Nanopore sequencing of forensic short tandem repeats using QNome of Qitan Technology.

Devices of nanopore sequencing can be highly portable and of low cost. Thus, nanopore sequencing is promising in in-field forensic applications. Previous investigations have demonstrated that nanopore sequencing is feasible for genotyping forensic short tandem repeats (STRs) by using sequencers of Oxford Nanopore Technologies. Recently, Qitan Technology launched a new portable nanopore sequencer and became the second supplier in the world. Here, for the first time, we assess the QNome (QNome-3841) for its accuracy in nanopore sequencing of STRs and compare with MinION (MinION Mk1B). We profile 54 STRs of 21 unrelated individuals and 2800M standard DNA. The overall accuracy for diploid STRs and haploid STRs were 53.5% (378 of 706) and 82.7% (134 of 162), respectively, by using QNome. The accuracies were remarkably lower than those of MinION (diploid STRs, 84.5%; haploid, 90.7%), with a similar amount of sequencing data and identical bioinformatics analysis. Although it was not reliable for diploid STRs typing by using QNome, the haploid STRs were consistently correctly typed. The majority of errors (58.8%) in QNome-based STR typing were one-repeat deviations of repeat units in the error from true allele, related with homopolymers in repeats of STRs.

The role of insulin-like growth factor 2 mRNA binding proteins in female reproductive pathophysiology.

Insulin-like growth factor 2 (IGF2) mRNA binding proteins (IMPs) family belongs to a highly conserved family of RNA-binding proteins (RBPs) and is responsible for regulating RNA processing including localization, translation and stability. Mammalian IMPs (IMP1-3) take part in development, metabolism and tumorigenesis, where they are believed to play a major role in cell growth, metabolism, migration and invasion. IMPs have been identified that are expressed in ovary, placenta and embryo. The up-to-date evidence suggest that IMPs are involved in folliculogenesis, oocyte maturation, embryogenesis, implantation, and placentation. The dysregulation of IMPs not only contributes to carcinogenesis but also disturbs the female reproduction, and may participate in the pathogenesis of reproductive diseases and obstetric syndromes, such as polycystic ovary syndrome (PCOS), pre-eclampsia (PE), gestational diabetes mellitus (GDM) and gynecological tumors. In this review, we summarize the role of IMPs in female reproductive pathophysiology, and hope to provide new insights into the identification of potential therapeutic targets.

The anticancer potential of the dietary polyphenol rutin: Current status, challenges, and perspectives.

Rutin is one of the most common dietary polyphenols found in vegetables, fruits, and other plants. It is metabolized by the mammalian gut microbiota and absorbed from the intestines, and becomes bioavailable in the form of conjugated metabolites. Rutin exhibits a plethora of bioactive properties, making it an extremely promising phytochemical. Numerous studies demonstrate that rutin can act as a chemotherapeutic and chemopreventive agent, and its anticancer effects can be mediated through the suppression of cell proliferation, the induction of apoptosis or autophagy, and the hindering of angiogenesis and metastasis. Rutin has been found to modulate multiple molecular targets involved in carcinogenesis, such as cell cycle mediators, cellular kinases, inflammatory cytokines, transcription factors, drug transporters, and reactive oxygen species. This review summarizes the natural sources of rutin, its bioavailability, and in particular its potential use as an anticancer agent, with highlighting its anticancer mechanisms as well as molecular targets. Additionally, this review updates the anticancer potential of its analogs, nanoformulations, and metabolites, and discusses relevant safety issues. Overall, rutin is a promising natural dietary compound with promising anticancer potential and can be widely used in functional foods, dietary supplements, and pharmaceuticals for the prevention and management of cancer.

LAWN TRIMMER-RELATED OPEN-GLOBE INJURIES IN TAIWAN.

