Anti-osteoporosis activity of casticin in ovariectomized rats.

Background: Postmenopausal osteoporosis (PMPO) is the most familiar type of osteoporosis, a silent bone disease. Casticin, a natural flavonoid constituent, improves osteoporosis in animal model. Nevertheless, the potential mechanism remains to be further explored. Methods: A model of PMPO was established in rats treated with ovariectomy (OVX) and RAW 264.7 cells induced with receptor activator of nuclear factor kappa-B ligand (RANKL). The effect and potential mechanism of casticin on PMPO were addressed by pathological staining, measurement of bone mineral density (BMD), three-point bending test, serum biochemical detection, filamentous-actin (F-actin) ring staining, TRAcP staining, reverse transcription quantitative polymerase chain reaction, western blot and examination of oxidative stress indicators. Results: The casticin treatment increased the femoral trabecular area, bone maturity, BMD, elastic modulus, maximum load, the level of calcium and estrogen with the reduced concentrations of alkaline phosphatase (ALP) and tumor necrosis factor (TNF)-α in OVX rats. An enhancement in the F-actin ring formation, TRAcP staining and the relative mRNA expression of NFATc1 and TRAP was observed in RANKL-induced RAW 264.7 cells, which was declined by the treatment of casticin. Moreover, the casticin treatment reversed the reduced the relative protein expression of Nrf2 and HO-1 and the concentrations of superoxide dismutase and glutathione peroxidase, and the increased content of malondialdehyde both in vivo and in vitro. Conclusion: Casticin improved bone density, bone biomechanics, the level of calcium and estrogen, the release of pro-inflammatory factor and oxidative stress to alleviate osteoporosis, which was associated with the upregulation of Nrf2/HO-1 pathway.

Long-read sequencing reveals the structural complexity of genomic integration of HPV DNA in cervical cancer cell lines.

BACKGROUND: Cervical cancer (CC) causes more than 311,000 deaths annually worldwide. The integration of human papillomavirus (HPV) is a crucial genetic event that contributes to cervical carcinogenesis. Despite HPV DNA integration is known to disrupt the genomic architecture of both the host and viral genomes in CC, the complexity of this process remains largely unexplored. RESULTS: In this study, we conducted whole-genome sequencing (WGS) at 55-65X coverage utilizing the PacBio long-read sequencing platform in SiHa and HeLa cells, followed by comprehensive analyses of the sequence data to elucidate the complexity of HPV integration. Firstly, our results demonstrated that PacBio long-read sequencing effectively identifies HPV integration breakpoints with comparable accuracy to targeted-capture Next-generation sequencing (NGS) methods. Secondly, we constructed detailed models of complex integrated genome structures that included both the HPV genome and nearby regions of the human genome by utilizing PacBio long-read WGS. Thirdly, our sequencing results revealed the occurrence of a wide variety of genome-wide structural variations (SVs) in SiHa and HeLa cells. Additionally, our analysis further revealed a potential correlation between changes in gene expression levels and SVs on chromosome 13 in the genome of SiHa cells. CONCLUSIONS: Using PacBio long-read sequencing, we have successfully constructed complex models illustrating HPV integrated genome structures in SiHa and HeLa cells. This accomplishment serves as a compelling demonstration of the valuable capabilities of long-read sequencing in detecting and characterizing HPV genomic integration structures within human cells. Furthermore, these findings offer critical insights into the complex process of HPV16 and HPV18 integration and their potential contribution to the development of cervical cancer.

Individual, family, and environmental correlates of fundamental motor skills among school-aged children: a cross-sectional study in China.

OBJECTIVE: This cross-sectional study examined the socio-ecological factors influencing fundamental motor skills (FMS) in Chinese school-aged children. METHODS: A total of 1012 parent-child pairs were randomly sampled between March-1st and April-15th, 2022. Based on the socio-ecological model of Children's FMS, three levels of factors: individual-level (e.g., demographic, physical, psychological, and behavioral characteristics of children), family-level (e.g., caregiver demographics, parental support, and socioeconomic status), and environmental factors (e.g., availability of physical activity equipment) were assessed using self-reported scales (e.g., the Self-perception Profile for Children, the Physical Activity Enjoyment Scale, and the 12-item Psychological Well-Being Scale for Children) and objective measures (e.g., ActiGraph GT3X, the Chinese National Student Physical Fitness Standard, and the Test of Gross Motor Development-Third Edition). Multi-level regression models were employed using SPSS. RESULTS: The results demonstrated that children's age, sex, physical fitness, parental support, and the quality of home and community physical activity environments consistently influenced all three types of FMS, including locomotor, ball, and composite skills. Additionally, seven individual-level factors (children's age, sex, body mass index, light physical activity, sleep duration, perceived motor competence, and physical fitness) were associated with different types of FMS. CONCLUSIONS: The findings underscore the multidimensional and complex nature of FMS development, with individual-level factors playing a particularly significant role. Future research should adopt rigorous longitudinal designs, comprehensive assessment tools covering various FMS skills, and objective measurement of parents' movement behaviors to better understand the strength and direction of the relationship between socio-ecological factors and children's FMS.

