Dietary emulsifier polysorbate 80 exposure accelerates age-related cognitive decline.

Gut microbial homeostasis is crucial for the health of cognition in elderly. Previous study revealed that polysorbate 80 (P80) as a widely used emulsifier in food industries and pharmaceutical formulations could directly alter the human gut microbiota compositions. However, whether long-term exposure to P80 could accelerate age-related cognitive decline via gut-brain axis is still unknown. Accordingly, in this study, we used the senescence accelerated mouse prone 8 (SAMP8) mouse model to investigate the effects of the emulsifier P80 intake (1 % P80 in drinking water for 12 weeks) on gut microbiota and cognitive function. Our results indicated that P80 intake significantly exacerbated cognitive decline in SAMP8 mice, along with increased brain pathological proteins deposition, disruption of the blood-brain barrier and activation of microglia and neurotoxic astrocytes. Besides, P80 intake could also induce gut microbiota dysbiosis, especially the increased abundance of secondary bile acids producing bacteria, such as Ruminococcaceae, Lachnospiraceae, and Clostridium scindens. Moreover, fecal microbiota transplantation from P80 mice into 16-week-old SAMP8 mice could also exacerbated cognitive decline, microglia activation and intestinal barrier impairment. Intriguingly, the alterations of gut microbial composition significantly affected bile acid metabolism profiles after P80 exposure, with markedly elevated levels of deoxycholic acid (DCA) in serum and brain tissue. Mechanically, DCA could activate microglial and promote senescence-associated secretory phenotype production through adenosine triphosphate-binding cassette transporter A1 (ABCA1) importing lysosomal cholesterol. Altogether, the emulsifier P80 accelerated cognitive decline of aging mice by inducing gut dysbiosis, bile acid metabolism alteration, intestinal barrier and blood brain barrier disruption as well as neuroinflammation. This study provides strong evidence that dietary-induced gut microbiota dysbiosis may be a risk factor for age-related cognitive decline.

The ecdysis triggering hormone system is essential for reproductive success in Mythimna separata (Walker).

Ecdysis triggering hormone (ETH) was originally discovered as a key hormone that regulates insect moulting via binding to its receptor, ETH receptor (ETHR). However, the precise role of ETH in moth reproduction remains to be explored in detail. ETH function was verified in vivo using Mythimna separata (Walker), an important cereal crop pest. RT-qPCR analysis revealed that transcriptional expression profiles of MsepETH showed evident sexual dimorphism in the adult stage. MsepETH expression increased in the females on day 3 and persisted thereafter till day 7, consistent with female ovarian maturation, and was merely detectable in males. Meanwhile, MsepETH expression levels were significantly higher in the trachea than in other tissues. MsepETHR-A and MsepETHR-B were expressed in both sexes and were significantly higher in the antennae than in other tissues. MsepETH and MsepETHR knockdown in females by RNA interference significantly reduced the expression of MsepETH, MsepETHR-A, MsepETHR-B, MsepJHAMT, and MsepVG, which delayed egg-laying and significantly reduced egg production. RNAi 20-hydroxyecdysone (20E) receptor (EcR) decreased MsepETH expression whereas injecting 20E restored egg production that had been disrupted by MsepETH interference. Meanwhile, RNAi juvenile hormone (JH) methoprene tolerant protein (Met) also decreased MsepETH expression and smearing JH analog methoprene (Meth) restored egg production. In conclusion, the reproduction roles of ETH, JH, and 20E were investigated in M. separata. These findings will lay the foundation for future research to develop an antagonist that reduces female reproduction and control strategies for pest insects.

A hepatocyte differentiation model reveals two subtypes of liver cancer with different oncofetal properties and therapeutic targets.

