2-week prevalence and associated factors of fever, diarrhea, and coexisting fever and diarrhea among children aged 6-23 months in rural Hunan Province.

Fever and diarrhea are key causes of malnutrition, growth and development disorders, and death among children. At present, most studies on the associated factors of fever and diarrhea in children are concentrated in African and South Asian countries, but relevant research in China is very limited. This study was aimed to analyze the two-week prevalence of fever, diarrhea, and coexisting fever and diarrhea among children aged 6-23 months in rural areas of Hunan Province and to explore the associated factors. The survey data of the Nutrition Improvement Program for Children in Poor Areas (NIPCPA) from 2016 to 2023 was used here. NIPCPA is a cross-sectional survey completed annually in Hunan to collect children's nutrition and health indicators. The two-week prevalence rates of fever, diarrhea, and coexisting fever and diarrhea among children aged 6-23 months were 12.2% (2066/16,985), 9.6% (1634/16,985), and 3.2% (542/16,985), respectively. Multivariate logistic regression analysis showed the risks of fever, diarrhea, and coexisting fever and diarrhea were higher among younger children. The high educational level of caregivers, effective consumption of Yingyangbao (a complementary food supplement containing iron, zinc, calcium, vitamins A, D, B1, B2, B12, folic acid, and other micronutrients), and complementary feeding meeting minimum dietary diversity and meeting minimum acceptable diet were protective factors against fever in children, with adjusted odds ratios (aORs) of 0.87 (95%CI: 0.78-0.98), 0.78 (0.69-0.87), 0.73 (0.65-0.82), and 0.74 (0.66-0.84), respectively. Effective consumption of Yingyangbao, and complementary feeding meeting the minimum dietary diversity and meeting minimum acceptable diet were protective factors against diarrhea in children, with aORs of 0.72 (95%CI: 0.63-0.83), 0.79 (0.70-0.91), and 0.80 (0.70-0.92), respectively. Effective consumption of Yingyangbao, and complementary feeding meeting the minimum dietary diversity and meeting minimum acceptable diet were protective factors against coexisting fever and diarrhea among children, with aORs of 0.53 (95%CI: 0.43-0.66), 0.71 (0.58-0.89), and 0.70 (0.56-0.88), respectively. Fever, diarrhea, and the coexisting fever and diarrhea affect one in eight, one in ten, and one in thirty children respectively in rural areas of Hunan. Effective interventions should be actively taken, such as improving the education level of caregivers, enhancing their scientific feeding skills for children, and promoting children's compliance with Yingyangbao consumption, to further reduce the prevalence of fever and diarrhea in children.

Silibinin, a commonly used therapeutic agent for non-alcohol fatty liver disease, functions through upregulating intestinal expression of fibroblast growth factor 15/19.

BACKGROUND AND PURPOSE: Silibinin is used to treat non-alcohol fatty liver disease (NAFLD) despite having rapid liver metabolism. Therefore, we investigated the role of the intestine in silibinin mechanism of action. EXPERIMENTAL APPROACH: NAFLD mice model was established by feeding them with a high-fat diet (HFD). Liver pathological were examined using H&E and oil red O staining. Tissue distribution of silibinin was detected by LC-MS/MS. SiRNA was employed for gene silencing and plasmid was used for gene overexpression. ChIP-qPCR assay was performed to detect the levels of histone acetylation. Recombinant adeno-associated virus 9-short hairpin-fibroblast growth factor (FGF)-15 and -farnesoid X receptor (FXR; NR1H4) were used to knockdown expression of FGF-15 and FXR. KEY RESULTS: Oral silibinin significantly reversed NAFLD in mice, although liver concentration was insufficient for reduction of lipid accumulation in hepatocytes. Among endogenous factors capable of reversing NAFLD, the expression of Fgf-15 was selectively up-regulated by silibinin in ileum and colon of mice. When intestinal expression of Fgf-15 was knocked down, protection of silibinin against lipid accumulation and injury of livers nearly disappeared. Silibinin could reduce activity of histone deacetylase 2 (HDAC2), enhance histone acetylation in the promoter region of FXR and consequently increase intestinal expression of FGF-15/19. CONCLUSION AND IMPLICATIONS: Oral silibinin selectively promotes expression of FGF-15/19 in ileum by enhancing transcription of FXR via reduction of HDAC2 activity, and FGF-15/19 enters into circulation to exert anti-NAFLD action. As the site of action is the intestine this would explain the discrepancy between pharmacodynamics and pharmacokinetics of silibinin.

