Celecoxib Blocks Vasculogenic Mimicry via an Off-Target Effect to Radiosensitize Lung Cancer Cells: An Experimental Study.

The resistance to radiotherapy in lung cancer can be attributed to vasculogenic mimicry (VM) to some extent. Celecoxib (CXB), a selective inhibitor of cyclooxygenase-2 (COX-2), is reported as a radiosensitizer in non-small cell lung cancer (NSCLC). However, whether CXB can regulate VM formation via an off-target effect to radiosensitize NSCLC remains unclear. This study aimed to elucidate the mechanism underlying the radiosensitizing effect of CXB on NSCLC, i.e., whether CXB can inhibit VM formation via binding to newly identified targets other than COX-2. CXB radiosensitivity assay was performed in BALB/c mice bearing H460 xenografts and C57 mice bearing Lewis lung cancer (LLC) xenografts, which were divided into the control, CXB, irradiation (IR) treatment, and IR plus CXB groups. VM formation was observed using 3D Matrigel, periodic acid solution (PAS) staining, and immunofluorescence staining. The potential off-targets of CXB were screened using Protein Data Bank (PDB) database, MGLTools 1.5.6, and AutoDock Vina 1.1.2 and confirmed by Western blotting, enzyme activity assay, and RNA interference in vitro experiments and by immunohistochemistry in vivo experiments. CXB treatment almost eliminated the enhancement of VM formation by IR in vitro and in vivo, partially due to COX-2 inhibition. Four potential off-targets were predicted by molecular docking. Among them, aminopeptidase N (APN) and integrin alpha-V (ITAV) were remarkably inhibited in protein expression and enzyme activity in vitro or in vivo, consistent with the remarkable reduction of VM formation in H460 xenografts in BALB/c mice. In conclusion, CXB dramatically blocked VM through inhibiting newly identified off-targets APN and ITAV, other than COX-2, then radiosensitizing NSCLC.

The incidence of placenta related disease after the hysteroscopic adhesiolysis in patients with intrauterine adhesions.

OBJECTIVE: To analyze the correlation between placenta related disease of pregnant women with antecedent hysteroscopic adhesiolysis due to intrauterine adhesions (IUA). MATERIALS AND METHODS: This is a single center, non-randomized, open-label, retrospective cohort Study. 74 patients who had adhesiolysis and hormone therapy for IUA and progressed into the third trimester were group A and 296 without IUA were group B. The main outcome measure is the incidence of placenta related disease including placenta accreta spectrum, placenta previa, placental abruption, intrauterine growth restriction (IUGR), and pregnancy-induced hypertension (PIH). The second outcome is the perinatal, and intrapartum complications. RESULTS: Patients in group A had a higher frequency of prior pregnancy times (2.51 ± 1.56 vs.1.84 ± 1.06, p = 0.001) and lower frequency of prior delivery times (0.20 ± 0.41 vs. 1.30 ± 0.51, p < 0.05) than group B at baseline. At delivery, there is no difference between the incidence of PIH and IUGR between two groups. However, a significantly higher frequency of placenta accreta (17.6% vs. 1.4%, OR = 15.56, 95% CI 4.91-49.34), placenta increta (5.4% vs. 0.7%, OR = 8.4, 95% CI 1.51-46.78), placenta previa (8.1% vs. 2.0%, OR = 4.265, 95%CI1.33-13.63) and postpartum hemorrhage (21.6% vs. 3.4%, OR = 7.890, 95% CI 3.41-18.26) were observed in group A than in group B. CONCLUSIONS: Compared to general population, the rates of placenta accreta, placenta increta, placenta previa, postpartum hemorrhage are higher among the IUA patients after hysteroscopic adhesiolysis, and special attention is needed at the termination of the pregnancy.

Recruitment of CD11b+Ly6C+ monocytes in non-small cell lung cancer xenografts challenged by anti-VEGF antibody.

