NK-92MI Cells Engineered with Anti-claudin-6 Chimeric Antigen Receptors in Immunotherapy for Ovarian Cancer.

Background: The application of chimeric antigen receptor (CAR) NK cells in solid tumors is hindered by lack of tumor-specific targets and inefficient CAR-NK cell efficacy. Claudin-6 (CLDN6) has been reported to be overexpressed in ovarian cancer and may be an attractive target for CAR-NK cells immunotherapy. However, the feasibility of using anti-CLDN6 CAR-NK cells to treat ovarian cancer remains to be explored. Methods: CLDN6 expression in primary human ovarian cancer, normal tissues and cell lines were detected by immunohistochemistry and western blot. Two types of third-generation CAR NK-92MI cells targeting CLDN6, CLDN6-CAR1 NK-92MI cells with domains containing self-activated elements (NKG2D, 2B4) and CLDN6-CAR2 NK-92MI cells with classical domains (CD28, 4-1BB) were constructed by lentivirus transfection, sorted by flow cytometry and verified by western blot and qPCR. OVCAR-3, SK-OV-3, A2780, Hey and PC-3 cells expressing the GFP and luciferase genes were transduced. Subcutaneous and intraperitoneal tumor models were established via NSG mice. The ability of CLDN6-CAR NK cells to kill CLDN6-positive ovarian cancer cells were evaluated in vitro and in vivo by live cell imaging and bioluminescence imaging. Results: Both CLDN6-CAR1 and CLDN6-CAR2 NK-92MI cells could specifically killed CLDN6-positive ovarian cancer cells (OVCAR-3, SK-OV-3, A2780 and Hey), rather than CLDN6 negative cell (PC-3), in vitro. CLDN6-CAR1 NK-92MI cells with domains containing self-activated elements (NKG2D, 2B4) exhibited stronger cytotoxicity than CLDN6-CAR2 NK-92MI cells with classical domains (CD28, 4-1BB). Furthermore, CLDN6-CAR1 NK cells could effectively eliminate ovarian cancer cells in subcutaneous and intraperitoneal tumor models. More importantly, CAR-NK cells combined with immune checkpoint inhibitors, anti-PD-L1, could synergistically enhance the antitumor efficacy of CLDN6-targeted CAR-NK cells. Conclusions: These results indicate that CLDN6-CAR NK cells possess strong antitumor activity and represent a promising immunotherapeutic modality for ovarian cancer.

The Behavioral Adaptations and Barriers of Patients Employing Non-Pharmacological Strategies for Cancer Pain Management-A Qualitative Study.

The use of non-pharmacological strategies to complement pharmacological approaches can enhance cancer pain management by promoting patient autonomy and increasing management effectiveness. This study aimed to explore the required behavioral adaptations and situational barriers that cancer patients encounter when utilizing non-pharmacological strategies to manage pain. We adopted an exploratory-descriptive qualitative research approach, purposive sampling, and semi-structured interview guidelines to conduct face-to-face interviews with 18 cancer patients experiencing moderate or severe levels of worst pain. Data were analyzed using inductive content analysis to explore patients' experiences. Five themes described the behavioral adaptations of patients using non-pharmacological strategies to deal with cancer pain: finding complementary therapies, utilizing assistive skills, adapting to assistive skills, diverting attention, and seeking help. Situational barriers faced by patients include being in the workplace or in a climate-affected environment. Behavioral adaptation is necessary for non-pharmacological strategies to coping with cancer pain. The behavioral skills can help the patients to overcome situational barriers to engagement with these strategies. Thus, health professionals are expected to help the patients acquire adequate behavioral adaptation and skills for self-pain management, and assess the effectiveness of the strategies.

Paris saponin â induce toxicity in zebrafish by up-regulation of p53 pathway and down-regulation of wnt pathway.

