Rapid in situ RNA imaging based on Cas12a thrusting strand displacement reaction.

RNA In situ imaging through DNA self-assembly is advantaged in illustrating its structures and functions with high-resolution, while the limited reaction efficiency and time-consuming operation hinder its clinical application. Here, we first proposed a new strand displacement reaction (SDR) model (Cas12a thrusting SDR, CtSDR), in which Cas12a could overcome the inherent reaction limitation and dramatically enhance efficiency through energy replenishment and by-product consumption. The target-initiated CtSDR amplification was established for RNA analysis, with order of magnitude lower limit of detection (LOD) than the Cas13a system. The CtSDR-based RNA in situ imaging strategy was developed to monitor intra-cellular microRNA expression change and delineate the landscape of oncogenic RNA in 66 clinic tissue samples, possessing a clear advantage over classic in situ hybridization (ISH) in terms of operation time (1 h versus 14 h) while showing comparable sensitivity and specificity. This work presents a promising approach to developing advanced molecular diagnostic tools.

Enhancement of sonodynamic treatment of ovarian cancer based on Pt-B-P ternary nanoparticles.

Sonodynamic therapy (SDT) can noninvasively focus sound energy to deep tumor tissues and activate sonosensitizer (such as chlorin e6(Ce6)) to produce antitumor effects. However, due to the hypoxic microenvironment of the tumor, the effect of sonodynamic therapy is limited. In this work, we successfully synthesized Platinum-Boron-Phosphorus ternary nanoparticles (Pt-B-P NPs) for the first time to efficiently catalyze the decomposition of hydrogen peroxide (H2O2) in tumor tissues to produce sufficient oxygen (O2) and improve the effect of sonodynamic treatment of ovarian cancer. In vitro studies, we found that compared with Platinum nanoparticles (Pt NPs), Pt-B-P NPs have the significantly increased ability to catalyze the decomposition of H2O2 to produce oxygen and thus the hypoxic environment of tumor cells could be improved efficiently. Meanwhile, the bio-distribution, therapeutic effect and bio-safety of Pt-B-P NPs in vivo were evaluated using BALB/c-nu mouse model of ovarian cancer and the desired result had been achieved.

A dual-signal electrochemical immunosensor for the detection of HPV16 E6 oncoprotein based on PdBP dendritic ternary nanospheres and MBSi-Chi nanocomposites.

Cervical cancer is a common cancer in women. HPV16 E6 oncoprotein is a reliable biomarker for cervical cancer. Although there are other methods for detecting E6 oncoprotein, the electrochemical method has more advantages, such as low cost, convenience and speed. In this study, a novel dual-signal electrochemical immunosensor for quick and sensitive detection of E6 oncoprotein based on a high efficiency catalyst and signal label was developed. Herein, to achieve quick detection, palladium-boron-phosphorus dendritic ternary nanospheres (PdBP NSs) not only acted as a catalyst to catalyze H2O2, but also as a support material to capture antibodies. Moreover, to realize sensitive detection, nanocomposites of mesoporous silica nanoparticles loaded with methylene blue and coated with chitosan (MBSi-Chi) were synthesized as a signal label, which can produce electrochemical signal. Under optimal conditions, the label-free immunosensor exhibited a linear range of 100 fg mL-1 to 4 ng mL-1 with a detection limit of 72.8 fg mL-1 (S/N = 3), and the sandwich-type immunosensor presented a linear range of 50 fg mL-1 to 4 ng mL-1 with a detection limit of 34.1 fg mL-1 (S/N = 3). The as-prepared dual-signal immunosensor had desirable specificity, stability and repeatability, implying its potential applications in clinical laboratory.

The establishment of an immunosensor for the detection of SPOP.