A series of antibodies against vascular endothelial growth factor (VEGF) have been developed for the treatment of various types of cancer, including non-small cell lung cancer (NSCLC) in recent years. However, tumors frequently demonstrate resistance to these strategies of VEGF inhibition. Efforts to better understand the mechanism underlying the acquired resistance to anti-VEGF antibodies are warranted. In the present study, in order to develop a xenograft model of acquired resistance to anti-VEGF antibody, xenografts of human adenocarcinoma A549 cells were generated through the successive inoculation of tumor tissue explants into first (F1), second (F2) and third (F3) generations of mice treated with the anti-VEGF antibody B20. Tumor growth rate and vessel-forming ability, assessed via cluster of differentiation (CD) 31 staining, were significantly lower in the F1, F2 and F3 groups compared with in the F0 control group (P<0.01), suggesting that drug resistance was not successfully acquired. The percentages of CD11b+ myeloid-derived suppressor cells and lymphocyte antigen 6C (Ly6C)+ subsets were significantly smaller in F1, F2 and F3 groups compared with in F0 (P<0.01). However, the ratio of Ly6C+ to CD11b+ cells was significantly higher in the F3 group compared with in F0 and F1 groups (P<0.01), indicating increasing recruitment of the Ly6C+ subset with successive challenges with the anti-VEGF antibody. In conclusion, the recruitment of CD11b+Ly6C+ monocytes increased with successive generations of NSCLC-xenografted mice challenged by B20, an anti-VEGF agent.

Neoascarophis sinensis n. sp. (Nematoda: Cystidicolidae), a parasite of Conger myriaster (Brevoort) (Anguilliformes: Congridae) in Chinese waters.

Neoascarophis sinensis n. sp. collected from the whitespotted conger Conger myriaster (Brevoort) (Anguilliformes: Congridae) in the Yellow and East China Seas, is described using both light and scanning electron microscopy. The new species is characterised mainly by the body size (8.5-10.5 mm in the males, 9.5-14.0 mm in the females), the location of the vulva (near equatorial region of the body), the non-bifurcate deirids, the lengths of the vestibule (40-50 µm in the males, 30-60 µm in the females) and glandular oesophagus (2.5-3.1 mm in the males, 3.1-3.5 mm in the females) and the morphology and length of the spicules (left spicule 400-410 µm, right spicule 130-150 µm). Neoascarophis sinensis n. sp. is the first species of Neoascarophis Machida, 1976 reported from the anguilliform fish and is also the only species of this genus found in the Chinese waters.

Yap1 promotes the survival and self-renewal of breast tumor initiating cells via inhibiting Smad3 signaling.

Tumor initiating cells (TICs) serve as the root of tumor growth. After identifying TICs in spontaneous breast tumors of the MMTV-Wnt1 mouse model, we confirmed the specific expression and activation of Yes-associated protein 1 (Yap1) within TICs. To investigate the role of Yap1 in the self-renewal of breast TICs and the underlying mechanism, we sorted CD49fhighEpCAMlow cells as breast TICs. Active Yap1 with ectopic expression in breast TICs promoted their colony formation in vitro (p< 0.01) and self-renewal in vivo (p< 0.01), and led to a 4-fold increase in TIC frequency (p< 0.05).A conditional knock-out mouse was reconstructed to generate Yap1 knock-out breast tumors. The loss of Yap1 led to a dramatic growth disadvantage of breast TICs in vitro (p< 0.01) and in vivo (p< 0.01), and it also led to an over 200-fold decrease in TIC frequency (p< 0.01). The expression of active Yap1 was negatively correlated with that of phosphorylated Smad3 (p-Smad3).Transforming growth factor ß (TGF-ß) served as a strong enhancer of Smad3 and an inhibitor of clonogenesis of TICs. The presence of SIS3, a specific inhibitor of Smad3, could rescue the TGF-ß -induced growth inhibition and reverse the Smad3 inhibition by Yap1. Analysis of a database containing 2,072 human breast cancer samples showed that higher expressions of Yap1 correlated with a poorer outcome of a 15-year survival rate and median overall survival (mOS)in patients, especially in those with basal breast tumors without estrogen receptor 1 (ER) expression. The findings indicate that active Yap1 promotes the self-renewal of breast TICs by inhibiting Smad3 signaling.

Essential role of miR-200c in regulating self-renewal of breast cancer stem cells and their counterparts of mammary epithelium.

