Live births from in vitro fertilization-embryo transfer following the administration of gonadotropin-releasing hormone agonist without gonadotropins: Two case reports.

BACKGROUND: The prevalence of female infertility between the ages of 25 and 44 is 3.5% to 16.7% in developed countries and 6.9% to 9.3% in developing countries. This means that infertility affects one in six couples and is recognized by the World Health Organization as the fifth most serious global disability. The International Committee for Monitoring Assisted Reproductive Technology reported that the global total of babies born as a result of assisted reproductive technology procedures and other advanced fertility treatments is more than 8 million. Advancements in controlled ovarian hyperstimulation procedures led to crucial accomplishments in human fertility treatments. The European Society for Human Reproduction and Embryology guideline on ovarian stimulation gave us valuable evidence-based recommendations to optimize ovarian stimulation in assisted reproductive technology. Conventional ovarian stimulation protocols for in vitro fertilization (IVF)-embryo transfer are based upon the administration of gonadotropins combined with gonadotropin-releasing hormone (GnRH) analogues, either GnRH agonists (GnRHa) or antagonists. The development of ovarian cysts requires the combination of GnRHa and gonadotropins for controlled ovarian hyperstimulation. However, in rare cases patients may develop an ovarian hyper response after administration of GnRHa alone. CASE SUMMARY: Here, two case studies were conducted. In the first case, a 33-year-old female diagnosed with polycystic ovary syndrome presented for her first IVF cycle at our reproductive center. Fourteen days after triptorelin acetate was administrated (day 18 of her menstrual cycle), bilateral ovaries presented polycystic manifestations. The patient was given 5000 IU of human chorionic gonadotropin. Twenty-two oocytes were obtained, and eight embryos formed. Two blastospheres were transferred in the frozen-thawed embryo transfer cycle, and the patient was impregnated. In the second case, a 37-year-old woman presented to the reproductive center for her first donor IVF cycle. Fourteen days after GnRHa administration, the transvaginal ultrasound revealed six follicles measuring 17-26 mm in the bilateral ovaries. The patient was given 10000 IU of human chorionic gonadotropin. Three oocytes were obtained, and three embryos formed. Two high-grade embryos were transferred in the frozen-thawed embryo transfer cycle, and the patient was impregnated. CONCLUSION: These two special cases provide valuable knowledge through our experience. We hypothesize that oocyte retrieval can be an alternative to cycle cancellation in these conditions. Considering the high progesterone level in most cases of this situation, we advocate freezing embryos after oocyte retrieval rather than fresh embryo transfer.

Tangeretin Synergizes with 5-Fluorouracil to Induce Autophagy through MicroRNA-21 in Colorectal Cancer Cells.

Combining innocuous natural products with cytotoxic agents may enhance the effectiveness of chemotherapy. Tangeretin is a citrus flavonoid that has antineoplastic properties, but its mechanism of action is still unknown. Here, we used a high throughput-screening (HTS) platform to screen for drugs that may synergize with tangeretin and confirmed the top hits against colorectal cancer (CRC) cells in vitro and in vivo. 5-Fluorouracil (5-FU) and PI3K/Akt inhibitors have come out as top hits that show a strong synergy effect with tangeretin by HTS. We further confirmed the synergistic effect of tangeretin with 5-FU against CRC cells in vitro and in vivo. Since 5-FU can increase microRNA-21 (miR-21) expression and activate PI3K/Akt signaling, we addressed if tangeretin acted at this level. In 5-FU treated cells, tangeretin inhibited miR-21 induction, rescued the expression of the target PTEN, reduced Akt activation, and induced autophagy. Together, our data indicated that a natural product, such as tangeretin, can modulate miR-21 expression and that this pathway might be a potential therapeutic target for CRC. Combining tangeretin with 5-FU may be useful in the clinic, since 5-FU is the current first line drug for treating CRC.

Integrating network pharmacology deciphers the action mechanism of Zuojin capsule in suppressing colorectal cancer.