Global integrative meta-analysis of the responses in soil organic carbon stock to biochar amendment.

Applying biochar to soil has been recognized as a promising practice of climate-smart agriculture, with considerable potential in enhancing soil organic carbon (SOC) sequestration. Previous studies showed that biochar-induced increases in SOC stock varied substantially among experiments, while the explanatory factors responsible for such variability are still not well assessed. Here, we conducted an integrative meta-analysis of the magnitude and efficiency of biochar-induced change in SOC stock, using a database including 476 field measurements at 101 sites across the globe. Biochar amendment increased SOC stock by 6.13 ± 1.62 (95% confidence interval, CI) and 7.01 ± 1.11 (95% CI) Mg C ha-1, respectively, compared to their unfertilized (R0) and mineral nitrogen (N) fertilized (Rn) references. Of which approx. 52% (R0) and 50% (Rn) were contributed directly by biochar-C input. Corresponding biochar carbon efficiencies in R0 and Rn datasets were estimated as 58.20 ± 10.37% and 65.58 ± 9.26% (95% CI), respectively. The change magnitude of SOC stock increased significantly (p < 0.01) with the increasing amount of biochar-C input, while carbon efficiency of biochar showed an opposite trend. Biochar amendment sequestered larger amounts of SOC with higher efficiency in acidic and loamy soils than in alkaline and sandy soils. Biochar amendments with higher C/N ratio caused higher SOC increase than those with lower C/N ratio. Random forest (RF) algorithm showed that accumulative biochar-C input, soil pH, and biochar C/N ratio were the three most-important factors regulating the SOC stock responses. Overall, these results suggest that applying high C/N ratio biochar in acidic soils is a recommendable agricultural practice from the perspective of enhancing organic carbon.

Evaporation Driven Hydrovoltaic Generator Based on Nano-Alumina-Coated Polyethylene Terephthalate Film.

Collecting energy from the ambient environment through green and sustainable methods is highly expected to alleviate pollution and energy problems worldwide. Here, we report a facile and flexible hydrovoltaic generator capable of utilizing natural water evaporation for sustainable electricity production. The generator was fabricated by coating nano-Al2O3 on a twistable polyethylene terephthalate film. An open circuit voltage of 1.7 V was obtained on a piece of centimeter-sized hydrovoltaic generator under ambient conditions. The supercapacitor charged by the hydrovoltaic device can power a mini-motor efficiently. Moreover, by expanding the size or connecting it in series/parallel, the energy output of the generator can be further improved. Finally, the influence factors and the mechanism for power generation were primarily investigated. Electrical energy is produced by the migration of water through charged capillary channels. The environmental conditions, the properties of the solution and the morphology of the film have important effects on the electrical performance. This study is anticipated to offer enlightenment into designing novel hydrovoltaic devices, providing diverse energy sources for various self-powered devices and systems.

Casticin promotes osteogenic differentiation of bone marrow stromal cells and improves osteoporosis in rats by regulating nuclear factor-κB/mitogen-activated protein kinase.

