CRISPR/Cas9-mediated knockout of Bsr-d1 enhances the blast resistance of rice in Northeast China.

KEY MESSAGE: The blast resistance allele of OsBsr-d1 does not exist in most japonica rice varieties of Jilin Province in China. The development of Bsr-d1 knockout mutants via CRISPR/Cas9 enhances broad-spectrum resistance to rice blast in Northeast China. Rice blast is a global disease that has a significant negative impact on rice yield and quality. Due to the complexity and variability of the physiological races of rice blast, controlling rice blast is challenging in agricultural production. Bsr-d1, a negative transcription factor that confers broad-spectrum resistance to rice blast, was identified in the indica rice cultivar Digu; however, its biological function in japonica rice varieties is still unclear. In this study, we analyzed the blast resistance allele of Bsr-d1 in a total of 256 japonica rice varieties from Jilin Province in Northeast China and found that this allele was not present in these varieties. Therefore, we generated Bsr-d1 knockout mutants via the CRISPR/Cas9 system using the japonica rice variety Jigeng88 (JG88) as a recipient variety. Compared with those of the wild-type JG88, the homozygous Bsr-d1 mutant lines KO#1 and KO#2 showed enhanced leaf blast resistance at the seedling stage to several Magnaporthe oryzae (M. oryzae) races collected from Jilin Province in Northeast China. Physiological and biochemical indices revealed that the homozygous mutant lines produced more hydrogen peroxide than did JG88 plants when infected with M. oryzae. Comparative RNA-seq revealed that the DEGs were mainly involved in the synthesis of amide compounds, zinc finger proteins, transmembrane transporters, etc. In summary, our results indicate that the development of Bsr-d1 knockout mutants through CRISPR/Cas9 can enhance the broad-spectrum resistance of rice in Northeast China to rice blast. This study not only provides a theoretical basis for disease resistance breeding involving the Bsr-d1 gene in Northeast China, but also provides new germplasm resources for disease-resistance rice breeding.

Prevalence and risk factors of postpartum depressive symptoms at 42 days among 2462 women in China.

BACKGROUND: Postpartum depression is a common and serious mental health problem that is affecting an increasing percentage of the world's population. We aimed to evaluate the prevalence of postpartum depressive symptoms in Beijing, China, during the COVID-19 pandemic and identify several potential risk factors. METHODS: This was a cross-sectional observational study conducted at Peking University First Hospital from 2020 to 2021. Women who delivered and had postpartum reviews at 42 days after delivery were invited to complete the Chinese version of the Edinburgh Postnatal Depression Scale (EPDS) under the guidance of trained nurses. A score of ≥10 was used as the threshold of postpartum depression. t-tests, chi-square tests or Mann-Whitney U tests were applied. A multivariate logistic regression analysis was conducted to assess the risk factors for postpartum depressive symptoms. RESULTS: A total of 2462 mothers were included in this study, 20.2 % of whom were considered to have postpartum depressive symptoms. Multivariate logistic regression analysis showed that formula feeding (OR = 2.219, 95 % CI: 1.300-3.786, P = 0.013), preterm birth (OR = 1.619, 95 % CI: 1.108-2.367, P = 0.013), cervical insufficiency (OR 3.022, 95 % CI: 1.200-7.615, P = 0.019) and history of depression (OR = 6.519, 95 % CI: 1.537-27.659, P = 0.011) were associated with a high prevalence of postpartum depressive symptoms. CONCLUSION: There is a high prevalence of postpartum depressive symptoms in developed regions of China during the COVID-19 pandemic. More attention should be given to mothers with risk factors for PPD, and follow-up care is needed.

Establishment of linkage phase, using Oxford Nanopore Technologies, for preimplantation genetic testing of Coffin-Lowry syndrome with a de novo RPS6KA3 mutation.