In this paper, we first synthesis three-dimensional jasmine-like Cu@L-aspartic acid(L-ASP) inorganic-organic hybrid nanoflowers to load palladium-platinum nanoparticles (Pd-Pt NPs) as the signal enhancer in order to quantify intracellular speckle-type POZ domain protein. Scanning electron microscope, fourier transform infrared, energy dispersive spectrometer, X-ray photoelectron spectroscopy analysis was used to characterize the newly synthesized materials. The newly formed Cu@L-Asp/Pd-PtNPs can catalyze the decomposition of hydrogen peroxide and exhibit excellent catalytic performance. When different concentration of speckle-type POZ domain protein is captured by speckle-type POZ domain protein antibody linked to the surface of Cu@L-Asp/Pd-Pt NPs, the current signal decreases with the increase concentration of speckle-type POZ domain protein. After optimization, the speckle-type POZ domain protein immunosensor exhibited a good linear response over a concentration range from 0.1-1 ng mL-1 with a low detection limit of 19 fg mL-1. The proposed sensor demonstrates good stability within 28 days, acceptable reproducibility (RSD = 0.52%) and selectivity to the speckle-type POZ domain protein in the presence of possible interfering substances and has potential application for detecting other intracellular macromolecular substances.

Hypoxia-induced SPOP attenuates the mobility of trophoblast cells through inhibition of the PI3K/AKT/GSK3ß pathway.

Placental hypoxia has been implicated in pregnancy pathologies such as pre-eclampsia and intrauterine growth restriction. However, the underlying mechanism by which the trophoblasts respond to hypoxia remains unclear. Speckle-type POZ protein (SPOP), an E3 ubiquitin ligase adapter, was previously reported to play important roles in various physiological and pathological processes. This study aims to investigate the expression and biological functions of SPOP after exposure to cobalt chloride (CoCl2 )-mimicked hypoxia conditions using human trophoblast-derived choriocarcinoma cell lines and extravillous cytotrophoblast. These data showed that SPOP protein was directly induced by CoCl2 -mimicked hypoxia and regulated by HIF-1α at the posttranscription level. CoCl2 treatment could dramatically influence the localization of SPOP in trophoblasts, especially the accumulation of SPOP into the nucleus. In addition, both CoCl2 -mimicked hypoxia and induction of endogenous SPOP expression by lentivirus transfection attenuated the migration and invasion abilities of trophoblasts. Furthermore, we demonstrated that SPOP was involved in CoCl2 -induced the inhibition of the PI3K/AKT/GSK3ß pathway in placental trophoblasts. Taken together, these data indicate that accumulation of HIF-1α augments the expression of SPOP in trophoblasts, which impairs trophoblastic mobility by targeting the PI3K/AKT/GSK3ß pathway. This potentially leads to insufficient uterine spiral artery remodeling and suboptimal placental perfusion, and thus the development of pregnancy-related complication.

SPOP attenuates migration and invasion of choriocarcinoma cells by promoting DHX9 degradation.

Speckle-type POZ protein (SPOP), a novel cancer- associated protein, was previously reported to function as a tumor suppressor or promoter in different malignant tumors. This research aims to investigate the biological functions and underlying molecular mechanisms of SPOP in choriocarcinoma. Our analysis of patient tissues and cell lines showed significantly decreased SPOP expression and highly expressed Nuclear DNA helicase II and RNA helicase A (DHX9), both of them are mainly located into the nucleus. Induction or depletion of endogenous SPOP with a lentivirus-based system correspondingly suppressed or promoted migration and invasion of choriocarcinoma cells. Mechanistically, we found that SPOP bound to DHX9 and induced the ubiquitination and degradation of DHX9 by recognizing a typical SPOP-binding motif in DHX9. SPOP-DHX9 interaction was demonstrated to play a critical role in regulating migration and invasion abilities of choriocarcinoma cells, the promotion of mobility ability in knocking down SPOP was partly counteracted by transfection with siRNA against DHX9. Taken together, our results suggest that SPOP suppresses migration and invasion of choriocarcinoma by promoting the ubiquitination and subsequent degradation of DHX9, which identifies the SPOP-DHX9 interaction may serve as a potential therapeutic target against choriocarcinoma.

CircRNA_0001449 disturbs phosphatidylinositol homeostasis and AKT activity by enhancing Osbpl5 translation in transient cerebral ischemia.