BACKGROUND AND PURPOSE: Zuojin capsule (ZJC), a classical prescription, is outstanding in improving the conditions of patients with gastrointestinal diseases and colorectal cancer (CRC). Although ZJC has multi-ingredient and multi-target characteristics, its pharmacological effect on colorectal cancer and the underlying mechanism remain unclear. METHOD: Here, the activity of ZJC against CRC was evaluated by the experiments with CRC cells and HCT-116 xenografted mice. The key genes of CRC were obtained from the cancer genome atlas (TCGA). The genes potentially targeted by ZJC were collected from traditional Chinese medicine systems pharmacology (TCMSP) database. The underlying pathways related to selected targets were analyzed through gene ontology (GO) and pathway enrichment analyses. Western blot (WB), cellular thermal shift assay (CETSA), molecular docking and quantitative real-time PCR (QRT-PCR) were carried out to confirm the validity of the targets. RESULTS: In vitro and in vivo results indicated that ZJC may inhibit CRC cells and tumor growth. The network pharmacological analysis indicated that 22 compounds, 51 targets and 20 pathways were involved in the compound-target-pathway network. Our results confirmed that ZJC inhibited cycle progression, migration and induced apoptosis by targeting candidate genes (CDKN1A, Bcl2, E2F1, PRKCB, MYC, CDK2, and MMP9). We found that ZJC could directly change the protein level by regulating the protein stability and transcriptional activity of the target. CONCLUSIONS: In summary, combined network pharmacology and biological experiments proved that the main ingredients of ZJC such as quercetin, (R)-Canadine, palmatine, rutaecarpine, evodiamine, beta-sitosterol and berberine can target CDKN1A, Bcl2, E2F1, PRKCB, MYC, CDK2 and MMP9 to combat colorectal cancer. The results of this study provide a basic theory for the clinical trials of Zuojin Capsules against colorectal cancer.

The Biological Function, Mechanism, and Clinical Significance of m6A RNA Modifications in Head and Neck Carcinoma: A Systematic Review.

Head and neck squamous cell carcinoma (HNSCC) is one of the most common cancers, yet the molecular mechanisms underlying its onset and development have not yet been fully elucidated. Indeed, an in-depth understanding of the potential molecular mechanisms underlying HNSCC oncogenesis may aid the development of better treatment strategies. Recent epigenetic studies have revealed that the m6A RNA modification plays important roles in HNSCC. In this review, we summarize the role of m6A modification in various types of HNSCC, including thyroid, nasopharyngeal, hypopharyngeal squamous cell, and oral carcinoma. In addition, we discuss the regulatory roles of m6A in immune cells within the tumor microenvironment, as well as the potential molecular mechanisms. Finally, we review the development of potential targets for treating cancer based on the regulatory functions of m6A, with an aim to improving targeted therapies for HNSCC. Together, this review highlights the important roles that m6A modification plays in RNA synthesis, transport, and translation, and demonstrates that the regulation of m6A-related proteins can indirectly affect mRNA and ncRNA function, thus providing a novel strategy for reengineering intrinsic cell activity and developing simpler interventions to treat HNSCC.

Dual role of RACK1 in airway epithelial mesenchymal transition and apoptosis.

Airway epithelial apoptosis and epithelial mesenchymal transition (EMT) are two crucial components of asthma pathogenesis, concomitantly mediated by TGF-ß1. RACK1 is the downstream target gene of TGF-ß1 shown to enhancement in asthma mice in our previous study. Balb/c mice were sensitized twice and challenged with OVA every day for 7 days. Transformed human bronchial epithelial cells, BEAS-2B cells were cultured and exposed to recombinant soluble human TGF-ß1 to induced apoptosis (30 ng/mL, 72 hours) and EMT (10 ng/mL, 48 hours) in vitro, respectively. siRNA and pharmacological inhibitors were used to evaluate the regulation of RACK1 protein in apoptosis and EMT. Western blotting analysis and immunostaining were used to detect the protein expressions in vivo and in vitro. Our data showed that RACK1 protein levels were significantly increased in OVA-challenged mice, as well as TGF-ß1-induced apoptosis and EMT of BEAS-2B cells. Knockdown of RACK1 (siRACK1) significantly inhibited apoptosis and decreased TGF-ß1 up-regulated EMT related protein levels (N-cadherin and Snail) in vitro via suppression of JNK and Smad3 activation. Moreover, siSmad3 or siJNK impaired TGF-ß1-induced N-cadherin and Snail up-regulation in vitro. Importantly, JNK gene silencing (siERK) also impaired the regulatory effect of TGF-ß1 on Smad3 activation. Our present data demonstrate that RACK1 is a concomitant regulator of TGF-ß1 induces airway apoptosis and EMT via JNK/Smad/Snail signalling axis. Our findings may provide a new insight into understanding the regulation mechanism of RACK1 in asthma pathogenesis.

