Blocking of SIRT7/FOXO3a axis by miR-152-3p enhances cisplatin sensitivity in breast cancer.

BACKGROUND: Cisplatin-based chemoresistance is major obstacle for breast cancer (BC) including Triple-negative breast cancer (TNBC). SIRT7 is reportedly involved in the progression of BC, the underlining mechanism in Cisplatin-based chemoresistance in BC remains unclear. This work is to elucidate effects of SIRT7 on cisplatin resistance in breast cancer regulated by miR-152-3p. METHODS: The RNA expression of SIRT7 and miRNAs in breast cancer were available from TCGA database. SIRT7-targeted miRNAs were predicted by TargetScan, miRanda, miRDB databases. The association of SIRT7 expression with predicted miRNA was validated by Luciferase assay. Cell apoptosis was determined by Flow cytometry. Cell viability was detected by CCK8 assay. The mRNA expression was measured by quantitative real-time polymerase chain reaction (qRT-PCR) assay. Protein expression was determined by Western blotting assay. RESULTS: SIRT7 mRNA levels were dramatically enhanced in BC tissues compared to para-carcinoma tissues, also increased in BC patients with Cisplatin-based chemotherapy containing TNBC compared with those without. The increase of SIRT7 expression was obviously relevant to shorter survive time of them. Importantly, SIRT7 inhibition facilitated Cisplatin-induced cell apoptosis of TNBC (MDA-MB-231 and MDA-MB-468) and non- TNBC (MCF-7). Notably, miR-152-3p was predicted as a negative regulator of SIRT7 by overlapping downregulated miRNAs in BC patients treated with Cisplatin-based chemotherapy and miRNAs to target SIRT7. Mechanically, miR-152-3p blocked SIRT7 to stimulate an activation of FOXO3a, cleaved PARP1 and Caspase-3, sensitizing Cisplatin-induced apoptosis of BC cells. CONCLUSIONS: Inhibition of SIRT7 by miR-152-3p may be a promising strategy against the resistance to cisplatin-based chemotherapy in BC containing TNBC.

Enhancing the therapeutic efficacy of gefitinib on subcutaneously transplanted SKOV3 ovarian cancer tumors in nude mice via ultrasound­stimulated microbubble cavitation.

The present study aimed to explore the effect of ultrasound-stimulated microbubble cavitation (USMC) on drug concentration and therapeutic efficacy of oral gefitinib in treating subcutaneously transplanted SKOV3 ovarian cancer tumors in nude mice. The present study employed the VINNO70 ultrasonic diagnostic and treatment integrated machine for USMC therapy. Firstly, the mechanical index was set at 0.25, and the therapeutic efficacy of USMC treatment was assessed at intervals of 5, 10 and 20 min. Briefly, 72 nude mice were randomized into the following four groups (n=18/group): Control group, USMC5 min group, USMC10 min group and USMC20 min group, and the therapeutic response to USMC treatment was evaluated by comparing pre-and post-intervention effects. Additionally, the combined therapeutic efficacy of USMC and gefitinib was investigated by randomly dividing 96 tumor-bearing mice into the following four groups (n=24/group): Control group, USMC group, gefitinib group and USMC + gefitinib group. Contrast-enhanced ultrasound, hematoxylin and eosin staining, western blotting, immunofluorescence staining, TUNEL staining, ELISA and liquid chromatography-mass spectrometry were performed in the present study. The results showed that USMC combined with gefitinib had the best treatment effect; the tumor inhibition rate was higher than that of gefitinib alone and the overall survival time was prolonged. In addition, the drug concentration in the tumor tissue obtained from the USMC + gefitinib group was revealed to be ~1.4 times higher than that detected in the group treated with gefitinib alone. The experimental results also confirmed that the strongest tumor inhibition rate and longest overall survival time was observed in the USMC + gefitinib group, followed by the gefitinib group and USMC group. STAT3 is an important signaling transducer and transcription factor, which, when phosphorylated, can lead to abnormal cell proliferation and malignant transformation. In addition, the upregulation of phosphorylated (p)-STAT3 is consider a reason for the poor efficacy of gefitinib in treating ovarian cancer. The present study revealed that ultrasound microbubble therapy could overcome this side effect. In conclusion, USMC improved the effects of oral gefitinib on subcutaneously transplanted SKOV3 ovarian cancer tumors in nude mice and increased drug penetration. In addition, USMC overcame the gefitinib-induced side effect of upregulated STAT3 phosphorylation and reduced the expression levels of p-STAT3 in the tumor.