[Association between infant and young child feeding indicators and the nutritional status of children aged 6-23 months in rural areas of Hunan Province in 2019].

OBJECTIVE: To estimate the status of complementary feeding among infants and young children aged 6-23 months in rural areas of Hunan Province. The association between infant and young child feeding indicators and child undernutrition were assessed. METHODS: A total of 1220 infants and young children aged 6-23 months from 24 investigated places of 6 cities in Hunan Province were selected by multi-stage stratified sampling for physical measurement, hemoglobin(Hb) test and caregiver interview. Complementary diet was analyzed according to the World Health Organization's definition of infant and young child feeding indicators. Z-scores were used to elevate nutrition status. Logistic regression models were used to explore the influencing factors of the nutritional status. RESULTS: The prevalence rates of underweight, stunting, wasting, overweight, obesity and anemia were 3.6%, 4.8%, 2.7%, 10.5%, 2.0% and 16.3%. The percentage of infants and young children aged 6-23 months in rural areas of Hunan Province who get minimum dietary diversity, minimum meal frequency, and minimum acceptable diet was 43.3%, 68.5% and 28.1%. None of the individual infant and young child feeding indicators showed significant association with undernutrition, except minimum meal frequency for obesity and anemia. CONCLUSION: The nutritional status of infants and young children in rural areas of Hunan Province has improved, but the anemia problem is still serious. Complementary feeding frequency is closely associated with anemia for infants and young children.

TAL1 hijacks MYCN enhancer that induces MYCN expression and dependence on mevalonate pathway in T-cell acute lymphoblastic leukemia.

A hallmark of T-cell acute lymphoblastic leukemia (T-ALL) is the dysregulated expression of oncogenic transcription factors (TFs), including TAL1, NOTCH1 and MYC. Rewiring of the transcriptional program disrupts the tightly controlled spatiotemporal expression of downstream target genes, thereby contributing to leukemogenesis. In this study, we first identify an evolutionarily conserved enhancer element controlling the MYCN oncogene (named enhMYCN) that is aberrantly activated by the TAL1 complex in T-ALL cells. TAL1-positive T-ALL cells are highly dependent on MYCN expression for their maintenance in vitro and in xenograft models. Interestingly, MYCN drives the expression of multiple genes involved in the mevalonate pathway, and T-ALL cells are sensitive to inhibition of HMG-CoA reductase (HMGCR), a rate-limiting enzyme of this pathway. Importantly, MYC and MYCN regulate the same targets and compensate for each other. Thus, MYCN-positive T-ALL cells display a dual dependence on the TAL1-MYCN and NOTCH1-MYC pathways. Together, our results demonstrate that enhMYCN-mediated MYCN expression is required for human T-ALL cells and implicate the TAL1-MYCN-HMGCR axis as a potential therapeutic target in T-ALL.

Real-Time Scene Text Detection With Differentiable Binarization and Adaptive Scale Fusion.

Recently, segmentation-based scene text detection methods have drawn extensive attention in the scene text detection field, because of their superiority in detecting the text instances of arbitrary shapes and extreme aspect ratios, profiting from the pixel-level descriptions. However, the vast majority of the existing segmentation-based approaches are limited to their complex post-processing algorithms and the scale robustness of their segmentation models, where the post-processing algorithms are not only isolated to the model optimization but also time-consuming and the scale robustness is usually strengthened by fusing multi-scale feature maps directly. In this paper, we propose a Differentiable Binarization (DB) module that integrates the binarization process, one of the most important steps in the post-processing procedure, into a segmentation network. Optimized along with the proposed DB module, the segmentation network can produce more accurate results, which enhances the accuracy of text detection with a simple pipeline. Furthermore, an efficient Adaptive Scale Fusion (ASF) module is proposed to improve the scale robustness by fusing features of different scales adaptively. By incorporating the proposed DB and ASF with the segmentation network, our proposed scene text detector consistently achieves state-of-the-art results, in terms of both detection accuracy and speed, on five standard benchmarks.

