp and coded for 333 amino acids. Hf-GAPDH was predicted to have an N-terminal NAD binding domain and a C-terminal glyceraldehyde dehydrogenase catalytic domain. The molecular structure of Hf-GAPDH was analyzed and the evolutionary relationship also established. The GAPDH protein sequence was conserved among ticks. The expression pattern of Hf-GAPDH, analyzed by real-time PCR, significantly differed among life phases, feeding stages, and tissues. As the ticks grew, the expression level of Hf-GAPDH was up-regulated. The expression levels of Hf-GAPDH in salivary glands and midguts from half-engorged ticks were lower than the same tissues from engorged ticks. This study will provide reference data for the follow-up verification of the GAPDH-related function and the feasibility as a potential anti-tick vaccine.

Ticks are obligate blood-sucking ectoparasites that infect a wide range of animals and humans, causing a variety of both human and animal diseases around the world. Ixodes nipponensis is the most commonly reported tick in Korea and Japan, but it is very rare in China. In this study, six I. nipponensis samples were collected from three black goats in Hunan province, China. Ticks identified morphologically as I. nipponensis were then examined by PCR with two different molecular markers: mitochondrial cox1 and the second internal transcribed spacer of ribosomal DNA genes. Sequence comparison and phylogenetic analysis of the cox1 sequences confirmed that all of the examined hard Ixodes ticks represented I. nipponensis. This finding indicates a potential risk of zoonotic I. nipponensis infection in humans and animals in China. To our knowledge, this is the first report documenting the occurrence of I. nipponensis infection in goats in China.

Circadian disruption-induced metabolic syndrome (CDIMS) involves body weight gain, changes in blood profile and gut microbiota. In this study, CDIMS was induced by shifted light dark cycle (SLDC) in C57BL/6J mice. Dietary intervention by oral administration of oat β-glucan (a polymeric prebiotic) alleviated CDIMS when compared to chicory inulin/fructan (an oligomeric prebiotic) and melatonin (a chronobiotic). Oat β-glucan reversed the increase in body weight, liver weight-to-body weight ratio and plasma leptin concentration as well as restored glucose tolerance. In altering gut microbiota, oat β-glucan increased the species richness, reversed the populations of 7 bacterial genera and increased butyrate producers including Ruminococcaceae and Lachnospiraceae which enhance gut barrier protection and regulate glucose homeostasis. Correlation analysis demonstrated the linking of the alleviation of CDIMS by prebiotics and melatonin with different microbial metabolic pathways involved in energy metabolism, biosynthesis of metabolites, metabolism of cofactors and vitamins and endocrine synthesis.

Argas persicus is an ectoparasite of poultry. The bacterial community structure and the pathogenic bacteria associated with different developmental stages of A. persicus have implications for control. Argas persicus were collected from chickens in the city of Jiuquan in Gansu, China. Bacterial DNA was extracted from the midgut contents of blood engorged larvae, nymphs and adult females. The V3-V4 hypervariable regions of 16S rRNA genes were sequenced using the IonS5TMXL platform. Identification of Rickettsia spp. and detection of Coxiella burnetii were performed using PCR on target genes. The bacterial diversity within larvae was the highest and the bacterial diversity within nymphs was greater than that of adults. At different classification levels, seven bacterial phyla were common phyla, 27 genera were common genera, and 18 species were common species in the three samples. At the phylum level, Proteobacteria showed a marked predominance in all samples. Rickettsia, Stenotrophomonas, Spiroplasma, and Coxiella were the dominant bacteria at the genus level. The Rickettsia species in A. persicus was identified as Rickettsia hoogstraalii and the Coxiella species was identified as a Coxiella-like endosymbiont. Additionally, some bacterial species such as Pseudomonas geniculata, Sphingomonas koreensis, and Acinetobacter haemolyticus are reported here for the first time in A. persicus.

In this primer, Cheng et al. outline what we know about the nature and control of differentiation of germline versus somatic nuclei in two groups of protozoa: the Ciliates and Foraminifera. This is shown to involve a remarkable variety of developmentally programmed phenomena such as genome editing mediated epigenetically by RNA, as well differential nuclear import.