PURPOSE: Work-related ocular trauma remains the leading cause of unilateral visual impairment worldwide. Many preventable work-related ocular injuries continue to occur, even at home. This study describes the characteristics, surgical techniques, and prognostic factors of lawn trimmer-related open-globe injuries in eastern Taiwan. METHODS: This was a retrospective, consecutive case series study. Slit-lamp biomicroscopy, dilated fundoscopy, and orbital computed tomography (CT) images were collected. RESULTS: Twenty-six eyes of 26 patients were enrolled in the study. Fifteen patients (57.7%) had an intraocular foreign body (IOFB). The IOFB was metallic in 13 cases and glass and stone in the other 2 cases. Seven IOFBs (46.7%) were retained in the anterior chamber, 7 (46.7%) in the posterior segment, and 1 (6.7%) in the intraconal space. Univariate analysis showed that the presence of IOFB trended toward the development of endophthalmitis; however, this was not statistically significant (hazard ratio, 2.25; 95% confidence interval 0.35-14.61; P = 0.658). Eleven patients had metallic IOFBs noted on CT scans with metal artifacts, whereas two patients had small metallic IOFBs without metal artifacts. One patient had a glass IOFB mimicking metal artifacts on the CT scan. In one case, CT failed to reveal the IOFB, and an intralenticular metallic foreign body was incidentally found intraoperatively. CONCLUSION: Our study provides a broad characterization of lawn trimmer-related open-globe injuries. The informative and diverse findings of IOFBs on CT scans will help clinicians detect and recognize IOFBs more precisely and perform the surgery without causing further damage.

Oxidative Transformation of Dihydroflavonols and Flavan-3-ols by Anthocyanidin Synthase from Vitis vinifera.

Twelve polyphenols from three distinct families (dihydroflavonols, flavan-3-ols, and flavanones) were studied as potential substrates of anthocyanidin synthase from Vitis vinifera (VvANS). Only flavan-3-ols of (2R,3S) configuration having either a catechol or gallol group on ring B are accepted as substrates. Only dihydroflavonols of (2R,3R) configuration are accepted as substrates, but a catechol or gallol group is not mandatory. Flavanones are not substrates of VvANS. HPLC and MS/MS analyses of the enzymatic products showed that the VvANS-catalyzed oxidative transformation of (+)-dihydroflavonols, such as dihydroquercetin, dihydrokaempferol and dihydromyricetin, leads only to the corresponding flavonols. Among the flavan-3-ols recognized as substrates, (+)-gallocatechin was only transformed into delphinidin by VvANS, whereas (+)-catechin was transformed into three products, including two major products that were an ascorbate-cyanidin adduct and a dimer of oxidized catechin, and a minor product that was cyanidin. Data from real-time MS monitoring of the enzymatic transformation of (+)-catechin suggest that its products are all derived from the initial C3-hydroxylation intermediate, i.e., a 3,3-gem-diol, and their most likely formation mechanism is discussed.

Forensic nanopore sequencing of STRs and SNPs using Verogen's ForenSeq DNA Signature Prep Kit and MinION.

The MinION nanopore sequencing device (Oxford Nanopore Technologies, Oxford, UK) is the smallest commercially available sequencer and can be used outside of conventional laboratories. The use of the MinION for forensic applications, however, is hindered by the high error rate of nanopore sequencing. One approach to solving this problem is to identify forensic genetic markers that can consistently be typed correctly based on nanopore sequencing. In this pilot study, we explored the use of nanopore sequencing for single nucleotide polymorphism (SNP) and short tandem repeat (STR) profiling using Verogen's (San Diego, CA, USA) ForenSeq DNA Signature Prep Kit. Thirty single-contributor samples and DNA standard material 2800 M were genotyped using the Illumina (San Diego, CA, USA) MiSeq FGx and MinION (with R9.4.1 flow cells) devices. With an optimized cutoff for allelic imbalance, all 94 identity-informative SNP loci could be genotyped reliably using the MinION device, with an overall accuracy of 99.958% (1 error among 2926 genotypes). STR typing was notably error prone, and its accuracy was locus dependent. We developed a custom-made bioinformatics workflow, and finally selected 13 autosomal STRs, 14 Y-STRs, and 4 X-STRs showing high consistency between nanopore and Illumina sequencing among the tested samples. These SNP and STR loci could be candidates for panel design for forensic analysis based on nanopore sequencing.

Actual mutational research of 19 autosomal STRs based on restricted mutation model and big data.