Advances in cognitive behavioral art therapy in the treatment of anxiety disorders in children and adolescents / 中国学校卫生

Abstract@#Cognitive behavior therapy (CBT) is one of the main psychotherapeutic methods used to treat anxiety disorders in children and adolescents. Art therapy, based on psychotherapy theory and methods, is rising in popularity abroad. At present, foreign scholars have started to apply cognitive behavioral art therapy (CBAT) for patients with anxiety disorders. Based on the current findings, children and adolescents with developing cognitive functions may also benefit from this therapy. This article summarizes the application status of CBAT in the treatment of anxiety disorder in patients at home and abroad, and innovatively explores its therapeutic potential for anxiety disorders in children and adolescents in China, thus providing a novel perspective that can address the shortcomings of CBT.

Prevalence, complete genome, and metabolic potentials of a phylogenetically novel cyanobacterial symbiont in the coral-killing sponge, Terpios hoshinota.

Terpios hoshinota is an aggressive, space-competing sponge that kills various stony corals. Outbreaks of this species have led to intense damage to coral reefs in many locations. Here, the first large-scale 16S rRNA gene survey across three oceans revealed that bacteria related to the taxa Prochloron, Endozoicomonas, SAR116, Ruegeria, and unclassified Proteobacteria were prevalent in T. hoshinota. A Prochloron-related bacterium was the most dominant and prevalent cyanobacterium in T. hoshinota. The complete genome of this uncultivated cyanobacterium and pigment analysis demonstrated that it has phycobiliproteins and lacks chlorophyll b, which is inconsistent with the definition of Prochloron. Furthermore, the cyanobacterium was phylogenetically distinct from Prochloron, strongly suggesting that it should be a sister taxon to Prochloron. Therefore, we proposed this symbiotic cyanobacterium as a novel species under the new genus Candidatus Paraprochloron terpiosi. Comparative genomic analyses revealed that 'Paraprochloron' and Prochloron exhibit distinct genomic features and DNA replication machinery. We also characterized the metabolic potentials of 'Paraprochloron terpiosi' in carbon and nitrogen cycling and propose a model for interactions between it and T. hoshinota. This study builds a foundation for the study of the T. hoshinota microbiome and paves the way for better understanding of ecosystems involving this coral-killing sponge.

Mask TextSpotter: An End-to-End Trainable Neural Network for Spotting Text with Arbitrary Shapes.

Unifying text detection and text recognition in an end-to-end training fashion has become a new trend for reading text in the wild, as these two tasks are highly relevant and complementary. In this paper, we investigate the problem of scene text spotting, which aims at simultaneous text detection and recognition in natural images. An end-to-end trainable neural network named as Mask TextSpotter is presented. Different from the previous text spotters that follow the pipeline consisting of a proposal generation network and a sequence-to-sequence recognition network, Mask TextSpotter enjoys a simple and smooth end-to-end learning procedure, in which both detection and recognition can be achieved directly from two-dimensional space via semantic segmentation. Further, a spatial attention module is proposed to enhance the performance and universality. Benefiting from the proposed two-dimensional representation on both detection and recognition, it easily handles text instances of irregular shapes, for instance, curved text. We evaluate it on four English datasets and one multi-language dataset, achieving consistently superior performance over state-of-the-art methods in both detection and end-to-end text recognition tasks. Moreover, we further investigate the recognition module of our method separately, which significantly outperforms state-of-the-art methods on both regular and irregular text datasets for scene text recognition.

Hepatitis E virus infection in 6-month-old pigs in Taiwan.