An overview of spirooxindole as a promising scaffold for novel drug discovery.

Introduction: Spirooxindole, a unique and versatile scaffold, has been widely studied in some fields such as pharmaceutical chemistry and synthetic chemistry. Especially in the application of medicine, quite a few compounds featuring spirooxindole motif have displayed excellent and broad pharmacological activities. Many identified candidate molecules have been used in clinical trials, showing promising prospects.Areas covered: This article offers an overview of different applications and developments of spirooxindoles (including the related natural products and their derivatives) in the process of drug innovation, including such as in anticancer, antimicrobial, anti-inflammatory, analgesic, antioxidant, antimalarial, and antiviral activities. Furthermore, the crucial structure-activity relationships, molecular mechanisms, pharmacokinetic properties, and main synthetic methods of spirooxindoles-based derivatives are also reviewed.Expert opinion: Recent progress in the biological activity profiles of spirooxindole derivatives have demonstrated their significant position in present-day drug discovery. Furthermore, we believe that the multidirectional development of novel drugs containing this core scaffold will continue to be the research hotspot in medicinal chemistry in the future.

MiR-451a suppressing BAP31 can inhibit proliferation and increase apoptosis through inducing ER stress in colorectal cancer.

The global morbidity and mortality of colorectal cancer (CRC) are ranked the third among gastrointestinal tumors in the world. MiR-451a is associated with several types of cancer, including CRC. However, the roles and mechanisms of miR-451a in CRC have not been elucidated. BAP31 is a predicted target gene of miR-451a in our suppression subtractive hybridization library. Its relationship with miR-451a and function in CRC are unclear. We hypothesized that miR-451a could induce apoptosis through suppressing BAP31 in CRC. Immunohistochemistry and real-time PCR were used to measure BAP31 expressions in CRC tissues and pericarcinous tissues from 57 CRC patients and CRC cell lines. Dual-luciferase reporter assay was used to detect the binding of miR-451a to BAP31. The expression of BAP31 protein in CRC tissues was significantly higher than that in pericarcinous tissues, which was correlated with distant metastasis and advanced clinical stages of CRC patients. The expression of BAP31 was higher in HCT116, HT29, SW620, and DLD cells than that in the normal colonic epithelial cell line NCM460. The expression of BAP31 was absolutely down-regulated when over-expressing miR-451a in HCT116 and SW620 cells compared with control cells. Mir-451a inhibited the expression of BAP31 by binding to its 5'-UTR. Over-expressing miR-451a or silencing BAP31 suppressed the proliferation and apoptosis of CRC cells by increasing the expressions of endoplasmic reticulum stress (ERS)-associated proteins, including GRP78/BIP, BAX, and PERK/elF2α/ATF4/CHOP, which resulted in increased ERS, cytoplasmic calcium ion flowing, and apoptosis of CRC cells. These changes resulting from over-expressing miR-451a were reversed by over-expressing BAP31 with mutated miR-451a-binding sites. Over-expressing miR-451a or silencing BAP31 inhibited tumor growth by inducing ERS. The present study demonstrated that miR-451a can inhibit proliferation and increase apoptosis through inducing ERS by binding to the 5'-UTR of BAP31 in CRC.

Use of cationic microbubbles targeted to P-selectin to improve ultrasound-mediated gene transfection of hVEGF165 to the ischemic myocardium.

Gene therapies have been applied to the treatment of cardiovascular disease, but their use is limited by the need to deliver them to the right target. We have employed targeted contrast ultrasound-mediated gene transfection (TCUMGT) via ultrasound-targeted microbubble destruction (UTMD) to transfer therapeutic genes to specific anatomic and pathological targets. Phospholipid microbubbles (MBs) with pcDNA3.1-human vascular endothelial growth factor 165 (pcDNA3.1-hVEGF165) plasmids targeted to P-selectin (MB+P+VEGFp) were created by conjugating monoclonal antibodies against P-selectin to the lipid shell. These microbubbles were divided into four groups: microbubble only (MB), microbubble+P-selectin (MB+P), microbubble+pcDNA3.1-hVEGF165 plasmid (MB+VEGFp), and microbubble+ P-selectin+pcDNA3.1-hVEGF165 plasmid (MB+P+VEGFp). The reverse transcription polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) results showed that the VEGF gene was successfully transfected by TCUMGT and the efficiency is increased with P-selectin targeting moiety. UTMD-mediated delivery of VEGF increased myocardial vascular density and improved cardiac function, and MB+P+VEGFp delivery showed greater improvement than MB+VEGFp. This study drew support from TCUGMT technology and took advantage of targeted ultrasound contrast agent to identify ischemic myocardium, release pcDNA3.1-hVEGF165 recombinant plasmid, and improve the myocardial microenvironment, so promoting the restoration of myocardial function.