Troxerutin dampened hypothalamic neuroinflammation via microglial IL-22/IL-22R1/IRF3 activation in dihydrotestosterone-induced polycystic ovary syndrome rats.

BACKGROUND: Polycystic ovary syndrome (PCOS) is the most common reproductive-endocrine condition in premenopausal women. Troxerutin, a common clinical anti-coagulant agent, was shown to work as a strong IL-22 boosting agent counteracting the hyperactivated gonadotrophin releasing hormone (GnRH) neurons and heightened GnRH release, the neuroendocrine origin of PCOS with unknown mechanism in rats. Exploring the off-label use of troxerutin medication for PCOS is thus sorely needed. METHODS: Serum IL-22 content and hypothalamic IL-22 protein were detected. Inflammatory factor levels in hypothalamo-pituitary were evaluated. Immunofluorescence staining was employed to determine the activation and M1/M2-prone polarization of microglia in arcuate hypothalamus and median eminence. RNA-sequencing and transcriptome analysis were applied to explore the potential driver of microglia M2-polarization in response to IL-22 bolstering effect. The function of microglial IL-22/IL-22R1/IRF3 system was further verified using in vivo knockdown of IL-22R1 and a potent IRF3 inhibitor in BV2 microglial cell lines in vitro. RESULTS: Troxerutin augmented serum IL-22 content, and its consequent spillover into the hypothalamus led to the direct activation of IL-22R1/IRF3 system on microglia, thereby promoted microglia M2 polarization in arcuate hypothalamus and median eminence, dampened hypothalamic neuroinflammation, inhibited hyperactive GnRH and rescued a breadth of PCOS-like traits in dihydrotestosterone (DHT) rats. The salutary effects of troxerutin treatment on hypothalamic neuroinflammation, microglial M1/2 polarization, GnRH secretion and numerous PCOS-like features were blocked by in vivo knockdown of IL-22R1. Moreover, evidence in vitro illustrated that IL-22 supplement to BV-2 microglia cell lines promoted M2 polarization, overproduction of anti-inflammatory marker and limitation of pro-inflammatory factors, whereas these IL-22 effects were blunted by geldanamycin, a potent IRF3 inhibitor. CONCLUSION: Here, the present study reported the potential off-label use of troxerutin medication, a common clinical anti-coagulant agent and an endogenous IL-22 enhancer, for multiple purposes in PCOS. The rational underlying the application of troxerutin as a therapeutic choice in PCOS derived from its activity as an IL-22 memetic agent targeting the neuro-endocrine origin of PCOS, and its promotive impact on microglia M2 polarization via activating microglial IL-22R1/IRF3 system in the arcuate hypothalamus and median eminence of DHT female rats.

Comparative diagnostic accuracy of the IOTA SRR and LR2 scoring systems for discriminating between malignant and Benign Adnexal masses by junior physicians in Chinese patients: a retrospective observational study.

BACKGROUND: The accuracy of ultrasound in distinguishing benign from malignant adnexal masses is highly correlated with the experience of ultrasound physicians. In China, most of ultrasound differentiation is done by junior physicians. PURPOSE: To compare the diagnostic performance of the International Ovarian Tumour Analysis (IOTA) Simple Rules Risk (SRR) and IOTA Logistic Regression Model 2 (LR2) scoring systems in Chinese patients with adnexal masses. METHODS: Retrospective analysis of ovarian cancer tumor patients who underwent surgery at a hospital in China from January 2016 to December 2021. Screening patients with at least one adnexal mass on inclusion and exclusion criteria. Two trained junior physicians evaluated each mass using the two scoring systems. A receiver operating characteristic curve was used to test the diagnostic performance of each system. RESULTS: A total of 144 adnexal masses were retrospectively collected. Forty masses were histologically diagnosed as malignant. Compared with premenopausal women, postmenopausal women had a much higher rate of malignant masses. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) of the SRR was 97.5% (95% CI: 86.8 -99.9%), 82.7% (95% CI: 74.0 -89.4%), 68.4% (95% CI: 58.7 -76.8%) and 98.9% (95% CI: 92.5 -99.8%). The sensitivity, specificity, PPV, NPV of the LR2 were 90.0% (95% CI: 76.5 -97.2%), 89.4% (95% CI: 81.9 -94.6%), 76.6% (95% CI: 65.0 -85.2%), and 95.9% (95% CI: 90.2 -98.3%). There was good agreement between two scoring systems, with 84.03% total agreement and a kappa value of 0.783 (95% CI: 0.70-0.864). The areas under the curve for predicting malignant tumours using SRR and LR2 were similar for all patients (P > 0.05 ). CONCLUSION: The two scoring systems can effectively distinguish benign from malignant adnexal masses. Both scoring systems have high diagnostic efficacy, and diagnostic efficacy is stable, which can provide an important reference for clinical decision making.

