Oteseconazole versus fluconazole for the treatment of severe vulvovaginal candidiasis: a multicenter, randomized, double-blinded, phase 3 trial.

Vulvovaginal candidiasis (VVC) is a common condition among women. Fluconazole remains the dominant treatment option for VVC. Oteseconazole is a highly selective inhibitor of fungal CYP51. This randomized, double-blinded, phase 3 trial was conducted to evaluate the efficacy and safety of oteseconazole compared with fluconazole in treating severe VVC. Female subjects presenting with vulvovaginal signs and symptoms score of ≥7 and positive Candida infection determined by potassium hydroxide test or Gram staining were randomly assigned to receive oteseconazole (600 mg on D1 and 450 mg on D2) or fluconazole (150 mg on D1 and D4) in a 1:1 ratio. The primary endpoint was the proportion of subjects achieving therapeutic cure [defined as achieving both clinical cure (absence of signs and symptoms of VVC) and mycological cure (negative culture of Candida species)] at D28. A total of 322 subjects were randomized and 321 subjects were treated. At D28, a statistically significantly higher proportion of subjects achieved therapeutic cure in the oteseconazole group than in the fluconazole group (66.88% vs 45.91%; P = 0.0002). Oteseconazole treatment resulted in an increased proportion of subjects achieving mycological cure (82.50% vs 59.12%; P < 0.0001) and clinical cure (71.25% vs 55.97%; P = 0.0046) compared with fluconazole. The incidence of treatment-emergent adverse events was similar between the two groups. No subjects discontinued study treatment or withdrew study due to adverse events. Oteseconazole showed statistically significant and clinically meaningful superiority over fluconazole for the treatment of severe VVC and was generally tolerated.

[Comparison of initiation of antihypertensive therapy strategies for primary prevention of cardiovascular diseases in Chinese population: A decision-analytic Markov modelling study].

OBJECTIVE: To evaluate the health benefits and intervention efficiency of different strategies of initiating antihypertensive therapy for the primary prevention of cardiovascular diseases in a community-based Chinese population from the Chinese electronic health records research in Yinzhou (CHERRY) study. METHODS: A decision-analytic Markov model was used to simulate and compare different antihypertensive initiation strategies, including: Strategy 1, initiation of antihypertensive therapy for Chinese adults with systolic blood pressure (SBP) ≥140 mmHg (2020 Chinese guideline on the primary prevention of cardiovascular diseases); Strategy 2, initiation of antihypertensive therapy for Chinese adults with SBP ≥130 mmHg; Strategy 3, initiation of antihypertensive therapy for Chinese adults with SBP≥140 mmHg, or with SBP between 130 and 140 mmHg and at high risk of cardiovascular diseases (2017 American College of Cardiology/American Heart Association guideline for the prevention, detection, evaluation, and management of high blood pressure in adults); Strategy 4, initiation of antihypertensive therapy for Chinese adults with SBP≥160 mmHg, or with SBP between 140 and 160 mmHg and at high risk of cardiovascular diseases (2019 United Kingdom National Institute for Health and Care Excellence guideline for the hypertension in adults: Diagnosis and management). The high 10-year cardiovascular risk was defined as the predicted risk over 10% based on the 2019 World Health Organization cardiovascular disease risk charts. Different strategies were simulated by the Markov model for ten years (cycles), with parameters mainly from the CHERRY study or published literature. After ten cycles of simulation, the numbers of quality-adjusted life years (QALY), cardiovascular events and all-cause deaths were calculated to evaluate the health benefits of each strategy, and the numbers needed to treat (NNT) for each cardiovascular event or all-cause death could be prevented were calculated to assess the intervention efficiency. One-way sensitivity analysis on the uncertainty of incidence rates of cardiovascular disease and probabilistic sensitivity analysis on the uncertainty of hazard ratios of interventions were conducted. RESULTS: A total of 213 987 Chinese adults aged 35-79 years without cardiovascular diseases were included. Compared with strategy 1, the number of cardiovascular events that could be prevented in strategy 2 increased by 666 (95% UI: 334-975), while the NNT per cardiovascular event prevented increased by 10 (95% UI: 7-20). In contrast to strategy 1, the number of cardiovascular events that could be prevented in strategy 3 increased by 388 (95% UI: 194-569), and the NNT per cardiovascular event prevented decreased by 6 (95% UI: 4-12), suggesting that strategy 3 had better health benefits and intervention efficiency. Compared to strategy 1, although the number of cardiovascular events that could be prevented decreased by 193 (95% UI: 98-281) in strategy 4, the NNT per cardiovascular event prevented decreased by 18 (95% UI: 13-37) with better efficiency. The results were consistent in the sensitivity analyses. CONCLUSION: When initiating antihypertensive therapy in an economically developed area of China, the strategy combined with cardiovascular risk assessment is more efficient than those purely based on the SBP threshold. The cardiovascular risk assessment strategy with different SBP thresholds is suggested to balance health benefits and intervention efficiency in diverse populations.