Clinical observation of the association between cancer aggressiveness and embryonic development stage implies the importance of developmental signals in cancer initiation and therapeutic resistance. However, the dynamic gene expression during organogenesis and the master oncofetal drivers are still unclear, which impeded the efficient elimination of poor prognostic tumors, including human hepatocellular carcinoma (HCC). In this study, human embryonic stem cells were induced to differentiate into adult hepatocytes along hepatic lineages to mimic liver development in vitro. Combining transcriptomic data from liver cancer patients with the hepatocyte differentiation model, the active genes derived from different hepatic developmental stages and the tumor tissues were selected. Bioinformatic analysis followed by experimental assays was used to validate the tumor subtype-specific oncofetal signatures and potential therapeutic values. Hierarchical clustering analysis revealed the existence of two subtypes of liver cancer with different oncofetal properties. The gene signatures and their clinical significance were further validated in an independent clinical cohort and The Cancer Genome Atlas database. Upstream activator analysis and functional screening further identified E2F1 and SMAD3 as master transcriptional regulators. Small-molecule inhibitors specifically targeting the oncofetal drivers extensively down-regulated subtype-specific developmental signaling and inhibited tumorigenicity. Liver cancer cells and primary HCC tumors with different oncofetal properties also showed selective vulnerability to their specific inhibitors. Further precise targeting of the tumor initiating steps and driving events according to subtype-specific biomarkers might eliminate tumor progression and provide novel therapeutic strategy.

The design and clinical application of a new appliance to treat impacted maxillary central incisors.

Maxillary central incisor impaction is one of the most common types of dental anomalies in children. Treatment of impacted central incisors is complicated and challenging given the position of the impacted central incisors, root development, and the complexity of the crown eruption direction. This study aimed to describe the use of a new multifunctional appliance for the treatment of impacted maxillary central incisors. This article reports the use of a novel appliance for the treatment of impacted maxillary central incisors. We describe the cases of two young patients with labial horizontally impacted maxillary central incisors. Both patients were treated using this novel appliance. Therapeutic effects were evaluated by comparing the pretreatment results, posttreatment cone-beam computed tomography images, and posttreatment clinical examination results. At the end of the treatment period using the novel appliance, the impacted central incisors had successfully been properly aligned in the dental arch, and the tooth roots had not resorbed. Both patients exhibited good dental alignment, with restored function and acceptable aesthetics. This article demonstrates that the new appliance was comfortable, convenient, safe, and effective in the treatment of impacted maxillary central incisors and that its clinical use should be promoted in the future.

Mesenchymal stromal cell treatment attenuates repetitive mild traumatic brain injury-induced persistent cognitive deficits via suppressing ferroptosis.

The incidence of repetitive mild traumatic brain injury (rmTBI), one of the main risk factors for predicting neurodegenerative disorders, is increasing; however, its underlying mechanism remains unclear. As suggested by several studies, ferroptosis is possibly related to TBI pathophysiology, but its effect on rmTBI is rarely studied. Mesenchymal stromal cells (MSCs), the most studied experimental cells in stem cell therapy, exert many beneficial effects on diseases of the central nervous system, yet evidence regarding the role of MSCs in ferroptosis and post-rmTBI neurodegeneration is unavailable. Our study showed that rmTBI resulted in time-dependent alterations in ferroptosis-related biomarker levels, such as abnormal iron metabolism, glutathione peroxidase (GPx) inactivation, decrease in GPx4 levels, and increase in lipid peroxidation. Furthermore, MSC treatment markedly decreased the aforementioned rmTBI-mediated alterations, neuronal damage, pathological protein deposition, and improved cognitive function compared with vehicle control. Similarly, liproxstatin-1, a ferroptosis inhibitor, showed similar effects. Collectively, based on the above observations, MSCs ameliorate cognitive impairment following rmTBI, partially via suppressing ferroptosis, which could be a therapeutic target for rmTBI.

In vitro antibacterial effects of photodynamic therapy against Enterococcus faecalis in root canals of deciduous teeth.