Short tandem repeat (STR) markers have been widely used in forensic paternity testing and individual identification, but the STR mutation might impact on the forensic result interpretation. Importantly, the STR mutation rate was underestimated due to ignoring the "hidden" mutation phenomenon in most similar studies. Considering this, we use Slooten and Ricciardi's restricted mutation model based on big data to obtain more accurate mutation rates for each marker. In this paper, the mutations of 20 autosomal STRs loci (D3S1358, D1S1656, D13S317, Penta E, D16S539, D18S51, D2S1338, CSF1PO, Penta D, TH01, vWA, D21S11, D6S1043, D7S820, D5S818, TPOX, D8S1179, D12S391, D19S433, and FGA; The restricted model does not include the correction factor of D6S1043, this paper calculates remaining 19 STR loci mutation rates) were investigated in 28,313 (Total: 78,739 individuals) confirmed parentage-testing cases in Chinese Han population. As a result, total 1665 mutations were found in all loci, including 1614 one-steps, 34 two-steps, 8 three-steps, and 9 nonintegral mutations. The loci-specific average mutation rates ranged from 0.00007700 (TPOX) to 0.00459050 (FGA) in trio's and 0.00000000 (TPOX) to 0.00344850 (FGA) in duo's. We analyzed the relationship between mutation rates of the apparent and actual, the trio's and duo's, the paternal and maternal, respectively. The results demonstrated that the actual mutation rates are more than the apparent mostly, and the values of µ1"/µ2"(apparent) are also greater than µ1/µ2 (actual) commonly (µ1", µ1; µ2", µ2 are the mutation rates of one-step and two-step). Therefore, the "hidden" mutations are identified. In addition, the mutations rates of trio's and duo's, the paternal and maternal, exhibit significant difference. Next, those mutation data are used to do a comparison with the studies of other Han populations in China, which present the temporal and regional disparities. Due to the large sample size, some rare mutation events, such as monozygotic (MZ) mutation and "fake four-step mutation", are also reported in this study. In conclusion, the estimation values of actual mutations are obtained based on big data, they can not only provide basic data for the Chinese forensic DNA and population genetics databases, but also have important significance for the development of forensic individual identification, paternity testing and genetics research.

Optimization and Characterization of Microwave-Assisted Hydro-Distillation Extraction of Essential Oils from Cinnamomum camphora Leaf and Recovery of Polyphenols from Extract Fluid.

In this study, the efficiency of microwave-assisted hydro-distillation (MAHD) to extract essential oil from Cinnamomum camphora leaf, and the recovery of polyphenols from extract fluid were investigated. The effects of microwave power, liquid-to-material ratio, and extraction time on the extraction efficiency were studied by a single factor test as well as the response surface methodology (RSM) based on the central composite design method. The optimal extraction conditions were a microwave power of 786.27 W, liquid-to-material ratio of 7.47:1 mL/g, and extraction time of 35.57 min. The yield of essential oil was 3.26 ± 0.05% (w/w), and the recovery of polyphenols was 4.97 ± 0.02 mg gallic acid equivalent/g dry weight under the optimal conditions. Furthermore, the comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS) was used to characterize the essential oils of fresh and fallen leaves, and 159 individual compounds were tentatively identified, accounting for more than 89.68 and 87.88% of the total contents, respectively. The main ingredients include sabinene, l-ß-pinene, ß-myrcene, α-terpineol, 3-heptanone, and ß-thujene, as well as δ-terpineol and 3-heptanone, which were first identified in C. camphora essential oil. In conclusion, the MAHD method could extract essential oil from C. camphora with high efficiency, and the polyphenols could be obtained from the extract fluid at the same time, improving the utilization of C. camphora leaf.

Antivirulence properties and related mechanisms of spice essential oils: A comprehensive review.

In recent decades, reduced antimicrobial effectiveness, increased bacterial infection, and newly emerged microbial resistance have become global public issues, leading to an urgent need to find effective strategies to counteract these problems. Strategies targeting bacterial virulence factors rather than bacterial survival have attracted increasing interest, since the modulation of virulence factors may prevent the development of drug resistance in bacteria. Spices are promising natural sources of antivirulence compounds owing to their wide availability, diverse antivirulence phytochemical constituents, and generally favorable safety profiles. Essential oils are the predominant and most important antivirulence components of spices. This review addresses the recent efforts of using spice essential oils to inhibit main bacterial virulence traits, including the quorum sensing system, biofilm formation, motility, and toxin production, with an intensive discussion of related mechanisms. We hope that this review can provide a better understanding of the antivirulence properties of spice essential oils, which have the potential to be used as antibiotic alternatives by targeting bacterial virulence.