Reduced levels of actin gamma 1 predict poor prognosis in ovarian cancer patients.

AIM: To investigate the expression and function of actin gamma 1 (ACTG1) in ovarian cancer. METHODS: We performed immunohistochemical staining of 176 ovarian cancer tissue samples in a human tissue microarray to detect expression of ACTG1. Staining intensity was examined in relation to clinicopathological parameters. To investigate the prognostic value of ACTG1, ACTG1 mRNA data from 300 ovarian cancer patients in The Cancer Genome Atlas database were examined. RESULTS: The immunohistochemical results demonstrated that levels of ACTG1 were reduced in the samples of human ovarian cancer tissue that were examined, and the levels negatively correlated with various clinicopathological parameters. Levels of ACTG1 mRNA also negatively correlated with clinical stage. In Kaplan-Meier survival analysis, higher levels of ACTG1 mRNA were associated with improved overall survival. In multivariate analysis by Cox regression, ACTG1 expression was identified as an independent prognostic marker of favorable overall survival. CONCLUSION: ACTG1 may represent a valuable marker for the prognosis of ovarian cancer, and further studies of ACTG1 are warranted.

MAP2K6-FP Enhances the Sensitiveness of Paclitaxel for Ovarian Cancer via Inducing Autophagy.

BACKGROUND: Paclitaxel is recommended as a first-line chemotherapeutic agent against ovarian cancer, but drug resistance becomes a major limitation. The key molecule or mechanism associated with paclitaxel resistance in ovarian cancer still remains unclear. Recent studies have revealed an association between autophagy and drug resistance. METHODS: We previously synthesized a MAPK kinase-recombinant fusion protein, MAP2K6-FP, that contains 3 domains: a protein transduction domain TAT, a human ovarian cancer HO8910 cell-specific binding peptide, and a potential antitumor effector domain MKK6(E). In this study, we investigated the effect of MAP2K6-FP on HO8910 cells treated with paclitaxel. RESULTS: The IC50 (concentration by which 50% cell growth was inhibited) was 20 µM for paclitaxel alone, 1.5 µg/mL for MAP2K6-FP alone, and 0.3 µg/mL for MAP2K6-FP and 15 µM for paclitaxel if combined, respectively. In addition, immunohistochemistry assay demonstrated that tumor tissues from ovarian cancer patients showed higher expression of LC-3, the autophagy-related protein, compared with normal ovarian tissues. MAP2K6-FP (0, 2.5, 5, 10, 20, and 40 µg/mL) dose-dependently increased the LC-3 expression in HO8910 cells. Immunofluorescence assay showed that paclitaxel alone increased the expression of LC-3 in HO8910 cells, which was further enhanced by the combination with MAP2K6-FP. Downregulation of LC-3 expression using LC-3 small interfering RNA inhibited the cytotoxicity effect of MAP2K6-FP. Furthermore, either MAP2K6-FP alone or in combination with paclitaxel increased the ratio of expressions of Beclin-1/Bcl-2, another autophagy-related markers, compared with paclitaxel alone. CONCLUSIONS: MAP2K6-FP enhanced the sensitiveness of paclitaxel for ovarian cancer via inducing autophagy.

Phage display screening identifies a novel peptide to suppress ovarian cancer cells in vitro and in vivo in mouse models.

BACKGROUND: Ovarian cancer is a possibly lethal gynecological malignancy and this study utilized phage display technology to screen and identify peptides that specifically bind to ovarian cancer cells and explored the effects of these peptides on ovarian cancer cells in vitro and in vivo. METHODS: The phage displayed peptide library was used to isolate the peptides binding to and internalizing into the ovarian carcinoma cells. Positive phage clones were characterized with DNA sequencing and bioinformatics analysis and then validated with immunofluorescence. Subsequently, the selected peptides were investigated for their cancer-related functions, including cell adhesion, spreading, motility, and invasion in vitro and in vivo. RESULTS: Peptide1 read as SWQIGGNwas the positive peptide and showed preferential binding to the target cells. Peptide 1 also inhibited cell proliferation, migration, invasion and adhesion of ovarian cancer HO8910 cells in vitro. In vivo, Peptide 1 led to a lower tumorigenicity of HO8910 cells, which was characterized by the inhibitory effect on tumor growth and metastasis of ovarian cells. CONCLUSION: These studies demonstrate that the phage display-identified tumor cell-binding peptide was able to control ovarian cancer cell viability, migration, invasion, and adhesion capacity in vitro as well as tumor growth and metastasis in vivo. Future studies will be aimed at evaluating the clinical efficacy of the peptide SWQIGGN in ovarian cancer patients.