We proposed a framework with parametric modeling to obtain biological relevant parameters from the total probiotic growth pattern and metabolite production curves. The lag phase, maximum increase rate, and maximum capacity were obtained via a 205-h exploratory In vitro fermentation of a library of 13 structural-characterized prebiotic candidates against an exclusively breastfed infant fecal inoculum. We also conducted 16S rRNA amplicon sequencing of the infant fecal inoculum. Moreover, we introduce a robust composite metabolite-based indicator that reflects the eubiosis/dysbiosis of microbiota to complement the conventional microbial markers. In terms of short-chain fatty acid, we discovered that polymeric beta-glucans from barley demonstrated potential as prebiotic candidates, while alpha-glucans as glycogen showed the least dissolved ammonia production. In terms of total probiotic, beta-glucans from oat and mushroom sclerotia of Pleurotus tuber-regium showed comparable sustainability when compared to alpha-glucans after 48 h. Being classical prebiotic, galacto-oligosaccharides gave the second-highest metabolite-based indicator, followed by lactose. While limited improvement could be made to lactose and oligosaccharides, polymeric beta-glucans from barley avails more capacity for novel prebiotic development, such as structural modification. We anticipate that more similar parallel screening with the element of time and parametric modeling will provide more novel insights.

DNA methylation and demethylation at CpG di-nucleotide sites plays important roles in cell fate specification of neural stem cells (NSCs). We have previously reported that DNA methyltransferases, Dnmt1and Dnmt3a, serve to suppress precocious astrocyte differentiation from NSCs via methylation of astroglial lineage genes. However, whether active DNA demethylase also participates in astrogliogenesis remains undetermined. In this study, we discovered that a Ten-eleven translocation (Tet) protein, Tet2, which was critically involved in active DNA demethylation through oxidation of 5-Methylcytosine (5mC), drove astrocyte differentiation from NSCs by demethylation of astroglial lineage genes including Gfap. Moreover, we found that an NSC-specific bHLH transcription factor Olig2 was an upstream inhibitor for Tet2 expression through direct association with the Tet2 promoter, and indirectly inhibited astrocyte differentiation. Our research not only revealed a brand-new function of Tet2 to promote NSC differentiation into astrocytes, but also a novel mechanism for Olig2 to inhibit astrocyte formation.

In endolysosomal networks, two hetero-hexameric tethers called HOPS and CORVET are found widely throughout eukaryotes. The unicellular ciliate Tetrahymena thermophila possesses elaborate endolysosomal structures, but curiously both it and related protozoa lack the HOPS tether and several other trafficking proteins, while retaining the related CORVET complex. Here, we show that Tetrahymena encodes multiple paralogs of most CORVET subunits, which assemble into six distinct complexes. Each complex has a unique subunit composition and, significantly, shows unique localization, indicating participation in distinct pathways. One pair of complexes differ by a single subunit (Vps8), but have late endosomal versus recycling endosome locations. While Vps8 subunits are thus prime determinants for targeting and functional specificity, determinants exist on all subunits except Vps11. This unprecedented expansion and diversification of CORVET provides a potent example of tether flexibility, and illustrates how ‘backfilling’ following secondary losses of trafficking genes can provide a mechanism for evolution of new pathways. This article has an associated First Person interview with the first author of the paper.