Phytomelatonin receptor PMTR1-mediated signaling regulates stomatal closure in Arabidopsis thaliana.

Melatonin has been detected in plants in 1995; however, the function and signaling pathway of this putative phytohormone are largely undetermined due to a lack of knowledge about its receptor. Here, we discovered the first phytomelatonin receptor (CAND2/PMTR1) in Arabidopsis thaliana and found that melatonin governs the receptor-dependent stomatal closure. The application of melatonin induced stomatal closure through the heterotrimeric G protein α subunit-regulated H2 O2 and Ca2+ signals. The Arabidopsis mutant lines lacking AtCand2 that encodes a candidate G protein-coupled receptor were insensitive to melatonin-induced stomatal closure. Accordingly, the melatonin-induced H2 O2 production and Ca2+ influx were completely abolished in cand2. CAND2 is a membrane protein that interacts with GPA1 and the expression of AtCand2 was tightly regulated by melatonin in various organs and guard cells. CAND2 showed saturable and specific 125 I-melatonin binding, with apparent Kd (dissociation constant) of 0.73 ± 0.10 nmol/L (r2  = .99), demonstrating this protein is a phytomelatonin receptor (PMTR1). Our results suggest that the phytomelatonin regulation of stomatal closure is dependent on its receptor CAND2/PMTR1-mediated H2 O2 and Ca2+ signaling transduction cascade.

Epigenetic inactivation of BRCA1 through promoter hypermethylation in ovarian cancer progression.

AIM: The BRCA1 promoter is hypermethylated in ovarian cancer patients. We postulated that this hypermethylation might be involved in ovarian cancer progression. METHODS: To confirm our hypothesis, tissue and serum samples were collected from ovarian carcinoma patients and categorized according to tumor stage. Healthy or benign ovarian disease tissue samples and corresponding serum samples were used as controls. Breast and ovarian cancer susceptibility gene 1 (BRCA1) promoter methylation levels were detected by real-time polymerase chain reaction (PCR). Real-time PCR was also used to evaluate BRCA1 gene expression, and Western blot was performed to assay the expression of BRCA1 protein. RESULTS: BRCA1 showed hypomethylation in 30 normal ovarian and 30 benign ovarian tumors, but showed hypermethylation or methylation in ovarian cancer patients. There was also a significant difference in the BRCA1 promoter methylation levels between different ovarian cancer stages. Compared to stage I and the control groups, there were higher BRCA1 promoter methylation frequencies in stage II and III ovarian cancers. BRCA1 methylation correlated with the loss of BRCA1 expression. BRCA1 promoter in stage I tumors showed hypomethylated. CONCLUSION: Promoter hypermethylation may act as a biomarker for sporadic ovarian cancer progression, but is unlikely to be helpful in the early diagnosis of ovarian cancer.

Heterocyclic Functionalized Donor-Acceptor Hole-Transporting Materials for Inverted Perovskite Solar Cells.

Hole transport materials (HTMs) with appropriate energy levels and comprehensive passivation effects help to obtain highly efficient and stable perovskite solar cells (PSCs). Electron-deficient character-induced HTMs can generate varying energy level alignments near the HTM/perovskite interface. Herein, we report the synthesis and investigation of two new dipolar HTMs, WWC103 and WWC105, based on 2-(1,1-dicyanomethylene)rhodamine and 4-cynophenylacetonitrile acceptors, enabling high-efficiency mixed-cation mixed-halide perovskite solar cells. Apart from having different acceptors, these HTMs are built on a heterocyclic frame, which can provide passivation effects and improve the morphology of the perovskite layer. As a result, these dopant-free HTM-based solar cells show a high open-circuit voltage and good power conversion efficiency. Among both, the solar cell based on the HTM with 2-(1,1-dicyanomethylene)rhodamine exhibits a high open-circuit voltage of 1.09 V with a champion power conversion efficiency of over 20.51%. The improved performance of WWC103 over WWC105 (19.74%) is attributed to the new acceptor, which, in addition to providing good energy-level alignments and hole mobility, also holds the ability to passivate the defects. The findings suggest a new acceptor unit for constructing dopant-free HTMs for efficient PSCs.