Background: This study aimed to perform preimplantation genetic testing (PGT) for a female Coffin-Lowry Syndrome (CLS) patient with a de novo mutation (DNM) in RPS6KA3. It was challenging to establish the haplotype in this family because of the lack of information from affected family members. Hence, we explored a new and reliable strategy for the detection of the DNM in PGT, using Oxford Nanopore Technologies (ONT) and the MARSALA platform. Methods: We performed whole-exome sequencing (WES) on the proband and confirmed the pathogenic mutation by Sanger sequencing. The proband then underwent PGT to prevent the transmission of the pathogenic mutation to her offspring. We diverged from the conventional methods and used long-read sequencing (LRS) on the ONT platform to directly detect the mutation and nearby SNPs, for construction of the haplotype in the preclinical phase of PGT. In the clinical phase of embryo diagnosis, the MARSALA method was used to detect both the SNP-based haplotype and chromosome copy number variations (CNVs), in each blastocyst. Finally, a normal embryo was selected by comparison to the haplotype of the proband and transferred into the uterus. Sanger sequencing and karyotyping were performed by amniocentesis, at 17 weeks of gestation, to confirm the accuracy of PGT. Results: Using WES, we found the novel, heterozygous, pathogenic c.1496delG (p.Gly499Valfs*25) mutation of RPS6KA3 in the proband. The SNP-based haplotype that was linked to the pathogenic mutation site was successfully established in the proband, without the need for other family members to be tested with ONT. Eight blastocysts were biopsied to perform PGT and were assessed with a haplotype linkage analysis (30 SNP sites selected), to give results that were consistent with direct mutation detection using Sanger sequencing. The results of PGT showed that three of the eight blastocysts were normal, without the DNM. Moreover, the patient had a successful pregnancy, after transfer of a normal blastocyst into the uterus, and delivered a healthy baby. Conclusion: The ONT platform, combined with the MARSALA method, can be used to perform PGT for DNM patients without the need for other samples as a reference.

Case report: Preimplantation genetic testing for X-linked alport syndrome caused by variation in the COL4A5 gene.

X-Linked Alport Syndrome (XLAS) is an X-linked, dominant, hereditary nephropathy mainly caused by mutations in the COL4A5 gene, found on chromosome Xq22. In this study, we reported a pedigree with XLAS caused by a COL4A5 mutation. This family gave birth to a boy with XLAS who developed hematuria and proteinuria at the age of 1 year. We used next-generation sequencing (NGS) to identify mutations in the proband and his parents and confirmed the results using Sanger sequencing. This testing showed there was a single nucleotide missense variation, c.3659G>A (p.Gly1220Asp) (NM_033380.3), in the COL4A5 gene. To prevent the inheritance of the syndrome, we used eight embryos for trophoblast biopsy after assisted reproductive technology treatment, and whole genome amplification (WGA) was performed using multiple annealing and looping-based amplification cycles (MALBAC). Embryos were subjected to Preimplantation Genetic Testing (PGT) procedures, including Sanger sequencing, NGS-based single nucleotide polymorphism (SNP) haplotype linkage analysis, and chromosomal copy number variation (CNV) analysis. The results showed that three embryos (E1, E2, and E4) were free of CNV and genetic variation in the COL4A5 gene. Embryo E1 (4AA) was transferred after consideration of the embryo growth rate, morphology, and PGT results. Prenatal diagnosis in the second trimester showed that the fetus had a normal karyotype and did not carry the COL4A5 mutation (c.3659G>A). Ultimately, a healthy boy was born and did not carry the pathogenic COL4A5 mutation, which indicated that PGT prevented the intergenerational transmission of the causative mutation of XLAS.

Effects of reduced follicle-stimulating hormone dosage before human chorionic gonadotropin trigger on in vitro fertilization outcomes.

