Development of a tetra-primer ARMS-PCR for identification of sika and red deer and their hybrids.

Accurate identification of deer-derived components is significant in food and drug authenticity. Over the years, several methods have been developed to authenticate these products; however, identifying whether female deer products are hybrids is challenging. In this study, the zinc finger protein X-linked (ZFX) gene sequences of sika deer (Cervus nippon), red deer (Cervus elaphus) and their hybrid offspring were amplified and sequenced, the X221 and X428 species-specific single nucleotide polymorphisms (SNP) loci were verified, and a tetra-primer amplification refractory mutation system (T-ARMS-PCR) assay was developed to identify the parent-of-origin of female sika deer, red deer, and their hybrid deer. The T-ARMS-PCR developed based on the X221 locus could identify sika deer, red deer, and their hybrid offspring according to the presence or absence of PCR product sizes of 486 bp, 352 bp, and 179 bp, respectively, just as X428 locus could identify sika deer, red deer, and their hybrid offspring according to the presence or absence of PCR product sizes of 549 bp, 213 bp, and 383 bp, respectively. Forty products labeled deer-derived ingredients randomly purchased were tested using this assay, and the results showed that the identification results based on the two SNP loci were utterly consistent with the actual sources. In addition, this method was found to be accurate, simple, convenient, and with high specificity, thus providing an essential technical reference for deer product species identification. It is also an important supplement to the identification methods of the original ingredients of existing deer products.

The Influence of Parents on Medication Adherence of Their Children in China: A Cross-Sectional Online Investigation Based on Health Belief Model.

Objective: To explore the influence of parents on the medication adherence of their children. Study Design: A cross-sectional online investigation. Methods: A questionnaire with 41 questions was designed based on the health belief model (HBM) distributed and collected online in 28 cities around China through multi-stage stratified sampling. The reliability of the questionnaire was assessed with Cronbach's α coefficient and split-half reliability, and its validity was evaluated with exploratory factor analysis and content validity index. The structural equation model (SEM) was constructed to explore the relationship between the parents' health beliefs and their children's medication adherence. Subgroup analysis was conducted to study the differences between parents with different demographic characteristics (male and female, rural and urban). Results: 573 questionnaires were included for analysis, with an effective rate of 62.97%. The Cronbach'α coefficient of the questionnaire was 0.821 > 0.6, the split-half reliability was 0.651 > 0.6, the I-CVI of each dimension were >0.78, and the S-CVI/AVE (I-CVI average) was 0.95 > 0.9. The result of the questionnaire exploratory factor analysis met the standard. According to the SEM, self-efficacy (λ = 0.177), perceived susceptibility (λ = -0.244), and perceived severity (λ = 0.243) were direct influencing factors of children's medication adherence. In the subgroup analysis, the model established by each subgroup was consistent with the model established by the overall sample. The absolute values of females' perceived susceptibility, severity, and self-efficacy for their children's medication adherence path coefficients were higher than males'. Conclusion: Parents' perceived severity and self-efficacy may positively impact on their children's medication adherence, while parents' susceptibility to children's medication non-adherence may negatively impact on children's medication adherence. Objective constraints, perceived barriers, and benefits may in directly impact on children's medication adherence. Women's health beliefs appear to have a more significant impact on their children's medication adherence than men's. It may be an effective strategy to increase their children's medication adherence by improving parents' health beliefs. Medical staff should explain medication adherence knowledge to the parents of children, and inform the children of the possible consequences of non-adherence with medication, to improve the subjective perception of parents on the severity of children's non-adherence with medication, and improve parents' self-efficacy in rational medication for children. In addition, attention should be paid to the mental health of the parents, and more social and psychological support.

ADARs act as potent regulators of circular transcriptome in cancer.