Toxic epidermal necrolysis induced by ritodrine in pregnancy: A case report.

BACKGROUND: Preterm birth accounts for about 12% of all pregnancies worldwide and is the leading cause of neonatal morbidity and mortality. In order to avoid premature birth and prolong gestational age, tocolytics are the first and the best choice. Ritodrine is the most commonly used tocolytic medication. However, side effects such as pulmonary edema, hypokalemia, and hyperglycemia are known. Here we report a rare but serious side effect-toxic epidermal necrolysis (TEN)-caused by ritodrine. CASE SUMMARY: A woman (31 years, gravida 4, para 2) was hospitalized because of premature contractions at 27 + 6 wk of gestation. A skin rash with pruritus appeared at 32 + 3 wk of gestation after administration of ritodrine, indomethacin, and dexamethasone, and it spread throughout the whole body in 3 d, particularly the four limbs. After 11 d' treatment, she was diagnosed with TEN. An emergency cesarean section was performed immediately to deliver the baby and intensive symptomatic treatment was promptly commenced after delivery. She recovered from the severe condition without any sequelae except for slight pigmentation after symptomatic treatment. CONCLUSION: When a skin rash appears during the administration of ritodrine, we are supposed to consider the risk of TEN.

Ganoderic acid D prevents oxidative stress-induced senescence by targeting 14-3-3ε to activate CaM/CaMKII/NRF2 signaling pathway in mesenchymal stem cells.

Stem cell senescence is an important cause of aging. Delaying senescence may present a novel way to combat aging and age-associated diseases. This study provided a mechanistic insight into the protective effect of ganoderic acid D (GA-D) against human amniotic mesenchymal stem cell (hAMSCs) senescence. GA-D, a Ganoderma lucidum-derived triterpenoid, markedly prevented hAMSCs senescence via activating the Ca2+ calmodulin (CaM)/CaM-dependent protein kinase II (CaMKII)/nuclear erythroid 2-related factor 2 (Nrf2) axis, and 14-3-3ε was identified as a target of GA-D. 14-3-3ε-encoding gene (YWHAE) knockdown in hAMSCs reversed the activation of the CaM/CaMKII/Nrf2 signals to attenuate the GA-D anti-aging effect and increase senescence-associated ß-galactosidase (SA-ß-gal), p16 and p21 expression levels, including reactive oxygen species (ROS) production, thereby promoting cell cycle arrest and decreasing differentiation potential. YWHAE overexpression maintained or slightly enhanced the GA-D anti-aging effect. GA-D prevented d-galactose-caused aging in mice by significantly increasing the total antioxidant capacity, as well as superoxide dismutase and glutathione peroxidase activity, and reducing the formation of malondialdehyde, advanced glycation end products, and receptor of advanced glycation end products. Consistent with the protective mechanism of GA-D against hAMSCs senescence, GA-D delayed the senescence of bone-marrow mesenchymal stem cells in this aging model in vivo, reduced SA-ß-gal and ROS production, alleviated cell cycle arrest, and enhanced cell viability and differentiation via regulating 14-3-3ε and CaM/CaMKII/Nrf2 axis. Therefore, GA-D retards hAMSCs senescence by targeting 14-3-3ε to activate the CaM/CaMKII/Nrf2 signaling pathway. Furthermore, the in vivo GA-D anti-aging effect may involve the regulation of stem cell senescence via the same signal axis.

Associations of Thyroid Function Tests with Lipid Levels and Adverse Pregnancy Outcomes During the First Trimester.