Evaluation of novel obesity- and lipid-related indices as predictors of abnormal glucose tolerance in Chinese women with polycystic ovary syndrome.

PURPOSE: We aimed to evaluate the performance of Chinese visceral adiposity index (CVAI), visceral adiposity index (VAI), lipid accumulation product (LAP), triglyceride glucose (TyG) as indices in screening abnormal glucose tolerance (AGT) in Chinese women with polycystic ovary syndrome (PCOS), using the oral glucose tolerance test (OGTT) as a reference test. In addition, we essentially compared the abilities of these indices with body mass index (BMI), waist circumference (WC), fasting plasma glucose (FPG). MATERIALS AND METHODS: All 1113 PCOS patients evaluated in this study underwent OGTTs. The 2-h post-oral glucose load (2 h-PG) level was used to categorize subjects into two groups: those having AGT or normal glucose tolerance (NGT) levels. RESULTS: A statistically significant positive correlation between levels of 2 h-PG and FPG, BMI, WC, LAP, VAI, CVAI, TyG, (P < 0.05), was observed. The strongest correlation was found between the levels of 2 h-PG and CVAI (r = 0.47). The CVAI provided the highest area under the receiver-operating characteristic curve (AUC) for AGT, followed by LAP, BMI, TyG, VAI, WC, and FPG. The CVAI of 32.61 (with AUC: 0.76, sensitivity: 73%, specificity: 70%, positive preductive value (PPV): 0.41, negative predictive value (NPV): 0.90) was found to be the cut-off point for AGT in Chinese women with PCOS. CONCLUSIONS: CVAI may not reliably detect AGT in Chinese women with PCOS. However, it is suitable as a first screening indicator to guide physicians to ordering OGTT.

Troxerutin attenuates insulin resistance via pancreatic IL-22/JAK1/STAT3 signaling activation in dihydrotestosterone-induced polycystic ovary syndrome rats.

Polycystic ovary syndrome (PCOS) is an extremely common endocrine-metabolic disorder and the main cause of infertility in premenopausal women, thus targeted treatments are sorely needed. Accumulative evidence showed that exogenous supplementation of IL-22 in PCOS mice may be of significant positive effect on insulin resistance (IR), a root causative factor for this condition, but much remained unknown about its mechanism. According to our previous study, troxerutin, a common anticoagulant and thrombolytic agent in clinic, alleviated various PCOS-like phenotypes in dihydrotestosterone (DHT)-treated rat model with unclear mechanism. Here, glucose tolerance tests (GTTs), insulin tolerance tests (ITTs), and homeostatic model assessment of insulin resistance (HOMA-IR) analyses revealed that troxerutin treatment in DHT-treated rats also significantly improved insulin resistance and enhanced serum IL-22 levels, which thereby activated IL-22R1/Janus kinase 1 (JAK1)/signal transducer and activator of transcription-3 (STAT3) signaling pathway in pancreatic islet. This protective effect of troxerutin on insulin resistance improvement was blocked by an inhibitor of p-STAT3, S3I-201. Troxerutin administration to DHT rats decreased the relative abundance of Bifidobacterium and enhanced secondary bile acid profiles, which were positively correlated with serum IL-22 concentration. Conclusively, the present study reported that troxerutin is an endogenous enhancer of IL-22 and the effect of troxerutin on insulin resistance improvement was via IL-22R1/JAK1/STAT3 signaling activation in a DHT-induced PCOS rat model. These insights may be translated into a primary therapeutic agent for PCOS with insulin resistance and hyperandrogenism.NEW & NOTEWORTHY Troxerutin decreased the relative abundance of Bifidobacterium, along with enhancement of secondary bile acids/IL-22 system, which thereby activated its downstream IL-22R1/JAK1/STAT3 signaling pathway in pancreatic ß cells, subsequently attenuated insulin resistance (IR), hyperandrogenism and PCOS-like phenotypes in DHT-induced PCOS rat models. Troxerutin is an endogenous IL-22 enhancer and may be of therapeutic value for PCOS with insulin resistance.