Delivery of Constitutively Active Mutant MKK6(E) With TAT-OSBP Induces Apoptosis in Human Ovarian Carcinoma HO8910 Cells.

Biologically active peptides and proteins are novel agents that show promise in the development of anticancer drugs. Their relatively low cell permeability and poor tumor selectivity, however, impede their widespread applicability. In this study, we evaluated the tumor selectivity, cellular internalization, and biological activity of a cell-permeable ovarian cancer cell-specific therapeutic protein consisting of TAT-OSBP and constitutively active MKK6(E), an upstream kinase of the p38 signaling pathway that mediates cellular apoptosis. OSBP, a 7-amino-acid peptide with high affinity for human ovarian cancer HO8910 cells, was conjugated to the cell-penetrating peptide (TAT) to form a tumor-selective peptide (TAT-OSBP), which was further conjugated with EGFP or MKK6(E). Flow cytometry and fluorescent microscopy were performed to evaluate the tumor-targeted penetration of TAT-OSBP-EGFP. The inhibitory effects of TAT-OSBP-MKK6(E) were determined by cell proliferation and apoptosis assays. The internalization efficiency of TAT-OSBP-EGFP was significantly higher than that of TAT-EGFP. TAT-OSBP-EGFP selectively penetrated HO8910 cells. TAT-OSBP-MKK6(E) fusion protein inhibited cancer cell growth to varying degrees, with the highest level of inhibition in HO8910 cells. Moreover, TAT-OSBP-MKK6(E) significantly induced apoptosis of HO8910 cells. However, there was no significant difference in apoptosis in the normal ovarian epithelial cells treated with either TAT-OSBP-MKK6(E) or TAT-MKK6(E). Our results demonstrate that TAT-OSBP-MKK6(E) is a novel artificially designed molecule, which induces apoptosis and selectively targets human ovarian carcinoma HO8910 cells. Our study provides novel insights that may aid in the development of a new generation of anticancer drugs.

[Screening of targeting peptides for highly metastatic human ovarian cancer cells and their effect on the biological behavior of ovarian cancer cells].

OBJECTIVE: To explore the effect of short peptides specifically binding to highly metastatic human ovarian cancer HO8910PM cells and their effect on the biological behavior of ovarian cancer cells. METHODS: The phage-displayed peptide library was used to isolate the peptides binding and internalizing into the HO8910PM cells. Positive phage clones were characterized with DNA sequencing and bioinformatics analysis. The positive phage clones specifically bound to HO8910 cells were validated with immunofluorescence detection and enzyme-linked immunosorbent assay (ELISA). Furthermore, selected peptides were investigated for their cancer-related functions, including cell adhesion, spreading, motility, and invasion in vitro and in nude mice in vivo. The apoptotic index was detected by TUNEL assay, and VEGF expression by immunohistochemistry. RESULTS: After 4 rounds of screening, apparent enrichment of phages was observed on the HO8910PM cells. ELISA assay showed that among the randomly selected 20 phage clones, 12 can specifically bind to HO8910PM cells. Immunofluorescence assay also showed that the selected positive phage clones can specifically bind to HO8910PM cells. The adherence test showed that the adherence rates of HO8910PM-peptide20, HO8910PM-peptide16 and HO8910PM cells were 49.0%, 96.8% and 100.0%, respectively. There was a significant difference between the cell adherence rates of HO8910PM-peptide20 and HO8910PM cells (P < 0.05). The peptide20 read as "THRVHLH" was a positive peptide and showed preferential binding to targeted cells. The peptide20 effectively inhibited tumor growth and metastasis in the nude mice, and the positive rates of VEGF protein in the tumor tissue of experimental, negative control and blank mice were 21.2%, 81.4% and 85.7%, respectively, showing that the positive rate of VEGF protein in the experimental group was significantly lower than that in the negative control and blank groups (P < 0.01), and the apoptotic index (AI) of the experimental group was (18.21 ± 2.49)%, significantly higher than the (3.76 ± 1.77)% in the negative control group and the (4.78 ± 1.57)% in the blank group (P < 0.01). CONCLUSIONS: A novel short peptide able to specifically bind to highly metastatic human ovarian cancer cells is successfully screened. It can effectively inhibit the growth, invasion and metastasis of ovarian cancer cells, and provides an ideal vector in targeted drug therapy for ovarian cancer.