Objective: The present study aims to evaluate the relationship of thyroid function during the first trimester of pregnancy with lipid levels and pregnancy outcomes. Methods: Women who delivered babies at the Shanghai General Hospital between March 2019 and December 2019 with a known pregnancy outcome and complete data were included in the present study (n = 1779). A retrospective cohort study of all subjects with available first-trimester thyroid function testing and lipid levels data was conducted, and the relationship of thyroid function with lipid levels and pregnancy outcomes was evaluated. The data were analyzed using the SPSS software for statistical correlation. Results: The proportion of caesarean sections was higher in women with hypothyroxinemia (HIA) and hypothyroidism than in women with euthyroidism. Hypothyroidism was shown to be related with polyhydramnios, preterm labor and hypertriglyceridemia. HIA was correlated with increased rates of gestational diabetes mellitus (GDM), preeclampsia, gestational hypertension and hypertriglyceridemia. Compared with the euthyroidism group, the hypothyroidism group had a higher apolipoprotein A1 (Apo A1) level and apolipoprotein B level; the subclinical hypothyroidism group had a higher total cholesterol (TC) level and low-density lipoprotein cholesterol level; the HIA group had higher triglyceride, high-density lipoprotein cholesterol levels and lower TC, Apo A1 levels. TC levels were positively correlated with the thyroid-stimulating hormone level and negatively correlated with free thyroxine (FT4) level, and free triiodothyronine and FT4 levels were positively correlated with GDM occurrence. Conclusion: Thyroid function in early pregnancy is associated with dyslipidemia and pregnancy outcomes; conventional screening of thyroid diseases in early pregnancy may help improve lipid levels and decrease adverse pregnancy outcomes.

Pomegranate peel-derived punicalagin: Ultrasonic-assisted extraction, purification, and its α-glucosidase inhibitory mechanism.

The pomegranate peel is a by-product of pomegranate fruit rich in polyphenols. In this study, pomegranate peel polyphenols were explored using LC-MS/MS, and punicalagin was the most abundant compound. The highest yield (505.89 ± 1.73 mg/g DW) of punicalagin was obtained by ultrasonic-assisted extraction (UAE) with the ethanol concentration of 53%, sample-to-liquid ratio of 1:25 w/v, ultrasonic power of 757 W, and extraction time of 25 min. Punicalagin was further purified by the macroporous resin D101 and prep-HPLC, reaching the purity of 92.15%. The purified punicalagin had the IC50 of 82 ± 0.02 µg/mL against α-glucosidase, similar to the punicalagin standard with IC50 of 58 ± 0.014 µg/mL, both exhibiting a mixed inhibitory mechanism. Molecular docking further revealed that a steric hindrance with the intermolecular energy of -7.99 kcal/mol was formed between punicalagin and α-glucosidase. Overall, pomegranate peel is a promising source of punicalagin to develop anti-diabetic functional foods.

Protective effects of low-molecular-weight components of adipose stem cell-derived conditioned medium on dry eye syndrome in mice.

The present study demonstrated the protective effects of low-molecular-weight adipose-derived stem cell-conditioned medium (LADSC-CM) in a mouse model of dry eye syndrome. Mice subjected to desiccating stress and benzalkonium chloride had decreased tear secretion, impaired corneal epithelial tight junction with microvilli, and decreased conjunctival goblet cells. Topical application of adipose-derived stem cell-conditioned medium (ADSC-CM) stimulated lacrimal tear secretion, preserved tight junction and microvilli of the corneal epithelium, and increased the density of goblet cells and MUC16 expression in the conjunctiva. The low-molecular-weight fractions (< 10 kDa and < 3 kDa) of ADSC-CM (LADSC-CM) provided better protections than the > 10 kDa or > 3 kDa fractions of ADSC-CM. In the in vitro study, desiccation for 10 min or hyperosmolarity (490 osmols) for 24 h caused decreased viability of human corneal epithelial cells, which were reversed by LADSC-CM. The active ingredients in the LADSC-CM were lipophobic and stable after heating and lyophilization. Our study demonstrated that LADSC-CM had beneficial effects on experimental dry eye. It is worthy of further exploration for the active ingredient(s) and the mechanism.

The role of microRNAs in ovarian cancer initiation and progression.