Circular RNAs (circRNAs) are produced by head-to-tail back-splicing which is mainly facilitated by base-pairing of reverse complementary matches (RCMs) in circRNA flanking introns. Adenosine deaminases acting on RNA (ADARs) are known to bind double-stranded RNAs for adenosine to inosine (A-to-I) RNA editing. Here we characterize ADARs as potent regulators of circular transcriptome by identifying over a thousand of circRNAs regulated by ADARs in a bidirectional manner through and beyond their editing function. We find that editing can stabilize or destabilize secondary structures formed between RCMs via correcting A:C mismatches to I(G)-C pairs or creating I(G).U wobble pairs, respectively. We provide experimental evidence that editing also favors the binding of RNA-binding proteins such as PTBP1 to regulate back-splicing. These ADARs-regulated circRNAs which are ubiquitously expressed in multiple types of cancers, demonstrate high functional relevance to cancer. Our findings support a hitherto unappreciated bidirectional regulation of circular transcriptome by ADARs and highlight the complexity of cross-talk in RNA processing and its contributions to tumorigenesis.

Topography of transcriptionally active chromatin in glioblastoma.

Molecular profiling of the most aggressive brain tumor glioblastoma (GBM) on the basis of gene expression, DNA methylation, and genomic variations advances both cancer research and clinical diagnosis. The enhancer architectures and regulatory circuitries governing tumor-intrinsic transcriptional diversity and subtype identity are still elusive. Here, by mapping H3K27ac deposition, we analyze the active regulatory landscapes across 95 GBM biopsies, 12 normal brain tissues, and 38 cell line counterparts. Analyses of differentially regulated enhancers and super-enhancers uncovered previously unrecognized layers of intertumor heterogeneity. Integrative analysis of variant enhancer loci and transcriptome identified topographies of transcriptional enhancers and core regulatory circuitries in four molecular subtypes of primary tumors: AC1-mesenchymal, AC1-classical, AC2-proneural, and AC3-proneural. Moreover, this study reveals core oncogenic dependency on super-enhancer-driven transcriptional factors, long noncoding RNAs, and druggable targets in GBM. Through profiling of transcriptional enhancers, we provide clinically relevant insights into molecular classification, pathogenesis, and therapeutic intervention of GBM.

Homocysteine induced a calcium-mediated disruption of mitochondrial function and dynamics in endothelial cells.

Homocysteine (Hcy) is a sulfur-containing amino acid that originated in methionine metabolism and the elevated level of Hcy in plasma is considered to be an independent risk factor for cardiovascular diseases (CVD). Endothelial dysfunction plays a major role in the development of CVD, while the potential mechanism of Hcy-induced endothelial dysfunction is still unclear. Here, in Hcy-treated endothelial cells, we observed the destruction of mitochondrial morphology and the decline of mitochondrial membrane potential. Meanwhile, the level of ATP was reduced and the reactive oxygen species was increased. The expressions of dynamin-related protein 1 (Drp1) and phosphate-Drp1 (Ser616) were upregulated, whereas the expression of mitofusin 2 was inhibited by Hcy treatment. These findings suggested that Hcy not only triggered mitochondrial dysfunction but also incurred an imbalance of mitochondrial dynamics in endothelial cells. The expression of mitochondrial calcium uniporter (MCU) was activated by Hcy, contributing to calcium transferring into mitochondria. Interestingly, the formation of mitochondria-associated membranes (MAMs) was increased in endothelial cells after Hcy administration. The inositol 1,4,5-triphosphate receptor (IP3R)-glucose-regulated protein 75 (Grp75)-voltage-dependent anion channel (VDAC) complex, which was enriched in MAMs, was also increased. The accumulation of mitochondrial calcium could be blocked by inhibiting with the IP3R inhibitor Xestospongin C (XeC) in Hcy-treated cells. Then, we confirmed that the mitochondrial dysfunction and the increased mitochondrial fission induced by Hcy could be attenuated after Hcy and XeC co-treatment. In conclusion, Hcy-induced mitochondrial dysfunction and dynamics disorder in endothelial cells were mainly related to the increase of calcium as a result of the upregulated expressions of the MCU and the IP3R-Grp75-VDAC complex in MAMs.

MNK1 and MNK2 enforce expression of E2F1, FOXM1, and WEE1 to drive soft tissue sarcoma.

