Abstract: Quinoa has been recognized as the sole “comprehensive nutritional crop”; however, it is susceptible to pre-harvest sprouting (PHS). While quantitative reverse transcription polymerase chain reaction (RT-qPCR) has been extensively employed for gene expression level detection, the selection of suitable reference genes is imperative to ensure precise gene expression quantification across diverse conditions. This study aims to identify stable reference genes in quinoa seeds under ABA and GA, in order to provide a basis for subsequent research on PHS. Seeds were subjected to different concentrations of ABA and GA (10 μM, 50 μM, 100 μM, and 200 μM). The most suitable treatment concentration was determined based on seed viability. Here, MON1, GAPDH, EIF3, EF1α, ACT, TUB1, and TUB6 were selected as candidate genes. The suitability of these reference genes under different conditions was assessed using various methods including Ct values, geNorm, NormFinder, BestKeeper, Delta Ct, and RefFinder. Based on the results obtained from the hormone experiments, it was observed that the application of 100 μM ABA and 200 μM GA yielded the most advantageous outcomes. Additionally, the most appropriate reference genes for different treatments are ACT and TUB1 (H2O treatment), EIF3 and MON1 (ABA, GA treatment and also for the combined data set of the three groups). However, GAPDH exhibited the least stability across all treatments. In summary, ACT is recommended as the reference gene for natural quinoa germination, while EIF3 and MON1 should be used for ABA and GA treatments.

Resumo: A quinoa foi reconhecida como a única “cultura nutricional abrangente”; no entanto, é suscetível à germinação na pré-colheita (BHS). Embora a reação em cadeia da polimerase com transcrição reversa quantitativa (RT-qPCR) tenha sido amplamente empregada para detecção do nível de expressão gênica, a seleção de genes de referência adequados é essencial para garantir a quantificação precisa da expressão gênica em diversas condições. Este estudo tem como objetivo identificar genes de referência estáveis em sementes de quinoa tratadas com ABA e GA, a fim de fornecer uma base para pesquisas subsequentes em BPC. As sementes foram submetidas a diferentes concentrações de ABA e GA (10 μM, 50 μM, 100 μM e 200 μM). A concentração de tratamento mais adequada foi determinada com base na viabilidade das sementes. MON1, GAPDH, EIF3, EF1α, ACT, TUB1 e TUB6 foram selecionados como genes candidatos. A adequação destes genes de referência sob diferentes condições foi avaliada utilizando vários métodos, incluindo valores Ct, geNorm, NormFinder, BestKeeper, Delta Ct e RefFinder. Com base nos resultados obtidos nos experimentos com hormônios, observou-se que a aplicação de 100 μM de ABA e 200 μM de GA produziu os resultados mais vantajosos. Além disso, os genes de referência mais apropriados para diferentes tratamentos são ACT e TUB1 (tratamento com H2O), EIF3 e MON1 (tratamento com ABA, GA e para o conjunto de dados combinados dos três grupos). No entanto, o GAPDH exibiu a menor estabilidade em todos os tratamentos. Em resumo, o ACT é recomendado como gene de referência para a germinação natural da quinoa, enquanto o EIF3 e o MON1 devem ser utilizados para os tratamentos com ABA e GA.

OBJECTIVES: Our purpose was to examine the associations of female genital infections and certain comorbidities with infertility. METHODS: The Taiwan National Health Research Database was searched for women with a new diagnosis of infertility between 2000 and 2013. Women without a diagnosis of infertility served as a control group and were matched with the infertility cases by age (±3 years) and index year. They were divided into two groups: ≤40 years old and >40 years old. Univariate and multivariate conditional logistic regression models were employed to identify the risk factors associated with infertility. RESULTS: A total of 18,276 women with a new diagnosis of infertility and 73,104 matched controls (mean cohort age, 31±6.2 years) were included. According to the adjusted multivariate analysis, pelvic inflammatory disease involving the ovary, fallopian tube, pelvic cellular tissue, peritoneum (odds ratio (OR)=4.823), and uterus (OR=3.050) and cervical, vaginal, and vulvar inflammation (OR=7.788) were associated with an increased risk of infertility in women aged ≤40 years. In women aged >40 years, pelvic inflammatory disease of the ovary, fallopian tube, pelvic cellular tissue, and peritoneum (OR=6.028) and cervical, vaginal, and vulvar inflammation (OR=6.648) were associated with infertility. Obesity, lipid metabolism disorders, dysthyroidism, abortion (spontaneous or induced), bacterial vaginosis, endometritis, and tubo-ovarian abscess were associated with an increased risk of infertility according to the univariate analysis but not the multivariate analysis. CONCLUSIONS: Female genital tract infections, but not the comorbidities studied here, are associated with an increased risk of infertility.

Rhein is a primary anthraquinone found in the roots of a traditional Chinese herb, rhubarb, and has been shown to have some anticancer effects. The aim of the present study was to investigate the effect of rhein on the apoptosis of the human gastric cancer line SGC-7901 and to identify the mechanism involved. SGC-7901 cells were cultured and treated with rhein (0, 50, 100, 150, and 200 µM) for 24, 48, or 72 h. Relative cell viability assessed by the MTT assay after treatment was 100, 99, 85, 79, 63% for 24 h; 100, 98, 80, 51, 37% for 48 h, and 100, 97, 60, 36, 15% for 72 h, respectively. Cell apoptosis was detected with TUNEL staining and quantified with flow cytometry using annexin FITC-PI staining at 48 h after 100, 200 and 300 µm rhein. The percentage of apoptotic cells was 7.3, 21.9, 43.5%, respectively. We also measured the mRNA levels of caspase-3 and -9 using real-time PCR. Treatment with 100 µM rhein for 48 h significantly increased mRNA expression of caspase-3 and -9. The levels of apoptosis-related proteins including Bcl-2, Bax, Bcl-xL, and pro-caspase-3 were evaluated in rhein-treated cells. Rhein increased the Bax:Bcl-2 ratio but decreased the protein levels of Bcl-xL and pro-caspase-3. Moreover, rhein significantly increased the expression of cytochrome c and apoptotic protease activating factor 1, two critical components involved in mitochondrial pathway-mediated apoptosis. We conclude that rhein inhibits SGC-7901 proliferation by inducing apoptosis and this antitumor effect of rhein is mediated in part by an intrinsic mitochondrial pathway.