BACKGROUND: Osteoporosis has influenced millions of people, especially postmenopausal women, which has become a big burden to the whole world. Although the diverse roles of casticin (CAS) on different diseases were identified, whether it was implicated with osteoporosis was unknown. METHODS: A rat model of osteoporosis was established through dexamethasone (DEX) treatment and a cell model reflecting the osteogenic and osteoclast induction was constructed in bone marrow stromal cells (BMSCs). The calcification at the late stage of induction was measured via Alizarin Red S staining. Western blot was applied to evaluate the levels of proteins. RESULTS: Hematoxylin and eosin staining revealed that the number of bone trabecular in DEX-induced osteoporosis rats was decreased, while increased doses of CAS treatment elevated the number of bone trabecular. CAS treatment alleviated DEX-induced osteoporosis in rats. Moreover, we found that CAS inhibited the nuclear factor-κB/mitogen-activated protein kinase (NF-κB/MAPK) pathway. In addition, CAS promoted osteogenic differentiation of BMSCs and reduced osteoclastogenesis of bone marrow monocytes. Finally, CAS was observed to retard the receptor activator of NFκ-B ligand-induced NF-κB/MAPK pathway. CONCLUSION: CAS promoted osteogenic differentiation of BMSCs and improved osteoporosis in rats by regulating the NF-κB/MAPK pathway. This might shed a light into using CAS as a drug treating osteoporosis in the future.

Study on the Relationship Between Diet, Physical Health and Gut Microflora of Chinese College Students.

Many elements of a modern lifestyle influence the gut microbiota but few studies have explored the effect of physical health level. This study was aimed to explore the relationship between diet, physical health and gut microbiota in Chinese college students. A total of 69 college students were recruited, including 27 college athletes (AS group) and 42 healthy controls (HC group). Fecal samples were collected for 16S rRNA sequencing. According to National Standards for Students' Physical Health (2014 revision), physical fitness measurements, dietary intake and health-related data were collected via questionnaires. ①According to the physical fitness scores, the physical fitness level of AS group was significantly higher than that of HC group (P < 0.05), there were no significant differences between the two groups in the frequency of intake of food. The frequency and duration of physical activity in the AS group were higher than those in the HC group (P < 0.05); ②The proportion and relative abundances of microorganism composition is varying at two groups: on the phylum level, AS group had mainly increased Firmicutes, Actinobacteria and reduced Bacteroidetes, Proteobacteria; on the genus level, AS group had mainly increased Faecalibacterium, Bifidobacterium and reduced Bacteroides; ③The associations with the 10 most abundant bacterial genera and physical fitness, dietary factors were investigated. Changes in the gut microbiota abundance can be sometimes reflective of a physical health status. Loss of the balance of gut microbial populations will lead to flora disorders and diseases. Therefore, further studies are needed to reveal the mechanisms behind the gut microbiota in its potential role.

Construction of Bioluminescent Sensors for Label-Free, Template-Free, Separation-Free, and Sequence-Independent Detection of both Clustered and Isolated Damage in Genomic DNA.

DNA damage induced by endogenous/exogenous factors may cause various diseases, and the genomic DNA damage has become an important biomarker for clinical diagnosis and risk assessment, but it remains a great challenge to accurately quantify both clustered and isolated damage because of their random locations, large diversity, and low abundance. Herein, we demonstrate the development of bioluminescent sensors for label-free, template-free, separation-free, and sequence-independent detection of both clustered and isolated damage in genomic DNA based on the base-excision repair (BER) pathway and terminal transferase (TdT)-initiated template-free isothermal cyclic amplification. The damaged bases are cleaved by DNA glycosylase to generate a new 3'-OH terminus, and subsequently, TdT catalyzes the repeated incorporation of dTTPs into the 3'-OH terminus to produce poly-T structures which can hybridize with the signal probe to form a poly-T sequence/signal probe duplex. Under the lambda exonuclease hydrolysis, a large number of adenosine monophosphate (AMP) molecules are produced to generate a high bioluminescence signal through the cyclic interconversion of AMP-adenosine triphosphate (ATP)-AMP in the presence of luciferin and firefly luciferase. Moreover, the introduction of APE1-induced cyclic cleavage signal amplification can greatly improve the detection sensitivity. The proposed strategy can detect both clustered and isolated damage in genomic DNA with extremely high sensitivity and excellent specificity, and it can even distinguish 0.001% DNA damage in the mixture. Importantly, it can detect the cellular DNA damage with a detection limit of 0.011 ng and further extend to measure various DNA damage with the integration of appropriate DNA repair enzymes.

Current Status of Probiotics as Supplements in the Prevention and Treatment of Infectious Diseases.