Phosphatidylinositol-3,4,5-trisphosphate [PI(3,4,5)P3] is a phosphorylated derivative of phosphatidylinositol 4-phosphate [PI(4)P] and phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2], which recruit and activate AKT in the plasma membrane (PM) to promote cellular survival. ORP5 anchors at the endoplasmic reticulum (ER)-PM contact sites and acts as a PI(4)P and PI(4,5)P2/phosphatidylserine (PS) exchanger. Here, a lipidomics analysis of the sensorimotor cortex revealed that transient middle cerebral artery occlusion (tMCAO) disturbs the homeostasis of phosphatidylinositols (PIs) and PS between the PM and ER. Conditional knockout mice showed that ORP5 contributes to this abnormal distribution. Abolishing the ORP5 gene significantly inhibited apoptosis and autophagy. RNA sequencing and RNA pull down analyses confirmed a competing endogenous RNA pathway in which circ_0001449 sponges miR-124-3p and miR-32-5p to promote Osbpl5 translation. Our data showed that circRNA_0001449 regulates membrane homeostasis via ORP5 and is involved in the AKT survival pathway.

SPOP Regulates The Biological Mechanism Of Ovarian Cancer Cells Through The Hh Signaling Pathway.

BACKGROUND: Ovarian cancer is characterized by high metastatic potential and high mortality. More than 80% of primary ovarian malignancies are epithelial ovarian cancers. There is increasing evidence that Speckle-type POZ protein (SPOP) is highly correlated with the development of various types of cancer. However, the effects of SPOP on epithelial ovarian cancer and the associated molecular mechanisms remain unclear. MATERIALS AND METHODS: We compared SPOP expression between epithelial ovarian cancer tissues and normal ovarian tissues by using immunohistochemical staining. To determine the role of SPOP in epithelial ovarian cancer cells, we overexpressed or knocked down SPOP in the epithelial ovarian cancer cell line OVCAR-3 using lentiviral vectors. RESULTS: Our results from the present study indicated that SPOP expression was significantly downregulated in human epithelial ovarian cancer and was associated with the FIGO stage and the histopathologic grading of the tumor. The overexpression and knockdown experiments revealed that SPOP inhibited proliferation while promoting apoptosis in ovarian cancer cells. Inhibition of SPOP mis-activated the Hedgehog (Hh) signaling pathway, thereby inhibiting apoptosis in ovarian cancer cells. CONCLUSION: SPOP suppresses proliferation and promotes apoptosis in human ovarian cancer cells by inhibiting the Hh signaling pathway, offering the possibility of new approaches for the treatment of ovarian cancer.

The application of antigen receptor gene rearrangement of BIOMED-2 in the pathologic diagnosis of 348 cases with non-Hodgkin lymphoma in a single institution in Southwest of China.

OBJECTIVE: To explore the clinical value of immunoglobulin (Ig) and T cell receptor (TCR) gene rearrangement in the diagnosis of non-Hodgkin lymphoma. METHODS: Using the standardized BIOMED-2 multiplex PCR strategy to detect IgH, IgK and TCR in 272 cases of mature B-cell lymphoma, 55 cases of mature T-cell lymphoma, 21 cases of extranodal NK/ T-cell lymphoma, nasal type, and 20 cases of lymphoid tissue reactive hyperplasia. RESULTS: Among all mature B-cell lymphomas, the sensitivity of Ig gene rearrangement was 91.18% (248/272), IgH and IgK gene rearrangement was 76.47% (208/272) and 75.00% (204/272), respectively, meanwhile the sensitivity of TCRγ rearrangement was 3.68% (10/272). In the 55 cases of mature T-cell lymphoma, the sensitivity of the detection of TCRγ was 76.36% (44/55), at the same time the sensitivity of Ig gene rearrangement was 14.55% (8/55), IgH and IgK gene rearrangement was 7.27% (4/55) and 12.73% (7/55), respectively. In 21 cases of extranodal NK/T cell lymphoma, nasal type, and 20 cases of reactive lymphoid hyperplasia, no gene rearrangement was found in the samples of IgH, IgK and TCR. The sensitivity of gene rearrangement in Ig/TCR in B and T-cell lymphoma was significantly different from that in the control group (P < 0.05). CONCLUSION: The Ig/TCR gene rearrangement of BIOMED-2 multiplex PCR strategy has important auxiliary value in the diagnosis of B/T-cell non-Hodgkin lymphoma respectively, however, a few B-cell lymphomas may company TCR gene rearrangement as well as a few T-cell lymphomas may accompany Ig gene rearrangement, it must be comprehensively judged with the combination of morphology, immunohistochemistry and clinical features.