Molecular mechanism of ferroptosis and its application in the treatment of clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is a common malignant tumor of the urinary tract, the incidence of which is continuously increasing and affects human health worldwide. Despite advances in existing treatments, treatment outcomes still need to be improved due to higher rates of postoperative recurrence, chemotherapy resistance, etc.; thus, there is an urgent need for innovative therapeutic approaches. Ferroptosis is a recently found type of regulated cell death that is characterized primarily by the buildup of lipid peroxidation products and fatal reactive oxygen species created by iron metabolism, which plays a crucial role in tumor progression and therapy.With the molecular mechanisms associated with ferroptosis being increasingly studied and refined, triggering ferroptosis by regulators that target ferroptosis and ccRCC may be the key to developing potential therapeutic strategies for ccRCC. Therefore, ferroptosis is expected to be a new breakthrough in treating ccRCC. This paper examines the mechanism of ferroptosis, the regulatory mechanism of ferroptosis in ccRCC, and the potential application of ferroptosis in combination with other therapies for the treatment of ccRCC. The goal is to offer novel perspectives for the research and clinical application of ferroptosis in the treatment of ccRCC.

[Effect and mechanism of gene therapy of lentivirus mediated RhoA shRNA on ovarian cancer xenograft in vivo].

OBJECTIVE: To investigate treatment effects of lentivirus mediated RhoA short hairpin RNA (shRNA) on xenograft tumor of ovarian cancer in nude mice in vivo and the underlying mechanism. METHODS: Human ovarian cancer cell line HO8910 were inoculated to establish subcutaneous xenograft model of human ovarian cancer. Tumor-bearing nude mice were assigned randomizely to three groups: Lenti-RhoA-sh group, Lenti- negative control (NC) group and phosphate buffered saline (PBS) group.lentivirus mediated RhoA shRNA, negative control lentivirus and PBS were respectively injected in the three groups. Effects of treatment were observed by tumor growth curve, tumor volume, tumor weight, and tumor inhibition rate. Xenograft tissues and liver, spleen, lung, and renal tissues were examined by hematoxylin and eosin (HE) staining or were detected by streptavidin-perosidase(SP)immunochemical method. The changes of RhoA gene expression in xenograft tissues after lentivirus mediated RhoA shRNA treated were also detected by real-time qPCR, immunochemistry and Western blot assay. Cell apoptosis in xenograft tissues were examined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method and apoptotic index (AI) were counted. RESULTS: Compared with Lenti-NC group and PBS group, the growth speed of xenograft in Lenti-RhoA-sh group delayed significantly after injection 9 days (P < 0.01) . Tumor volume (338 ± 114) mm(3) decreased significantly in the Lenti-RhoA-sh group when compared with those in Lenti-NC group (1190 ± 332) mm(3) and PBS group (1101 ± 396) mm(3) (P < 0.01) . Tumor weight (0.23 ± 0.11) g decreased significantly in the Lenti-RhoA-sh group when compared with Lenti-NC group (0.79 ± 0.19) g and PBS group (0.74 ± 0.17) g (P < 0.01) . Real-time qPCR result shown that the expression of RhoA mRNA (0.30 ± 0.05) decreased significantly in the Lenti-RhoA-sh group compared with Lenti-NC group (0.95 ± 0.06) and PBS group(1.00 ± 0.11; P < 0.01) .Western blot result showed that the expression level of RhoA protein decreased significantly in the Lenti-RhoA-sh group (0.14 ± 0.06) compared with those in Lenti-NC group(0.78 ± 0.14) and PBS group (0.75 ± 0.13;P < 0.01). TUNEL staining displayed that AI significantly increased in the Lenti-RhoA-sh group (20.9 ± 3.4) % compared with those in Lenti-NC group (5.2 ± 2.0) % and PBS group (6.0 ± 2.1) % (P < 0.01). CONCLUSION: Lentivirus mediated RhoA shRNA may be effectively down-regulate of the expression of RhoA, inhibit the growth of subcutaneous xenograft tumor of ovarian cancer in nude mice by increasing the cell apoptosis.

Goethite/montmorillonite adsorption coupled with electrocoagulation for improving fluoride removal from aqueous solutions.