OBJECTIVE: To determine whether a reduced dose of follicle-stimulating hormone (FSH) before human chorionic gonadotropin (hCG) trigger during ovarian stimulation can affect in vitro fertilization (IVF) outcomes. METHODS: This study included 347 patients with a normal ovarian response who received a reduced dose of FSH before hCG trigger for 2-3 days (Group A) and 671 patients who did not receive a reduced dose (Group B) from a university-affiliated IVF center between January 2021 and December 2022. The primary endpoint was estrogen (E2) and progesterone (P) levels on the day of hCG trigger, fresh embryo transfer cycles, laboratory outcomes, and clinical outcomes between the two groups. RESULTS: On the day of hCG trigger, Group A had significantly lower E2 and P levels than those in Group B (3454.95 ± 1708.14 pg/mL versus 3798.70 ± 1774.26 pg/mL, p = 0.003; and 1.23 ± 0.53 ng/mL versus 1.37 ± 0.59 ng/mL, p < 0.001, respectively). The proportion of patients with P levels ≥ 1.5 ng/mL was 22.48% in Group A compared to 34.58% in Group B (p < 0.001), while the proportion of patients with E2 ≥ 5000 pg/mL was 15.27% in Group A compared to 25.93% in Group B (p < 0.001). The fresh embryo-transfer cycle rate in Group A was higher than that in group B (54.47% and 32.64%, respectively; p < 0.001). Despite the reduction in FSH dosage, there were no significant differences between groups regarding the number of oocytes retrieved, total number of mature oocytes, normal fertilization rate, cleavage rate, Day 3 top-quality rate, implantation rate, pregnancy rate per cycle, and early pregnancy loss rate. CONCLUSION: While a reduced dose of FSH prior to hCG trigger during ovarian stimulation did not significantly affect IVF outcomes, it was associated with lower E2 and P levels, resulting in fewer cycles with E2 ≥ 5000 pg/mL and P ≥ 1.5 ng/mL on the day of the hCG trigger.

One-pot synthesis of Ca-based magnetic hydrochar derived from consecutive hydrothermal and pyrolysis processing of bamboo for high-performance scavenging of Pb(â ¡) and tetracycline from water.

Ca-based magnetic bamboo-derived hydrochar described as Ca-MBHC was synthesized by one-pot pyrolysis, and was applied to remediation of lead (Pb) and tetracycline (TC) polluted water. Characterizations not only attested the loading of CaCO3 and Fe0 onto the hydrochar, but also demonstrated the magnetism of Ca-MBHC. Adsorption kinetic experiments showed that the Ca-MBHC could eliminate Pb(II) and TC during a wide range of pH, and appeared rapid uptake equilibrium within 240 and 60 min for Pb(II) and TC, severally. Adsorption isotherm experiments showed that the Ca-MBHC possessed highest adsorption of 475.58 mg/g concerning Pb(II), and heterogeneous uptake of 142.44 mg/g for TC. Furthermore, the Ca-MBHC could achieve Pb(II) binding owing to complexation, reduction, ion exchange and electrostatic attraction, whereas the TC uptake might be related to π-π stacking reciprocities, pore filling and hydrogen bonding. Overall, the Ca-MBHC could be viewed as an excellent adsorbent for scavenging Pb(II) and tetracycline from water.

[Effect of community rehabilitation skill on patients with depression].

OBJECTIVE: To explore the effect of community depression rehabilitation skill on depressive symptoms and social function of patients with different diagnoses. METHODS: A total of 74 patients(20 males, 54 females)with depression in Beijing primary care center were divided into rehabilitation group(49 cases, average 51.80 years) and control group(25 cases, average 54.42 years) from August to November 2019.A total of 70 patients in two groups completed the final follow-up study. Patients in the rehabilitation group received depression community rehabilitation skill training for 8 weeks, while patients in the control group received daily community services. The two groups were assessed by the scale at baseline and 8 weeks later. The scales are as follow: basic information form and Hamilton depression scale(HAMD-17) and social disability screening schedule(SDSS), nurses global assessment of suicide risk(NGASR), morisky medication aderence scale(MMAS-8). For counting data χ~2inspection; The t-test for measurement data. RESULTS: Before the intervention, There was no significant difference in HAMD-17, NGASR, MMAS-8 and SDSS total score and each factor score of SDSS between the two groups(P>0.05). Paired t-test showed that after intervention, HAMD-17 total score, NGASR total score, and SDSS factors of social withdrawal, social activities outside the family, activities within the family, family functions, self-care of personal life, relationship and interest to the outside world, sense of responsibility and planning(P<0.05) in rehabilitation group were all significantly lower than before intervention. CONCLUSION: The rehabilitation skills of depression can improve the depressive symptoms and social function of different kinds of patients, and is suitable for promotion in the community.