A label-free immunosensor for the detection of nuclear matrix protein-22 based on a chrysanthemum-like Co-MOFs/CuAu NWs nanocomposite.

In this study, a new, simple, and label-free electrochemical immunosensor was presented for the detection of nuclear matrix protein-22 (NMP-22). In order to accurately monitor very small amounts of NMP-22, it was advantageous to use highly efficient nanomaterials as signals. For this reason, we synthesized a chrysanthemum-like nanocomposite (Co-MOFs/CuAu NWs), using Co-based metal-organic frameworks (Co-MOFs) as carriers and copper gold nanowires (CuAu NWs) wrapped around their surface, which was applied for modifying a glassy carbon electrode (GCE). The Co-MOFs/CuAu NWs possessed outstanding catalytic capabilities, which served as signal materials and simultaneously carried the anti-NMP-22 antibody (Ab). When different concentrations of the NMP-22 antigen (Ag) were specifically attached to the immunosensor, the current responses decreased by varying degrees. The designed biosensor used the principle to establish a linear regression equation and achieve an accurate quantification of NMP-22. After optimization, the NMP-22 sensor exhibited a good linear response over a concentration range from 0.1 pg mL-1 to 1 ng mL-1, with a lower detection limit of 33 fg mL-1 (based on S/N = 3). The proposed biosensor demonstrated the advantages of ultra-sensitivity, high specificity and acceptable reproducibility, suggesting that the proposed strategy has the potential for the quantification of NMP-22 in human urine samples. Moreover, the novel nanocomposite Co-MOFs/CuAu NWs are promising materials for electrochemical sensors to detect other biomolecules.

Retrospective analysis of the association between human epidermal growth factor receptor 2 amplification and chromosome enumeration probe 17 status in patients with breast cancer.

The aim of the present study was to identify potential human epidermal growth factor receptor 2 (HER2) amplification, according to American Society of Clinical Oncology and the College of American Pathologists (ASCO/CAP) 2013 HER2 testing guidelines, in patients previously determined not to possess HER2 amplification, in accordance with previous 2007 guidelines. Potential discrepancies may arise from chromosome enumeration probe 17 (CEP17) amplification, deletion, polysomyor monosomy. HER2, CEP17, tumor protein p53 (TP53) and retinoic acid receptor α (RARA) genes from 67 patient specimens with suspected amplification, polysomy or monosomy of CEP17 were analyzed using fluorescence in situ hybridization. HER2 status was interpreted using 2007 and 2013 ASCO HER2 test guidelines as well as the reference genes TP53 and RARA. According to ASCO/CAP2007 HER2 guidelines, 20 patients exhibited HER2 amplification (29.85%), 41 were without HER2 amplification (including 25 with polysomy, 15 with monosomy and 1 with suspected monosomy plus co-amplification of HER2 and CEP17) and the remaining 6 patients were equivocal. Using ASCO/CAP 2013 HER2 guidelines, 49 patients exhibited HER2 gene amplification (73.1%). The 29-patient increase included 6 originally at equivocal levels but now demonstrating amplification, 22 originally with polysomy but now revealing co-amplification, and 1 with suspected monosomy plus co-amplification of HER2 and CEP17. According to TP53 and RARA, HER2 was amplified in 43 patients (64.1%). Using the revised guidelines, HER2, originally identified as amplified in 6 patients, was not amplified following the introduction of TP53 and RARA control genes. Among these 6, 4 possessed normal TP53 and RARA. The incidence of co-amplification of HER2 and CEP17 was 1.4% (21/1,518). RARA and TP53 are suitable control genes to evaluate HER2 status.

Negative genic switch of HER-2 in the primary tumor instead of the synchronous metastatic nodal lesions after neoadjuvant chemotherapy in a patient with primary HER2-positive breast cancer.