With the increasing problem of fluoride pollution, it is urgent to find an efficient method to remove fluoride (F-). In this study, a new material goethite-montmorillonite-sorbent (GMS) was prepared and added into the electrocoagulation (EC) reaction to form a new pathway (EC/GMS) for the removal of fluoride. Scanning electron microscopy (SEM), Brunauer-Emmett-Teller (BET), Fourier-transform infrared (FT-IR), X-ray photoelectron spectroscopy (XPS) and other characterization methods were used to analyze the properties of GMS. The fluoride removal performance and mechanism of EC/GMS was studied. The results showed that GMS could provide numerous adsorption sites. EC/GMS could achieve a high removal efficiency of 95.98% and lower energy consumption of 0.58 kW h m-3 for 60 min. EC/GMS could achieve a removal efficiency of 99.47% after optimization by single-factor experiments and RSM-BBD optimal experiments. Meantime, the removal rate of the EC/GMS still reached over 87% after six cycles. The kinetic analysis indicated that the degradation pathways could also achieve a high removal rate for high fluoride-containing concentration solutions within a short time. The stretching vibration of C-F and C-O and the existence of F- revealed that the electrophoresis of the electrodes, adsorption of GMS, and co-precipitation of flocs were the main removal pathways, and the accelerating effect between the electrocoagulation and adsorption process was addressed. This study provides a new pathway for removing fluoride from aqueous environments.

Role of Toll-like receptor 4 in inflammation-induced preterm delivery.

The aim of the present study was to investigate the potential role of Toll-like receptor 4 (TLR4) in lipopolysaccharide (LPS)-induced preterm delivery. Intraperitoneal injection of LPS in the presence or absence of previous TLR4 blockade was performed to establish a murine model of preterm delivery. The incidences of preterm delivery and fetal death were calculated. Flow cytometry was performed to examine the percentages of blood CD45(+)CD86(+), CD3(+)CD69(+), CD19(+)CD69(+) and CD49b(+)CD69(+) cell subsets, and the percentages of placenta CD45(+)CD86(+), CD45(+)CD49b(+) and CD49b(+)CD69(+) cell subpopulations. In our study, an inflammation-induced preterm delivery model was established by intraperitoneal injection of LPS. Blocking TLR4 significantly decreased LPS-induced preterm delivery and fetal death. LPS treatment markedly up-regulated the percentages of blood CD45(+)CD86(+), CD3(+)CD69(+) and CD49b(+)CD69(+) cells, and of placenta CD45(+)CD86(+), CD45(+)CD49b(+) and CD49b(+)CD69(+) cells. TLR4 blockade almost completely abrogated LPS-induced elevated cell proportions. These data demonstrate that TLR4 plays a critical role in inflammation-induced preterm delivery.

Up-regulation of apoptosis by gonadotrophin-releasing hormone agonist in cultures of endometrial cells from women with symptomatic myomas.

BACKGROUND: The aim of the present study was to evaluate the effect of gonadotrophin-releasing hormone agonist (GnRH-a), which is widely used in the medical treatment of symptomatic myomas, on the rate of endometrial cell apoptosis in cultures from women with symptomatic myomas. METHODS: The study included 36 women with symptomatic myomas without endometrial hyperplasia or endometrial carcinoma, and 22 controls. Endometrial biopsy specimens were obtained from all subjects. Levels of apoptosis were examined in epithelial endometrial cell cultures before and after incubation with GnRH-a (triptorelin). The percentage of apoptotic cells was evaluated using the terminal deoxynucleotidyl transferase-mediated d-UTP nick end labeling assay and flow cytometry was used to evaluate Annexin V levels. RESULTS: Levels of spontaneous apoptosis were significantly lower in endometrial cultures from patients with symptomatic myomas than in those from control subjects (P < 0.01). Concentrations as low as 10(-7) M GnRH-a enhanced apoptosis in endometrial cultures from patients with symptomatic myomas (3.48% +/- 0.27% apoptotic cells in untreated samples and 25.45 +/- 0.95% in cells treated with 10(-7) M GnRH-a; P <0.01). The percentage of apoptotic cells also increased when cultures from control women were treated with GnRH-a (8.10 +/- 0.18% in untreated samples and 15.29 +/- 2.30% in treated samples; P <0.01). Levels of apoptosis were dependent on both dose of GnRH-a and time of treatment. CONCLUSIONS: GnRH-a stimulates apoptosis in endometrial cells from patients with symptomatic myomas and this could, at least in part, account for the therapeutic action of GnRH-a.

Efficacy of gonadotropin-releasing hormone agonist and an extended-interval dosing regimen in the treatment of patients with adenomyosis and endometriosis.