Paris saponin I, II, and VII are three important components in Paris polyphylla, which have been widely studied as tumor cytotoxic drugs, but their safety in vivo has not been reported. Therefore, this study evaluated the safety of these three drugs based on the zebrafish model. Firstly, the lethality curves and lethal concentration of 50% (LC50) values of the three saponins were determined and the results showed the values of LC50 of Paris saponin I, II, and VII were 122.2, 210.7, 566.2 ng/mL, respectively. And then our data revealed that Paris saponin I, II and VII had definite hepatotoxicity, as shown by their significant reduction in the liver area and fluorescence intensity of zebrafish. Besides, Paris saponin Ⅰ affected the heart rate of zebrafish obviously, suggesting its cardiovascular toxicity. Afterwards, we found Paris saponin Ⅰ and Ⅶ reduced the area and fluorescence intensity of kidney in zebrafish, and had mild nephrotoxicity. And when treated with Paris saponin I, the pathological section of liver tissue in zebrafish showed vacuoles, severe necrosis of hepatocytes, and then the apoptosis of hepatocytes could be observed by TUNEL staining. Eventually, we found that the genes expression of p53, Bax and ß-catenin changed significantly in the administration group of Paris saponin I. In general, our study proved Paris saponin Ⅰ was the most toxic of the three saponins, and the most definite toxic target sites were liver and cardiovascular. And it was further inferred that the totoxicity of Paris saponin Ⅰ may be related to the regulation of p53 pathway and Wnt pathway. These results above showed the toxicity of the three saponins in zebrafish, suggesting their safety should be paid more attention in the future.

Genetic Signature of Pinctada fucata Inferred from Population Genomics: Source Tracking of the Invasion in Mischief Reef of Nansha Islands.

Among the anthropogenic stresses that marine ecosystems face, biological invasions are one of the major threats. Recently, as a result of increasingly intense anthropogenic disturbance, numerous marine species have been introduced to their non-native ranges. However, many introduced species have uncertain original sources. This prevents the design and establishment of methods for controlling or preventing these introduced species. In the present study, genomic sequencing and population genetic analysis were performed to detect the geographic origin of the introduced Pinctada fucata population in the Mischief Reef of the South China Sea. The results of population genetic structure analysis showed a close relationship between the Mischief Reef introduced population and the Lingshui population, indicating that Lingshui may be the potential geographical origin. Furthermore, lower heterozygosity and nucleotide diversity were observed in the introduced population in Mischief Reef, indicating lower genetic diversity than in other native populations. We also identified some selected genomic regions and genes of the introduced population, including genes related to temperature and salinity tolerance. These genes may play important roles in the adaptation of the introduced population. Our study will improve our understanding of the invasion history of the P. fucata population. Furthermore, the results of the present study will also facilitate further control and prevention of invasion in Mischief Reef, South China Sea.

Remodelling of tumour microenvironment by microwave ablation potentiates immunotherapy of AXL-specific CAR T cells against non-small cell lung cancer.

The complex immunosuppressive tumour microenvironment (TME) and lack of tumour-specific targets hinder the application of chimeric antigen receptor (CAR) T cells in the treatment of solid tumours. Combining local treatment with CAR T cell immunotherapy may regulate the TME and enhance the killing potency of CAR T cells in solid tumours. Here, we show that AXL, which is highly expressed in non-small cell lung cancer (NSCLC) but not in normal tissues, might be a target for CAR T cell therapy. AXL-CAR T cells alone cause moderate tumour regression in subcutaneous and pulmonary metastatic lung cancer cell-derived xenograft models. Combination of microwave ablation (MWA) and AXL-CAR T cells have superior antitumour efficacy. MWA enhances the activation, infiltration, persistence and tumour suppressive properties of AXL-CAR T cells in AXL-positive NSCLC patient-derived xenograft tumours via TME remodelling. The combination therapy increases the mitochondrial oxidative metabolism of tumour-infiltrating CAR T cells. Combination treatment induces significant tumour suppression without observed toxicities in humanized immunocompetent mice. The synergistic therapeutic effect of MWA and AXL-CAR T cells may be valuable for NSCLC treatment.

Transcriptomic analysis of male three-spot swimming crab (Portunus sanguinolentus) infected with the parasitic barnacle Diplothylacus sinensis.