BACKGROUND: A few retrospective studies have indicated that neoadjuvant chemotherapy (NAC) in breast cancer may change biomarker profiles of the primary tumor. Little is known about the status of HER-2 gene of the synchronous nodal metastases when that of the residual tumor undergoes negative conversion in a neoadjuvant setting. CASE PRESENTATION: We describe a female patient with left breast cancer (T2N2M0) who underwent negative conversion of HER-2 in the primary tumor instead of the synchronous nodal lesions after NAC. Core needle biopsy showed invasive ductal carcinoma with HER2 immunohistochemistry (IHC) (2+) and amplified HER-2 gene determined by fluorescence in situ hybridization (FISH). Then, the patient underwent 4 cycles of anthracycline- and taxane-based NAC and subsequent left modified radical mastectomy. Postoperative pathology showed invasive ductal carcinoma involving 4 of 12 surgically excised axillary lymph nodes with HER2 IHC (1+) and FISH negative (HER2 gene not amplified) in the residual tumor of the breast specimen. Due to the negative genic switch of HER2 after NAC, the patient rejected to accept trastuzumab. Under the patient's consent, the synchronous nodal lesions were further investigated and showed HER2 IHC(-) but FISH positive (HER-2 gene amplified). Therefore, the patient agreed to accept adjuvant trastuzumab treatment every 3 weeks for 1 year. CONCLUSIONS: We propose further assessment of HER2 gene in the synchronous nodal metastases, especially when negative genic switch of HER-2 occurs in the primary tumor after NAC in order to tailor the systemic regimens for breast cancer patients.

Enhanced Depth Imaging Optical Coherence Tomography: A New Way Measuring Choroidal Thickness in Pregnant Women.

The body changes markedly during pregnancy; each system behaves differently from a nonpregnant state. As the eyes are the only windows to see directly what is going on in the internal environment, more and more researches have been done to explain the association between ocular changes and the physiological and pathological changes during pregnancy. The choroid is one of the critical parts of the eye, providing nutrition. And abnormal choroid may result in ocular dysfunction and visual problems. As the optical coherence tomography develops, a rapid, direct, noninvasive, and nontoxic way is available to obtain the choroid situation of pregnant women, which may explain the mechanism of pregnancy-related eye diseases. This review would summarize relevant original articles published from January 1, 2008 to December 1, 2016 to assess the changes of choroidal thickness (CT) with enhanced depth imaging optical coherence tomography (EDI-OCT) during pregnancy. And the relationship between choroidal thickness changes and pregnancy remains uncertain. To our knowledge, this is the first review of EDI-OCT in assessing the choroidal thickness of the pregnant women.

Primary central nervous system diffuse large B-cell lymphoma shows an activated B-cell-like phenotype with co-expression of C-MYC, BCL-2, and BCL-6.

Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma, whose main prognostic factor is closely related to germinal center B-cell-like subtype (GCB- DLBCL) or activated B-cell-like type (non-GCB-DLBCL). The most common type of primary central nervous system lymphoma is diffuse large B-cell type with poor prognosis and the reason is unclear. This study aims to stratify primary central nervous system diffuse large B-cell lymphoma (PCNS-DLBCL) according to the cell-of-origin (COO) and to investigate the multiple proteins expression of C-MYC, BCL-6, BCL-2, TP53, further to elucidate the reason why primary central nervous system diffuse large B-cell lymphoma possesses a poor clinical outcome as well. Nineteen cases of primary central nervous system DLBCL were stratified according to immunostaining algorithms of Hans, Choi and Meyer (Tally) and we investigated the multiple proteins expression of C-MYC, BCL-6, BCL-2, TP53. The Epstein-Barr virus and Borna disease virus infection were also detected. Among nineteen cases, most (15-17 cases) were assigned to the activated B-cell-like subtype, highly expression of C-MYC (15 cases, 78.9%), BCL-2 (10 cases, 52.6%), BCL-6 (15 cases, 78.9%). Unfortunately, two cases were positive for PD-L1 while PD-L2 was not expressed in any case. Two cases infected with BDV but no one infected with EBV. In conclusion, most primary central nervous system DLBCLs show an activated B-cell-like subtype characteristic and have multiple expressions of C-MYC, BCL-2, BCL-6 protein, these features might be significant factor to predict the outcome and guide treatment of PCNS-DLBCLs.

Prospective evaluation of FISH for detecting upper tract urothelial carcinoma in voided urine specimens.