Feasibility of Computer-Aided Design in Limb Lengthening Surgery: Surgical Simulation and Guide Plates.

OBJECTIVE: To evaluate the feasibility and utility of computer-aided design (CAD) in surgical treatment of leg length discrepancy (LLD) using monorail external fixators. METHODS: In the present case series, we retrospectively analyzed seven patients diagnosed with LLD who were surgically treated using a monorail external fixator between June 2018 and August 2020. A personalized surgical emulation of each patient was designed using CAD based on preoperative CT scans to measure limb parameters. Through reverse engineering, a surgical guide plate was then designed to assist with correcting the limb deformity. Patient general information and clinical history, leg length, mechanical lateral distal femoral angle (mLDFA), anatomical anterior distal tibial angle (aADTA), and surgical parameters were recorded during the perioperative period. Three months after external fixator removal, distraction-consolidation time (DCT), healing index (HI), and lower extremity function score (LEFS) were calculated, and statistically analyzed by paired T-test. RESULTS: The mean limb lengthening achieved was 6.41 ± 2.54 (range, 3.30-10.54) cm with either varus or valgus correction. The mean operative duration was 151 ± 41.87 (84-217) minutes and mean blood loss was 53.58 ± 22.51(25-87) ml. The mean distraction-consolidation time was 3.67 ± 1.13 (range, 2.5-6.0) months and mean external fixator duration was 11 ± 2.45 (range, 8-14) months. The mean healing index (HI) was 18.11 ± 3.58 (range, 12.8-22.7) days/cm. Mean LEFS scores improved postoperatively from 32.17 ± 8.57 (range, 24-45) to 61.17 ± 6.68 (range, 50-67) with a significant difference (T = -14.26,P < 0.001). CONCLUSIONS: Simultaneous length and angular correction can be achieved by incorporating CAD into the surgical treatment of patients with LLD, without compromising postoperative lower limb function. CAD demonstrates utility in the surgical treatment of LLD by improving the functionality of monorail external fixators.

Evaluation of Novel Obesity and Lipid-Related Indices as Indicators for the Diagnosis of Metabolic Syndrome and Premetabolic Syndrome in Chinese Women with Polycystic Ovary Syndrome.

OBJECTIVES: According to the International Diabetes Federation (IDF) criteria, previous studies in Chinese women with polycystic ovary syndrome (PCOS) reported a low prevalence of metabolic syndrome (MS); however, the same population predisposed to developing pre-MS. Early identification and treatment of individuals with MS and pre-MS are imperative to prevent their adverse consequences. Moreover, fasting plasma glucose (FPG) was not accurate in detecting pathoglycemia in women with PCOS as they have shown characteristically postprandial abnormalities in the carbohydrate metabolism. Therefore, we aimed to compare the discriminative performance of various indices for identifying MS and pre-MS/MS (pre-MS and MS) using the updated Chinese Diabetes Society (uCDS) criteria in Chinese women with PCOS. METHODS: 1083 Chinese women with PCOS were included in this study. We measured and evaluated 8 indices in all individuals. Based on the uCDS criteria for MS, patients who had no less than two components of MS but did not meet the criteria for the diagnosis of MS were considered as having pre-MS. Receiver operating characteristic (ROC) curves and the area under ROC curves (AUCs) levels were used to assess the accuracy of each index in detecting MS and pre-MS/MS. RESULTS: Among the 8 indices assessed, the lipid accumulation product (LAP) provided the highest AUCs for detecting MS and pre-MS/MS, followed by CVAI, WTI, VAI, TyG, TG/HDL, WC, and BMI. The optimal cutoff points determined for LAP were 45.13 (sensitivity 88.0%, specificity 88.4%, and Youden index 0.764) for MS and 28.01 (sensitivity 87.5%, specificity 80.7%, and Youden index 0.681) for pre-MS/MS, respectively. CONCLUSIONS: uCDS criteria are reasonably more suitable for detecting MS and pre-MS in Chinese women with PCOS. Based on this criterion, LAP is the best index for the diagnosis of MS and pre-MS/MS in Chinese women with PCOS, out of the 8 obesity and lipid-related indices assessed.

Elevated Circulating Levels of Carnal ENST00000550337.1 Are Associated with Polycystic Ovary Syndrome in Chinese Women.