Diplothylacus sinensis is reported as an intriguing parasitic barnacle that can negatively affect the growth, molting, reproduction in several commercially important portunid crabs. To better understand the molecular mechanisms of host-parasite interactions, we characterized the gene expression profiles from the healthy and D. sinensis infected Portunus sanguinolentus by high-through sequence method. Totally, the transcriptomic analysis generated 52, 266, 600 and 51, 629, 604 high quality reads from the infected and control groups, respectively. The clean reads were assembled to 90,740 and 69,314 unigenes, with the average length of 760 bp and 709 bp, respectively. The expression analysis showed that 18,959 genes were significantly changed by the parasitism of D. sinensis, including 4769 activated genes and 14,190 suppressed genes. The differentially expressed genes were categorized into 258 KEGG pathways and 647 GO terms. The GO analysis mapped 13 DEGs related to immune system process and 32 DEGs related to immune response, respectively, suggesting a potential alteration of transcriptional expression patterns in complement cascades of P. sanguinolentus. Additionally, 4 representative molting-related genes were down-regulated in parasitized group, indicating D. sinensis infection appeared to suppress the producing of ecdysteroid hormones. In conclusion, the present study improves our understanding on parasite-host interaction mechanisms, which focuses the function of Ecdysone receptor, Toll-like receptor and cytokine receptor of crustacean crabs infestation with rhizocephalan parasites.

Phototherapeutic effect of transformable peptides containing pheophorbide a on colorectal cancer.

Photodynamic therapy (PDT) and photothermal therapy (PTT) have attracted research interest for their noninvasive nature and selective treatment of tumor tissues. They are effective through the generation of reactive oxygen species (ROS) or heat. Nevertheless, several problems, including low bioavailability and long-lasting cutaneous photosensitivity, have limited their clinical application. In this study, we reported an in situ self-assembly strategy that could improve various biological properties of the photosensitizer in vivo. A photosensitizer connected to a receptor-mediated smart peptide can self-assemble into nanoparticles (NPs) under the force of hydrophobic interaction and then transform into a nanofibrillar network after attaching to the tumor cell surface with the help of the ß-sheet-forming peptide KLVFF. The supramolecular structural changes deeply affected the PDT and PTT properties of the photosensitizer on tumors. After being aggregated into the nanostructure, the water solubility and targeting ability of the photosensitizer was ameliorated. Moreover, the improvement of the photothermal conversion efficiency, ROS generation, and tumor retention followed the formation of nanofibrils (NFs). This self-assembly strategy showed the ability of supramolecular nanofibrils to improve the bioavailability of photosensitizers, which provides a new potential treatment avenue for various cancer therapies.

Renal Cancer Detection: Fusing Deep and Texture Features from Histopathology Images.

Histopathological images contain morphological markers of disease progression that have diagnostic and predictive values, with many computer-aided diagnosis systems using common deep learning methods that have been proposed to save time and labour. Even though deep learning methods are an end-to-end method, they perform exceptionally well given a large dataset and often show relatively inferior results for a small dataset. In contrast, traditional feature extraction methods have greater robustness and perform well with a small/medium dataset. Moreover, a texture representation-based global approach is commonly used to classify histological tissue images expect in explicit segmentation to extract the structure properties. Considering the scarcity of medical datasets and the usefulness of texture representation, we would like to integrate both the advantages of deep learning and traditional machine learning, i.e., texture representation. To accomplish this task, we proposed a classification model to detect renal cancer using a histopathology dataset by fusing the features from a deep learning model with the extracted texture feature descriptors. Here, five texture feature descriptors from three texture feature families were applied to complement Alex-Net for the extensive validation of the fusion between the deep features and texture features. The texture features are from (1) statistic feature family: histogram of gradient, gray-level cooccurrence matrix, and local binary pattern; (2) transform-based texture feature family: Gabor filters; and (3) model-based texture feature family: Markov random field. The final experimental results for classification outperformed both Alex-Net and a singular texture descriptor, showing the effectiveness of combining the deep features and texture features in renal cancer detection.