Soft tissue sarcoma (STS) is a heterogeneous disease that arises from connective tissues. Clinical outcome of patients with advanced tumors especially de-differentiated liposarcoma and uterine leiomyosarcoma remains unsatisfactory, despite intensive treatment regimens including maximal surgical resection, radiation, and chemotherapy. MAP kinase-interacting serine/threonine-protein kinase 1 and 2 (MNK1/2) have been shown to contribute to oncogenic translation via phosphorylation of eukaryotic translation initiation factor 4E (eIF4E). However, little is known about the role of MNK1/2 and their downstream targets in STS. In this study, we show that depletion of either MNK1 or MNK2 suppresses cell viability, anchorage-independent growth, and tumorigenicity of STS cells. We also identify a compelling antiproliferative efficacy of a novel, selective MNK inhibitor ETC-168. Cellular responsiveness of STS cells to ETC-168 correlates positively with that of phosphorylated ribosomal protein S6 (RPS6). Mirroring MNK1/2 silencing, ETC-168 treatment strongly blocks eIF4E phosphorylation and represses expression of sarcoma-driving onco-proteins including E2F1, FOXM1, and WEE1. Moreover, combination of ETC-168 and MCL1 inhibitor S63845 exerts a synergistic antiproliferative activity against STS cells. In summary, our study reveals crucial roles of MNK1/2 and their downstream targets in STS tumorigenesis. Our data encourage further clinical translation of MNK inhibitors for STS treatment.

Mosquito Larvicidal Potential of Tropical Seaweeds: Acetylcholinesterase Inhibitory Effects of Bryopsis pennata, Padina australis and Sargassum binderi on Aedes aegypti (L.) and Aedes albopictus Skuse

@#Introduction: Inhibition of the cholinesterase’s function leads to paralysis and death. This mechanism is served as a common mode of action of insecticide. The three tropical seaweeds, namely Bryopsis pennata, Padina australis and Sargassum binderi were reported for its potential mosquito larvicidal effect. In the present study, these seaweeds were evaluated for their potential as a cholinesterase inhibitor in the mechanism of larvicidal action. Methods: Acetylcholinsterase (AChE) inhibition assay was carried out based on the colorimetric method using a microplate reader. Phytochemical content of the seaweed extracts was screened by using liquid chromatography-mass spectroscopy (LC-MS). Results: Green seaweed B. pennata showed the strongest inhibition effect towards in vitro AChE by using tissue homogenates of Aedes aegypti (IC50 value = 0.84 mg mL-1) and Aedes albopictus as the enzyme source (IC50 value = 0.92 mg mL-1). The pattern of Lineweaver-Burk plots revealed that B. pennata was a mixed type inhibitor of AChE, as the readings of Km, Vmax, Ki and Ki’, indicates that it had a strong inhibition ability with high binding affinity towards both free enzyme and enzyme-substrate complex. Conclusion: These findings suggest the compound(s) in B. pennata extract serves as a promising source that could be developed into a mosquito larvicidal agent with AChE inhibition effect.

Bromodomain and extraterminal proteins foster the core transcriptional regulatory programs and confer vulnerability in liposarcoma.

Liposarcomas (LPSs) are a group of malignant mesenchymal tumors showing adipocytic differentiation. Here, to gain insight into the enhancer dysregulation and transcriptional addiction in this disease, we chart super-enhancer structures in both LPS tissues and cell lines. We identify a bromodomain and extraterminal (BET) protein-cooperated FUS-DDIT3 function in myxoid LPS and a BET protein-dependent core transcriptional regulatory circuitry consisting of FOSL2, MYC, and RUNX1 in de-differentiated LPS. Additionally, SNAI2 is identified as a crucial downstream target that enforces both proliferative and metastatic potentials to de-differentiated LPS cells. Genetic depletion of BET genes, core transcriptional factors, or SNAI2 mitigates consistently LPS malignancy. We also reveal a compelling susceptibility of LPS cells to BET protein degrader ARV-825. BET protein depletion confers additional advantages to circumvent acquired resistance to Trabectedin, a chemotherapy drug for LPS. Moreover, this study provides a framework for discovering and targeting of core oncogenic transcriptional programs in human cancers.