NUDCD1 promotes metastasis through inducing EMT and inhibiting apoptosis in colorectal cancer.

Colorectal cancer (CRC) is the third most commonly diagnosed cancer and the third leading cause of cancer death in both men and women. NudC domain containing 1 (NUDCD1) was identified as an oncoprotein which was activated or over-expressed in various human cancers. We aimed to investigate the effects and mechanisms of NUDCD1 in human CRC. The expression of NUDCD1 in CRC and pericarcinous tissues from 70 CRC patients were determined by real-time PCR, western blotting, and immunohistochemistry. The correlation between the expression of NUDCD1 and clinical characteristics was analyzed. The expression of NUDCD1 in five CRC cell lines and normal colon mucosal epithelial cell line was measured by real-time PCR. Then we knock down NUDCD1 in HCT116 and HT 29 cells. The cell viability assay, scratch assay, migration and invasion assay and flow cytometry were used to analyze NUDCD1's effects on the proliferation, migration, invasion, cell cycle and apoptosis of CRC cells. NUDCD1's effects on CRC xenografts of nude mice was also determined. Results showed that the expression of NUDCD1 was much higher in CRC tissues than that in pericarcinous tissues. Over-expression of NUDCD1 in human CRC tissues was significantly associated with lymph node metastasis, distant metastasis, and advanced stages. The expression of NUDCD1 was higher in all of the CRC cell lines than that in normal colon epithelial mucosal cells. To knockdown NUDCD1 resulted in significant decreases in cell viability and proliferation, decreased protein expression of N-cadherin and increased protein expression of E-cadherin which were biomarkers of EMT, arrested the cell cycle and increased apoptosis via down-regulated cyclin D1, Bcl2, and up-regulated cleaved-caspase3. Furthermore, to knockdown NUDCD1 inactivated IGF1R-ERK1/2 signaling pathway in vitro and in vivo, and suppressed the xenografts of CRC. In conclusion, NUDCD1 promotes the carcinogenesis and metastasis of CRC through inducing EMT and inhibiting apoptosis, which suggests NUDCD1 be a potential biomarker for CRC.

Suppression subtractive hybridization identified differentially expressed genes in colorectal cancer: microRNA-451a as a novel colorectal cancer-related gene.

To investigate differentially expressed genes regulated by microRNA-451a in colorectal cancer. We detected expression of microRNA-451a in colorectal cancer samples and normal pericarcinous tissues from 68 colorectal cancer patients and the correlation between microRNA-451a and clinical features of these patients. Then, the expression of microRNA-451a in HCT116, SW620, HT29, SW480, and DLD cells was also measured. The suppression subtractive hybridization method was used with two HCT116 cell lines with overexpressing or underexpressing microRNA-451a, respectively. The most highly increased genes were screened. Their functions were predicted by gene ontology analysis. The expression ratio of microRNA-451a in colorectal cancer to pericarcinous tissues was 0.37. Expression of microRNA-451a was decreased in HCT116, SW620, HT29, SW480, and DLD cells. In our suppression subtractive hybridization library, expression of seven genes was the most highly increased when underexpressing microRNA-451a. They were BCAP31, EEF1A1, CDC20, WDR6, TUFM, RPL13, and RPL7A. Expression of DKK1, PSME1, NDUFA3, and GNB2 was most highly increased when overexpressing microRNA-451a. Gene ontology analysis showed that the main functions of these genes were associated with translational elongation, protein localization to the endoplasmic reticulum, translation, poly(A) RNA binding, negative regulations of Wnt signaling pathway, and so on. MicroRNA-451a was demonstrated to be downregulated in colorectal cancer patient tissues whose target genes were analyzed and functions were predicted by suppression subtractive hybridization.