OBJECTIVES: Polycystic ovary syndrome (PCOS) is a common endocrine and metabolic disease. Some studies reported that the development of PCOS may be closely related to insulin resistance (IR). Interestingly, the long noncoding RNA (lncRNA) ENST00000550337.1 in peripheral blood is mainly involved in glucose metabolism. Therefore, the purpose of our study was to explore the relationship between lncRNA ENST00000550337.1 level and PCOS patients. MATERIALS AND METHODS: Seventy-five PCOS patients and 72 healthy controls were enrolled in this study. We used qRT-PCR to detect the expression level of lncRNA ENST00000550337.1 in peripheral blood leukocytes from patients with PCOS. We also investigated potential relationships between lncRNA ENST00000550337.1 and the endocrine parameters in PCOS. RESULTS: We observed that the expression of lncRNA ENST00000550337.1 in PCOS patients was significantly higher than that in the control subjects and positively correlated with PCOS occurrence, waist circumference, waist-hip ratio, IR, fasting insulin levels, and blood glucose. The expression of lnc RNA ENST00000550337.1 was positively correlated with PCOS (p = 0.003). There were independent correlations between IR and expression of lncRNA ENST00000550337.1 in patients with PCOS. Patients with elevated lncRNA ENST00000550337.1 expression had significantly increased PCOS risk after adjusting for age and BMI. LncRNA ENST00000550337.1 expression level provided a sensitivity of 81.3% and a specificity of 78.1% with a threshold value of 6.4648 for the prediction of PCOS. The area under the ROC was 0.813. LIMITATIONS: There are some limitations to this study. First, the sample size was limited and the causal relationship between lncRNA ENST00000550337.1 and PCOS was not investigated due to the cross-sectional study design. Second, HOMA-IR does not fully accurately reflect the IR of patients. CONCLUSIONS: The present study indicated that lnc RNA ENST00000550337.1 was related to PCOS occurrence, and elevated levels may be a risk factor for PCOS women. In addition, lncRNA ENST00000550337.1 might promote PCOS development partially by increasing IR and can be used as a potential molecular marker in patients with PCOS.

Correction to: Troxerutin protects against DHT-induced polycystic ovary syndrome in rats.

An amendment to this paper has been published and can be accessed via the original article.

Troxerutin protects against DHT-induced polycystic ovary syndrome in rats.

The exact pathogenesis of polycystic ovary syndrome (PCOS), the most common neuroendocrine disorder in women of reproductive age, has not been fully elucidated. Recent studies suggested that chronic inflammation and neurotransmitter disorder involved in the progress of PCOS. Troxerutin, a natural flavonoid, was reported to possess neuroprotective effect in several disease models by inhibiting inflammation or enhancing neurotrophic factor. In this study, we investigated the possible protective effect and mechanism of troxerutin in a dihydrotestosterone (DHT)-induced rat model of PCOS. The PCOS rat models were treated with troxerutin at a dose of 150 mg/kg or 300 mg/kg for up to 4 weeks. Results showed that 300 mg/kg troxerutin significantly decreased the body weight gain and improved the pathological changes of ovary induced by DHT. Meanwhile, the elevated gonadotrophin-releasing hormone (GnRH), gonadotrophin and testosterone in the serum of PCOS rats were reduced with the treatment of troxerutin. The expression of kisspeptin and NKB in arcuate nucleus and their receptors kiss1r and NK3r in GnRH positive neurons of median eminence were markedly decreased in troxerutin-treated rats. Of note, the GnRH inhibitory regulator GABA and stimulatory regulator glutamate were also restored to the normal level by troxerutin. The present study indicated that troxerutin may exhibit a protective effect in PCOS rat model via regulating neurotransmitter release.

Outcomes of the Chimney Technique for Endovascular Repair of Aortic Dissection Involving the Arch Branches.