Retrospective analysis of crescent score in clinical prognosis of IgA nephropathy.

The scoring of crescents (Cs) was recently added to the Oxford classification for IgA nephropathy (IgAN). Because of the short-term use of the C score in clinical practice, its validity and applicability need to be verified. We, retrospectively, analyzed the clinicopathological data of 144 primary IgAN patients diagnosed at our hospital from March 2017 to March 2019 and with complete ≥6-month follow-up data. We found that the C score was positively correlated with the Lee's classification in the assessment of renal pathological changes and significantly correlated with increased proteinuria and decreased estimated glomerular filtration rate. Univariate Cox regression analysis showed an association of C formation with IgAN prognosis, and multivariate Cox regression indicated Cs as an independent prognosis factor. The optimal proportion of Cs for prognosis prediction by the receiver operating characteristic curve was 11%. Kaplan-Meier survival curve revealed a significantly decreased renal survival rate in patients with C proportions ≥11%. Further multivariate Cox regression analysis confirmed that the C proportion ≥11% is an independent risk factor for poor prognosis of IgAN patients. Our findings demonstrate that Cs are independently related to the prognosis of patients with IgAN, and the proportion of Cs ≥11% is an independent risk factor for poor outcomes.

CAR NK-92 cells targeting DLL3 kill effectively small cell lung cancer cells in vitro and in vivo.

Small cell lung cancer (SCLC) is characterized by a high relapse rate, drug tolerance, and limited treatment choices. Chimeric antigen receptor (CAR)-modified NK cells represent a promising immunotherapeutic modality for cancer treatment. However, their potential applications have not been explored in SCLC. Delta-like ligand 3 (DLL3) has been reported to be overexpressed in SCLC and may be a rational target for CAR NK immunotherapy. In this study, we developed DLL3-specific NK-92 cells and explored their potential in the treatment of SCLC. A coculture of DLL3+ SCLC cell lines with DLL3-CAR NK-92 cells exhibited significant in vitro cytotoxicity and cytokine production. DLL3-CAR NK-92 cells induced tumor regression in an H446-derived pulmonary metastasis tumor model under a good safety threshold. The potent antitumor activities of DLL3-CAR NK-92 cells were observed in subcutaneous tumor models of SCLC. Moreover, obvious tumor-infiltrated DLL3-CAR NK-92 cells were detected in DLL3+ SCLC xenografts. These findings indicate that DLL3-CAR NK-92 cells might be a potential strategy for the treatment of SCLC.

Construction of a Multifunctional Nano-Scale Metal-Organic Framework-Based Drug Delivery System for Targeted Cancer Therapy.

The antitumor activity of triptolide (TP) has received widespread attention, although its toxicity severely limits its clinical application. Therefore, the design of a targeted drug delivery system (TDDS) has important application prospects in tumor treatment. Metal-organic frameworks (MOFs), with high drug-carrying capacity and good biocompatibility, have aroused widespread interest for drug delivery systems. Herein, folic acid (FA) and 5-carboxylic acid fluorescein (5-FAM) were used to modify Fe-MIL-101 to construct a functionalized nano-platform (5-FAM/FA/TP@Fe-MIL-101) for the targeted delivery of the anti-tumor drug triptolide and realize in vivo fluorescence imaging. Compared with Fe-MIL-101, functionalized nanoparticles not only showed better targeted therapy efficiency, but also reduced the systemic toxicity of triptolide. In addition, the modification of 5-FAM facilitated fluorescence imaging of the tumor site and realized the construction of an integrated nano-platform for fluorescence imaging and treatment. Both in vitro and in vivo studies of functionalized nanoparticles have demonstrated excellent fluorescence imaging and synergistic targeting anticancer activity with negligible systemic toxicity. The development of functional nano-platform provides new ideas for the design of MOF-based multifunctional nano-drug delivery system, which can be used for precise treatment of tumor.

Development of mesothelin-specific CAR NK-92 cells for the treatment of gastric cancer.

