Secretomes reveal several novel proteins as well as TGF-β1 as the top upstream regulator of metastatic process in breast cancer

Purpose

Metastatic breast cancer is resistant to many conventional treatments and novel therapeutic targets are needed. We previously isolated subsets of 4T1 murine breast cancer cells which metastasized to liver (4TLM), brain (4TBM), and heart (4THM). Among these cells, 4TLM is the most aggressive one, demonstrating mesenchymal phenotype. Here we compared secreted proteins from 4TLM, 4TBM, and 4THM cells and compared with that of hardly metastatic 67NR cells to detect differentially secreted factors involved in organ-specific metastasis.

Method and results

Label-free LC–MS/MS proteomic technique was used to detect the differentially secreted proteins. Eighty-five of over 500 secreted proteins were significantly altered in metastatic breast cancer cells. Differential expression of several proteins such as fibulin-4, Bone Morphogenetic Protein 1, TGF-β1 MMP-3, MMP-9, and Thymic Stromal Lymphopoietin were further verified using ELISA or Western blotting. Many of these identified proteins were also present in human metastatic breast carcinomas. Annexin A1 and A5, laminin beta 1, Neutral alpha-glucosidase AB were commonly found at least in three out of six studies examined here. Ingenuity Pathway Analysis showed that proteins differentially secreted from metastatic cells are involved primarily in carcinogenesis and TGF-β1 is the top upstream regulator in all metastatic cells.

Conclusions

Cells metastasized to different organs displayed significant differences in several of secreted proteins. Proteins differentially altered were fibronectin, insulin-like growth factor-binding protein 7, and Procollagen-lysine, 2-oxoglutarate 5-dioxygenase 1. On the other hand, many exosomal proteins were also common to all metastatic cells, demonstrating involvement of key universal factors in distant metastatic process.

     

The Impact of laparoscopic surgery of peritoneal endometriosis and endometrioma on the outcome of ICSI cycles

Our objective was to assess the role of laparoscopic removal of ovarian endometriomas and ablation of peritoneal endometriosis on the outcome of intracytoplasmic sperm injection (ICSI) - Embryo Transfer cycles by comparing with the results of patients with untreated endometriomas and tubal factor without underlying endometriosis confirmed by laparoscopy. For this purpose, between 2002 and 2015, outcomes of 257 ICSI cycles of 150 patients, including 91 cycles of 48 patients in minimal endometriosis, 57 cycles of 25 patients in endometrioma removal, 65 cycles of 53 patients in non-operated endometrioma, and 44 cycles of 24 patients in tubal factor groups were retrospectively analyzed. Basal ovarian reserve was significantly lower, mean starting and total gonadotropin consumption was significantly higher, and mean serum E2 on the day of hCG injection, number of dominant follicles, number of retrieved total, and MII oocytes were all significantly lower in the endometrioma cystectomy group. Fertilization and embryo cleavage rates were also significantly the lowest in the endometrioma cystectomy group, whereas clinical pregnancy and live birth rates were comparable among all groups. The number of transferred embryos and duration of infertility were the most significant predictors of clinical pregnancy and live birth. Basal antral follicle count was also significant in predicting live birth.     

Wound healing potential of Sambucus ebulus L. leaves and isolation of an active component,quercetin 3-O-glucoside

Ethnopharmacological relevance

The leaves of Sambucus ebulus L. are used in Turkish folk medicine for treatment of high fever, rheumatic pains, snake bites and wounds.

Aim of the study

Four different solvent extracts by using n-hexane, diethyl ether, ethyl acetate and methanol were prepared from the leaves of Sambucus ebulus in order to investigate the claimed wound healing activity in vivo and isolation of active component(s) from the active extract through the bioassay-guided fractionation procedures.

Materials and methods

Linear incision and circular excision wound models on rats and mice were employed. The methanol extract was fractionated by successive chromatographic techniques. Wound healing activity of each fraction was investigated following the bioassay-guided fractionation procedures. Moreover, the tissue samples were examined histopathologically.

Results

Significant wound healing activity was observed for the ointment prepared with methanol extract at 1% concentration. The methanol extract treated groups of animals showed 84.3% contraction in circular excision model, which was close to contraction value of the reference drug Madecassol^® (100%). On the other hand, a significant increase (43.7%) in the wound tensile strength was determined with the same extract on incision wound model. Subfractions showed significant but reduced wound healing activity on both in vivo wound models. A flavonoid derivative “quercetin 3-O-glucoside” was isolated and determined as one of the active component of active final subfraction. The results of histopathological examination supported the outcome of linear incision and circular excision wound models.

Conclusion

The experimental data revealed that the methanolic extract of Sambucus ebulus leaves displayed remarkable wound healing activity.     

Is bacterial vaginosis associated with a pro-inflammatory cytokine profile in endometrial secretions of women undergoing IVF?