OBJECTIVE: This study aimed at evaluating the in vitro antibacterial efficacy of photodynamic therapy (PDT) on planktonic E. faecalis and its biofilm in the root canal of infected deciduous teeth. METHODS: Forty root canals of maxillary deciduous anterior teeth were enlarged up to #35 K-file and inoculated with E. faecalis for 21 days. The root canals were randomly assigned into four groups (n = 10): The normal saline group (control), 1% NaClO group, PDT group, and the 1% NaClO + PDT group. Paper point samples were obtained at baseline (S1) and after treatment (S2). The colony-forming units (CFU) were counted, and the bacterial growth rate calculated. From each subgroup, 5 samples were randomly selected after treatment and a scanning laser confocal microscope (CLSM) used to determine the distribution of dead / living bacteria on the biofilm surface of each subgroup. A scanning electron microscope (SEM) was used to observe bacterial morphologies in the root canal walls of the remaining 5 samples in each subgroup. The Kruskal-Wallis test and Dunn test with boferroni adjustment were used to analyze the effect of the different treatment techniques on the E. faecalis in root canals. RESULTS: Compared to the saline group, PDT significantly reduced bacterial counts in the root canal (p < 0.05). The CFU counts were lowest (p < 0.05) in the 1% NaClO and in 1% NaClO + PDT groups. The rate of bacterial death on the surface of the biofilm in the PDT group was significantly increased after treatment (p < 0.05), and the rate of bacterial death was highest in 1%NaClO group and 1%NaClO + PDT group (p < 0.05). CONCLUSION: PDT has an antibacterial activity against E. faecalis in the root canal of deciduous teeth. Its activity against planktonic E. faecalis is better than the activity on the intact biofilm. The antibacterial activity of PDT on E. faecalis in root canals of deciduous teeth is lower compared to that of 1% NaClO.

Triptolide delivery: Nanotechnology-based carrier systems to enhance efficacy and limit toxicity.

Triptolide (TP) possesses a wide range of biological and pharmacological activities involved in the treatment of various diseases. However, widespread usages of TP raise the urgent issues of the severe toxicity, which hugely limits its further clinical application. The novel functional nanostructured delivery system, which is of great significance in enhancing the efficacy, reducing side effects and improving bioavailability, could improve the enrichment, penetration and controlled release of drugs in the lesion location. Over the past decades, considerable efforts have been dedicated to designing and developing a variety of TP delivery systems with the intention of alleviating the adverse toxicity effects and enhancing the bioavailability. In this review, we briefly summarized and discussed the recent functionalized nano-TP delivery systems for the momentous purpose of guiding further development of novel TP delivery systems and providing perspectives for future clinical applications.

Demography of Mythimna separata (Lepidoptera: Noctuidae) at outdoor fluctuating temperatures.

The oriental armyworm Mythimna separata (Walker) (Lepidoptera: Noctuidae) is a major migratory pest of cereal crops in East Asia, South Asia and Australia. To comprehensively understand the ecological tolerance of M. separata, we collected life table data of individuals from four consecutive generations reared under outdoor natural fluctuating temperatures from 15 April to 17 October 2018 in Yangling, Shaanxi, China. The results showed that the immature stage in early summer and summer were shorter than in spring and autumn. High mortality in late larval instar and pupal stages was observed in the summer generation. The adult pre-oviposition period in autumn was longer than the other seasons. The population in the earlier two seasons had heavier pupae and higher fecundity than the population in the latter two seasons. The intrinsic rate of increase and the finite rate of increase was the highest in early summer (r = 0.1292 day-1, λ = 1.1391 day-1), followed by spring (r = 0.1102 day-1, λ = 1.1165 day-1), and was the lowest in summer (r = 0.0281 day-1, λ = 1.0293 day-1). The results of this study would be useful to predict the population dynamics of M. separata and deepen our standing of the adaptiveness of this migratory pest in natural fluctuating ambient environments.

Development of an oncogenic dedifferentiation SOX signature with prognostic significance in hepatocellular carcinoma.