Background: The application of chimeric antigen receptor (CAR) NK cells in solid tumors is hindered by lack of tumor-specific targets and inefficient CAR NK cell efficacy. It has been reported that mesothelin (MSLN) may be an ideal immunotherapy target for gastric cancer. However, the feasibility of using anti-MSLN CAR NK cells to treat gastric cancer remains to be studied. Methods: MSLN expression in primary human gastric cancer, normal tissues and cell lines were detected. MSLN and CD19 targeted CAR NK-92 (MSLN- and CD19-CAR NK) cells were constructed, purified and verified. N87, MKN-28, AGS and Huh-7 cells expressing the GFP and luciferase genes were transduced. Cell- and patient-derived xenograft (PDX) were established via NSG mice. The ability of MSLN-CAR NK cells to kill MSLN-positive gastric cancer cells were evaluated in vitro and in vivo. Results: MSLN-CAR NK cells can specifically kill MSLN-positive gastric cancer cells (N87, MKN-28 and AGS), rather than MSLN negative cell (Huh-7), in vitro. Moreover, compared with parental NK-92 cells and CD19-CAR NK cells, stronger cytokine secretions were secreted in MSLN-CAR NK cells cocultured with N87, MKN-28 and AGS. Furthermore, MSLN-CAR NK cells can effectively eliminate gastric cancer cells in both subcutaneous and intraperitoneal tumor models. They could also significantly prolong the survival of intraperitoneally tumor-bearing mice. More importantly, the potent antitumor effect and considerable NK cell infiltration were observed in the patient-derived xenograft treated with MSLN-CAR NK cells, which further warranted the therapeutic effects of MSLN-CAR NK cells to treat gastric cancer. Conclusion: These results demonstrate that MSLN-CAR NK cells possess strong antitumor activity and represent a promising therapeutic approach to gastric cancer.

IL-7 and CCL19-secreting CAR-T cell therapy for tumors with positive glypican-3 or mesothelin.

Although chimeric antigen receptor (CAR)-engineered T cells have shown great success in the treatment of B cell malignancies, this strategy has limited efficacy in patients with solid tumors. In mouse CAR-T cells, IL-7 and CCL19 expression have been demonstrated to improve T cell infiltration and CAR-T cell survival in mouse tumors. Therefore, in the current study, we engineered human CAR-T cells to secrete human IL-7 and CCL19 (7 × 19) and found that these 7 × 19 CAR-T cells showed enhanced capacities of expansion and migration in vitro. Furthermore, 7 × 19 CAR-T cells showed superior tumor suppression ability compared to conventional CAR-T cells in xenografts of hepatocellular carcinoma (HCC) cell lines, primary HCC tissue samples and pancreatic carcinoma (PC) cell lines. We then initiated a phase 1 clinical trial in advanced HCC/PC/ovarian carcinoma (OC) patients with glypican-3 (GPC3) or mesothelin (MSLN) expression. In a patient with advanced HCC, anti-GPC3-7 × 19 CAR-T treatment resulted in complete tumor disappearance 30 days post intratumor injection. In a patient with advanced PC, anti-MSLN-7 × 19 CAR-T treatment resulted in almost complete tumor disappearance 240 days post-intravenous infusion. Our results demonstrated that the incorporation of 7 × 19 into CAR-T cells significantly enhanced the antitumor activity against human solid tumor. Trial registration: NCT03198546. Registered 26 June 2017, https://clinicaltrials.gov/ct2/show/NCT03198546?term=NCT03198546&draw=2&rank=1.

Feasibility and safety of fine positioning needle-mediated breathing control in CT-guided percutaneous puncture of small lung/liver nodules adjacent to diaphragm.