Hepatitis E virus (HEV) is the causative agent of acute hepatitis E. Genotype 3 (G3) and 4 (G4) HEV have recently been identified in and isolated from swine as the main HEV genotypes worldwide. However, there is limited information on HEV infection status among pigs in Taiwan, especially pigs in the stage before transportation to the slaughterhouse. To determine the frequency of HEV infection among pigs in Taiwan, we detected and quantified HEV RNA contained in 295 fecal specimens collected from 6-month-old pigs bred in 30 pig farms located in 8 counties. We found that 25.1% (74/295) of the fecal specimens were positive for HEV RNA by a quantitative real-time reverse transcription-polymerase chain reaction, and the copy number ranged from 2.3 × 103 to 2.08 × 107 copies/g. Amplification of a 338 bp sequence in ORF2 was achieved in 16 of 74 HEV RNA-positive samples, and their nucleotide sequences were determined. Two HEV sequences appeared to belong to subtype 3a of G3 and the remaining 14 HEV sequences belonged to subtype 4b of G4 (G4b). The entire genome sequence of two G4b HEVs was obtained by next-generation sequence analyses, and the phylogenetic analyses indicated that unique G4b HEVs were circulating in pig farms in Taiwan. In the present study, we found that both G3 and G4 HEVs were circulating in Taiwanese pig farms and G4b was the predominant subtype. In addition, the relatively high detection frequency of HEV RNA in the 6-month-old pigs indicated that Taiwanese pigs just before transportation to the slaughterhouse are at risk of carrying HEVs, and thus thorough cooking or heating of pork meat or organs is needed before consumption in Taiwan and possibly in other countries as well.

Anemia prevalence, severity and associated factors among children aged 6-71 months in rural Hunan Province, China: a community-based cross-sectional study.

BACKGROUND: In recent 10 years, with the rapid socioeconomic development and the extensive implementation of children nutrition improvement projects, the previous epidemiological data cannot reflect the actual level of anemia among children in China, especially in rural areas. Therefore, this study analyzed the prevalence, severity and associated factors of anemia among children aged 6-71 months in rural Hunan Province. METHODS: A community-based cross-sectional study was conducted. Through multistage stratified cluster sampling, 5229 children aged 6 to 71 months and their caregivers were randomly selected from 72 villages across 24 towns in 12 counties from rural Hunan. The demographic characteristics of children and their caregivers, feeding practice, nutritional status of children, caregivers' anemia-related feeding knowledge, and gestational conditions of mothers were acquired by using a unified questionnaire. Peripheral blood from the left-hand middle fingertip was sampled from each child, and hemoglobin concentration was measured using a HemoCue301 portable hemoglobin analyzer (Sweden). Associated factors analyses involving overall anemia and anemia severities were conducted on multivariate logistic regression models. RESULTS: The overall anemia prevalence was 8.8%, and the prevalence of mild, moderate and severe anemia was 6.3, 2.5 and 0.1%, respectively. Children age groups of 6-11 months, 12-23 months and 36-47 months, exclusive breast-feeding within 6 months after birth, and maternal moderate/severe anemia were significantly associated with an increased risk of overall anemia in children. Children age groups of 6-11 months and 12-23 months were significantly associated with an increased risk of mild anemia in children. Children age groups of 6-11 months, 12-23 months and 36-47 months, low caregivers' anemia-related feeding knowledge level, and maternal moderate/severe anemia were significantly associated with an increased risk of moderate/severe anemia in children. Children who underwent regular physical examination were less likely to have moderate/severe anemia. The common protective factor for overall, mild and moderate/severe anemia in children was high family income. CONCLUSIONS: The anemia status of preschool children in rural Hunan Province was a mild public health problem and associated with children age group, feeding practice, regular physical examination, family income, caregivers' anemia-related feeding knowledge level, and maternal moderate/severe anemia.

TextBoxes++: A Single-Shot Oriented Scene Text Detector.

Scene text detection is an important step of scene text recognition system and also a challenging problem. Different from general object detection, the main challenges of scene text detection lie on arbitrary orientations, small sizes, and significantly variant aspect ratios of text in natural images. In this paper, we present an end-to-end trainable fast scene text detector, named TextBoxes++, which detects arbitrary-oriented scene text with both high accuracy and efficiency in a single network forward pass. No post-processing other than an efficient non-maximum suppression is involved. We have evaluated the proposed TextBoxes++ on four public datasets. In all experiments, TextBoxes++ outperforms competing methods in terms of text localization accuracy and runtime. More specifically, TextBoxes++ achieves an f-measure of 0.817 at 11.6fps for 1024 × 1024 ICDAR 2015 Incidental text images, and an f-measure of 0.5591 at 19.8fps for 768 × 768 COCO-Text images. Furthermore, combined with a text recognizer, TextBoxes++ significantly outperforms the stateof-the-art approaches for word spotting and end-to-end text recognition tasks on popular benchmarks.