BACKGROUND: The results of different chimney techniques in different zones of aortic arch were analyzed, so as to provide clues to decrease the complications of chimney thoracic endovascular repair (cTEVAR). METHODS: Between April 2012 and April 2017, 234 patients with aortic dissection involving arch branches received cTEVAR. Among these patients, 156 (66.7%) received single chimney (SC), 48 (20.5%) received double chimneys (DCs), and 30 (12.8%) received triple chimneys (TCs). A total of 342 chimney grafts (CGs) were used. RESULTS: All chimney grafts were successfully implanted, and no migration or occlusion was observed during follow-up. Mortality of this cohort was 1.7% (4/234). Three (3/234, 1.3%) patients suffered from cerebral vascular events. Seventy-five (75/234, 32.1%) had intraoperative type I endoleak. Twenty-seven (27/75, 36.0%) of them were found to have automatically disappeared in the follow-up period. The false lumens of 33 (33/75, 44.0%) were found to be stable, and 15 (15/75, 20%) were found to have expanded and were successfully retreated by endoleak embolization. The proximal tear located in zone 0 had higher instant endoleak rate than zone 1, zone 2, and zone 3 (P = 0.041; P = 0.042; P = 0.009). TC were found to have more instant endoleak than SC (P < 0.001) and DC (P = 0.012). But in the follow-up period, there was no significant difference. CONCLUSIONS: TCs and the proximal tear locating in zone 0 were found to be with higher instant endoleak rate, and it may need more rigorous follow-up. Some of the endoleak after cTEVAR could automatically disappear and some could be completely re-treated by gutter embolization procedure.

Repetitive Position Change Improves Gastric Cleanliness for Magnetically Controlled Capsule Gastroscopy.

BACKGROUND AND AIMS: Good gastric preparation is essential for magnetically controlled capsule gastroscopy (MCCG) examination. This study aims to determine if repetitive position change after dimethicone premedication could further improve gastric cleanliness for MCCG. METHODS: Consecutive patients referred for MCCG in our center from May 7 to May 31, 2018 were prospectively enrolled and randomized to undergo repetitive position change for 15 min (position change group) or not (conventional group) after ingesting dimethicone. Primary outcome was gastric cleanliness score and secondary outcomes were detection rate of positive findings, number of lesions per patient, gastric examination time, and safety of MCCG. RESULTS: Totals of 43 and 40 were included in the position change and conventional groups, respectively. Gastric cleanliness score in the position change group was significantly higher than in the conventional group (21.2 ± 1.0 vs. 18.6 ± 2.0, P  < 0.001), as was the proportion of acceptable gastric cleanliness (gastric cleanliness score ≥ 18) (100% vs. 72.5%, P  < 0.001). There was no statistical difference in detection rate of positive findings between the two groups (27.9% vs. 27.5%, P  = 0.97). In the position change group, the gastric examination time was significantly reduced (13.2 ± 4.0 vs. 15.3 ± 5.1, P = 0.043). No adverse events were observed. CONCLUSIONS: Repetitive position change after dimethicone premedication significantly improves gastric cleanliness for MCCG examination. Clinical Trial Registration ClinicalTrials.gov, ID: NCT03514966.

Variation of DNA Methylome of Zebrafish Cells under Cold Pressure.

DNA methylation is an essential epigenetic mechanism involved in multiple biological processes. However, the relationship between DNA methylation and cold acclimation remains poorly understood. In this study, Methylated DNA Immunoprecipitation Sequencing (MeDIP-seq) was performed to reveal a genome-wide methylation profile of zebrafish (Danio rerio) embryonic fibroblast cells (ZF4) and its variation under cold pressure. MeDIP-seq assay was conducted with ZF4 cells cultured at appropriate temperature of 28°C and at low temperature of 18°C for 5 (short-term) and 30 (long-term) days, respectively. Our data showed that DNA methylation level of whole genome increased after a short-term cold exposure and decreased after a long-term cold exposure. It is interesting that metabolism of folate pathway is significantly hypomethylated after short-term cold exposure, which is consistent with the increased DNA methylation level. 21% of methylation peaks were significantly altered after cold treatment. About 8% of altered DNA methylation peaks are located in promoter regions, while the majority of them are located in non-coding regions. Methylation of genes involved in multiple cold responsive biological processes were significantly affected, such as anti-oxidant system, apoptosis, development, chromatin modifying and immune system suggesting that those processes are responsive to cold stress through regulation of DNA methylation. Our data indicate the involvement of DNA methylation in cellular response to cold pressure, and put a new insight into the genome-wide epigenetic regulation under cold pressure.

Three-Component Functionalized Dihydropyridine Synthesis via a Formal Inverse Electron-Demand Hetero-Diels-Alder Reaction.

A mild three-component synthetic approach to versatile 2-amino-1,4-dihydropyridines from terminal alkynes, sulfonyl azides, and N-sulfonyl-1-aza-1,3-butadienes was successfully developed and relied on the in situ generation of metalated ynamide intermediates Ib to achieve a formal inverse electron-demand hetero-Diels-Alder reaction. Experimental results suggest that alkali metal cations (Li(+) and Cs(+) ions) might play a critical role to achieve the cycloaddition process.