Background: Although particular matter (PM) increases incidence and severity of idiopathic pulmonary fibrosis, the underlying mechanism remains elusive. Methods: The effects of PM were evaluated in a murine model of bleomycin-induced pulmonary fibrosis. Mice were divided into four groups, receiving: (1) Saline (control), (2) bleomycin, (3) PM, or (4) bleomycin plus PM (Bleo+PM). Additional groups of Bleo+PM mice were treated with sivelestat (an inhibitor of neutrophil elastase) or reparixin (a C-X-C motif chemokine receptor 2 antagonist), or were genetically modified with keratinocyte chemoattractant (KC) deletion. Results: Pulmonary fibrosis was not observed in the control or PM groups. Bleomycin induced pulmonary fibrosis within 14 days. The Bleo+PM group showed worse pulmonary fibrosis when compared to the bleomycin group. Analyses of immune cell profile and chemokine/cytokine concentrations at day 2-bronchoalveolar lavage fluid (BALF) revealed that the Bleo+PM group had increased neutrophil number and elastase level and KC concentration compared to the bleomycin group. Neutrophil elastase activated the Smad2/Smad3/&alpha, SMA pathway to induce collagen deposition, while sivelestat abrogated the increased severity of pulmonary fibrosis caused by PM. Chemotaxis assay revealed that BALF of the Bleo+PM group recruited neutrophil, which was dependent on KC. Further, genetic KC deletion or pharmaceutical inhibition of KC binding to CXCR2 with reparixin ameliorated the PM-induced increased severity of pulmonary fibrosis. Conclusions: These data provide evidence that the PM-induced increased severity of pulmonary fibrosis depends on KC-mediated neutrophil chemotaxis and give additional mechanic insight that will aid in the development of therapeutic strategies.

Allergic rhinitis (AR) is a worldwide highly prevalent nasal inflammatory disease with elusive mechanisms about the regulation of innate immune response. The roles and mechanisms of NLRP3, a typical inflammasome, in AR development remain unclear. Here we investigate the roles of NLRP3 inflammasome activation in the development and progression of AR and try to uncover its potential mechanisms underlying. Wildtype and NLRP3 knockout mice were applied to construct the ovalbumin (OVA)-induced AR model. Caspase-1 specific inhibitor Belnacasan and inflammasome activator ATP were used for adjuvant stimulation of AR-model mice respectively. We found that the production of IL-1β and the activation of inflammasome were increased in both patients and mice with AR. NLRP3 deficiency markedly suppressed AR progression with reduced inflammatory response and epithelium pyroptosis in mice with AR. Furthermore, Caspase-1 inhibitor treatment in vivo ameliorated the development and progression of AR with favorable outcomes. Mechanistically, inflammation augments and nasal mucosa injury during AR were partially due to ASC-specks accumulation and subsequent cell pyroptosis. Our study reveals the previously unknown roles of NLRP3 inflammasome in promoting the development and progression of AR via enhancing inflammatory response and epithelium pyroptosis and thus provides a potential clue for allergic disease interventions.

Scant information is available regarding the proteins involved in blood meal processing in ticks. Here, we aimed to highlight the midgut proteins involved in preventing blood meal coagulation, and in facilitating intracellular digestion in the tick Haemaphysalis flava. Proteins were extracted from the midgut contents of fully engorged and partially engorged ticks. We used liquid chromatography tandem-mass spectrometry (LC-MS/MS) analysis to identify 131 unique peptides, and 102 proteins. Of these, 15 proteins, each with at least two unique peptides, were recognized with high confidence. We also retrieved 18 unigenes from our previous published transcriptomic libraries of the midguts and salivary glands of H. flava, and inferred the primary structures of nine proteins and fragments of five proteins. There were 23 and 21 unique proteins in the midgut contents of fully engorged and partially engorged ticks, respectively. We detected 58 shared proteins in the midgut contents of both fully engorged and partially engorged ticks. Of these, seven were significantly differentially expressed between fully engorged and partially engorged ticks: actin, calmodulin, elongation factor-1α, hsp90, multifunctional chaperone, tubulin α, and tubulin β. Our results demonstrated that the proteome of the midgut contents, combined with the transcriptome of the midgut, was a viable method for the reinforcement of protein identification. This method will facilitate further study of blood meal processing by ticks, as well as the identification of clues for tick infestation control. The existence of numerous proteins detected in the midgut contents also highlight the complexity of blood digestion in ticks; this area is in need of further investigation.