To assess the efficacy, safety, and feasibility of a separate inserted positioning fine needle-mediated breathing-control technique applied to computed tomography (CT)-guided percutaneous puncture for biopsy or microwave ablation (MWA) of small lung/liver nodules near diaphragm. Total 46 patients with pulmonary/liver small nodules (≤ 3 cm in size) near diaphragm(nodule within 1 cm distance to the diaphragm)were undergone percutaneous biopsy ( n = 15) or MWA (n = 31) under the guidance of CT, and a separate positioning fine needle-mediated breathing-control technique was applied for the precise punctures. CT plain scan was performed to monitor the complications after the procedure. The patient baseline data, operation details, successful rate, major complications as well as radiation dose during the procedure were recorded and analyzed. With the assistance of a fine positioning needle insertion for controlling the breathing, the puncture success rate for biopsy or MWA reached 91.30% (42/46). For biopsy, the mean nodule diameter, nodule distance to the diaphragm, puncture time and radiation dose during CT scan were 2.27 cm ± 0.74, 0.61 cm ± 0.24, 18.67 min ± 6.23, 28.84 mSv ± 6.99, respectively; For MWA, the mean nodule diameter, nodule distance to the diaphragm, puncture time and CT radiation dose were 2.35 cm ± 0.64, 0.69 cm ± 0.23, 38.71 min ± 13.65, 33.02 mSv ± 8.77, respectively. Totally, there were three and four cases found minimal puncture-related hemoptysis and pneumothorax needed no additional treatments, respectively. We recently developed and verified a feasible, safe and highly effective puncture technique with reasonable radiation dose for CT-guided biopsy or MWA for small nodules abutting diaphragm, therefore worthy of extensive application to similar clinical situations.

Ginsenoside Rb1 Attenuates Triptolide-Induced Cytotoxicity in HL-7702 Cells via the Activation of Keap1/Nrf2/ARE Pathway.

Triptolide (TP) is the major bioactive compound extracted from Tripterygium wilfordii Hook F. It exerts anti-inflammatory, antirheumatic, antineoplastic, and neuroprotective effects. However, the severe hepatotoxicity induced by TP limits its clinical application. Ginsenoside Rb1 has been reported to possess potential hepatoprotective effects, but its mechanism has not been fully investigated. This study was aimed at investigating the effect of ginsenoside Rb1 against TP-induced cytotoxicity in HL-7702 cells, as well as the underlying mechanism. The results revealed that ginsenoside Rb1 effectively reversed TP-induced cytotoxicity in HL-7702 cells. Apoptosis induced by TP was suppressed by ginsenoside Rb1 via inhibition of death receptor-mediated apoptotic pathway and mitochondrial-dependent apoptotic pathway. Pretreatment with ginsenoside Rb1 significantly reduced Bax/Bcl-2 ratio and down-regulated the expression of Fas, cleaved poly ADP-ribose polymerase (PARP), cleaved caspase-3, and -9. Furthermore, ginsenoside Rb1 reversed TP-induced cell cycle arrest in HL-7702 cells at S and G2/M phase, via upregulation of the expressions of cyclin-dependent kinase 2 (CDK2), cyclin E, cyclin A, and downregulation of the expressions of p53, p21, and p-p53. Ginsenoside Rb1 increased glutathione (GSH) and superoxide dismutase (SOD) levels, but decreased the reactive oxygen species (ROS) and malondialdehyde (MDA) levels. Pretreatment with ginsenoside Rb1 enhanced the expression levels of nuclear factor-erythroid 2-related factor 2 (Nrf2), total Nrf2, NAD(P)H: quinone oxidoreductases-1 (NQO-1), heme oxygenase-1 (HO-1), and Kelch-like ECH-associated protein 1 (Keap1)/Nrf2 complex. Therefore, ginsenoside Rb1 effectively alleviates TP-induced cytotoxicity in HL-7702 cells through activation of the Keap1/Nrf2/ARE antioxidant pathway.

Use of chimeric antigen receptor NK-92 cells to target mesothelin in ovarian cancer.