Two mutations in TUBB8 cause developmental arrest in human oocytes and early embryos.

RESEARCH QUESTION: How can the effect of genetic mutations that may cause primary female infertility be evaluated? DESIGN: Patients and their family members underwent whole-exome sequencing and Sanger sequencing to detect the infertility-causing gene and inheritance pattern. To study the function of mutant proteins in vitro, vectors containing wild-type or mutant TUBB8 cDNA were constructed for transient expression in HeLa cells, and in-vitro transcribed mRNA were used for microinjection in germinal vesicle-stage mouse oocytes. Immunofluorescence staining was used to observe the microtubule structure in HeLa cells or meiotic spindle in mouse oocytes. RESULTS: A maternally inherited TUBB8 (Tubulin beta 8 class VIII) mutation (NM_177987.2: c. 959G>A: p. R320H) and a previously reported (NM_177987.2: c. 161C>T: p. A54V) recessive mutation from two infertile female patients were identified. The oocytes from the patient carrying p.A54V mutation failed fertilization, whereas oocytes with p.R320H mutation could be fertilized but showed heavy fragmentation during early development. In vitro, functional assays showed that p. A54V mutant disrupted the microtubule structure in HeLa cells (49.3% of transfected cells) and caused large polar body extrusion in mouse oocytes (27.5%), whereas the p.R320H mutant caused a higher abnormal rate (69.7%) in cultured cells and arrested mouse oocytes at meiosis I (38.7%). CONCLUSION: Two TUBB8 mutations (p.A54V and p.R320H) were identified and their pathogeny was confirmed by in-vitro functional assays.

Erratum: miR-424 suppresses proliferation and promotes apoptosis of human ovarian granulosa cells by targeting Apelin and APJ expression.

[This corrects the article on p. 3660 in vol. 12, PMID: 32774725.].

Microwave-assisted synthesis of ß-cyclodextrin functionalized celluloses for enhanced removal of Pb(II) from water: Adsorptive performance and mechanism exploration.

Herein, ß-cyclodextrin (ß-CD) was efficiently grafted onto rice husk-based celluloses using different cross-linking agents of epichlorohydrin (EPI) and glutaraldehyde (GA). By feat of microwave irradiation, the functionalization procedure was completed in 17 min, and the synthesized RHEPIMWß-CD and RHGAMWß-CD exhibited fast adsorption equilibrium for Pb(II) within 20 min, excellent monolayer adsorption capacities of 216.06 and 279.08 mg g-1 across an extensive pH scope of 3.0-6.0, unaffected affinity to Pb(II) during the existence of co-existing ions, superior reusability with over 81% and 87% of Pb(II) uptake sustained for four adsorption-desorption cycles. Thermodynamic parameters implied that the uptake process of Pb(II) occurred spontaneously (-ΔG0) with an endothermic characteristic (+ΔH0). Furthermore, electrostatic attraction and complexation were demonstrated to enhance the Pb(II) uptake onto the RHEPIMWß-CD and RHGAMWß-CD. In fix-bed columns, these two adsorbents also efficiently eliminated Pb(II) under various flow rates with experimental breakthrough curves well simulated by Thomas and Yoon-Nelson models. Significantly, the RHEPIMWß-CD and RHGAMWß-CD could effectively purify acid battery effluent containing Pb(II) for meeting regulatory requirement. Overall, the fast fabrication, excellent adsorption and recycling performance facilitate the development of tailored adsorbents for Pb(II) elimination in wastewater.

Pooled analysis of the oncological outcomes in robotic gastrectomy versus laparoscopic gastrectomy for gastric cancer.