Uremic toxins, such as indoxyl sulfate (IS) and kynurenine, accumulate in the blood in the event of kidney failure and contribute to further bone damage. To maintain the homeostasis of the skeletal system, bone remodeling is a persistent process of bone formation and bone resorption that depends on a dynamic balance of osteoblasts and osteoclasts. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates the toxic effects of uremic toxins. IS is an endogenous AhR ligand and is metabolized from tryptophan. In osteoclastogenesis, IS affects the expression of the osteoclast precursor nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) through AhR signaling. It is possible to increase osteoclast differentiation with short-term and low-dose IS exposure and to decrease differentiation with long-term and/or high-dose IS exposure. Coincidentally, during osteoblastogenesis, through the AhR signaling pathway, IS inhibits the phosphorylation of ERK, and p38 reduces the expression of the transcription factor 2 (Runx2), disturbing osteoblastogenesis. The AhR antagonist resveratrol has a protective effect on the IS/AhR pathway. Therefore, it is necessary to understand the multifaceted role of AhR in CKD, as knowledge of these transcription signals could provide a safe and effective method to prevent and treat CKD mineral bone disease.

The effect of DNA topology on transfection efficiency of mammalian cells has been widely tested on plasmids smaller than 10 kb, but little is known for larger DNA vectors carrying intact genomic DNA containing introns, exons, and regulatory regions. Here, we demonstrate that circular BACs transfect more efficiently than covalently closed linear BACs. We found up to 3.1- and 8.9- fold higher eGFP expression from circular 11 kb and 100 kb BACs, respectively, compared to linear BACs. These findings provide insights for improved vector development for gene delivery and expression studies of large intact transgenes in mammalian cells.

Background: The Beijing lineage of Mycobacterium tuberculosis (MTB) is the most predominant MTB strain in Asian countries and is spreading worldwide, however, the East AfricanâIndian (EAI) lineage is also particularly prevalent in many tropical Asian countries. The evolutionary relationships among MTB EAI isolates from Taiwan and those of tropical Asian countries remain unknown. Methods: The EAI strains collected from patients in Taiwan were analyzed using spacer oligonucleotide typing and mycobacterial interspersed repetitive unitâvariable number of tandem repeats (MIRU-VNTR) typing, and compared with published profiles from Cambodia and Singapore to investigate potential epidemiological linkages. Results: Among the three countries, the EAI lineage was most prevalent in Cambodia (60%; Singapore, 25.62%; and Taiwan, 21.85%), having also the highest rates of multidrug resistance and lowest rates of clustering of MTB isolates. We describe a convenient method using seven selected MIRU-VNTR loci for first-line typing to discriminate Beijing and EAI lineages. A potential epidemiological linkage in these tropical Asian countries is also discussed based on a minimum-spanning tree constructed using 24 MIRU-VNTR loci of MTB EAI strains. Conclusion: This study identified evolutionary relationships among MTB EAI isolates from Taiwan and those of two other tropical Asian countries, Cambodia and Singapore. Keywords: East AfricanâIndian family, Mycobacterium tuberculosis, tropical Asian countries

Background Growing evidence indicates that oxidative stress (OS) plays a pivotal role in Diabetic nephropathy (DN). In a previous study we demonstrated that ALA/LA protected HK-2 cells against high glucose-induced cytotoxicity. So we aimed to establish the glucose injury model of HK-2 cells and investigate the beneficial effects of ALA/LA on high glucose-induced excessive production of TGF-β1 and the possible mechanisms mediating the effects. Methods The expression of OS markers in high glucose-induced HK-2 cells treated with ALA/LA., including the antioxidant enzymes and reactive oxygen species (ROS) production, as well as the apoptosis rate were assayed by ELISA and flow cytometry. The p38/transforming growth factor β1 (TGF-β1) signal pathway were measured by real-time RT-PCR and western blot. Results The modeling condition of glucose toxicity on HK-2 cells was at the glucose concentration of 40.9 mM. ALA/LA can significantly increase the activities of antioxidant enzymes and decrease ROS production stimulated by high glucose. The study also found that ALA/LA caused a decrease in the apoptosis rate and TGF-β1 level of HK-2 cells under high glucose stress through the ROS/p38 pathway. Conclusions ALA/LA exerts protective effects in vitro through inhibition of ROS generation, down regulation of the activation of the p38MAPK pathway and the expression of TGF-β1 in HK-2 cells.