Probiotics play an important role against infectious pathogens via their effects on the epithelium, the production of antimicrobial compounds, and competitive exclusion. Administration of probiotic supplements may reduce the risk of infectious diseases and the use of antibiotics, hence contributing to a reduction or a delay of the development of multi-resistant bacteria. Infection is a constant concern for people who experience recurrent infections, and antibiotic treatment usually fails due to antibiotic resistance. Therefore, an infection can lead to severe illness and hospitalization if left untreated. A growing number of studies have demonstrated promising results for a variety of probiotic strains used to prevent or treat acute and recurrent infectious diseases, but additional standardized clinical research is needed.

Oropharyngeal Probiotic ENT-K12 Prevents Respiratory Tract Infections Among Frontline Medical Staff Fighting Against COVID-19: A Pilot Study.

Healthcare workers at the frontline are facing a substantial risk of respiratory tract infection during the COVID-19 outbreak due to an extremely stressful work schedule and public health event. A well-established first-line defense on oropharyngeal microbiome could be a promising strategy to protect individuals from respiratory tract infections including COVID-19. The most thoroughly studied oropharyngeal probiotic product which creates a stable upper respiratory tract microbiota capable of preventing upper respiratory tract infections was chosen to evaluate the safety and efficacy on reducing episodes of upper respiratory tract infections for COVID-19 healthcare workers. To our knowledge to date, this is the very first study describing the beneficial effects of oropharyngeal probiotic been administered by healthcare workers during the COVID-19 pandemic. In this randomized controlled trial, we provided the probiotics to frontline medical staff who work in the hospitals in Wuhan and had been in close contact with hospitalized COVID-19 patients for prophylactic use on a daily basis. Our finding suggests that oropharyngeal probiotic administration significantly reduced the incidence of respiratory tract infections by 64.8%, reduced the time experiencing respiratory tract infections and oral ulcer symptoms by 78%, shortened the days absent from work by 95.5%, and reduced the time under medication where there is no record of antibiotic and anti-viral drug intake in the probiotic group. Furthermore, medical staff treated with Bactoblis experienced sustained protection from respiratory tract infections since the 10th day of oropharyngeal probiotic administration resulting in an extremely low incidence rate of respiratory tract infections.

D-Dimer and Prothrombin Time Are the Significant Indicators of Severe COVID-19 and Poor Prognosis.

OBJECTIVE: To investigate the value of coagulation indicators D-dimer (DD), prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), and fibrinogen (Fg) in predicting the severity and prognosis of COVID-19. METHODS: A total of 115 patients with confirmed COVID-19, who were admitted to Tianyou Hospital of Wuhan University of Science and Technology between January 18, 2020, and March 5, 2020, were included. The dynamic changes of DD, PT, APTT, and Fg were tested, and the correlation with CT imaging, clinical classifications, and prognosis was studied. RESULTS: Coagulation disorder occurred at the early stage of COVID-19 infection, with 50 (43.5%) patients having DD increased and 74 (64.3%) patients having Fg increased. The levels of DD and Fg were correlated with clinical classification. Among 23 patients who deceased, 18 had DD increased at the first lab test, 22 had DD increased at the second and third lab tests, and 18 had prolonged PT at the third test. The results from ROC analyses for mortality risk showed that the AUCs of DD were 0.742, 0.818, and 0.851 in three times of test, respectively; PT was 0.643, 0.824, and 0.937. In addition, with the progression of the disease, the change of CT imaging was closely related to the increase of the DD value (P < 0.01). CONCLUSIONS: Coagulation dysfunction is more likely to occur in severe and critically ill patients. DD and PT could be used as the significant indicators in predicting the mortality of COVID-19.

Magnetic and magneto-transport studies of two-dimensional ferromagnetic compound Fe3GeTe2.

We have systematically reported the magnetic and magneto-transport properties of two-dimensional itinerant ferromagnetic compound Fe3GeTe2 at high magnetic fields of 58 T and demonstrated the correlation between its transport and magnetism. Anomalous two-steps magnetic ordering and antiferromagnetic-like transitions in zero field-cooling (ZFC) curves for H ∥ ab-plane are observed. Additionally, we find that intrinsic negative magnetoresistances in bulk Fe3GeTe2 single crystal are mainly attributed to the suppression of spin-fluctuations in low magnetic fields. Complex evolutions of temperature dependent high field magnetoresistances are detected under different magnetic field and current configurations, which can be explained as a result of the competition between spin-fluctuations, the magnon-scatterings and classical cyclotronic effects.

The effects of external knowledge source heterogeneity on enterprise process and product innovation performance.