BACKGROUND: Breast cancer stem cells (BCSCs) have been reported as the origin of breast cancer and the radical cause of drug resistance, relapse and metastasis in breast cancer. BCSCs could be derived from mutated mammary epithelial stem cells (MaSCs). Therefore, comparing the molecular differences between BCSCs and MaSCs may clarify the mechanism underlying breast carcinogenesis and the targets for gene therapy. Specifically, the distinct miRNome data of BCSCs and MaSCs need to be analyzed to find out the key miRNAs and reveal their roles in regulating the stemness of BCSCs. METHODS: MUC1(-)ESA(+) cells were isolated from normal mammary epithelial cell line MCF-10A by fluorescence-activated cell sorting (FACS) and tested for stemness by clonogenic assay and multi-potential differentiation experiments. The miRNA profiles of MaSCs, BCSCs and breast cancer MCF-7 cells were compared to obtain the candidate miRNAs that may regulate breast tumorigenesis. An miRNA consecutively upregulated from MaSCs to BCSCs to MCF-7 cells, miR-200c, was chosen to determine its role in regulating the stemness of BCSCs and MaSCs in vitro and in vivo. Based on bioinformatics, the targets of miR-200c were validated by dual-luciferase report system, western blot and rescue experiments. RESULTS: In a 2-D clonogenic assay, MUC1(-)ESA(+) cells gave rise to multiple morphological colonies, including luminal colonies, myoepithelial colonies and mixed colonies. The clonogenic potential of MUC1(-)ESA(+) (61.5 ± 3.87 %) was significantly higher than that of non-stem MCF-10A cells (53.5 ± 3.42 %) (P < 0.05). In a 3-D matrigel culture, MUC1(-)ESA(+) cells grew into mammospheres with duct-like structures. A total of 12 miRNAs of interest were identified, 8 of which were upregulated and 4 downregulated in BCSCs compared with MaSCs. In gain- and lost-of-function assays, miR-200c was sufficient to inhibit the self-renewal of BCSCs and MaSCs in vitro and the growth of BCSCs in vivo. Furthermore, miR-200c negatively regulated programmed cell death 10 (PDCD10) in BCSCs and MaSCs. PDCD10 could rescue the tumorigenesis inhibited by miR-200c in BCSCs. DISCUSSION: Accumulating evidence shows that there is a milignant transformation from MaSCs into BCSCs. The underlying mechanism remains unclear. In present study, miRNA profiles between MaSCs and BCSCs were obtained. Then miRNA-200c, downregulated in both MaSCs and BCSCs, were verified as anti-oncogene, and played essential role in regulating self-renewal of both kinds of stem-like cells. These findings reveal a novel insights of breast tumorigenesis. CONCLUSIONS: PDCD10 is a target gene of miR-200c and also a possible mechanism by which miR-200c plays a role in regulating the stemness of BCSCs and MaSCs.

MicroRNA profile of tumorigenic cells during carcinogenesis of lung adenocarcinoma.

To obtain microRNA (miRNA) profile and clarify their biological function in tumorigenic Sca-1(+) CD34(+) cells during carcinogenesis of lung adenocarcinoma. After intranasal infection with recombinant Adeno-Cre viruses (AdV-Cre), lung adenocarcinoma was identified pathologically in Lox-stop-lox Kras (LSL-Kras) G12D mice. Sca-1(+) CD34(+) cells were sorted by flow cytometry and tested for tumor-initiating ability, self-renewal and tumorigenicity. MiRNA profiles were obtained using microarray and further confirmed by real-time RT-PCR (qRT-PCR). MiRNA functions were predicted bioinformatically, and miR-294 function was verified to explore its role in tumor migration and invasion. Lung adenocarcinoma was induced in LSL-Kras G12D mice within 30 days. In vivo, the tumorigenicity of Sca-1(+) CD34(+) cells was 25 times stronger than Sca-1(-) CD34(-) cells. During tumorigenesis of lung adenocarcinoma, the expression of 145 miRNAs in Sca-1(+) CD34(+) cells increased and 72 miRNAs decreased (P < 0.01). Four successively up-regulated miRNAs (miR-15a*, miR-203, miR-294 and miR-295*) and three successively down-regulated ones (miR-19b, miR-483 and miR-615-5p) were identified. Among them, miR-294 could constitutively bind to 3'-UTR of matrix metalloproteinase 3 (MMP3), and down-regulate MMP3 protein expression. MiR-294 also significantly inhibited migration and invasion of Lewis lung cancer cells. MiRNAs are characteristically expressed in tumor-initiating Sca-1(+) CD34(+) cells of lung adenocarcinoma, and may play important roles during the carcinogenesis of lung adenocarcinoma.