Runt-related transcription factor 1(RUNX1), a key factor in hematopoiesis that mediates specification and homeostasis of hematopoietic stem and progenitor cells (HSPCs), is also overexpressed in several solid human cancers, and correlated with tumor progression. However, the expression and function of RUNX1 in pancreatic ductal adenocarcinoma were still unclear. Here, we show that RUNX1 is highly expressed in pancreatic adenocarcinoma tissues and knocking down of RUNX1 attenuated aggressiveness in pancreatic cell lines. Moreover, we found that RUNX1 could negatively regulate the expression of miR-93. Bioinformatics method showed that there are two binding sites in the the promotor region of miR-93 precursor and through ChIP-qPCR and firefly luciferase reporter assay, we vertified that these two binding sites each have transcriptive activity in one pancreatic cell lines. This result supported our presumption that RUNX1 regulate miR-93 through binding to the promotor region of miR-93. Besides, the expression and function of miR-93 is quite the opposite, miR-93 overexpression suppresses migration and invasiveness in pancreatic cell lines supporting that RUNX1 negatively regulated miR-93. Our findings provided evidence regarding the role of RUNX1 as an oncogene through the inhibition of miR-93. Targeting RUNX1 can be a potential therapeutic strategy in pancreatic cancer.

The ciliate protozoan Cryptocaryon irritans parasitizes marine fish and causes lethal white spot disease. Sporadic infections as well as large-scale outbreaks have been reported globally and the parasite's broad host range poses particular threat to the aquaculture and ornamental fish markets. In order to better understand C. irritans' population structure, we sequenced and compared mitochondrial cox-1, SSU rRNA, and ITS-1 sequences from 8 new isolates of C. irritans collected in China, Japan, and Taiwan. We detected two SSU rRNA haplotypes, which differ at three positions, separating the isolates into two main groups (I and II). Cox-1 sequences also support the division into two groups, and the cox-1 divergence between these two groups is unexpectedly high (9.28% for 1582 nucleotide positions). The divergence is much greater than that detected in Ichthyophthirius multifiliis, the ciliate protozoan causing freshwater white spot disease in fish, where intraspecies divergence on cox-1 sequence is only 1.95%. ITS-1 sequences derived from these eight isolates and from all other C. irritans isolates (deposited in the GenBank) not only support the two groups, but further suggest the presence of a third group with even greater sequence divergence. Finally, a small Ka/Ks ratio estimated from cox-1 sequences suggests that this gene in C. irritans remains under strong purifying selection. Taken together, the C. irritans species may consists of many subspecies and/or syngens. Further work is needed to determine if there is reproductive isolation between the groups we have defined.

Glioblastomas are among the most fatal brain tumors; however, the molecular determinants of their tumorigenic behavior are not adequately defined. In this study, we analyzed the role of KMT2A in the glioblastoma cell line U-87 MG. KMT2A knockdown promoted cell proliferation. Moreover, it increased the DNA methylation of NOTCH1 and NOTCH3 and reduced the expression of NOTCH1 and NOTCH3. NOTCH1 or NOTCH3 activation inhibited U-87 MG cell proliferation, whereas NOTCH1 and NOTCH3 inhibition by shRNAs induced cell proliferation, thus demonstrating the tumor-suppressive ability of NOTCH1 and NOTCH3 in U-87 MG cells. The induced cell proliferation caused by KMT2A knockdown could be nullified by using either constitutively active NOTCH1 or constitutively active NOTCH3. This result demonstrates that KMT2A positively regulates NOTCH1 and NOTCH3 and that this mechanism is essential for inhibiting the U-87 MG cell proliferation. The role of KMT2A knockdown in promoting tumor growth was further confirmed in vivo by transplanting U-87 MG cells into the brains of zebrafish larvae. In conclusion, we identified KMT2A-NOTCH as a negative regulatory cascade for glioblastoma cell proliferation, and this result provides important information for KMT2A- or NOTCH-targeted therapeutic strategies for brain tumors.