Prenatal androgen excess enhances stimulation of the GNRH pulse in pubertal female rats.

In adolescent girls with polycystic ovary syndrome (PCOS), neuroendocrine derangements manifest after the onset of puberty, characterized by rapid LH pulse frequency. The early mechanism underlying the pubertal regulation of the GNRH/LH pulsatile release in adolescents with PCOS remains uncertain. To determine the effects of prenatal androgen exposure on the activation of GNRH neurons and generation of LH pulse at puberty, we administrated 5α-dihydrotestosterone to pregnant rats and observed serum LH levels and expression of hypothalamic genes in female offspring from postnatal 4 to 8 weeks. The 6-week-old prenatally androgenized (PNA) female rats exhibited an increase in LH pulse frequency. The hypothalamic expression of neurokinin B (Nkb (Tac2)) and Lepr mRNA levels in PNA rats increased remarkably before puberty and remained high during puberty, whereas elevated Kiss1 mRNA levels were detected only after the onset of puberty. Exogenous kisspeptin, NK3R agonist, and leptin triggered tonic stimulation of GNRH neurons and increased LH secretion in 6-week-old PNA rats. Leptin upregulated Kiss1 mRNA levels in the hypothalamus of pubertal PNA rats; however, pretreatment with a kisspeptin antagonist failed to suppress the elevated serum LH stimulated by leptin, indicating that the stimulatory effects of leptin may be conveyed indirectly to GNRH neurons via other neural components within the GNRH neuronal network, rather than through the kisspeptin-GPR54 pathway. These findings validate the hypotheses that NKB and leptin play an essential role in the activation of GNRH neurons and initiation of increased LH pulse frequency in PNA female rats at puberty and that kisspeptin may coordinate their stimulatory effects on LH release.

Synthesis, antifungal activities and qualitative structure activity relationship of carabrone hydrazone derivatives as potential antifungal agents.

Aimed at developing novel fungicides for relieving the ever-increasing pressure of agricultural production caused by phytopathogenic fungi, 28 new hydrazone derivatives of carabrone, a natural bioactive sesquisterpene, in three types were designed, synthesized and their antifungal activities against Botrytis cinerea and Colletotrichum lagenarium were evaluated. The result revealed that all the derivatives synthesized exhibited considerable antifungal activities in vitro and in vivo, which led to the improved activities for carabrone and its analogues and further confirmed their potential as antifungal agents.

Prenatal androgen excess programs metabolic derangements in pubertal female rats.

Owing to the heterogeneity in the clinical symptoms of polycystic ovary syndrome (PCOS), the early pathophysiological mechanisms of PCOS remain unclear. Clinical, experimental, and genetic evidence supports an interaction between genetic susceptibility and the influence of maternal environment in the pathogenesis of PCOS. To determine whether prenatal androgen exposure induced PCOS-related metabolic derangements during pubertal development, we administrated 5α-dihydrotestosterone (DHT) in pregnant rats and observed their female offspring from postnatal 4 to 8 weeks. The prenatally androgenized (PNA) rats exhibited more numerous total follicles, cystic follicles, and atretic follicles than the controls. Fasting glucose, insulin, leptin levels, and homeostatic model assessment for insulin resistance were elevated in the PNA rats at the age of 5-8 weeks. Following intraperitoneal glucose tolerance tests, glucose and insulin levels did not differ between two groups; however, the PNA rats showed significantly higher 30- and 60-min glucose levels than the controls after insulin stimulation during 5-8 weeks. In addition, prenatal DHT treatment significantly decreased insulin-stimulated phosphorylation of AKT in the skeletal muscles of 6-week-old PNA rats. The abundance of IR substrate 1 (IRS1) and IRS2 was decreased in the skeletal muscles and liver after stimulation with insulin in the PNA group, whereas phosphorylation of insulin-signaling proteins was unaltered in the adipose tissue. These findings validate the contribution of prenatal androgen excess to metabolic derangements in pubertal female rats, and the impaired insulin signaling through IRS and AKT may result in the peripheral insulin resistance during pubertal development.

Preparation of lipid aspirin sustained-release pellets by solvent-free extrusion/spheronization and an investigation of their stability.