As a global manufacturing centre, China is transitioning from a 'Made in China' to 'Create in China' perspective. An ever-increasing number of companies are developing new competitive advantages and improving their innovation levels by acquiring external knowledge. Yet, studies rarely discuss the influence of various sources of knowledge on process and product innovation performance in China's manufacturing enterprises. Based on the Pavitt industry classification, we use a bivariate Probit model to investigate the influence of external knowledge sources on innovation performance, and test it by using Enterprise Survey data for China, published by the World Bank in 2013. Our empirical analysis indicates that external sources of knowledge, with the exception of suppliers, have a significantly positive influence on process and product innovation performance. Specifically, in the process of technological innovation, peers have a positive effect on enterprise process improvement, especially in the science-based sector. In product innovation, close technical cooperation with users accelerates the commercial manifestation of products, especially in the specialized supplier sector.

Dehydrocostus lactone inhibits cell proliferation and induces apoptosis by PI3K/Akt/Bad and ERS signalling pathway in human laryngeal carcinoma.

The anti-cancer effect of dehydrocostus lactone (DHL) derived from Saussurea costus (Falc.) Lipech against laryngeal carcinoma was assessed. The cytotoxic activity of DHL against laryngeal carcinoma is still obscure. Therefore, our study investigated the role of DHL in the growth inhibition of laryngeal carcinoma in vitro and in vivo, and the molecular mechanism of DHL-induced apoptosis in cancer cells of the larynx. The results showed that DHL inhibits the viability, migration and proliferation of Hep-2 and TU212 cells with little toxic effects on human normal larynx epithelial HBE cell line. Flow cytometry analysis (FAC) analysis and staining assay (Hoechst 33258) indicated that DHL stimulated Hep-2 and TU212 cell apoptosis in a dose-dependent manner. Mechanistically, DHL is capable of inhibiting Hep-2 and TU212 cell viability via promoting p53 and P21 function, meanwhile DHL dose-dependently induces Hep-2 and TU212 cells apoptosis via activating mitochondrial apoptosis by inhibiting PI3K/Akt/Bad pathway and stimulating endoplasmic reticulum stress-mediated apoptosis pathway. In vivo, DHL inhibited the growth of the Hep-2 nude mouse xenograft model and observed no significant signs of toxicity in the organs of nude mice. In vivo experiments further confirmed the anti-cancer effect of DHL on laryngeal carcinoma cells in vitro, and DHL-treated nude mice can reduce the volume of tumours. Together, our study indicated that DHL has the potential to inhibit human laryngeal carcinoma via activating mitochondrial apoptosis pathway by inhibiting PI3K/Akt/Bad signalling pathway and stimulating endoplasmic reticulum stress-mediated apoptosis pathway, providing a strategy for the treatment of human laryngeal carcinoma.

Germacrone Inhibits Cell Proliferation and Induces Apoptosis in Human Esophageal Squamous Cell Carcinoma Cells.

Germacrone, a natural 10-membered monocyclic sesquiterpene with three double bonds and a ketone, was isolated from the roots of traditional Chinese medicine Saussurea costus (SC). The pharmacological value and intrinsic mechanism of germacrone in the treatment of esophageal squamous cell carcinoma (ESCC) are still unclear. Therefore, in this study, we further explored the internal molecular mechanism by which germacrone exerts its antiproliferation and antimigration ability against ESCC. 3-(4,5-Dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assays showed that germacrone dose-dependently inhibited the proliferation of ESCC cells. Flow cytometry analysis (FACS) and wound healing experiments on germacrone treated ESCC cells showed that germacrone could induce apoptosis and inhibit the migration of ESCC cells in a dose-dependent manner. In the study on the mechanism of action of germacrone in antiesophageal cancer, we found that germacrone increased the ratio of Bax/Bcl-2 in the cytoplasm of ESCC, resulting in the activation of Caspase-9 and Caspase-3 and decreased the expression of Grp78, thereby reducing the inhibition of Caspase-12 and Caspase-7. In addition, we found that germacrone also inhibited STAT3 phosphorylation in a dose-dependent manner. In conclusion, we determined that germacrone exerted an antiesophageal effect through intrinsic apoptotic signaling pathways and by inhibiting STAT3 activity in ESCC cells.

Study of the Relationship between Microbiome and Colorectal Cancer Susceptibility Using 16SrRNA Sequencing.