Enhanced aerobic glycolysis of nasopharyngeal carcinoma cells by Epstein-Barr virus latent membrane protein 1.

Latent membrane protein 1 (LMP1) is a principal viral oncoprotein in Epstein-Barr virus (EBV)-associated malignancies, including nasopharyngeal carcinoma (NPC), which acts through regulating tumorigenesis and metabolic reprogramming of cancers. In the presence of oxygen, we demonstrated that glucose consumption, lactate production and lactate dehydrogenase (LDH) activity were significantly increased upon LMP1 expression in NPC cells and in a LMP1 variant derived from NPC patients-transformed BALB/c-3T3 cells. The amounts of the α subunit of hypoxia-inducible factor-1 (HIF-1α), a key regulator of aerobic glycolysis, and its targets, pyruvate dehydrogenase kinase 1 (PDK1) and the pyruvate kinase M2 (PKM2) isoform, were also consistently elevated by LMP1. Moreover, in parallel with reductions in the oxygen consumption rate and mitochondrial membrane potential in cells, an augmented extracellular lactate concentration was observed due to LMP1 induction. In conclusion, our results proved facilitation of the Warburg effect by LMP1 through alteration of mitochondrial function in NPC cells.

Novel G9 rotavirus strains co-circulate in children and pigs, Taiwan.

Molecular epidemiologic studies collecting information of the spatiotemporal distribution of rotavirus VP7 (G) and VP4 (P) genotypes have shown evidence for the increasing global importance of genotype G9 rotaviruses in humans and pigs. Sequence comparison of the VP7 gene of G9 strains identified different lineages to prevail in the respective host species although some of these lineages appear to be shared among heterologous hosts providing evidence of interspecies transmission events. The majority of these events indicates the pig-to-human spillover, although a reverse route of transmission cannot be excluded either. In this study, new variants of G9 rotaviruses were identified in two children with diarrhea and numerous pigs in Taiwan. Whole genome sequence and phylogenetic analyses of selected strains showed close genetic relationship among porcine and human strains suggesting zoonotic origin of Taiwanese human G9 strains detected in 2014-2015. Although the identified human G9P[19] and G9P[13] rotaviruses represented minority strains, the repeated detection of porcine-like rotavirus strains in Taiwanese children over time justifies the continuation of synchronized strain surveillance in humans and domestic animals.

Cytotoxicity of 11-epi-Sinulariolide Acetate Isolated from Cultured Soft Corals on HA22T Cells through the Endoplasmic Reticulum Stress Pathway and Mitochondrial Dysfunction.

Natural compounds from soft corals have been increasingly used for their antitumor therapeutic properties. This study examined 11-epi-sinulariolide acetate (11-epi-SA), an active compound isolated from the cultured soft coral Sinularia flexibilis, to determine its potential antitumor effect on four hepatocellular carcinoma cell lines. Cell viability was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and the results demonstrated that 11-epi-SA treatment showed more cytotoxic effect toward HA22T cells. Protein profiling of the 11-epi-SA-treated HA22T cells revealed substantial protein alterations associated with stress response and protein synthesis and folding, suggesting that the mitochondria and endoplasmic reticulum (ER) play roles in 11-epi-SA-initiated apoptosis. Moreover, 11-epi-SA activated caspase-dependent apoptotic cell death, suggesting that mitochondria-related apoptosis genes were involved in programmed cell death. The unfolded protein response signaling pathway-related proteins were also activated on 11-epi-SA treatment, and these changes were accompanied by the upregulated expression of growth arrest and DNA damage-inducible protein (GADD153) and CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP), the genes encoding transcription factors associated with growth arrest and apoptosis under prolonged ER stress. Two inhibitors, namely salubrinal (Sal) and SP600125, partially abrogated 11-epi-SA-related cell death, implying that the protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK)-activating transcription factor (ATF) 6-CHOP or the inositol-requiring enzyme 1 alpha (IRE1α)-c-Jun N-terminal kinase (JNK)-cJun signal pathway was activated after 11-epi-SA treatment. In general, these results suggest that 11-epi-SA exerts cytotoxic effects on HA22T cells through mitochondrial dysfunction and ER stress cell death pathways.