A novel solvent-free extrusion/spheronization technique was investigated for preparing stable aspirin sustained-release pellets. Lipids as binders and the matrix in this technique were extruded below their melting points, and spheronized in a thermomechanical process. Four types of lipids (adeps solidus, Compritol(®) 888 ATO, Precirol(®) ATO5 and Compritol(®) HD5 ATO) and their admixture in different ratios were used to obtain spherical and extended-release pellets. Pellets containing 80% aspirin, 15% adeps solidus and 5% Compritol(®) 888 ATO had the best spherical geometry and met the dissolution requirements of aspirin extended-release tablets in USP 31. Storage stability studies showed that the content of free salicylic acid increased sharply in the traditional pellets produced by wet extrusion/spheronization, from 1.91 to 7.84%, whereas there was little increase in the lipid pellets (from 0.48 to 1.08%). The dissolution rate from the optimal pellets (F11) stored at 26°C did not change, but became faster at 40°C/RH75% after 5 months. Powder X-ray diffraction, scanning electron microscopy (SEM) and differential scanning calorimetry were used to investigate the physical properties of the pellets during stability testing. The increase in the rate of drug release from aged pellets (40°C/RH75%) may result from the partially melted adeps solidus observed in SEM photographs. This study suggests that it is possible to prepare sustained-release pellets by solvent-free extrusion/spheronization using an appropriate mixture of lipids with high stability. In particular, this novel technique is excellent for hygroscopic drugs.

[Effects of PI3K/Akt/NF-κB signal pathway on FSH facilitation on cell proliferation and invasion by human epithelial ovarian cancer].

OBJECTIVE: To explore the effects of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/nuclear factor-kappa B (NF-κB) signal pathway on the process of follicle-stimulating hormone (FSH) facilitating cell proliferation and invasion in human epithelial ovarian cancer. METHODS: Ovarian cancer cell lines SKOV3 and 3AO were cultured to exponential phase, then assigned to control group, FSH group, LY294002 group and FSH + LY294002 group, respectively. Cells were treated with different concentration of FSH and LY294002, respectively. The effects of FSH on cell proliferation were observed by methylthiazolyl tetrazolium (MTT). Morphological changes were observed by phase contrast microscope. The ability of cell invasion was investigated by transwell invasion assay. The expression of FSH receptor (FSHR), Akt1/2, phosphorylated-Akt (p-Akt) and NF-κB p65 protein were detected by western blot. RESULTS: (1) FSH could promote the proliferation of SKOV3 and 3AO cells. When the cells were treated with 40 U/L FSH for 48 hours (SKOV3) and 24 hours (3AO), compared with those in control groups, they reached the highest proliferation rate (P < 0.05), respectively. (2) The morphology of SKOV3 and 3AO cells in four groups:in control group, SKOV3 cells were short spindle and 3AO cells were long spindle, the nuclei of them were both roundness or oval, the cytoplasm were bright. In FSH group, the cells changed to slightly longer or polygonal, they were full in shape, meanwhile, the cell intensity were higher than control group. In LY294002 group, some cells changed from spindle to round, and began to shrink. The cell intensity diminished. The morphology of FSH + LY294002 group was similar with control group, but the cell intensity was lower than that in FSH group. (3) The number of SKOV3 cell that passed through the membrane in control group, FSH group, LY294002 group and FSH + LY294002 group was (26 ± 6), (118 ± 19), (18 ± 5) and (38 ± 7), respectively. The number of 3AO cell was (19 ± 4), (134 ± 20), (12 ± 3) and (58 ± 11), respectively. The results showed that the number of cells in FSH group was significantly higher than that in control group (P < 0.05), while the number of cell in FSH + LY294002 group was significantly fewer than that in FSH group (P < 0.05). (4) There was no significant difference in the expression of FSHR and Akt1/2 between FSH group and control group (P > 0.05), but FSH increased the expression of p-Akt and the ratio of NF-κB p65 in the nucleus versus cytoplasm in SKOV3 and 3AO cells, there were significant differences compared with control group (P < 0.05). LY294002 reversed the effects of FSH on increasing the expression of p-Akt and the ratio of NF-κB p65 in the nucleus versus cytoplasm, there were significant differences among LY294002 group, FSH + LY294002 group and FSH group (P < 0.05). CONCLUSION: The effects of FSH on proliferation and invasion of ovarian cancer cell lines SKOV3 and 3AO may be realized by regulating the activity of NF-κB in PI3K/Akt signal pathway.