AIMS: To determine the effects of gonadotropin-releasing hormone agonist (GnRH-a) and an extended-interval dosing regimen in the treatment of patients with adenomyosis and endometriosis. METHODS: This was a prospective observational study in the setting of a hospital outpatient clinic. Seventy women suffering from adenomyosis and endometriosis were randomly divided into 2 groups: extended-interval dosing (experimental group) and conventional dosing (control group). METHODS: Patients in the experimental group received a 4-dose regimen (triptorelin 3.75 mg by intramuscular injection every 6 weeks for a total of 4 doses). The patients in the control group received a conventional regimen (1 injection every 4 weeks for a total of 6 doses). The main outcome measures were relief and recurrence of dysmenorrhea and related climacteric symptoms, reduction of uterine volume, and serum levels of 17-beta-oestradiol (E(2)), luteinizing hormone (LH), and follicle-stimulating hormone (FSH). RESULTS: The reliving rate of dysmenorrhea was 100% in patients treated with both the new regimen and the convention regimen after 6 months. The uterine volume was reduced 37.6% and 39.2%, respectively. And the levels of LH, FSH and E(2) were decreased significantly (p < 0.001). The E(2 )levels were reduced to the postmenopausal level. The hormone profile of the experimental group was similar to that of the control group (p > 0.05). CONCLUSION: The use of the extended-interval dosing regimen of triptorelin depot in patients with adenomyosis or endometriosis results in a consistent hypo-oestrogenised state, which is similar to that achieved by the conventional regimen. The new regimen reduces the cost of treatment.

Effect of first, second, and third trimester placental factors on CD4, CCR5, and CXCR4 expression in human peripheral blood lymphocytes.

OBJECTIVE: To investigate effect of first, second, and third trimester placental factors (PF) on CD4, CCR5, and CXCR4 expression in human peripheral blood lymphocytes (PBLs), and to explore their influence on human immunodeficiency virus (HIV) vertical transmission. METHODS: Human peripheral blood mononuclear cells (PBMCs) were treated with first, second, and third trimester PF (concentration 25%) respectively for 24 hours. The expression of CD4, CCR5, and CXCR4 in PBLs, and the percentages of CCR5(+), CXCR4(+)ìand CCR5(+)CXCR4(+) cells in peripheral blood CD4(+) lymphocytes were determined with flow cytometry. RESULTS: All trimester PFs reduced CCR5 expression in PBLs. The efficiency of the first trimester PF was higher than that of the second and third trimester PF. The percentage of CCR5(+) cells in peripheral blood CD4(+) lymphocytes of PF groups was significantly lower than that of the control group, and the percentage of CCR5(+) cells in peripheral blood CD4(+) lymphocytes of the first trimester PF group was significantly lower than that of the second and third trimester group. The percentages of CCR5(+)CXCR4(+) cells in peripheral blood CD4(+) lymphocytes of PF groups were significantly decreased as compared with the control group, and the percentage of CCR5(+)CXCR4(+) cells in peripheral blood CD4(+) lymphocytes of the first trimester PF group was significantly lower than that of the third trimester PF group. CONCLUSION: PF can reduce the expression of CCR5 in human PBLs and peripheral blood CD4(+) lymphocytes, indicating that PF might reduce R5 virus infection via preventing HIV entry, and might play an important role in reducing R5 virus intrauterine infection.

Protective effects of rutin on liver injury in type 2 diabetic db/db mice.

The aim of this study was to evaluate the protective effect of rutin on the liver of type 2 diabetic mice and explore the correlation mechanism. The db/db mice, selected as the type 2 diabetes mellitus (T2DM) models, have random blood glucose (RBG) and glucose level after 2 h of oral glucose loading of more than 16.7 mmol/L. After administration of 120 mg/kg or 60 mg/kg rutin, to T2DM mice, RBG, oral glucose tolerance, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum, and advanced glycation end products (AGEs) in vivo and vitro of different groups were detected. The liver pathological changes were observed under light and electron microscopy. Western blotting was used to detect the protein expression of insulin receptor substrate 2 (IRS-2) and phosphorylation of phosphatidylinositol 3 kinase (PI3K) on p85, Akt on Ser473, glycogen synthase kinase 3ß (GSK-3ß) on Ser9, real-time quantitative PCR was used to detect IRS-2 mRNA expression. Moreover, dynamically observing the effect of rutin on the generation of AGEs in non-enzymatic protein glycosylated system, Cell Counting Kit-8 (CCK-8) method was used to detect the effect of rutin on proliferation activity of HepG2 liver cells. The results showed that RBG and glucose levels of oral glucose tolerance test (OGTT) of mice in model group were significantly higher than that of normal group, which were significantly reduced after the rutin treatment. Rutin could reduce the ALT, AST activities and AGEs level in serum and potentiate the expression of IRS-2, P-PI3K (p85), P-Akt (Ser473), P-GSK-3ß (Ser9) protein and IRS-2 mRNA in the liver tissue of db/db mice. Moreover, rutin could significantly alleviate the structure disorder of liver, reduce the degeneration and necrosis of liver cells and formation of collagen fibers of db/db mice. The results in vitro also showed that rutin could obviously inhibit the generation of AGEs, and promoted the proliferation activity of high glucose-stimulating HepG2 cells. In general, the protective effect of rutin on the liver of T2DM may be mediated by facilitating signal transduction and activated state of insulin IRS-2/PI3K/Akt/GSK-3ß signal pathway, promoting hepatocyte proliferation, reducing blood glucose level and generation of AGEs.