Metal-Organic Framework Supported on Processable Polymer Matrix by In Situ Copolymerization for Enhanced Iron(III) Detection.

Metal-organic frameworks (MOFs) represent a promising class of porous materials. However, MOFs show poor processability that impedes their full potential in applications. This work develops a composite strategy to skillfully load MOFs on a polymer plate to afford processability for these powder materials. A predesigned mesoporous MOF with active -NH2 groups around the pore walls was prepared and its copolymerization with the -NCO groups of macromonomers (polyurethane acrylate) could be facilely induced by an initiator under mild conditions. Notably, the target MOF-polymer composite is transparent, elastic, and shows enhanced Fe3+ detection compared with that of the individual MOF functional component. This result can be ascribed to the synergistic effect of the composite with newly formed chemical bonds between the MOF particle and polymer matrix.

[Values of the sperm deformity index, acrosome abnormity rate, and sperm DNA fragmentation index of optimized sperm in predicting IVF fertilization failure].

OBJECTIVE: To investigate the values of the sperm deformity index (SDI), acrosome abnormity rate (AAR), and DNA fragmentation index (DFI) of optimized sperm in the prediction of fertilization failure (fertilization rate < 25%) in conventional in vitro fertilization (IVF). METHODS: We selected 695 cycles of conventional IVF for pure oviductal infertility in this study, including 603 cycles of normal fertilization and 92 cycles of fertilization failure. On the day of oocyte retrieval, we examined sperm morphology, acrosome morphology, and DNA fragmentation using the Diff-Quik, PSA-FITC and SCD methods. We established the joint predictor (JP) by logistic equation and analyzed the values of different parameters in predicting fertilization failure with the receiver operating characteristic (ROC) curve. RESULTS: The fertilization rate was negatively correlated with SDI (r = - 0.07; P = 0.03), AAR (r = -0.49; P < 0.01), and DFI (r = -0. 21; P < 0.01). The SDI, AAR, and DFI in the normal fertilization group were 1.24 ± 0.20, (7.75 ± 2.28)%, and (7.87 ± 3.15)%, and those in the fertilization failure group were 1.42 ± 0.15, (12.02 ± 3.06)%, and (13.32 ± 4.13)%, respectively, all with statistically significant differences between the two groups (P < 0.05). SDI, AAR, and DFI were all risk factors of fertilization failure ( OR = 2.68, 14.11, and 3.85; P = 0.01, < 0.01, and < 0.01). The areas under the ROC curves for SDI, AAR, DFI, and JP were 0.651 ± 0.033, 0.895 ± 0.019, 0.789 ± 0.022, and 0.915 ± 0.017, respectively. According to the Youden index, the optimal cut-off values of SDI, AAR, and DFI obtained for the prediction of fertilization failure were approximately 1.45, 10%, and 12%. CONCLUSION: The SDI, AAR and DFI of optimized sperm are closely associated with the fertilization rate, and all have the value for predicting fertilization failure in IVF. The AAR is more valuable than the other single predictors, but JP is more effective than the AAR.

Dehydroandrographolide enhances innate immunity of intestinal tract through up-regulation the expression of hBD-2.

BACKGROUND: Dehydroandrographolide (DA) is one of major active components in the well-known oriental herbal medicine Andrographis paniculata (Burm.f) Nees which belongs to the Acanthaceae family. DA is used for the treatment of infections in China. However, DA has not been found to significantly inhibit bacterial and viral growth directly. The current study investigates the effect of DA on the expression of human ß -defensin-2 (hBD-2) in human intestinal epithelial cells and the possible signaling pathways. METHODS: Human intestinal epithelial HCT-116 cells were incubated with 1-100 µM DA for 2-24 h. RT-PCR and Western blot were used to assess the expression of hBD-2. The specific inhibitors were used and the levels of phosphorylation of signaling molecules were detected for dissecting the signaling pathways leading to the induction of hBD-2. RESULTS: MTT assay showed there was no obvious cytotoxicity for HCT-116 cells by 1-100 µM DA treatment. RT-PCR and Western blot assays showed that DA (1-100 µM) could up-regulate the expression of hBD-2, and the effect lasted longer than 24 h. By using SB203580 and SB202190 (inhibitors of p38), the enhancement of hBD-2 expression were significantly attenuated. However, inhibitor of ERK and inhibitor of JNK could not block the effect of DA. Furthermore, Western blot found activation of p38 but not ERK and JNK in DA-treated HCT-116 cells. CONCLUSION: The results suggested that DA enhanced innate immunity of intestinal tract by up-regulating the expression of hBD-2 through the p38 MAPK pathways.