Insecticide susceptibility status and resistance mechanism of Anopheles cracens Sallum and Peyton and Anopheles maculatus Theobald (Family: Culicidae) from knowlesi malaria endemic areas in Peninsular Malaysia

Objective: To assess the insecticide susceptibility status of Anopheles cracens (An. cracens) and Anopheles maculatus (An. maculatus) from knowlesi malaria endemic areas in Peninsular Malaysia towards DDT, malathion and deltamethrin and to determine the resistance mechanism involved. Methods: Adult and larval mosquitos were collected for surveillance. Susceptibility status of Anopheles was determined using the standard WHO adult bioassay, larval bioassay and biochemical enzyme assay. Results: WHO adult bioassay results indicated An. cracens collected from Kampung Sungai Ular, Pahang was resistant towards 4% DDT, while An. maculatus collected from Kampung Sokor, Kelantan and Kampung Sungai Lui, Selangor exhibited resistance towards 4% DDT. However, the enzyme activity profiles varied according to strains and species. The resistance ratio of larval bioassay, showed that all strains and species tested were susceptible to malathion and temephos. Conclusions: Since only a few anopheline strains exhibited low level of insecticide resistance towards malathion, DDT and temephos. These insecticides are still considered effective for vector control program towards An. cracens and An. maculatus.

Targetable BET proteins- and E2F1-dependent transcriptional program maintains the malignancy of glioblastoma.

Competitive BET bromodomain inhibitors (BBIs) targeting BET proteins (BRD2, BRD3, BRD4, and BRDT) show promising preclinical activities against brain cancers. However, the BET protein-dependent glioblastoma (GBM)-promoting transcriptional network remains elusive. Here, with mechanistic exploration of a next-generation chemical degrader of BET proteins (dBET6), we reveal a profound and consistent impact of BET proteins on E2F1- dependent transcriptional program in both differentiated GBM cells and brain tumor-initiating cells. dBET6 treatment drastically reduces BET protein genomic occupancy, RNA-Pol2 activity, and permissive chromatin marks. Subsequently, dBET6 represses the proliferation, self-renewal, and tumorigenic ability of GBM cells. Moreover, dBET6-induced degradation of BET proteins exerts superior antiproliferation effects compared to conventional BBIs and overcomes both intrinsic and acquired resistance to BBIs in GBM cells. Our study reveals crucial functions of BET proteins and provides the rationale and therapeutic merits of targeted degradation of BET proteins in GBM.

Experimental study on the mechanism of evaporative dry eye treated by Qingxuan decoction in rabbits / 国际眼科杂志(Guoji Yanke Zazhi)

·AIM: To study the mechanism and effect of Qingxuan decoction on evaporative dry eye in rabbit model. ·METHODS: Totally 25 healthy male Japanese white rabbits were randomly divided into 5 groups: control group, model group, western medicine group, high dose of Qingxuan decoction group, low dose of Qingxuan decoction group. The blank control group did not do any treatment. The improved dry eye model of rabbit was prepared by the improved method of glandular burning of the eyelid plate. The high and low dose group were given daily 27. 2mg/kg, 6. 8mg/kg Qingxuan decoction by gavage. The model group was intragastric with the same amount of normal saline every day. The western medicine group with tobramycin and dexamethasone ophthalmic ointment 1 drops, once a day. The treatment were administered continuously for 28d. At 14d before the experiment, 7d before the experiment, 7d after the model, and 14d after the model, all the rabbits were tested by Schirmer Ⅰ test ( SⅠt) and break-up time (BUT). On the 15d after modeling, the animals were sacrificed by excessive anaesthesia. Rabbit ocular surface tissue sections were prepared. Hematoxylin - eosin staining method was used to observe the corneal morphological changes in each group. The concentrations of TNF-α, IL-1 and IL-6 in the ocular surface of rabbits were detected by ELISA. ·RESULTS: (1) BUT, SⅠt: 7d after the model had been prepared, BUT and SⅠt of the model group and the western medicine group, high dose and low dose of Qingxuan decoction group was improved ( P< 0. 05 );Those of western medicine group, high dose and low dose of Qingxuan Decoction group compared with the model group, were significantly different (P<0. 05). (2) TNF-α, IL-1, IL-6: The ELISA assay showed that TNF-α and IL-1, IL-6 concentration in the model group rabbits was significantly higher than those of the control group, TNF-α and IL-1, IL-6 concentration in western medicine group and high dose group of rabbits was significantly lower than those in the model group, the differences were statistically significant (P<0. 05), and in high dose group the effect was better than that of Western medicine group. ( 3 ) Histopathological examination: on the 14d after the model, the corneal epithelium in the blank control group was stratified well. The cells in the base were columnar, near the surface, the cornea epithelium showed a squamous change. Conjunctiva showed complete epithelial layer and subconjunctival tissue layer, and goblet cells arranged closely. The number of corneal epithelial cells in model group was reduced or even stripped, and the matrix layer was disorder; Irregular loss of conjunctival epithelial cell layer and a large decrease in goblet cells. The corneal morphology of the rabbits in the western medicine group and the high dose group was close to the normal group, and the number of conjunctival goblet cells was not significantly different from that in the blank control group. ·CONCLUSION: The expression of Qingxuan decoction can inhibit the inflammatory reaction through down -regulation of TNF-α and IL-1, IL-6 and in evaporative dry eye rabbit cornea and conjunctiva, so as to improve the ocular symptoms, increase tear secretion, prolong the time of BUT.