AIM: Robotic gastrectomy (RG) is more and more widely used in the treatment of gastric cancer. However, the long-term oncological outcomes of RG have not been well evaluated. The aim of this study was to evaluate the long-term oncological outcomes of RG and laparoscopic gastrectomy (LG) in the treatment of gastric cancer. MATERIALS AND METHODS: PubMed, China National Knowledge Infrastructure, Cochrane Library and EMBASE electronic databases were searched until August 2019. Eligible studies were analysed for comparison of oncological outcomes between RG and LG in patients with gastric cancer. RESULTS: Eleven retrospective comparative studies, which included 1347 (32.52%) patients in the RG group and 2795 (67.48%) patients in the LG group, were selected for the analysis. Meta-analysis of the 11 included studies showed that there was no statistically significant difference in the OS between the RG and LG groups (hazard ratios [HRs] = 0.97, 95% confidence intervals [CIs] = 0.80-1.19, P = 0.80). Six studies evaluated disease-free survival (DFS), and pooled analysis showed that there was no statistically significant difference in DFS between RG group and LG group (HR = 0.94, 95% CIs = 0.72-1.23, P = 0.65). According to the odds ratio (OR) analysis, there was no significant difference in 3-year OS, 5-year OS, 3-year DFS and 5-year DFS between the RG and LG groups. Nine articles reported the recurrence rate, and the meta-analysis showed that there was no statistically significant difference between the RG and LG groups (OR = 0.88, 95% CIs = 0.69-1.12, P = 0.31). CONCLUSIONS: This meta-analysis indicated that the long-term oncological outcomes in the RG group were similar to that in the LG group.

miR-424 suppresses proliferation and promotes apoptosis of human ovarian granulosa cells by targeting Apelin and APJ expression.

OBJECTIVE: Polycystic ovary syndrome (PCOS) is associated with alteration of Apelin signaling in ovarian granulosa cells (GCs). However, the molecular mechanisms regulating Apelin expression remain poorly understood. This study aims to investigate the role of miR-424 in modulating Apelin expression and GC functions. METHODS: miRNA expression in GCs was altered by transfection with specific miR-424 mimics and inhibitors. Apelin level was determined by ELISA. miR-424 and mRNA expression were analyzed by quantitative RT-PCR. Protein abundance was measured by western blotting. Genomic sequence targeted by miR-424 was validated by dual-luciferase reporter assay. Apelin gene was overexpressed by transfection of LV-003 vector carrying its cDNA. GC proliferation was analyzed by MTS method, and its cell cycle progression and apoptosis were measured by flow cytometry. RESULTS: Apelin concentration was increased in serum and follicular fluid from PCOS patients, accompanied by upregulated APJ (Apelin receptor) expression and suppressed miR-424 expression in GCs. miR-424 mimics suppressed Apelin and APJ expression in KGN cells by targeting 3' UTR of Apelin and APJ, whereas miR-424 inhibitors had the opposite effects. miR-424 inhibited KGN cell proliferation and cell cycle progression by down-regulating Cyclin-D/E expression. Moreover, miR-424 promoted KGN cell apoptosis by increasing truncated Caspase-3 level. The regulation of KGN cell proliferation and apoptosis by miR-424 was mediated by directly suppressing Apelin gene expression, instead of inhibiting Apelin peptide activity. CONCLUSION: miR-424 suppresses proliferation and promotes apoptosis of human ovarian granulosa cells by directly targeting and inhibiting Apelin and APJ expression.

Long-term outcomes of laparoscopic sleeve gastrectomy versus Roux-en-Y gastric bypass for morbid obesity: Results from a meta-analysis of randomized controlled trials.