Mesothelin (MSLN) has been reported to be overexpressed in ovarian cancer and may be an ideal target for immunotherapy. Recent studies have suggested that natural killer (NK) cells may be better chimeric antigen receptor (CAR) drivers because of their favorable innate characteristics, such as directly recognizing and killing tumor cells, resulting in a graft-versus-tumor effect but irresponsible for graft-versus-host disease (GVHD). The therapeutic effects of CAR-engineered NK cells targeting MSLN in ovarian cancer have not been evaluated. In this study, MSLN- and CD19-targeted CAR NK-92 (MSLN- and CD19-CAR NK) cells were constructed. Both MSLN- and CD19-CAR molecules were highly expressed on the surface of NK-92 cells following lentiviral gene transduction. MSLN-CAR NK cells specifically killed MSLN-positive ovarian cancer cells (OVCAR-3 and SK-OV-3), rather than MSLN-negative cells (SK-HEP-1), in vitro. Moreover, compared with parental NK-92 cells and CD19-CAR NK cells, stronger cytokine secretion was detected in MSLN-CAR NK cells cocultured with OVCAR-3 and SK-OV-3. Furthermore, MSLN-CAR NK cells effectively eliminated ovarian cancer cells in both subcutaneous and intraperitoneal tumor models; these cells also significantly prolonged the survival of intraperitoneally tumor-bearing mice. These results demonstrate that MSLN-CAR NK cells have robust specific antitumor activity, both in vitro and in vivo, suggesting that mesothelin could be a potential target for CAR NK cells and could be applied in the treatment of ovarian cancer.

Insulin Reduces Reaction of Follicular Granulosa Cells to FSH Stimulation in Women With Obesity-Related Infertility During IVF.

CONTEXT: Women with obesity usually need larger doses of FSH for ovarian stimulation, resulting in poor outcomes; however, the mechanism is still unclear. OBJECTIVE: To investigate the molecular regulation of FSH receptor (FSHR) expression associated with obesity. DESIGN: Case-control study to improve in vitro fertilization (IVF) outcomes. PATIENTS: Women with obesity (82) and women who were overweight (457) undergoing IVF and 1790 age-matched controls with normal weight from our reproductive medicine center. MAIN OUTCOME MEASURES: FSHR expression was decreased in parallel with body mass index (BMI), whereas the estradiol (E2) level on the human chorionic gonadotropin (hCG) trigger day was significantly lower. RESULTS: FSHR expression in human granulosa cells (hGCs), both mRNA (P = 0.02) and protein (P = 0.001) levels, was decreased in women who were overweight or obese. Both insulin (P < 0.001) and glucose (P = 0.0017) levels were positively correlated with BMI in fasting blood and follicle fluids (FFs) but not with FFs leptin level. We treated human granulosa-like tumor cells (KGN) cells with insulin; E2 production was compromised; the level of phosphorylated (p)-protein kinase B (p-Akt2) decreased, whereas p-glycogen synthase kinase 3 (GSK3) increased; and there were similar changes in hGCs from women with obesity. Stimulated hGCs from women with obesity with compound 21 (CP21), an inhibitor of GSK3ß, resulted in upregulated ß-catenin activation and increased FSHR expression. CP21 also increased the expression of insulin receptor substrate 1 and phosphatidylinositol 3-kinase (PI3K), as well as p-Akt2. CONCLUSIONS: Women with obesity in IVF were associated with reduced FSHR expression and E2 production caused by a dysfunctional insulin pathway. Decreased FSHR expression in hGCs from women with obesity and insulin-treated KGN cells could be rescued by an inhibitor of GSK3ß, which might be a potential target for the improvement of the impaired FSH-stimulation response in women with obesity.

The complete mitochondrial genome of milk shark, Rhizoprionodon acutus (Ruppell 1837).

The complete mitogenome sequence of the milk shark, of which Rhizoprionodon acutus was determined using long PCR. The genome was 16,693 bp in length and it contained 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes, and 1 D-loop region. The overall base composition of the heavy strand is A (31.63%), C (23.91%), T (31.38%), and G (13.08%). However, the base composition of A (31.38%), C (13.08%), T (31.63%), and G (23.91%) was in the light strand. Except for that COXI began with GTG, all the protein initiation codons were ATG. What's more, all the protein termination codons were TAA, in addition to ND2, ND3, and Cytb ended with TAG, COXII and ND4 finished only with T, and ND6 with AGG. Moreover, based on thirteen concatenated PCGs nucleotide datasets, the phylogenetic analysis showed high value support for the following sister clade among the two genera (Rhizoprionodon acutus, Rhizoprionodon; Scoliodon macrorhynchos and Scoliodon laticaudus, Scoliodon).