BACKGROUND: Gradual loss of terminal differentiation markers and gain of stem cell-like properties is a major hall mark of cancer malignant progression. The stem cell pluripotent transcriptional factor SOX family play critical roles in governing tumor plasticity and lineage specification. This study aims to establish a novel SOX signature to monitor the extent of tumor dedifferentiation and predict prognostic significance in hepatocellular carcinoma (HCC). METHODS: The RNA-seq data from The Cancer Genome Atlas (TCGA) LIHC project were chronologically divided into the training (n = 188) and testing cohort (n = 189). LIRI-JP project from International Cancer Genome Consortium (ICGC) data portal was used as an independent validation cohort (n = 232). Kaplan-Meier and multivariable Cox analyses were used to examine the clinical significance and prognostic value of the signature genes. RESULTS: The SOX gene family members were found to be aberrantly expressed in clinical HCC patients. A five-gene SOX signature with prognostic value was established in the training cohort. The SOX signature genes were found to be closely associated with tumor grade and tumor stage. Liver cancer dedifferentiation markers (AFP, CD133, EPCAM, and KRT19) were found to be progressively increased while hepatocyte terminal differentiation markers (ALB, G6PC, CYP3A4, and HNF4A) were progressively decreased from HCC patients with low SOX signature scores to patients with high SOX signature scores. Kaplan-Meier survival analysis further indicated that the newly established SOX signature could robustly predict patient overall survival in both training, testing, and independent validation cohort. CONCLUSIONS: An oncogenic dedifferentiation SOX signature presents a great potential in predicting prognostic significance in HCC, and might provide novel biomarkers for precision oncology further in the clinic.

Detwinning through migration of twin boundaries in nanotwinned Cu films under in situ ion irradiation.

The mechanism of radiation-induced detwinning is different from that of deformation detwinning as the former is dominated by supersaturated radiation-induced defects while the latter is usually triggered by global stress. In situ Kr ion irradiation was performed to study the detwinning mechanism of nanotwinned Cu films with various twin thicknesses. Two types of incoherent twin boundaries (ITBs), so-called fixed ITBs and free ITBs, are characterized based on their structural features, and the difference in their migration behavior is investigated. It is observed that detwinning during radiation is attributed to the frequent migration of free ITBs, while the migration of fixed ITBs is absent. Statistics shows that the migration distance of free ITBs is thickness and dose dependent. Potential migration mechanisms are discussed.

Characterization of Integrons and qnr Genes in Proteeae from a Teaching Hospital in China.

BACKGROUND: Proteeae isolates displaying multidrug-resistance (MDR) are the second most common causes of hospital-associated infections. The aim of this study was to screen class 1-3 integrons and plasmid-mediated quinolone resistance (PMQR) genes in Proteeae isolates from the First Affiliated Hospital of the Wenzhou Medical University. MATERIALS AND METHODS: 176 Proteeae isolates were collected from clinical specimens of inpatients between January 2011 and December 2013. Susceptibility testing was determined by the agar dilution method. Class 1-3 integrons and PMQR genes were amplified by polymerase chain reaction, and the variable regions of integrons were determined by restriction fragment length polymorphisms. RESULTS: 68.2% Proteeae isolates exhibited MDR phenotypes: 46.6 and 10.8% Proteeae isolates were positive for intI1 and intI2, respectively. The resistance rate of integron-positive isolates to aminoglycosides, fluoroquinolones, and trimethoprim/sulfamethoxazole was significantly higher than integron-negative isolates. Sequence analysis revealed that dfrA1-sat2-aadA1, dfrA1-catB2-sat2-aadA1, and sat2-aadA1 were first detected in Morganella morganii strains isolated from China. PMQR was determined by qnrD in 40 strains (22.7%). CONCLUSION: Our results indicate that class 1 and 2 integrons are common among Proteeae isolates. Meanwhile, qnrD are highly prevalent in Proteeae isolated from our hospital.

Bioengineering Novel Chimeric microRNA-34a for Prodrug Cancer Therapy: High-Yield Expression and Purification, and Structural and Functional Characterization.