The objective of this prospective cohort study was to elucidate whether bacterial vaginosis (BV) is associated with a pro-inflammatory endometrial secretion cytokine profile and whether there is a relationship between BV and the concentrations of a number of key regulatory cytokines, chemokines and growth factors. A total of 198 women undergoing IVF treatment were included. Prior to embryo transfer, participants underwent screening for BV according to Nugent criteria by a Gram-stained cervical smear. The concentrations of 17 soluble mediators of human implantation were measured by multiplex immunoassay in endometrial secretions aspirated prior to embryo transfer. Seventeen (8.6%) women had BV (Nugent score >6). Multivariable logistic regression showed a significant positive association between interleukin-β and the presence of BV (P = 0.011; Nugent score >6 versus ?6) and a significant negative association between eotaxin and BV (P = 0.003). No significant differences were found in the ratios of distinct pro- and anti-inflammatory cytokines in endometrial secretions from women with or without BV. In conclusion, BV is associated with higher concentrations of interleukin-β in endometrial secretions compared with women without BV. However, no distinct difference in pro- and anti-inflammatory profiles is present. An effect on endometrial receptivity is unlikely.     

Macrophages eliminate circulating tumor cells after monoclonal antibody therapy

The use of monoclonal antibodies (mAbs) as therapeutic tools has increased dramatically in the last decade and is now one of the mainstream strategies to treat cancer. Nonetheless, it is still not completely understood how mAbs mediate tumor cell elimination or the effector cells that are involved. Using intravital microscopy, we found that antibody-dependent phagocytosis (ADPh) by macrophages is a prominent mechanism for removal of tumor cells from the circulation in a murine tumor cell opsonization model. Tumor cells were rapidly recognized and arrested by liver macrophages (Kupffer cells). In the absence of mAbs, Kupffer cells sampled tumor cells; however, this sampling was not sufficient for elimination. By contrast, antitumor mAb treatment resulted in rapid phagocytosis of tumor cells by Kupffer cells that was dependent on the high-affinity IgG-binding Fc receptor (FcγRI) and the low-affinity IgG-binding Fc receptor (FcγRIV). Uptake and intracellular degradation were independent of reactive oxygen or nitrogen species production. Importantly, ADPh prevented the development of liver metastases. Tumor cell capture and therapeutic efficacy were lost after Kupffer cell depletion. Our data indicate that macrophages play a prominent role in mAb-mediated eradication of tumor cells. These findings may help to optimize mAb therapeutic strategies for patients with cancer by helping us to aim to enhance macrophage recruitment and activity.     

Fate and function of hepatitis-C-virus-specific T-cells during peginterferon-alpha2b therapy for acute hepatitis C

BACKGROUND: Strong hepatitis C virus (HCV)-specific T-cell responses are associated with spontaneous clearance of acute hepatitis C. However, recent studies described a decline in HCV-specific CD8+ T-cells during interferon treatment, suggesting that the success of acute HCV therapy might be independent of adaptive immunity. METHODS: T-cell responses of eight human leukocyte antigen (HLA)-A2-positive, acutely infected patients treated with peginterferon-alpha2b were studied by ELISPOT and proliferation assays and flow cytometry analysis using HCV-specific tetramers. RESULTS: HCV-specific T-cells predominately declined during therapy. However, diverse patterns of CD4+ and CD8+ T-cell kinetics were observed. In patients with sustained virological response chemokine receptor 3 (CXCR-3) expression of HCV-specific CD8+ T-cells was upregulated, indicating homing to the liver. Low levels of T-cells remained detectable throughout treatment and follow up. In contrast, T-cells of a relapse patient did not upregulate CXCR-3 but displayed a higher staining for annexin-V, followed by a complete loss of peripheral virus-specific CD8+ T-cells by week 12. CONCLUSIONS: Kinetics of HCV-specific T-cell responses are heterogeneous in interferon-treated patients with acute hepatitis C. The decline of T-cells might be a consequence of both apoptosis and homing. The balance between cell death and regulation of chemokine receptors might lead to different long-term outcomes.     

Turkish adolescents' knowledge on and attitude toward emergency contraception

ObjectiveAdolescents tend to be at risk for unwanted pregnancies, so detecting their level of knowledge on emergency contraception and providing them information is important to prevent such pregnancies. Hence, in two faculties at Gazi University, this study aimed to detect freshman students' level and need of knowledge on emergency contraception and to evaluate their attitude towards emergency contraception.Materials and MethodsThe study was performed with freshman students of the Occupational Education Faculty and the Technical Education Faculty. A questionnaire including questions about demographic properties, obstetrical history, status of contraceptive use, level of knowledge and opinions on emergency contraception was administered to the students. Data was analyzed statistically with the computer program EPI Info 6.0.ResultsA total number of 385 adolescents were included in the studys; 157 of whom were males (40.8%) and 228 of whom were females (59.2%). To the question “is there any way to prevent a possible pregnancy after an unprotected sexual intercourse?”, 166 students replied “yes” (50.5%), 39 “no” (11.9%) and 124 “I do not know” (37.7%). Of 166 students replying “yes,” 114 (68.7%) listed a possible contraceptive method. The mostly cited method was “morning after pills” (n = 62; 54.4%), followed by curettage (n = 15; 13.2%). Among all students, 158 (49.8%) informed us that they were aware of the presence of “morning after pills” whereas 159 (50.2%) claimed they were not. Eighty-six male students (70.5%) and 115 female students (72%) emphasized that they would use emergency contraception upon necessity.ConclusionHalf of the participants were familiar with various options to prevent pregnancy after an unprotected sexual intercourse episode, but they lacked specific knowledge about possible methods and ways to use them. Thus, it is essential that information about emergency contraception be included in adolescents' educational programs and that adolescents be provided with easily accessible medical services.