A lot of previous studies have recently reported that the gut microbiota influences the development of colorectal cancer (CRC) in Western countries, but the role of the gut microbiota in Chinese population must be investigated fully. The goal of this study was to determine the role of the gut microbiome in the initiation and development of CRC. We collected fecal samples of 206 Chinese individuals: 59 with polyp (group P), 54 with adenoma (group A), 51 with colorectal cancer (group CC), and 42 healthy controls (group HC).16S ribosomal RNA (rRNA) was used to compare the microbiota community structures among healthy controls, patients with polyp, and those with adenoma or colorectal cancer. Our study proved that intestinal flora, as a specific indicator, showed significant differences in its diversity and composition. Sobs, Chao, and Ace indexes of group CC were significantly lower than those of the healthy control group (CC group: Sobs, Chao, and Ace indexes were 217.3 ± 69, 4265.1 ± 80.7, and 268.6 ± 78.1, respectively; HC group: Sobs, Chao, and Ace indexes were 228.8 ± 44.4, 272.9 ± 58.6, and 271.9 ± 57.2, respectively). When compared with the healthy individuals, the species richness and diversity of intestinal flora in patients with colorectal cancer were significantly reduced: PCA and PCoA both revealed that a significant separation in bacterial community composition between the CC group and HC group (with PCA using the first two principal component scores of PC1 14.73% and PC2 10.34% of the explained variance, respectively; PCoA : PC1 = 14%, PC2 = 9%, PC3 = 6%). Wilcox tests was used to analyze differences between the two groups, it reveals that Firmicutes (P=0.000356), Fusobacteria (P=0.000001), Proteobacteria (P=0.000796), Spirochaetes (P=0.013421), Synergistetes (P=0.005642) were phyla with significantly different distributions between cases and controls. The proportion of microorganism composition is varying at different stages of colon cancer development: Bacteroidetes (52.14%) and Firmicutes (35.88%) were enriched in the healthy individuals; on the phylum level, the abundance of Bacteroidetes (52.14%-53.92%-52.46%-47.06%) and Firmicutes (35.88%-29.73%-24.27%-25.36%) is decreasing with the development of health-polyp-adenomas-CRC, and the abundance of Proteobacteria (9.33%-12.31%-16.51%-22.37%) is increasing. PCA and PCOA analysis showed there was no significant (P < 0.05) difference in species similarity between precancerous and carcinogenic states. However, the composition of the microflora in patients with precancerous lesions (including patients with adenoma and polyp) was proved to have no significant disparity (P < 0.05). Our study provides insights into new angles to dig out potential biomarkers in diagnosis and treatment of colorectal cancer and to provide scientific advice for a healthy lifestyle for the sake of gut microbiota.

A single quantum dot-based nanosensor with multilayer of multiple acceptors for ultrasensitive detection of human alkyladenine DNA glycosylase.

Base excision repair (BER) is an important DNA repair pathway involved in the maintenance of genome stability. As the initiator of BER, DNA glycosylase can remove a damaged base from DNA through cleaving the N-glycosidic bond between the sugar moiety and the damaged base. Accurate quantification of DNA glycosylase is essential for the early diagnosis of various human diseases. However, conventional methods for DNA glycosylase assay usually suffer from poor sensitivity and complex probe design. Herein, we develop a single quantum dot-based nanosensor with multilayer of multiple acceptors for ultrasensitive detection of human alkyladenine DNA glycosylase (hAAG) using apurinic/apyrimidinic endonuclease 1 (APE1)-assisted cyclic cleavage-mediated signal amplification in combination with the DNA polymerase-assisted multiple cyanine 5 (Cy5)-mediated fluorescence resonance energy transfer (FRET). The presence of hAAG induces the cleavage of the hairpin substrate, generating a trigger. The resultant trigger can hybridize with a probe modified with an AP site, initiating the APE1-mediated cyclic cleavage to produce a large number of primers. The primers can subsequently initiate the polymerase-mediated signal amplification with a biotin-modified capture probe as the template, generating the biotin-/multiple Cy5-labeled double-stranded DNAs (dsDNAs). The resultant dsDNAs can assemble onto the QD surface to form the QD-dsDNA-Cy5 nanostructure, leading to efficient FRET from the QD to Cy5 under the excitation of 405 nm. In contrast to the typical QD-based FRET approaches, the assembly of multilayer of multiple Cy5 molecules onto a single QD significantly amplifies the FRET signal. We further verify the FRET model with one donor and multilayered acceptors theoretically and experimentally. This single QD-based nanosensor can sensitively detect hAAG with a detection limit of as low as 4.42 × 10-12 U µL-1. Moreover, it can detect hAAG even in a single cancer cell, and distinguish the cancer cells from the normal cells. Importantly, this single QD-based nanosensor can be used for the kinetic study and inhibition assay, and it may become a universal platform for the detection of other DNA repair enzymes by designing appropriate DNA substrates.