Development of anticancer treatments based on microRNA (miRNA/miR) such as miR-34a replacement therapy is limited to the use of synthetic RNAs with artificial modifications. Herein, we present a new approach to a high-yield and large-scale biosynthesis, in Escherichia coli using transfer RNA (tRNA) scaffold, of chimeric miR-34a agent, which may act as a prodrug for anticancer therapy. The recombinant tRNA fusion pre-miR-34a (tRNA/mir-34a) was quickly purified to a high degree of homogeneity (>98%) using anion-exchange fast protein liquid chromatography, whose primary sequence and post-transcriptional modifications were directly characterized by mass spectrometric analyses. Chimeric tRNA/mir-34a showed a favorable cellular stability while it was degradable by several ribonucleases. Deep sequencing and quantitative real-time polymerase chain reaction studies revealed that tRNA-carried pre-miR-34a was precisely processed to mature miR-34a within human carcinoma cells, and the same tRNA fragments were produced from tRNA/mir-34a and the control tRNA scaffold (tRNA/MSA). Consequently, tRNA/mir-34a inhibited the proliferation of various types of human carcinoma cells in a dose-dependent manner and to a much greater degree than the control tRNA/MSA, which was mechanistically attributable to the reduction of miR-34a target genes. Furthermore, tRNA/mir-34a significantly suppressed the growth of human non-small-cell lung cancer A549 and hepatocarcinoma HepG2 xenograft tumors in mice, compared with the same dose of tRNA/MSA. In addition, recombinant tRNA/mir-34a had no or minimal effect on blood chemistry and interleukin-6 level in mouse models, suggesting that recombinant RNAs were well tolerated. These findings provoke a conversation on producing biologic miRNAs to perform miRNA actions, and point toward a new direction in developing miRNA-based therapies.

Chimeric MicroRNA-1291 Biosynthesized Efficiently in Escherichia coli Is Effective to Reduce Target Gene Expression in Human Carcinoma Cells and Improve Chemosensitivity.

In contrast to the growing interests in studying noncoding RNAs (ncRNAs) such as microRNA (miRNA or miR) pharmacoepigenetics, there is a lack of efficient means to cost effectively produce large quantities of natural miRNA agents. Our recent efforts led to a successful production of chimeric pre-miR-27b in bacteria using a transfer RNA (tRNA)-based recombinant RNA technology, but at very low expression levels. Herein, we present a high-yield expression of chimeric pre-miR-1291 in common Escherichia coli strains using the same tRNA scaffold. The tRNA fusion pre-miR-1291 (tRNA/mir-1291) was then purified to high homogeneity using affinity chromatography, whose primary sequence and post-transcriptional modifications were directly characterized by mass spectrometric analyses. Chimeric tRNA/mir-1291 was readily processed to mature miR-1291 in human carcinoma MCF-7 and PANC-1 cells. Consequently, recombinant tRNA/mir-1291 reduced the protein levels of miR-1291 target genes, including ABCC1, FOXA2, and MeCP2, as compared with cells transfected with the same doses of control methionyl-tRNA scaffold with a sephadex aptamer (tRNA/MSA). In addition, tRNA-carried pre-miR-1291 suppressed the growth of MCF-7 and PANC-1 cells in a dose-dependent manner, and significantly enhanced the sensitivity of ABCC1-overexpressing PANC-1 cells to doxorubicin. These results indicate that recombinant miR-1291 agent is effective in the modulation of target gene expression and chemosensitivity, which may provide insights into high-yield bioengineering of new ncRNA agents for pharmacoepigenetics research.

Rapid production of novel pre-microRNA agent hsa-mir-27b in Escherichia coli using recombinant RNA technology for functional studies in mammalian cells.