Positive regulation of HIF-1A expression by EBV oncoprotein LMP1 in nasopharyngeal carcinoma cells.

Latent membrane protein 1 (LMP1) is a pivotal viral oncoprotein that contributes to the carcinogenesis of Epstein-Barr virus (EBV)-associated malignancies, including nasopharyngeal carcinoma (NPC). We investigated the regulation of hypoxia-inducible factor 1-α (HIF-1α) by LMP1. In NPC cells, we found that LMP1 significantly enhanced the HIF-1α mRNA level, and not only the protein amount as described previously. Mechanistically, the stability of the HIF-1α transcript was remarkably prolonged by LMP1 via reduced expressions of RNA-destabilizing proteins tristetraprolin (TTP) and pumilio RNA-binding family member 2 (PUM2) through C-terminal activation region 1 (CTAR1) and CTAR3 interaction with the ERK1/2 and STAT3 signaling pathways, respectively, in parallel with hindrance of PUM2 binding to the HIF-1α mRNA 3'-untranslated region (3'-UTR). On the other hand, HIF-1A promoter activity was also obviously facilitated by the LMP1 CTAR1-recruited ERK1/2/NF-κB pathway. Intriguingly, in this scenario, augmented HIF-1α further exhibited positive auto-regulation of its own gene transcription. Our results showed the first time that LMP1 directly up-regulates HIF-1A transcription and post-transcription in NPC cells, in addition to providing evidence of an increase in the HIF-1α mRNA level caused by a tumor-associated virus under normoxic conditions.

Charactering the ZFAND3 gene mapped in the sex-determining locus in hybrid tilapia (Oreochromis spp.).

Zinc finger AN1-type domain 3 (ZFAND3) is essential for spermatogenesis in mice. However, its function in teleosts remains unclear. In this study, we characterized the ZFAND3 gene (termed as OsZFAND3) in an important food fish, tilapia. The OsZFAND3 cDNA sequence is 1,050 bp in length, containing an ORF of 615 bp, which encodes a putative peptide of 204 amino acid residues. Quantitative real-time PCR revealed that the OsZFAND3 transcripts were exclusively expressed in the testis and ovary. In situ hybridization showed that the high expression of OsZFAND3 transcripts was predominantly localized in the spermatocyte and spermatid. These results suggest that OsZFAND3 is involved in male germ cell maturation. Three single nucleotide polymorphisms (SNPs) were detected in the introns of OsZFAND3. The OsZFAND3 gene was mapped in the sex-determining locus on linkage group 1 (LG1). The three SNPs in the OsZFAND3 gene were strictly associated with sex phenotype, suggesting that the OsZFAND3 gene is tightly linked to the sex-determining locus. Our study provides new insights into the functions of the OsZFAND3 gene in tilapia and a foundation for further detailed analysis of the OsZFAND3 gene in sex determination and differentiation.

11-epi-Sinulariolide acetate reduces cell migration and invasion of human hepatocellular carcinoma by reducing the activation of ERK1/2, p38MAPK and FAK/PI3K/AKT/mTOR signaling pathways.