Efficacy and safety of endostar® combined with chemotherapy in patients with advanced soft tissue sarcomas.

BACKGROUND: Soft tissue sarcomas (STS) are a heterogeneous group of tumors, and approximately 40-50% of patients with STS develop metastatic disease. The median overall survival of those patients was 12 months and their 5-year survival rate was 8%. Therefore, study on more effective treatment, especially the targeting therapies, is urgently needed. OBJECTIVE: To evaluate the efficacy and safety of Endostar® combined with chemotherapy in patients with advanced STS. METHODS: A retrospective case-series study was conducted in Cancer Institute of PLA, Xinqiao Hospital. A total of 71 patients suffering from advanced STS (IIB - IV) were included, of whom 49 cases treated with chemotherapy alone were defined as the control group and the rest 22 cases treated with the traditional chemotherapy combined with Endostar® were defined as the test group. The short-term therapeutic effects including objective response rate (ORR), disease control rate (DCR) and safety were evaluated in the two groups. In the follow-up, progression-free survival (PFS) and overall survival (OS) were also observed. RESULTS: In the test and control groups, the ORR was 18.2% and 12.2%, respectively (P = 0.767), and the DCR was 86.4% and 61.2%, respectively (P=0.034). The median time to progression in the test and control groups was 120 days and 70 days with significant difference (P = 0.017), while the median overall survival was 452 days and 286 days without significant difference (P = 0.503). The one-year survival rate in the test group and control group was 56.2% and 35.4%, respectively, while the two-year survival rate was 30.2% and 26.5%, respectively. No significant difference in the side effects was found between the two groups. CONCLUSIONS: Endostar® combined with chemotherapy resulted in a higher DCR and longer PFS in the patients with advanced STS, and the toxicity was tolerable.

Morphological and molecular characterisation of Heliconema hainanensis sp. nov. (Spirurina: Physalopteridae) from congers in the South China Sea, with a key to the species of Heliconema

Heliconema hainanensis sp. nov. collected from Uroconger lepturus (Richardson) (Anguilliformes: Congridae), Muraenesox cinereus (Forsskål) and Congresox talabonoides (Bleeker) (Anguilliformes: Muraenesocidae) in the South China Sea was described using light and scanning electron microscopy. The new species differs from its congeners by the following morphology: pseudolabia, the number and arrangement of caudal papillae (4 pairs of pedunculate precloacal papillae arranged in 2 groups of 2 and 2 pairs and 6 pairs of pedunculate postcloacal papillae arranged in 4 groups of 1, 2, 1 and 2 pairs), the length of spicules [left spicule 0.51-0.69 mm, right spicule 0.20-0.27 mm, spicule (right:left) ratio 1:2.20-2.69] and the morphology of the female tail tip. In addition, specimens of the new species collected from the three different hosts and specimens of an unidentified species of Heliconema collected from U. lepturus were characterised using molecular methods by sequencing the internal transcribed spacer (ITS) of ribosomal DNA. Analyses and comparison of the ITS sequence of H. hainanensis sp. nov. with Heliconema sp. support the validity of the new species based on morphological observations. An identification key to the species of Heliconema is also provided.

Neural stem cells enhance nerve regeneration after sciatic nerve injury in rats.