Anti-proliferation effects of isorhamnetin on lung cancer cells in vitro and in vivo.

BACKGROUND: Isorhamnetin (Iso), a novel and essential monomer derived from total flavones of Hippophae rhamnoides that has long been used as a traditional Chinese medicine for angina pectoris and acute myocardial infarction, has also shown a spectrum of antitumor activity. However, little is known about the mechanisms of action Iso on cancer cells. OBJECTIVES: To investigate the effects of Iso on A549 lung cancer cells and underlying mechanisms. MATERIALS AND METHODS: A549 cells were treated with 10~320 µg/ml Iso. Their morphological and cellular characteristics were assessed by light and electronic microscopy. Growth inhibition was analyzed by MTT, clonogenic and growth curve assays. Apoptotic characteristics of cells were determined by flow cytometry (FCM), DNA fragmentation, single cell gel electrophoresis (comet) assay, immunocytochemistry and terminal deoxynucleotidyl transferase nick end labeling (TUNEL) . Tumor models were setup by transplanting Lewis lung carcinoma cells into C57BL/6 mice, and the weights and sizes of tumors were measured. RESULTS: Iso markedly inhibited the growth of A549 cells with induction of apoptotic changes. Iso at 20 µg/ml, could induce A549 cell apoptosis, up-regulate the expression of apoptosis genes Bax, Caspase-3 and P53, and down-regulate the expression of Bcl-2, cyclinD1 and PCNA protein. The tumors in tumor-bearing mice treated with Iso were significantly smaller than in the control group. The results of apoptosis-related genes, PCNA, cyclinD1 and other protein expression levels of transplanted Lewis cells were the same as those of A549 cells in vitro. CONCLUSIONS: Iso, a natural single compound isolated from total flavones, has antiproliferative activity against lung cancer in vitro and in vivo. Its mechanisms of action may involve apoptosis of cells induced by down-regulation of oncogenes and up-regulation of apoptotic genes.

Correlation of over-expressions of miR-21 and Notch-1 in human colorectal cancer with clinical stages.

AIM: The aim of the study was to identify expressions of Notch-1, microRNA-21 (miR-21), and phosphatase and tensin homolog (PTEN) in colorectal cancer (CRC), and to explore their relationship with clinical stages and metastatic status of CRC. MAIN METHODS: 102 CRC patients were enrolled and clinical data were analyzed. Expressions of Notch-1 and miR-21 in CRC and adjacent non-tumor tissues of these patients were measured by real time-PCR. Protein expressions of Notch-1 and PTEN of 12 paired tissues were determined by Western blot and immunohistochemistry. The correlations between gene expressions in different clinical stages as well as metastatic status were evaluated by linear regression. KEY FINDINGS: Notch-1 was over-expressed in 86.27% (88/102) CRC tissues, particularly in advanced stages, while miR-21 expression was increased to 74.51% (76/102) in CRC tissues compared with matched adjacent non-tumor tissues. The expressions of Notch-1 and miR-21 were positively correlated with CRC development, especially in advanced-stages (r(2)=0.3839, p<0.01). Expressions of PTEN were significantly down-regulated in CRC tissues and negatively correlated with expressions of Notch-1 (r(2)=0.5207, p<0.01) and miR-21 (r(2)=0.6996, p<0.01). SIGNIFICANCE: These data indicate that the crosstalk between Notch-1 and miR-21 is involved in CRC development through degradation of PTEN.

NRP-1 expression in bladder cancer and its implications for tumor progression.

Neuropilin-1 (NRP-1) overexpression has been reported in a variety of human cancers. However, the role of NRP-1 in bladder cancer (BC) remains unclear. The aim of present study was to analyze NRP-1 protein expression in BC tissues and to assess its prognostic significance for BC. NRP-1 messenger ribonucleic acid (mRNA) and protein expression were determined by real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and immunohistochemistry in specimens of primary cancer and their adjacent noncancerous tissues in BC patients. Additionally, NRP-1 protein expression in 139 archived paraffin-embedded BC samples was analyzed by immunohistochemistry and correlated with clinicopathological characteristics and survival. Student's t test, Spearman's rank correlation, Kaplan-Meier plots, and Cox's proportional hazards regression model were used to analyze the data. By qRT-PCR and immunohistochemistry, the levels of NRP-1 mRNA and protein were significantly higher in BC, compared to that in adjacent noncancerous tissues (P<0.001). High expression of NRP-1 was significantly associated with histologic grade (P=0.016) and tumor stage (P=0.001). Multivariate analysis showed that high expression of NRP-1 was an independent prognostic factor for overall survival. Our study suggests that overexpression of NRP-1 may play an important role in the progression of BC, and NRP-1 expression may serve as a biomarker for poor prognosis for BC.