Noncoding microRNAs (miRNAs or miRs) have been revealed as critical epigenetic factors in the regulation of various cellular processes, including drug metabolism and disposition. However, research on miRNA functions is limited to the use of synthetic RNA and recombinant DNA agents. Herein, we show that novel pre-miRNA-27b (miR-27b) agents can be biosynthesized in Escherichia coli using recombinant RNA technology, and recombinant transfer RNA (tRNA)/mir-27b chimera was readily purified to a high degree of homogeneity (>95%) using anion-exchange fast protein liquid chromatography. The tRNA-fusion miR-27b was revealed to be processed to mature miRNA miR-27b in human carcinoma LS-180 cells in a dose- and time-dependent manner. Moreover, recombinant tRNA/miR-27b agents were biologically active in reducing the mRNA and protein expression levels of cytochrome P450 3A4 (CYP3A4), which consequently led to lower midazolam 1'-hydroxylase activity. These findings demonstrate that pre-miRNA agents can be produced by recombinant RNA technology for functional studies.

[Antimicrobial resistance characteristics of and disinfectant-resistant gene distribution in Staphylococcus aureus isolates from male urogenital tract infection].

OBJECTIVE: To study the antibiotic- and disinfectant-resistance features of and disinfectant-resistant gene distribution in Staphylococcus aureus (Sa) isolated from the urogenital tract of male patients with urogenital tract infection (UTI). total of 152 Sa isolates were collected from the urethral discharge specimens from male UTI patients. The minimum inhibition concentration (MIC) of antimicrobial agents and disinfectants commonly used against Sa were tested by standard ager dilution; the methicillin-resistant Sa (MRSA) isolates detected by cefoxitin disk diffusion and mecA gene amplification; Staphylococcal cassette chromosome mec (SCCmec) genotyping performed by multiplex PCR; the disinfectants gene qac (quaternary ammonium compound) amplified by PCR; and the clonal relatedness of qacA/B-positive MRSA isolates investigated by pulsed-field gel electrophoresis (PFGE). RESULTS: Out of the 152 Sa isolates, 91 (59.9%) were found to be MRSA. SCCmec genotyping showed SCCmec V to be the main type, accounting for 63.7% (58/91), with 8 (8.8%) isolates of SCCmec I, 2 (2.2%) isolates of SCCmec II, 19 (20.9%) isolates of SCCmec III, and 4 (4. 4%) isolates of SCCmec IV. The Sa isolates exhibited high rates of non-susceptibility to penicillin (95.4%) , erythromycin (72.4% ) , ciprofloxacin (42. 8%), and levofloxacin (44.7%), and a fairly high sensitivity to nitrofurantoin, teicoplanin, linezolid, and vancomycin. The MIC in the Sa isolates was 0. 25 -16 microg/ml for chlorhexidine; MIC50 and MIC90 were 2.0 and 4.0 microg/ml respectively for MRSA strains and both 1.0 microg/ml for MSSA strains. Out of the 152 Sa isolates, 72 (47.4%) harbored the qacA/B gene, 6 (3.9%) the smar (qacC + qacD) gene, 9 (5.9%) the qacE delta 1 gene, and 2 (1.3%) the qacH gene, but no qacG and qacJ genes were detected. PFGE analysis showed that the qacA/B-positive MRSA isolates were distributed CONCLUSION: Clinical Sa isolates exhibited varied degrees of resistance to commonly used antibiotics, and in a polyclonal manner. some showed a robust tolerance to chlorhexidine. The main disinfectant-resistant gene is qacA/B. Antimicrobial agents and disinfectants should be used rationally according to clinicians.

Change in oral health-related behaviours of children before and after dental treatments under general anaesthesia.