Pregnancy outcomes of PCOS overweight/obese patients after controlled ovarian stimulation with the GnRH antagonist protocol and frozen embryo transfer.

BACKGROUND: Overweight/obese women with polycystic ovary syndrome (PCOS) are at increased risk of subfertility and complications of pregnancy, compared with normal-weight women. To implement controlled ovarian hyperstimulation (COH), the improved efficacy of the gonadotrophin-releasing hormone antagonist (GnRH-ant) protocol has been demonstrated, as well as frozen embryo transfer (FET). OBJECTIVE: This retrospective study evaluated the pregnancy outcomes after combined GnRH-ant protocol and FET in overweight/obese women with PCOS, with reference to that of normal-weight women with PCOS. METHODS: Women with PCOS (n = 398) who underwent the GnRH-ant protocol for COH followed by FET, were stratified as normal-weight (BMI < 24 kg/m2) or overweight/obese (BMI ≥24 kg/m2). The outcomes of pregnancy were compared. RESULTS: The overweight/obese patients had significantly lower rates of embryo implantation (47.7%), live birth (47.8%), and live births of twins (10.9%) compared with the normal-weight group (58.4%, 60.8%, and 30.0%, respectively; P = 0.006, 0.015, and 0.000), while the rate of late abortion was significantly higher (11.0% cf. 3.8%, P = 0.030). BMI was the only significant factor affecting the probability of live birth. CONCLUSION: The pregnancy outcomes of overweight/obese women with PCOS after COH via the GnRH-ant protocol and FET remained at a significant deficit compared with that of normal-weight women with PCOS.

Nucleolin mediated pro-angiogenic role of Hydroxysafflor Yellow A in ischaemic cardiac dysfunction: Post-transcriptional regulation of VEGF-A and MMP-9.

Hydroxysafflor Yellow A (HSYA), a most representative ingredient of Carthamus tinctorius L., had long been used in treating ischaemic cardiovascular diseases in China and exhibited prominently anticoagulant and pro-angiogenic activities, but the underlying mechanisms remained largely unknown. This study aimed to further elucidate the pro-angiogenic effect and mechanism of HSYA on ischaemic cardiac dysfunction. A C57 mouse model of acute myocardial infarction (AMI) was firstly established, and 25 mg/kg HSYA was intraperitoneally injected immediately after operation and given once, respectively, each morning and evening for 2 weeks. It was found that HSYA significantly improved ischaemia-induced cardiac haemodynamics, enhanced the survival rate, alleviated the myocardial injury and increased the expressions of CD31, vascular endothelial growth factor-A (VEGF-A) and nucleolin in the ischaemic myocardium. In addition, HSYA promoted the migration and tube formation of human umbilical vein endothelial cells (HUVECs), enhanced the expressions of nucleolin, VEGF-A and matrix metalloproteinase-9 (MMP-9) in a dose- and time-dependent manner. However, down-regulation of nucleolin expression sharply abrogated the effect mentioned above of HSYA. Further protein-RNA coimmunoprecipitation and immunoprecipitation-RT-PCR assay showed that nucleolin binded to VEGF-A and MMP-9 mRNA and overexpression of nucleolin up-regulated the mRNA expressions of VEGF-A and MMP-9 in the HUVECs through enhancing the stability of VEGF-A and MMP-9 mRNA. Furthermore, HSYA increased the mRNA expressions of VEGF-A and MMP-9 in the extract of antinucleolin antibody-precipitated protein from the heart of AMI mice. Our data revealed that nucleolin mediated the pro-angiogenic effect of HSYA through post-transcriptional regulation of VEGF-A and MMP-9 expression, which contributed to the protective effect of HSYA on ischaemic cardiac dysfunction.