Compound danshen tablet ameliorated aß25-35-induced spatial memory impairment in mice via rescuing imbalance between cytokines and neurotrophins.

BACKGROUND: Compound Danshen Tablet (CDT), a Traditional Chinese Medicine, has recently been reported to improve spatial cognition in a rat model of Alzheimer's disease. However, in vivo neuroprotective mechanism of the CDT in models of spatial memory impairment is not yet evaluated. The present study is aimed to elucidate the cellular mechanism of CDT on Aß25-35-induced cognitive impairment in mice. METHODS: Mice were randomly divided into 5 groups: the control group (sham operated), the Aß25-35 treated group, the positive drug group, and large and small dosage of the CDT groups, respectively. CDT was administered at a dose of 0.81 g/kg and 0.405 g/kg for 3 weeks. The mice in the positive drug group were treated with 0.4 mg/kg of Huperzine A, whereas the mice of the control and Aß25-35 treated groups were administrated orally with equivalent saline. After 7 days of preventive treatment, mice were subjected to lateral ventricle injection of Aß25-35 to establish the mice model of Alzheimer's disease. Spatial memory impairment was evaluated by Morris water maze test. Choline acetyltransferase (ChAT) contents in hippocampus and cortex were quantified by ELISA. The levels of cytokines, receptor of activated protein kinase C1 (RACK1) and brain-derived neurotrophic factor (BDNF) in hippocampus were measured by RT-PCR and ELISA. RESULTS: The results showed that Aß25-35 caused spatial memory impairment as demonstrated by performance in the Morris water maze test. CDT was able to confer a significant improvement in spatial memory, and protect mice from Aß25-35-induced neurotoxicity. Additionally, CDT also inhibited the increase of TNF-α and IL-6 level, and increased the expression of choline acetyltransferase (ChAT), receptor of activated protein kinase C1 (RACK1) and brain-derived neurotrophic factor (BDNF) in brain as compared to model mice. CONCLUSION: These findings strongly implicate that CDT may be a useful treatment against learning and memory deficits in mice by rescuing imbalance between cytokines and neurotrophins.

Apoptosis induced by ß,ß-dimethylacrylshikonin is associated with Bcl-2 and NF-κB in human breast carcinoma MCF-7 cells.

ß,ß-dimethylacrylshikonin (DA) is a natural naphthoquinone derivative compound of Lithospermum erythrorhizon with various biological activities. The present study aimed to investigate the inhibitory effects and underlying mechanisms of DA in human breast carcinoma MCF-7 cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that DA inhibited the proliferation of MCF-7 cells in a dose- and time-dependent manner. The half maximal inhibitory concentration of DA with regard to the proliferation of MCF-7 cells was 0.050±0.016 mM. The characteristics of cell apoptosis, including cell shrinkage, nuclear pyknosis and chromatin condensation, were all observed in DA-treated cells. DA decreased the expression levels of Bcl-2 and increased the expression of Bax and caspase-3 compared with those in the control. DA inhibited the activity of the nuclear factor (NF)-κB pathway, by downregulating the expression of the p65 subunit, and inhibited the Iκb phosphorylation. DA inhibits the proliferation of MCF-7 cells in vitro by inducing apoptosis through the downregulation of Bcl-2, upregulation of Bax and partial inactivation of the NF-κB pathway.

[Study on leuprorelin acetate in treatment of uterine adenomyosis with infertility].