Knockdown of RhoA expression alters ovarian cancer biological behavior in vitro and in nude mice.

RhoA regulates cell proliferation, migration, angiogenesis and gene expression. Altered RhoA activity contributes to cancer progression. The present study investigated the effects of RhoA knockdown on the regulation of ovarian cancer biological behavior in vitro and in nude mice. The expression of RhoA was knocked down using a lentivirus carrying RhoA short hairpin RNA (shRNA) in ovarian cancer cells and was confirmed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. The altered ovarian cancer biological behaviors were assayed by cell viability, terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling (TUNEL), migration, invasion, and nude mice tumorigenicity assays, while the altered gene expression was detected by RT-qPCR and western blot analysis. The results showed that lentivirus-carrying RhoA shRNA significantly suppressed RhoA expression in ovarian cancer cells, which suppressed tumor cell viability, migration, invasion and adhesion in vitro. RhoA silencing also inhibited the tumorigenicity of ovarian cancer cells in nude mice, which was characterized by the suppression of tumor xenograft formation and growth and induction of tumor cell apoptosis. The results of the present study demonstrated that knockdown of RhoA expression had a significant antitumor effect on ovarian cancer cells in vitro and in nude mice, suggesting that RhoA may be a target for the development of a novel therapeutic strategy in the control of ovarian cancer.

TAT-OSBP-1-MKK6(E), a novel TAT-fusion protein with high selectivity for human ovarian cancer, exhibits anti-tumor activity.

To improve the selectivity of TAT-fusion proteins for targeted cancer therapy, we developed a novel TAT-based target-specific fusion protein, TAT-OSBP-1-MKK6(E), and evaluated its selectivity and anti-tumor activity in vitro and in vivo. The fusion protein containing TAT-OSBP-1-MKK6(E) has three functional domains: (1) the protein transduction domain of TAT, (2) the human ovarian cancer HO8910 cell-specific binding peptide (OSBP-1) and (3) the potential anti-tumor effector domain of MKK6(E). The transduction efficiency, selectivity, cytotoxicity and apoptotic effect of TAT-OSBP-1-MKK6(E) were examined using immunofluorescence, CCK8 assay and flow cytometry. The in vivo anti-tumor efficacy and target specificity of the fusion protein were evaluated using a nude mouse model with subcutaneous xenografts of human ovarian cancer HO8910 cells. Tumor-bearing mice were divided into three treatment groups that received tail vein injections of TAT-OSBP-1-MKK6(E), TAT-OSBP-1 or normal saline. Tumor growth inhibition was determined by tumor volume, weight and morphology. The distribution and apoptotic effect of TAT-OSBP-1-MKK6(E) were assessed by immunohistochemical staining and TUNEL assays. TAT-OSBP-1-MKK6(E) can be selectively internalized into human ovarian cancer HO8910 cells, rather than normal ovarian OSE cells. In vivo, the fusion protein was mainly expressed in the tumor xenograft, but not in ovary or liver tissues. As a result, TAT-OSBP-1-MKK6(E) significantly induced growth inhibition and apoptosis of tumor cells in vitro and in vivo, with limited effects in normal cells and tissues. TAT-OSBP-1-MKK6(E) treatment can selectively target HO8910 cells in vitro and in vivo, leading to growth inhibition and apoptosis of tumor cells. As such, TAT-OSBP-1-MKK6(E) may be a potential approach for ovarian cancer target therapy.

Mitogen-activated Protein Kinase Kinase 6-fusion Protein (MAP2K6-FP) Potentiates the Anti-tumor effects of Paclitaxel in Ovarian Cancer.