BACKGROUND: Laparoscopic sleeve gastrectomy and laparoscopic Roux-en-Y gastric bypass (LRYGB) are 2 widely used procedures performed in bariatric surgery. However, their long-term weight loss effects have not been well compared. OBJECTIVES: To evaluate the long-term outcome of 2 procedures for treating morbid obesity. SETTING: The gastrointestinal surgery center, Eastern Hospital, Sichuan Academy of Medical Sciences & Sichuan Provincial People's Hospital. METHODS: Three electronic databases, PubMed, CNKI, and EMBASE, were searched ending in September 2018. Eligible studies were prospective randomized controlled trials. RESULTS: Of 15 randomized controlled trials studies, 1381 cases were included, 697 were randomly divided into the laparoscopic sleeve gastrectomy group and 684 to the LRYGB group. A statistically significant percent excess weight loss reduction in weighted mean difference was observed in LRYGB group at 5 years (weighted mean difference = -8.9, 95% confidence interval [CI] = -13.08 to -4.73, P < .0001), and at 3 years (weighted mean difference = -11.96, 95% CI = -17.62 to -6.30, P < .0001). However, there was not any statistically difference between the 2 procedures in <3 years. The total complication of the LSC group were less than that of the LRYGB groups (odds ratio = .52, 95%CI = .35-.76, P = .0007). This meta-analysis showed that the LRYGB procedure could reduce gastrointestinal reflux disease (odds ratio = .26, 95%CI = .11-.61, P = .002); however, no statistically significant differences were found in type 2 diabetes, hypertension, dyslipidemia, and sleep apnea. CONCLUSIONS: This meta-analysis showed that significantly greater in percent excess weight loss after 5 years of LRYGB treatment; however, it seems to have a higher incidence of total complications.

The Role of Stents as Bridge to Surgery for Acute Left-Sided Obstructive Colorectal Cancer: Meta-Analysis of Randomized Controlled Trials.

BACKGROUND: The role of self-expanding metallic stents (SEMS) as a bridge to surgery for acute left-sided obstructive colorectal cancer has remained controversial. OBJECTIVE: To study the efficacy of this approach, we performed a meta-analysis at the gastrointestinal surgery center of Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital (Eastern Hospital). METHODS: Two PubMed and science-direct electronic databases were searched up to December 30, 2017. Eligible studies were randomized controlled trials (RCTs). RESULTS: We selected 8 RCTs articles, which included 497 cases. The directly stoma rates were significantly lower in the stent group (odds ratio [OR] = 0.46, 95% confidence intervals [CIs] = 0.30-0.70, p = 0.0003). The successful primary anastomosis rates were significantly higher in the stent group (OR = 2.29, 95% CIs = 1.52-3.45, p < 0.0001). The post-procedural complication rates were significantly lower in the stent group (OR = 0.39, 95% CI = 0.18-0.82, p = 0.01). However, tumor recurrence rates were significantly higher in the stent group (OR = 1.79, 95% CI = 1.09-2.93, p = 0.02). Conclusions This meta-analysis confirms that SEMS placement could reduce direct stoma rate and increases the successful primary anastomosis rate; however, it was associated with a seemingly higher incidence of tumor recurrence.

Seminal plasma cell-free mitochondrial DNA copy number is associated with human semen quality.

OBJECTIVE: Previous studies have shown that mitochondrial DNA (mtDNA) copy number in human sperm is associated with semen quality. This study examines whether there is association between seminal cell-free mtDNA copy number and human semen parameters. STUDY DESIGN: Seminal cell-free mtDNA copy number was measured in 205 men. Semen reactive oxygen species (ROS) level was also measured. Sperm quality was assessed using World Health Organization criteria, while sperm motility parameters were determined by computer-aided sperm analysis. RESULTS: Comparing with normozoospermia, patients with asthenozoospermia and oligoasthenozoospermia had a significant decrease in cell-free mtDNA copy number and an increase in ROS level in seminal plasma (P < 0.05 for all). Seminal cell-free mtDNA copy number positively correlated with sperm concentration, motility, morphology and motion characteristics (r for 0.247 to 0.673, P < 0.05 for all). Semen ROS level negatively correlated with sperm concentration, motility, and motion characteristics (r for -0.261 to -0.676, P < 0.05 for all). There is a negative relationship between seminal cell-free mtDNA copy number and ROS level (r= -0.573, P < 0.05). CONCLUSION: The results suggested that seminal cell-free mtDNA copy number is associated with semen parameters, which may serve as a novel diagnostic marker of semen quality.

Impact of multidisciplinary collaborative pharmaceutical care on knowledge, adherence, and efficacy of hormone therapy in climacteric women.