Endo-siRNAs regulate early embryonic development by inhibiting transcription of long terminal repeat sequence in pig†.

Activity of some endogenous retroviruses (ERVs) has been proven to be important for development of early mammalian embryo. However, abnormal activation of ERVs can also cause genetic diseases due to their ability to retrotranspose, so the regulatory mechanism to limit transcription of ERVs needs to be clarified. Endogenous small interfering RNA (endo-siRNA) has been reported to protect cells against transposable elements (TEs). Here, we determined the role of ERVs long terminal repeat sequences (LTRs) derived endo-siRNAs (LTR-siRNAs) on inhibition of the activity of ERVs during early embryonic development in pig. Seven most highly expressed LTR-siRNAs were identified in porcine zygote by high-throughput small RNA sequencing. We verified that the biogenesis of the LTR-siRNAs was DICER-dependent and they were generated from double-stranded RNA (dsRNA) formed by sense and antisense transcripts of LTRs. And, the expression of sense and antisense of LTRs might be due to the loss of DNA methylation at some LTR loci. Furthermore, we showed that the LTR-siRNAs could regulate early embryonic development by repression of LTRs expression at a post-transcriptional level. So, we propose here, during early embryonic development when epigenetic reprogramming occurs, the endo-siRNA pathway acts as a sophisticated balance of regulatory mechanism for ERV activity.

Toxicity and bioaccumulation of three hexabromocyclododecane diastereoisomers in the marine copepod Tigriopus japonicas.

The three major hexabromocyclododecane (HBCD) diastereoisomers, i.e. α-, ß- and γ-HBCD, have distinct physical and chemical properties that may potentially result in different levels of bioaccumulation and toxicity in aquatic organisms. To assess the impact of diastereomeric variation in HBCDs, the marine copepod Tigriopus japonicus was exposed to α-, ß- and γ-HBCD in isolation. Results showed that all the three diastereoisomers had a similar potency to cause growth delay in T. japonicas. Variation was observed in the overall survival rate with exposure to α- and ß-HBCD, and this resulted in significantly higher lethal toxicity in T. japonicas than that with exposure to γ-HBCD. Exposure to α-, ß- and γ-HBCD led to the generation of ROS in T. japonicas, a possibly toxic mechanism. Both α- and ß-HBCD showed a higher potential to induce oxidative stress, which may be a factor in the higher lethal toxicity observed with α- and ß-HBCD exposure. It is of note that T. japonicus was found to be more sensitive to all three diastereoisomers in the F1 generation than in the F0 generation. The bioconcentration potential of HBCD diastereoisomers can be ranked in the order α-HBCD>γ-HBCD>ß-HBCD and was found to be higher in T. japonicus than has been reported for fish species.

Endo-siRNA deficiency results in oocyte maturation failure and apoptosis in porcine oocytes.

Both microRNAs (miRNAs) and endogenous small interfering RNAs (endo-siRNAs) play key regulatory roles in gene expression. Some studies have demonstrated that the function of miRNA is suppressed in mouse oocytes, suggesting that endo-siRNA, not miRNA, is essential for female meiosis. This finding has yet to be confirmed in other species. In this study, by knockdown of DICER1, DROSHA and its cofactor DiGeorge syndrome critical region 8 (DGCR8) in porcine oocytes, we found that the proportion of oocytes with DICER1 deficiency that developed to meiosis II (MII) stage was significantly lower than oocytes with DROSHA and DGCR8 deficiency (39.23 versus 68.71 and 71.25% respectively; P<0.05). Oocytes lacking DROSHA and DGCR8 formed a barrel-shaped metaphase I spindle, with chromosomes tightly aligned at the metaphase plate whereas most oocytes (87%) lacking DICER1 showed spindle abnormalities during oocyte in vitro maturation. Furthermore, DICER1 deficiency also resulted in oocyte apoptosis. These results indicate that endo-siRNAs are essential for oocyte maturation in pigs.