This study evaluated the behavioural changes pertaining to children's oral health before and after the dental general anaesthesia (DGA), with particular focus on the factors associated with these changes. The records were collected for the children who received DGA from July 2015 to November 2016, and relevant questionnaires were obtained from their parents/guardians for the information prior to and after the DGA. The questionnaire included Early Childhood Oral Health Impact Scale (ECOHIS) and Dental Subscale of Children's Fear Survey Schedule (CFSS-DS) to investigate the changes in Oral Health-related Quality of Life (OHRQoL) and dental fear. The DGA impact on children's oral hygiene habits and oral health-related behaviours was assessed by analysing the data. The chi-square test and Mann-Whitney test were employed to evaluate the differences. Total of 141 patients (89 before DGA and 77 after DGA, 25 being common) participated in this study. There were 60 children below 5 years and 29 over 5 years before DGA, while 41 children below 5 years and 36 over 5 years after DGA. Most parents/guardians were educated above undergraduate level (59.6% before DGA, 55.8% after DGA). More children lived with grandparents (61.8% before DGA, 54.5% after DGA) than only with parents (20.2% before DGA, 26.0% after DGA). In total, 73.0% (65/89) children before DGA brushed teeth more than twice a day. This proportion increased to 90.9% after DGA (70/77, p = 0.03). The eating difficulty decreased after DGA according to ECOHIS (p = 0.01). CFSS-DS score also decreased after DGA (p < 0.05). After DGA, children's oral hygiene habits and oral health-related quality of life (OHRQoL) improved, children fear for dental treatment decreased, and parents became more attentive towards children oral health.

Effect of short neuropeptide F signaling on larval feeding in Mythimna separata.

Mythimna separata is a notorious phytophagous pest which poses serious threats to cereal crops owing to the gluttony of the larvae. Because short neuropeptide F (sNPF) and its receptor sNPFR are involved in a diversity of physiological functions, especially in functions related to feeding in insects, it is a molecular target for pest control. Herein, an sNPF and 2 sNPFRs were identified and cloned from M. separata. Bioinformatics analysis revealed that the sNPF and its receptors had a highly conserved RLRFamide C-terminus and 7 transmembrane domains, respectively. The sNPF and its receptor genes were distributed across larval periods and tissues, but 2 receptors had distinct expression patterns. The starvation-induced assay elucidated that sNPF and sNPFR expression levels were downregulated under food deprivation and recovered with subsequent re-feeding. RNA interference knockdown of sNPF, sNPFR1, and sNPFR2 by injection of double-stranded RNA into larvae not only suppressed food consumption and increased body size and weight, but also led to decrease of glycogen and total lipid contents, and increase of trehalose compared with double-stranded green fluorescent protein injection. Furthermore, molecular docking was performed on the interaction mode between sNPFR protein and its ligand sNPF based on the 3-dimensional models constructed by AlphaFold; the results indicated that both receptors were presumably activated by the mature peptide sNPF-2. These results revealed that sNPF signaling played a considerably vital role in the feeding regulation of M. separata and represents a potential control target for this pest.

Stimuli-responsive incremental DNA machine auto-catalyzed CRISPR-Cas12a feedback amplification permits ultrasensitive molecular diagnosis of esophageal cancer-related microRNA.

Development of new diagnostic methods is essential for disease diagnosis and treatment. In this work, we present a stimuli-responsive incremental DNA machine auto-catalyzed CRISPR-Cas12a (SRI-DNA machine/CRISPR-Cas12a) feedback amplification for ultrasensitive molecular detection of miRNA-21, which is an important biomarker related closely to the initiation and development of cancers, such as esophageal cancer. Strategically, the powerful SRI-DNA machine and efficient trans-cleavage activity of the CRISPR-Cas12a system are ingeniously integrated via a rationally designed probe termed as stem-elongated functional hairpin probe (SEF-HP). The SRI-DNA machine begins with the target miRNA, the trigger of the reaction, binding complementarily to the SEF-HP, followed by autonomously performed mechanical strand replication, cleavage, and displacement circuit at multiple sites. This conversion process led to the amplified generation of numerous DNA activators that are complementary with CRISPR RNA (CrRNA). Once formed the DNA activator/CrRNA heteroduplex, the trans-cleavage activity of the CRISPR-Cas12a was activated to nonspecific cleavage of single-stranded areas of a reporter probe for fluorescence emission. Under optimal conditions, the target miRNA can be detected with a wide linear range and an excellent specificity. As a proof-of-concept, this SRI-DNA machine/CRISPR-Cas12a feedback amplification system is adaptable and scalable to higher-order artificial amplification circuits for biomarkers detection, highlighting its promising potential in early diagnosis and disease treatment.