Ticks and tick-borne-diseases have serious public health implications, and screening feasible protein candidates for vaccines development is identified to be an effective alternative to control of tick infestations. In current study, we focused on cloning the full-length gene encoding a heat shock cognate protein 70 (Hsc70), a molecular chaperone of critical functional roles belonging to heat shock protein 70 (HSP70) family, in salivary glands of Haemaphysalis flava, namely Hf-Hsc70, and analyzing the expression of Hf-Hsc70 in different life phases, organs and ambient temperatures. Rapid amplification of cDNA ends (RACE) was performed to amplify the 5′ and 3′ ends of Hf-Hsc70. The expression profiles of Hf-Hsc70 were studied by semi-quantitative real-time PCR (RT-PCR). The full-length of Hf-Hsc70 was 2363 bp, and contained an ORF of 1965 bp encoding a protein of 648 amino acids. The expression levels of Hf-Hsc70 at different life phases were in the order of female larvae < female fully engorged nymphs < male adult ticks < female full engorged adult ticks < female half engorged adult ticks. The relative expression of Hf-Hsc70 in salivary glands was steadily higher than that in midguts (p

Neural crest cells are multipotent progenitors that migrate extensively and differentiate into numerous derivatives. The developmental plasticity and migratory ability of neural crest cells render them an attractive model for studying numerous aspects of cell progression. We observed that zebrafish rgs2 was expressed in neural crest cells. Disrupting Rgs2 expression by using a dominant negative rgs2 construct or rgs2 morpholinos reduced GTPase-activating protein activity, induced the formation of neural crest progenitors, increased the proliferation of nonectomesenchymal neural crest cells, and inhibited the formation of ectomesenchymal neural crest derivatives. The transcription of pparda (which encodes Pparδ, a Wnt-activated transcription factor) was upregulated in Rgs2-deficient embryos, and Pparδ inhibition using a selective antagonist in the Rgs2-deficient embryos repaired neural crest defects. Our results clarify the mechanism through which the Rgs2–Pparδ cascade regulates neural crest development; specifically, Pparδ directly binds to the promoter and upregulates the transcription of the neural crest specifier sox10. This study reveals a unique regulatory mechanism, the Rgs2–Pparδ–Sox10 signaling cascade, and defines a key molecular regulator, Rgs2, in neural crest development.

Background The tuberculosis (TB) pandemic remains a leading cause of human morbidity and mortality, despite widespread use of the only licensed anti-TB vaccine, bacille Calmette-Guerin (BCG). The protective efficacy of BCG in preventing pulmonary TB is highly variable; therefore, an effective new vaccine is urgently required. Methods In the present study, we assessed the ability of novel recombinant BCG vaccine (rBCG) against Mycobacterium tuberculosis by using modern immunological methods. Results Enzyme-linked immunospot assays demonstrated that the rBCG vaccine, which coexpresses two mycobacterial antigens (Ag85B and CFP10) and human interleukin (IL)-12 (rBCG2) elicits greater interferon-γ (IFN-γ) release in the mouse lung and spleen, compared to the parental BCG. In addition, rBCG2 triggers a Th1-polarized response. Our results also showed that rBCG2 vaccination significantly limits M. tuberculosis H37Rv multiplication in macrophages. The rBCG2 vaccine surprisingly induces significantly higher tumor necrosis factor-α (TNF-α) production by peripheral blood mononuclear cells that were exposed to a nonmycobacterial stimulus, compared to the parental BCG. Conclusion In this study, we demonstrated that the novel rBCG2 vaccine may be a promising candidate vaccine against M. tuberculosis infection.

Bone metastasis is a deadly consequence of cancers, in which osteoclast forms a vicious cycle with tumor cells. Bone metastasis attenuation by clinical usage of osteoclast inhibitors and in our osteopetrotic mouse genetic models with β-catenin constitutive activation or peroxisome proliferator-activated receptor γ deficiency fully support the important role of osteoclast in driving the bone metastatic niche. However, the mechanisms for this "partnership in crime" are underexplored. Here we show that osteoclasts reprogram their lipid secretion to support cancer cells. Metabolomic profiling reveals elevated prometastatic arachidonic acid (AA) but reduced antimetastatic lysophosphatidylcholines (LPCs). This shift in lipid osteoclastokines synergistically stimulates tumor cell proliferation, migration, survival, and expression of prometastatic genes. Pharmacologically, combined treatment with LPCs and BW-755C, an inhibitor of AA signaling via blocking lipoxygenase and cyclooxygenase, impedes breast cancer bone metastasis. Our findings elucidate key paracrine mechanisms for the osteoclast-cancer vicious cycle and uncover important therapeutic targets for bone metastasis.