The aim of the present study was to estimate the diagnostic value of fluorescence in situ hybridization (FISH) analysis of tumor cells in voided urine specimens for detecting upper tract urothelial carcinoma (UTUC). Cytology and FISH analyses were conducted on voided urine collected in the morning from 125 patients with suspected UTUC. During follow-up, ureteroscopy with biopsy and histopathology were used to confirm the presence of tumors. The average follow-up time was 23.8 months (range, 6-36 months). A total of 8 patients who could not be contacted until the last follow-up were excluded from the study. Of the remaining 117 patients, 19 were histologically demonstrated to have UTUC, of whom, 3 patients had stage pTis disease, 6 had stage pTa disease, 5 had stage pT1 disease and 5 had stage pT2 disease (7 G1, 8 G2 and 4 G3). The overall sensitivity of FISH to detect UTUCs in voided urine specimens was 84.21% (16/19), whereas that of cytology was 42.11% (8/19) (P<0.05). The overall specificity of FISH to detect UTUCs in voided urine specimens was 89.80% (88/98), compared with 94.90% (93/98) of cytology (P>0.05). The positive predictive value (PPV) and negative predictive value of FISH were 80.00% (16/20) and 97.78% (88/90), respectively, whereas those of cytology were 100.00% (8/8) (P>0.05) and 90.29% (93/103) (P>0.05), respectively. The present data indicated that FISH was a method capable of detecting UTUCs in voided urine specimens with good sensitivity and specificity, although it exhibited a high rate of false positivity and low PPV.

Speckle-type POZ (pox virus and zinc finger protein) protein gene deletion in ovarian cancer: Fluorescence in situ hybridization analysis of a tissue microarray.

The aim of the present study was to investigate the status of speckle-type POZ (pox virus and zinc finger protein) protein (SPOP) gene located on chromosome 17q21 in ovarian cancer (OC). The present study evaluated a tissue microarray, which contained 90 samples of ovarian cancer and 10 samples of normal ovarian tissue, using fluorescence in situ hybridization (FISH). FISH is a method where a SPOP-specific DNA red fluorescence probe was used for the experimental group and a centromere-specific DNA green fluorescence probe for chromosome 17 was used for the control group. The present study demonstrated that a deletion of the SPOP gene was observed in 52.27% (46/88) of the ovarian cancer tissues, but was not identified in normal ovarian tissues. Simultaneously, monosomy 17 was frequently identified in the ovarian cancer tissues, but not in the normal ovarian tissues. Furthermore, the present data revealed that the ovarian cancer histological subtype and grade were significantly associated with a deletion of the SPOP gene, which was assessed by the appearance of monosomy 17 in the ovarian cancer samples; the deletion of the SPOP gene was observed in a large proportion of serous epithelial ovarian cancer (41/61; 67.21%), particularly in grade 3 (31/37; 83.78%). In conclusion, deletion of the SPOP gene on chromosome 17 in ovarian cancer samples, which results from monosomy 17, indicates that the SPOP gene may serve as a tumor suppressor gene in ovarian cancer.

SPOP Regulates Endometrial Stromal Cell Decidualization in Mice.

Ubiquitination is a regulatory mechanism that occurs after protein translation. To date, few studies have reported on ubiquitination during embryo implantation. We used real-time quantitative polymerase chain reaction, immunohistochemistry, and Western blotting analyses to analyze the expression of speckle-type pox virus and zinc finger (POZ) protein (SPOP; an adapter of E3 ligases of ubiquitination) in mouse uteri during early pregnancy and pseudopregnancy using an artificially induced decidualization model and a steroid hormone-processing model. At the same time, we established an artificially induced decidualization in vitro model. We observed that SPOP regulates endometrial stromal cell decidualization in mice and that hormones regulate the expression of SPOP. This study suggests that ubiquitination may be involved in embryonic implantation.

Grb7 gene amplification and protein expression by FISH and IHC in ovarian cancer.