Cancer metastasis is one of the major causes of death in cancer. An active compound, 11-epi-sinulariolide acetate (11-epi-SA), isolated from the cultured soft coral Sinularia flexibilis has been examined for potential anti-cell migration and invasion effects on hepatocellular carcinoma cells (HCC). However, the molecular mechanism of anti-migration and invasion by 11-epi-SA on HCC, along with their corresponding effects, remain poorly understood. In this study, we investigated anti-migration and invasion effects and the underlying mechanism of 11-epi-SA in HA22T cells, and discovered by trans-well migration and invasion assays that 11-epi-SA provided a concentration-dependent inhibitory effect on the migration of human HCC HA22T cells. After treatment with 11-epi-SA for 24 h, there were suppressed protein levels of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) and urokinase-type plasminogen activator (uPA) in HA22T cells. Meanwhile, the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase-2 (TIMP-2) were increased in a concentration-dependent manner. Further investigation revealed that 11-epi-SA suppressed the phosphorylation of ERK1/2 and p38MAPK. The 11-epi-SA also suppressed the expression of the phosphorylation of FAK/PI3K/AKT/mTOR pathways.

High-throughput gender identification of penguin species using melting curve analysis.

Most species of penguins are sexual monomorphic and therefore it is difficult to visually identify their genders for monitoring population stability in terms of sex ratio analysis. In this study, we evaluated the suitability using melting curve analysis (MCA) for high-throughput gender identification of penguins. Preliminary test indicated that the Griffiths's P2/P8 primers were not suitable for MCA analysis. Based on sequence alignment of Chromo-Helicase-DNA binding protein (CHD)-W and CHD-Z genes from four species of penguins (Pygoscelis papua, Aptenodytes patagonicus, Spheniscus magellanicus, and Eudyptes chrysocome), we redesigned forward primers for the CHD-W/CHD-Z-common region (PGU-ZW2) and the CHD-W-specific region (PGU-W2) to be used in combination with the reverse Griffiths's P2 primer. When tested with P. papua samples, PCR using P2/PGU-ZW2 and P2/PGU-W2 primer sets generated two amplicons of 148- and 356-bp, respectively, which were easily resolved in 1.5% agarose gels. MCA analysis indicated the melting temperature (Tm) values for P2/PGU-ZW2 and P2/PGU-W2 amplicons of P. papua samples were 79.75°C-80.5°C and 81.0°C-81.5°C, respectively. Females displayed both ZW-common and W-specific Tm peaks, whereas male was positive only for ZW-common peak. Taken together, our redesigned primers coupled with MCA analysis allows precise high throughput gender identification for P. papua, and potentially for other penguin species such as A. patagonicus, S. magellanicus, and E. chrysocome as well.

Sinulariolide induced hepatocellular carcinoma apoptosis through activation of mitochondrial-related apoptotic and PERK/eIF2α/ATF4/CHOP pathway.

Sinulariolide, an active compound isolated from the cultured soft coral Sinularia flexibilis, has potent anti-microbial and anti-tumorigenesis effects towards melanoma and bladder cancer cells. In this study, we investigated the effects of sinulariolide on hepatocellular carcinoma (HCC) cell growth and protein expression. Sinulariolide suppressed the proliferation and colony formation of HCC HA22T cells in a dose-dependent manner and induced both early and late apoptosis according to flow cytometry, Annexin V/PI stain and TUNEL/DAPI stain analyses. A mechanistic analysis demonstrated that sinulariolide-induced apoptosis was activated through a mitochondria-related pathway, showing up-regulation of Bax, Bad and AIF, and down- regulation of Bcl-2, Bcl-xL, MCl-1 and p-Bad. Sinulariolide treatment led to loss of the mitochondrial membrane potential, release of mitochondrial cytochrome c to the cytosol, and activation of both caspase-9 and caspase-3. Sinulariolide-induced apoptosis was significantly blocked by the caspase inhibitors Z-VAD-FMK and Z-DEVD-FMK. The increased expression of cleaved PARP also suggested that caspase-independent apoptotic pathway was involved. In the western blotting; the elevation of ER chaperones GRP78; GRP94; and CALR; as well as up-regulations of PERK/eIF2α/ATF4/CHOP; and diminished cell death with pre-treatment of eIF2α phosphatase inhibitor; salubrinal; implicated the involvement of ER stress-mediated PERK/eIF2α/ATF4/CHOP apoptotic pathway following sinulariolide treatment in hepatoma cells. The current study suggested sinulariolide-induced hepatoma cell cytotoxicity involved multiple apoptotic signal pathways. This may implicate that sinulariolide is a potential compound for the treatment of hepatocellular carcinoma.