OBJECTIVE: To investigate the antitumor effects of a mitogen-activated protein kinase (MAPK) kinase fusion protein, TAT-OSBP-MKK6E (MAP2K6-FP), and paclitaxel as single agents and in combination against HO8910 human ovarian cancer cells. METHODS: We previously synthesized a MAPK kinase-recombinant fusion protein, MAP2K6-FP, that contains three domains: a protein transduction domain TAT, a human ovarian cancer HO8910 cell-specific binding peptide (OSBP), and a potential anti-tumor effector domain MKK6 (E). The HO8910 cells were exposed to MAP2K6-FP, paclitaxel, or both for 24 h. The antiproliferative effects were determined using the Cell Counting Kit-8 assay. Antitumor synergy was determined by computing the combination index. The in vivo antitumor effects of both drugs as single agents and in combination were tested using HO8910 cells implanted subcutaneously in female BALBC/c nude mice. TUNEL assay, immunohistochemical evaluation, and western blotting were performed to investigate the mechanism of action. RESULTS: A synergistic anti-proliferative effect was observed between MAP2K6-FP and paclitaxel at multiple drug concentrations, resulting in combination indices ranging from 0.3-0.85. In vivo testing against HO8910 cells in a xenograft tumor model indicated that both drugs were effective as single agents and that MAP2K6-FP and paclitaxel in combination had a synergistic antitumor effect. The combination treatment resulted in significantly altered caspase-3, vascular endothelial growth factor (VEGF), and proliferating cell nuclear antigen expression compared to treatment with the single agents (P<0.05). CONCLUSION: Both MAP2K6-FP and paclitaxel can inhibit cell proliferation and induce apoptosis in ovarian cancer HO8910 cells. Interestingly, the combination of MAP2K6-FP and paclitaxel had a synergistic antitumor effect on HO8910 cells, which induced apoptosis by increasing caspase-3 expression and decreasing VEGF expression.

[Comparison between lentivirus and plasmid as shRNA vector targeting RhoA gene of ovary cancer cell line HO8910].

OBJECTIVE: To compare the different effects of lentivirus and plasmid as shRNA vector targeting RAS homologue gene-family member A (RhoA) of ovary cancer cell line HO8910. METHODS: Lentivirus and plasmid vectors carrying siRNA targeting RhoA gene were respectively transferred into HO8910 cells. The transferred cells were screened with puromycin for the ones stably silencing RhoA gene. As passaging went on, the two types of vectors were compared in the ability of silencing RhoA gene expression by means of fluorescence microscopy, real-time quantitative PCR and Western blotting, and in the inhibitory effects on the invasion and metastasis of cells by scratch wound migration assay and tube formation assay. RESULTS: Two groups of stable RhoA-knockdown cell lines were established with lentivirus and plasmid as RNAi vectors, respectively. Detected by fluorescence microscopy, the expression rate of GFP decreased in the plasmid group as the culture generation increased; it was 70% and 45% at the 15th and 25th generations respectively. The expression rate of GFP in the lentivirus group maintained above 95%. Both real-time quantitative PCR and Western blotting indicated that the expression of RhoA mRNA and protein had no significant difference in the two groups at the 3rd generation; but as the culture generation increased, the expression of RhoA mRNA and protein in the lentivirus group kept at the lower level, while it increased in the plasmid group. The scratch wound migration assay and tube formation assay revealed that lentivirus as RNAi vector suppressed more stably and persistently the invasion and metastasis of ovary cancer cell line HO8910 as compared with the plasmid group (P<0.05). CONCLUSION: Lentivirus as RNAi vector can suppress the RhoA gene expression more stably as compared with the plasmid; the plasmid is more suitable for transient transfection studies.

[Screening of short peptides that bind specifically to ovarian cancer cell line through whole-cell subtraction biopanning].

OBJECTIVE: To obtain short peptides that bind specifically to ovarian carcinoma cell line HO8910 by whole-cell subtraction biopanning as an ideal vector for targeted drug delivery in ovarian cancer therapy. METHODS: With the HO8910 ovarian carcinoma cells as the target cells and human normal ovarian epithelial cells as the adherent cells, 4 rounds of panning from a PH.D-C7C phage-display peptide library were carried out. Individual phage clones were selected and identified by ELISA. Positive phage clones were characterized with DNA sequencing and bioinformatics analysis. The specific binding of the positive phage clones to HO8910 cells was tested with immunofluorescence cytochemistry. RESULTS: After 4 rounds of biopanning, phage clones showed preferential binding to the target cells. ELISA identified 12 positve phage clones from 20 randomly selected ones. Heptapentide (SWQIGGN) that bound specifically to ovarian cancer cells was obtained by DNA sequencing. The results of immunofluorescence cytochemistry indicated that phage1 could be specifically bound to HO8910 cells. CONCLUSION: By means of whole-cell subtraction biopanning, we found a novel heptapentide which was able to bind specifically to ovarian cancer cells.