A new naphthol derivative, [F-18]FEONM, has been designed to be a new Tau protein PET imaging molecule for detecting Tau tangle status of Alzheimer's disease in a transgenic mouse model. In order to synthesize this molecule, 2-acetyl-6-methoxynaphthalene was the starting compound and needed five steps preparation to bonding functional and leaving group for preparing the precursor. Radiofluorination on the precursor was carried out by a modified [F-18]FDG autosynthesizer. After half an hour processing, it yielded more than 99% chemical and radiochemical purity product at up to 60% yield. Radiofluorination kinetic study showing 80% [F-18]FEONM would be produced within the first 5 min. The radiofluorination gap function factor is 0.122 for 150 min reaction. Recovery yield of crude [F-18]FEONM could be up to 60% by semi-preparative HILIC HPLC column purification with 95% ethanol elution. Brain glioma tumor mice PET imaging were performed after tail vein injecting [F-18]FEONM. Binding in mice brain tumor reached steady state after 30 min injection and shows 30–70% higher uptake compared to normal mouse. The static brain tumor uptake showed the summation accumulation binding rate was 60% faster than normal mouse. In vitro postmortem AD patient brain slice binding showed 70% higher uptake than normal brain. The uptake of spherical stem cells also showed higher association with neuronal differentiation and PET imaging of Tau and ABeta transgenic mice also showed visualized difference. The specific binding ratio of hippocampus to cerebellum of control mouse is 46.56%, cortex to cerebellum is 11.08% in a 12-month-old P301S Tau transgenic mouse; hippocampus to cerebellum is 106.66%, cortex to cerebellum is 82.48%. Relative ratio of hippocampus to cerebellum of a P301S mouse to control mouse is 2.29, cortex to cerebellum is 7.44.

Ciliates are unicellular eukaryotes known for their cellular complexity and wide range of natural habitats. How they adapt to their niches and what roles they play in ecology remain largely unknown. The genus Tetrahymena is among the best-studied groups of ciliates and one particular species, Tetrahymena thermophila, is a well-known laboratory model organism in cell and molecular biology, making it an excellent candidate for study in protist ecology. Here, based on cytochrome c oxidase subunit I (COX1) gene barcoding, we identify a total of 19 different putative Tetrahymena species and two closely related Glaucoma lineages isolated from distinct natural habitats, of which 13 are new species. These latter include 11 Tetrahymena species found in the bladder traps of Utricularia plants, the most species-rich and widely distributed aquatic carnivorous plant, thus revealing a previously unknown but significant symbiosis of Tetrahymena species living among the microbial community of Utricularia bladder traps. Additional species were collected using an artificial trap method we have developed. We show that diverse Tetrahymena species may live even within the same habitat and that their populations are highly dynamic, suggesting that the diversity and biomass of species worldwide is far greater than currently appreciated.

The heat shock protein 70 (HSPA) family and their genes have been studied in ticks and are considered as possible antigen candidates for the development of anti-tick vaccines. However, knowledge about their members, structure and function in ticks is incomplete. Based on our transcriptomic data, the full length of four HSPA genes in Haemaphysalis flava (Acari: Ixodidae) was cloned via rapid amplification of cDNA ends. The open reading frame of HSPA2A, HSPA2B, HSPA5 and HSPA9 was 1920, 1911, 1983 and 2088 bp in length, respectively. Three family signatures and one localization motif were in the encoding proteins. HSPA2A and HSPA2B were predicted to be located at cytoplasm/nucleus, whereas HSPA5 and HSPA9 were at endoplasmic reticulum and mitochondria, respectively. In silico simulation demonstrated that those proteins had distinct numbers of α-helixes, extended strands and coils, and different antigenic epitopes. Expression of HSPA5 and HSPA9 in the salivary gland was significantly higher in partially-engorged female adult ticks than the fully-engorged (P < 0.01) as shown by a quantitative polymerase chain reaction. Our data indicated that H. flava ticks had at least four HSPA genes encoding proteins with different cellular locations, structures and expression profiles, suggesting their diverse roles in tick biology.