BACKGROUND: The objective of this study was to evaluate the impact of pharmaceutical care on the knowledge, adherence, and efficacy of hormone therapy in climacteric women participated in multidisciplinary collaborative clinic, launched by Peking University First Hospital (Beijing, People's Republic of China) in 2012. METHODS: A total of 296 patients were recruited (intervention group n=150, control group n=146). The patients in the intervention group visited the multidisciplinary collaborative clinic for their initial hormone therapy, receiving individualized pharmaceutical care (PC), whereas the control group visited only the general clinic without PC. The pill count method, knowledge assessment questionnaire (Cronbach's α=0.80), and the modified Kupperman Index were used to assess the knowledge, adherence, and efficacy at months 3, 6, and 12. RESULTS: The intervention group, which received PC, showed significantly higher and better knowledge, adherence, and efficacy than the control group, demonstrating the effectiveness of the PC provided. The knowledge scores of the intervention and control groups at months 3, 6, and 12 were (73.12 vs 59.28), (77.63 vs 66.19), and (80.81 vs 66.64); the data for adherence were (90.97% vs 82.17%), (93.21% vs 87.79%), and (95.81% vs 93.38%); and the values of the modified Kupperman Index were (17.15 vs 24.05), (13.22 vs 22.01), and (12.21 vs 23.15), respectively. CONCLUSION: PC improved the knowledge, adherence, and efficacy of hormone therapy in climacteric women. Therefore, the multidisciplinary collaborative model investigated in our study should be advocated in other health care institutions for the benefit of more patients. Further large-sample and long-term studies should be conducted to evaluate the effects of PC on patient clinical outcomes, including its impact on the safety and efficacy of long-term use of hormone therapy, as well as the economic benefits.

Overexpression of a New Zinc Finger Protein Transcription Factor OsCTZFP8 Improves Cold Tolerance in Rice.

Cold stress is one of the most important abiotic stresses in rice. C2H2 zinc finger proteins play important roles in response to abiotic stresses in plants. In the present study, we isolated and functionally characterized a new C2H2 zinc finger protein transcription factor OsCTZFP8 in rice. OsCTZFP8 encodes a C2H2 zinc finger protein, which contains a typical zinc finger motif, as well as a potential nuclear localization signal (NLS) and a leucine-rich region (L-box). Expression of OsCTZFP8 was differentially induced by several abiotic stresses and was strongly induced by cold stress. Subcellular localization assay and yeast one-hybrid analysis revealed that OsCTZFP8 was a nuclear protein and has transactivation activity. To characterize the function of OsCTZFP8 in rice, the full-length cDNA of OsCTZFP8 was isolated and transgenic rice with overexpression of OsCTZFP8 driven by the maize ubiquitin promoter was generated using Agrobacterium-mediated transformation. Among 46 independent transgenic lines, 6 single-copy homozygous overexpressing lines were selected by Southern blot analysis and Basta resistance segregation assay in both T1 and T2 generations. Transgenic rice overexpressing OsCTZFP8 exhibited cold tolerant phenotypes with significantly higher pollen fertilities and seed setting rates than nontransgenic control plants. In addition, yield per plant of OsCTZFP8-expressing lines was significantly (p < 0.01) higher than that of nontransgenic control plants under cold treatments. These results demonstrate that OsCTZFP8 was a C2H2 zinc finger transcription factor that plays an important role in cold tolerance in rice.

Effects of dehydroepiandrosterone supplementation on mice with diminished ovarian reserve.

Dehydroepiandrosterone (DHEA) has been used to improve the pregnancy rate in women with diminished ovarian reserve(DOR) during in vitro fertilization. We aimed to validate the effects of DHEA and identify the possible mechanisms. We constructed a mice model with DOR and analyzed the hormone parameters and follicle counts. In vivo experiment, FSH and LH concentrations in the serum were significantly elevated in the DOR group. However, the FSH and LH concentrations were partially reversed in the DOR + DHEA group. The E2, AMH and INHB were down-regulated in the DOR group and reversed in the DOR + DHEA group. Our study supported evidences that DHEA might modulate the hormone receptors in the ovary and hormone secretions to the peripheral circulation to regulate the ovary reserve functions.