OBJECTIVE: Overexpression of growth factor receptor-bound protein 7 (Grb7) has been found in numerous human cancers. The aim of this study was to evaluate the correlation between Grb7 gene amplification and protein expression in ovarian cancer (OC). METHODS: We use Tissue Microarray (TMA) respectively to detect the gene amplification and protein expression of Grb7 in 90 cases OC and 10 control specimens of normal ovarian tissues by IHC and FISH. RESULTS: The Grb7 protein expression by IHC analysis was observed in 52/90 (57.8%) OC with 3 cases (3.3%) scored 3(+) and 9 cases (10%) scored 2(+) Grb7 gene amplification by FISH analysis was successfully detectable in 6 specimens with a positive rate of 6.8% (6/88) in which immunostaining 3(+), 2(+) and negative (1(+)/0) expressions of Grb7 were 100.0% (3/3), 11.1% (1/9) and 2.6% (2/76), respectively. Our data exhibited that the IHC and FISH results had a good consistency between Grb7 gene amplification and Grb7 protein expression (Kappa = 0.651, P < 0.001). Both the results of IHC and FISH revealed that Grb7 did not seem to have a role in OC clinicopathology. CONCLUSION: There is a close relationship between Grb7 gene amplification and GRB7 protein overexpression in human OC. IHC might have limited diagnostic value especially in these tumors and especially in characterizing genetically diverse borderline cases, FISH could be superior to IHC.

The PI3K/Akt signaling pathway exerts effects on the implantation of mouse embryos by regulating the expression of RhoA.

The aim of this study was to investigate whether the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway affects the implantation of mouse embryos by regulating the expression of RhoA. The expression of PI3K, Akt, phosphorylated (p-)Akt, phosphatase and tensin homolog (PTEN) and RhoA in the uterus of mice on day 5 of pregnancy (D5) and in pseudopregnant mice was examined by quantitative reverse transcription polymerase chain reaction (qRT-PCR), immunohistochemistry and western blot analysis. A functional analysis of these genes was also performed by the intrauterine injection with the PI3K inhibitor, LY294002, on day 2 of pregnancy (D2). The expression levels of PI3K, p-Akt, RhoA at the implantation site were higher than those at the inter-implantation site in the endometrium; however, opposite effects were observed for PTEN expression. The expression levels of the above genes in the pseudopregnant group and in the group injected with the PI3K/Akt inhibitor, LY294002, were markedly lower than those in the pregnant group. Functional experiments revealed that the number of implantation sites had been significantly decreased (P<0.05) following the intrauterine injection of the PI3K inhibitor, LY294002, on day 2 of gestation compared with the contralateral injection of phosphate-buffered saline (PBS). These results suggest that the PI3K/Akt signaling pathway affects embryo implantation by regulating the expression of RhoA.

Protective effects of ginsenosides against Bisphenol A-induced cytotoxicity in 15P-1 Sertoli cells via extracellular signal-regulated kinase 1/2 signalling and antioxidant mechanisms.

Numerous studies have demonstrated that Bisphenol A (BPA) can cause reproductive toxicity. Ginseng has wide range of pharmacological actions and, more importantly, has proven its worth with respect to reproductive function in several reports. We have suggested that ginsenosides, the main active components of ginseng, may protect against BPA-induced cell damage. Therefore, an in vitro culture model of 15P-1 Sertoli cells was employed to investigate whether ginsenosides have protective effects on BPA-stimulated 15P-1 Sertoli cells. The results revealed that ginsenosides (75 µg/ml) significantly inhibited BPA-induced decreases in cell viability and increases in apoptosis. Immunofluorescence staining showed that BPA exposure-induced collapse of vimentin intermediate filaments was prevented by the application of ginsenosides. Ginsenosides also inhibited extracellular signal-regulated kinase (ERK1/2) phosphorylation and BPA-induced alterations of Bcl-2 and Bax protein expression in 15P-1 Sertoli cells. Furthermore, the alterations of T-AOC, superoxide dismutase, glutathione peroxidase, glutathione reductase, glutathione and malondialdehyde levels in BPA-stimulated cells were partially prevented with pre-treatment with ginsenosides. Taken together, these results suggest that ginsenosides have protective effects against BPA-induced cell damage and that these effects are mediated by preventing ERK1/2 phosphorylation and through the enhancement of cellular antioxidant capacity. Ginsenosides may therefore be beneficial in the prevention of environmental BPA-induced, reproduction-related toxicity.