Epithelial ovarian cancer (EOC) has the highest mortality rate of all gynaecological cancers. One of the greatest impediments to improving outcome is an incomplete understanding of the molecular underpinnings of EOC pathogenesis and progression. Recent studies suggest that microRNAs (miRNAs) are involved in ovarian tumorigenesis and cancer development. Several miRNA profiling studies have identified some consensus aberrantly expressed miRNAs in EOC tissues, and these EOC-related miRNAs may play critical roles in the pathogenesis and progression of EOC. Moreover, some of the miRNAs may have diagnostic or prognostic significance. In this review, recent progress to reveal the role of miRNAs in EOC will be addressed, and a model for miRNA functions in ovarian cancer initiation and progression will be proposed.

Aberrant methylation of breast and ovarian cancer susceptibility gene 1 in chemosensitive human ovarian cancer cells does not involve the phosphatidylinositol 3'-kinase-Akt pathway.

Methylation is an important silencing mechanism of breast and ovarian cancer susceptibility gene 1 (BRCA1) expression in sporadic ovarian cancer. However, the role of BRCA1 methylation in chemotherapy in sporadic ovarian cancer and the related pathways have not been understood completely. This study has determined the roles of BRCA1 hypermethylation in chemotherapy of sporadic ovarian cancer and its related signaling pathways. We used bisulfite sequencing, real-time polymerase chain reaction, and western blotting to check the methylation state and expression levels of BRCA1 of the following cell lines: platinum-sensitive human ovarian cancer cell line COC1, platinum-resistant cell line COC1/DDP, SKOV-3, and 5-Aza-dC treated COC1. The cisplatin sensitivity of ovarian cancer cells was examined by MTS (methyl-thiazol tetrazolium) assay. Tumorigenicity in vivo and DDP-based chemosensitivity were compared among the above cells. Phosphatidylinositol 3'-kinase (PI3K)-Akt pathway activation in ovarian cancer cells was studied by western blotting. The frequency of BRCA1 methylation in the COC1 cell line was higher than in COC1/DDP and SKOV-3 cell lines, whereas the mRNA and protein expression of BRCA1 were lower than in the COC1/DDP and SKOV-3 cell lines. DNA demethylation decreased the chemosensitivity of COC1 cells and partially increased the expression levels of BRCA1. The activation of the PI3K-Akt pathway was low in ovarian cancer cells. Our results indicate that hypermethylation of BRCA1 might play an important role in the chemosensitivity of ovarian cancer, and that the PI3K-Akt pathway is not involved in this response.

Screening and Spontaneous Mutation of Pickle-Derived Lactobacillus plantarum with Overproduction of Riboflavin, Related Mechanism, and Food Application.

Riboflavin, also known as vitamin B2, plays an important role in human cell metabolism and participates in various redox reactions and in energy utilization. In this study, 90 riboflavin-producing lactic acid bacteria (LAB) were screened out from pickle juices. The yields of riboflavin in these LAB were about 0.096-0.700 mg/L, and one strain, Lactobacillus plantarum RYG-YYG-9049, was found to produce the highest riboflavin content. Next, roseoflavin was used to induce the spontaneous mutation of RYG-YYG-9049, and selected roseoflavin-resistant colonies generally produced higher riboflavin contents, ranging from 1.013 to 2.332 mg/L. The No. 10 mutant, L. plantarum RYG-YYG-9049-M10, had the highest riboflavin content. Next, the molecular mechanism of enhancing riboflavin production in RYG-YYG-9049-M10 was explored, leading to the finding that roseoflavin treatment did not change the rib operons including the ribA, ribB, ribC, ribH, and ribG genes. Unexpectedly, however, this mechanism did induce an insertion of a 1059-bp DNA fragment in the upstream regulatory region of the rib operon, as compared to the wild-type RYG-YYG-9049. To the best of our knowledge, this is the first report that roseoflavin could induce an insertion of DNA fragment in LAB to increase riboflavin content, representing a new mutation type that is induced by roseoflavin. Finally, in order to fortify riboflavin content in soymilk, RYG-YYG-9049 and RYG-YYG-9049-M10 were used to ferment soymilk, and several fermentation parameters were optimized to obtain the fermented soymilk with riboflavin contents of up to 2.920 mg/L. In general, roseoflavin induction is an economical and feasible biotechnological strategy to induce riboflavin-overproducing LAB, and this strategy can be used to develop LAB-fermented functional foods that are rich in riboflavin.