Reduced Klotho expression contributes to poor survival rates in human patients with ovarian cancer, and overexpression of Klotho inhibits the progression of ovarian cancer partly via the inhibition of systemic inflammation in nude mice.

Klotho is a recently discovered anti­aging gene, which has been reported as a tumor suppressor in numerous human malignancies; however, the role of Klotho in human ovarian cancer remains to be elucidated. The aim of the present study was to detect the expression of Klotho and evaluate its association with the progression of human ovarian cancer. A clinical follow­up study of 120 patients with ovarian cancer and 78 normal controls was conducted. The expression levels of Klotho were determined by western blotting and immunohistochemistry. The results demonstrated that high Klotho expression levels were detected in all normal controls, whereas the positive rate of Klotho was 61.6% in the ovarian cancer group, which was significantly decreased compared with in the control group (P<0.01). Furthermore, reduced Klotho expression was significantly correlated with decreased survival rates in patients with ovarian cancer (P=0.025). Subsequently, Klotho levels were detected in seven human ovarian cancer cell lines by western blotting. The results demonstrated that the highest levels of Klotho were detected in CaOV3 cells, medium levels of Klotho were detected in CaOV4 and SKOV­3 cells, and almost no Klotho was detected in the other four cell lines: OVCA 432, OVCAR­5, OVCAR­8 and A2780 cells. The association between Klotho levels and cell proliferation was determined by MTT assay, and the results indicated that higher levels of Klotho inhibited the proliferation of A2780 and OVCAR­5 cells, whereas reduced Klotho expression promoted cell growth of CaOV3 and SKOV­3 cells. In addition, the plasma levels of inflammatory cytokines in tumor­bearing mice and normal control mice were detected by enzyme­linked immunosorbent assay, and plasma interleukin (IL)­6 and IL­1ß levels were elevated in all tumor­bearing mice. Notably, the mRNA expression levels of IL­6 were significantly higher in the liver, ovaries and kidneys of Klotho­/­ mice compared with in wild type mice (P<0.01), thus indicating that aberrant Klotho expression may contribute to systemic inflammation in Klotho­/­ mice. Finally, the in vivo antitumor role of aberrant Klotho expression was determined in a nude mice model. A2780 cells were transfected with pCMV6­Klotho and the stably transfected cells were screened; the mice were injected with the stably transfected cells. The results indicated that tumor volume and tumor weight were significantly decreased in the pCMV6­Klotho group compared with in the pCMV6 vector group (P<0.01). These findings suggest that overexpression of Klotho may suppress tumor growth in animal models. In conclusion, Klotho was demonstrated to act as a potent tumor suppressor in human ovarian cancer cells. Reduced Klotho expression was detected in the specimens of patients with ovarian cancer, and overexpression of Klotho significantly inhibited cell proliferation of human ovarian cancer cells. Therefore, Klotho may be considered a useful key target for the molecular therapy of human ovarian cancer.

Puerarin Protects Human Neuroblastoma SH-SY5Y Cells against Glutamate-Induced Oxidative Stress and Mitochondrial Dysfunction.

Glutamate, the principal excitatory neurotransmitter, plays a central role in brain metabolism; however, aberrant neurotransmission of glutamate has been linked to neurodegenerative diseases. Therefore, the effective agents that target at glutamate-induced cell injury will be useful for prevention and treatment of neurodegenerative diseases. In this study, the neuroprotective effect of puerarin, an active isoflavone extracted from the Chinese herb Radix puerariae, against glutamate-induced cell injury in human neuroblastoma SH-SY5Y cells was evaluated for the first time. The results showed that the pretreatment of puerarin could attenuate glutamate-induced cell injury in a dose-dependent manner. This protective effect was mediated through inhibiting reactive oxygen species generation, attenuating the upregulation of Bax and downregulation of Bcl-2, preserving mitochondrial membrane potential (MMP), preventing cytochrome c release, and reducing caspase activity. These findings may significantly contribute to a better understanding of the neuroprotective effect of puerarin and provide new insights into its application toward neurodegenerative diseases in the future.