Larvicidal activity, inhibition effect on development, histopathological alteration and morphological aberration induced by seaweed extracts in Aedes aegypti (Diptera: Culicidae)

OBJECTIVE@#To investigate the larvicidal activity, inhibition effect on development, histopathological alteration and morphological aberration induced by the extracts derived from seaweeds Bryopsis pennata (B. pennata), Sargassum binderi (S. binderi) and Padina australis in Aedes aegypti (Ae. aegypti) larvae and to characterize the phytochemical components of the three seaweeds.@*METHODS@#Larvicidal activity of the seaweeds towards the larvae of Ae. aegypti was determined according to WHO. The inhibition effect of seaweeds was assessed by determining the mortality, adult emergence rate, larval and pupa duration of the treated larvae. Histopathological effect on midgut epithelium of larvae and morphological aberration induced by the methanol extracts were examined. Phytochemical analysis was done to determine the presence of alkaloids, saponins, steroids and terpenoids in the seaweeds.@*RESULTS@#Chloroform partition of B. pennata extract exhibited the strongest larvicidal activity (LC50 = 82.55 μg/mL), followed by methanol extract of B. pennata (LC50 = 160.07 μg/mL) and chloroform partition of S. binderi extract (LC50 = 192.43 μg/mL). The methanol extract of S. binderi exhibited the strongest effect on prolongation of larval period (1.5-fold longer as compared to control) and resulted in strongest inhibition effect in adult emergence (98.67%). The histopathological study showed that larvae treated with seaweed extracts had cytopathological alteration of the midgut epithelium. The morphological observation revealed that the anal papillae and terminal spiracles of larvae were the common sites of aberrations.@*CONCLUSIONS@#The study provided information on various effects of seaweed extracts on Ae. aegypti. Further investigation on identifying the active compounds and their mechanisms of action is recommended.

Larvicidal activity, inhibition effect on development, histopathological alteration and morphological aberration induced by seaweed extracts in Aedes aegypti (Diptera: Culicidae)

Objective: To investigate the larvicidal activity, inhibition effect on development, histopathological alteration and morphological aberration induced by the extracts derived from seaweeds Bryopsis pennata (B. pennata). Sargassum binderi (S. binderi) and Padina australis in Aedes aegypti (Ae. aegypti) larvae and to characterize the phytochemical components of the three seaweeds. Methods: Larvicidal activity of the seaweeds towards the larvae of Ae. aegypti was determined according to WHO. The inhibition effect of seaweeds was assessed by determining the mortality, adult emergence rate, larval and pupa duration of the treated larvae. Histopathological effect on midgut epithelium of larvae and morphological aberration induced by the methanol extracts were examined. Phytochemical analysis was done to determine the presence of alkaloids, saponins, steroids and terpenoids in the seaweeds. Results: Chloroform partition of B. pennata extract exhibited the strongest larvicidal activity (LC