OBJECTIVE: To study clinical efficacy of leuprorelin acetate in treatment of uterine adenomyosis with infertility. METHODS: From January 1,2011 to March 31,2012, 166 cycles in 166 infertile patients combined with uterine adenomyosis undergoing in vitro fertilization embryo transplant (IVF-ET) with long protocol ovum induction by leuprorelin acetate in centre of medical reproduction, ningbo women and children's hospital were studied retrospectively. In the mean time, 200 cycles in 200 infertile patients with tubal factors were enrolled as control group.The volume of uterus and outcome of IVF-ET were compared and studied between two groups. RESULTS: (1) Volume of uterus:in adenomyosis group, after 2-6 cycles of injecting leuprorelin acetate (3.75 mg/28 days), the mean uterine volume was shrinked from (180 ± 73) cm(3) to (86 ± 67) cm(3) (P < 0.05). (2) Outcome of IVF-ET:the rate of embryo implantation was 39.1% in adenomyosis group and 35.8% in control group. The rate of clinical pregnancy was 54.2% in adenomyosis group and 53.7% in control group. The rate of abortion was 4.7% in adenomyosis group and 4.2% in control group. They all did not show statistical differences (P > 0.05). (3) In adenomyosis group, the rate of fertilization, two pronuclear (2PN) and superior embryo were 67.2%(319/475), 60.8% (289/475) and 52.9% (162/306) in patients with failed pregnancy and 74.2% (423/570), 67.7% (386/570) and 62.1% (256/412) in patients with successful pregnancy after IVF-ET, which reached significant difference (P < 0.05). CONCLUSION: Leuprorelin acetate could improve volume of uterine adenomyosis and outcome of pregnancy in patients undergoing IVF-ET.

Alterations in the frequency of trinucleotide repeat dynamic mutations in offspring conceived through assisted reproductive technology.

STUDY QUESTION: How does the frequency of trinucleotide repeat dynamic mutations in offspring conceived through assisted reproductive technology (ART) compare with the frequency of these mutations in control offspring conceived from spontaneous pregnancies? SUMMARY ANSWER: There is a slight increase in dynamic mutation instability in offspring conceived through ART compared with the naturally conceived offspring. WHAT IS KNOWN ALREADY: There is evidence to suggest that ART can increase the risk of birth defects and karyotypic abnormalities. However, the accumulating evidence of an association between ART and de novo genetic aberrations is controversial. STUDY DESIGN, SIZE, DURATION: A prospective clinical observational study was performed on 246 families recruited from an in vitro fertilisation (IVF) centre at a tertiary-care, university-affiliated teaching hospital from 2008 to 2012. The study included 147 ART families [75 IVF and 72 intracytoplasmic sperm injection (ICSI)] in the study group and 99 natural-conception families in the control group. PARTICIPANTS, SETTING, METHODS: Parental, umbilical cord and infant peripheral blood samples were collected, and the trinucleotide repeats of the ATN1, AR, ATXN1, ATXN3, Huntington, DMPK and FMR-1 genes were investigated between the generations; these genes were chosen due to their ability to undergo dynamic mutation. The frequencies and sizes of the mutational repeats, as well as the intergenerational instability, were measured. MAIN RESULTS AND THE ROLE OF CHANCE: In 2466 transmissions identified in the ART offspring, 2.11% (n = 52/2466) of the alleles were unstable upon transmission, while in the control group offspring, the frequency of dynamic mutation was 0.77% (n = 10/1300); this difference was statistically significant (P < 0.01). The unstable transmission alleles were detected in 32 (2.48%) of the 1288 alleles from the IVF offspring and in 20 (1.70%) of the 1178 alleles from the ICSI offspring; both of these frequencies were significantly different from that of naturally conceived offspring (0.77%) (P < 0.01 and P < 0.05, respectively). However, there were no significant differences in the sizes of the mutational repeats or in the rates of expansion or contraction among the three groups (P > 0.05). The repeat copy numbers of the examined genes were found to be within the normal ranges in all parents and infants. LIMITATIONS, REASONS FOR CAUTION: One strength of our study is the relatively large sample size; we were able to detect mutations in seven common dynamic genes, and this large sample size allowed us to detect unstable alleles. Although we observed a clear alteration in the frequency of dynamic mutation in the ART offspring compared with controls, further studies are urgently needed to confirm this observation and determine the cause of this phenomenon. WIDER IMPLICATIONS OF THE FINDINGS: DNA microsatellite analysis provides an important tool to assess genomic instability. In this study, we report an association between ART and the frequency of dynamic mutation. The instability could be a reflection of the core infertility problem, the controlled ovarian hyperstimulation and/or the in vitro culture conditions.