With the development of tissue engineering and the shortage of autologous nerve grafts in nerve reconstruction, cell transplantation in a conduit is an alternative strategy to improve nerve regeneration. The present study evaluated the effects and mechanism of brain-derived neural stem cells (NSCs) on sciatic nerve injury in rats. At the transection of the sciatic nerve, a 10-mm gap between the nerve stumps was bridged with a silicon conduit filled with 5 × 10(5) NSCs. In control experiments, the conduit was filled with nerve growth factor (NGF) or normal saline (NS). The functional and morphological properties of regenerated nerves were investigated, and expression of hepatocyte growth factor (HGF) and NGF was measured. One week later, there was no connection through the conduit. Four or eight weeks later, fibrous connections were evident between the proximal and distal segments. Motor function was revealed by measurement of the sciatic functional index (SFI) and sciatic nerve conduction velocity (NCV). Functional recovery in the NSC and NGF groups was significantly more advanced than that in the NS group. NSCs showed significant improvement in axon myelination of the regenerated nerves. Expression of NGF and HGF in the injured sciatic nerve was significantly lower in the NS group than in the NSCs and NGF groups. These results and other advantages of NSCs, such as ease of harvest and relative abundance, suggest that NSCs could be used clinically to enhance peripheral nerve repair.

Comparative study of transvaginal ultrasonographic and diagnostic hysteroscopic findings in postmenopausal breast cancer patients treated with tamoxifen.

BACKGROUND: There is an association between postmenopausal tamoxifen therapy and endometrial pathologies. We investigated the usefulness of diagnostic hysteroscopy and transvaginal ultrasonography (TVS) and estimated whether diagnostic hysteroscopy improves detection of endometrial pathologies in postmenopausal breast cancer patients on tamoxifen. METHODS: Ninety-seven postmenopausal breast cancer patients who had been taking tamoxifen 20 mg/d for ≥ 6 months went through TVS, diagnostic hysteroscopy, and endometrial biopsy examinations. The presence of endometrial histopathologic features with abnormal TVS and diagnostic hysteroscopic findings were correlated. RESULTS: No endometrial cancer was found in any of the 97 patients. Fifty-three patients (54.6%) developed endometrial polyps as diagnosed histopathologically. Fifty-nine patients (60.8%) tested positive in TVS exams, of whom 43 had polyps, four had hyperplasia, and 12 atrophy. Thirty-eight patients (39.2%) tested negative in TVS exams, of whom 10 had polyps, three hyperplasia, and 25 atrophy. TVS exams presented 63.6% specificity, 81.8% sensitivity, 72.9% positive-predictive value, and 73.7% negative-predictive value, whereas the corresponding values of diagnostic hysteroscopy were 100%, 98.1%, 100%, and 97.8% respectively. The correct ratio of hysteroscopy was significantly higher than that of TVS (P = 0.000). CONCLUSIONS: In postmenopausal breast cancer patients treated with tamoxifen, TVS alone is not sufficient for the detection of endometrial pathologies. Additional use of diagnostic hysteroscopy considerably improves the detection of polyps, thus significantly reducing the rate of false-negative findings of endometrial pathologies.

[Expression of B7-H1/PD-1 in mucosa of chronic rhinosinusitis].

OBJECTIVE: To explore the expression of B7-H1/PD-1 and the localization of these proteins in human sinus mucosa of chronic rhinosinusitis (CRS). METHODS: Twenty-eight patients with CRS (17 with nasal polyps (CRSwNP) and 11 without (CRSsNP)) and 11 control patients were recruited. B7-H1/PD-1 expressions in sinus mucosa among these 3 groups were detected by immunofluorescent staining, real-time quantitative PCR and Western blot. Flow cytometry was also employed to identify the type of B7-H1 or PD-1 positive cells in nasal polyps. RESULTS: Compared with normal mucosa, B7-H1/PD-1 mRNA was highly expressed in sinonasal mucosa of CRSwNP and CRSsNP (P < 0.01). There was no difference between CRSwNP and CRSsNP (P > 0.05). The protein level of B7-H1/PD-1 was similar to mRNA level. As indicated by immunofluorescent assay, B7-H1 was mainly expressed in sinonasal epithelial cells, submucosal gland cells and inflammatory cells within the subepithelial layer while PD-1 was expressed in inflammatory cells within the subepithelial layer. The B7-H1 or PD-1 positive cells in nasal polyps were partially CD19+ B cells and CD3+ T cells respectively. CONCLUSION: B7-H1/PD-1 is up-regulated in sinus mucosa of CRS. B7-H1/PD-1 may play an important role in the pathogenesis of CRS.