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Establishment of a model of sentinel lymph node metastasis using immunodeficient swine.

ABSTRACT: Lymph node metastasis occurs via the migration of cancer cells through the lymphatic system. Sentinel lymph node (SLN) biopsy is a common diagnostic strategy. SLNs have been studied using healthy rodents and large animals without metastasis. Here we used immunodeficient swine to establish a model of lymph node metastasis. We used RAG2-knockout immunodeficient swine. A431 human epithelial carcinoma cells expressing green fluorescent protein were injected subcutaneously into the posterior sides of the auricle, forelimb and hindlimb of knockout swine. Indigo carmine dye was injected subcutaneously 8 weeks after tumour cell transplantation. SLNs were extracted, observed using a stereoscopic fluorescence microscope and analysed histologically using haematoxylin and eosin staining, and immunohistochemistry. Lymphoid follicles were found in wild-type swine, and a few aggregated lymphocytes and immature lymphoid follicles were observed in knockout swine. Fluorescence in the lymph nodes indicated metastasis of tumour cells to the lymph nodes. Tumour cells replaced lymph node architectures, showed high-grade nuclear atypia and formed irregular tumour nests. Our model may be useful for the preclinical validation of diagnostic methods and minimally invasive treatment of metastatic cancer.

SUBMITTER: Kurihara T

PROVIDER: S-EPMC6538626 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Establishment of a model of sentinel lymph node metastasis using immunodeficient swine.

Kurihara Toshiaki T Matsuda Sachiko S Nakamura Yuki Y Suzuki Shunichi S Fuchimoto Daiichiro D Onishi Akira A Saeki Kohei K Nakagawa Takayuki T Fujiwara Reina R Kamata Masatoshi M Kuramoto Junko J Kameyama Kaori K Sekino Masaki M Kusakabe Moriaki M Hayashida Tetsu T Jinno Hiromitsu H Kitagawa Yuko Y

Scientific reports 20190528 1

Lymph node metastasis occurs via the migration of cancer cells through the lymphatic system. Sentinel lymph node (SLN) biopsy is a common diagnostic strategy. SLNs have been studied using healthy rodents and large animals without metastasis. Here we used immunodeficient swine to establish a model of lymph node metastasis. We used RAG2-knockout immunodeficient swine. A431 human epithelial carcinoma cells expressing green fluorescent protein were injected subcutaneously into the posterior sides of ...[more]

PMID: 31138877

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Project description:BackgroundSentinel lymph node metastasis is a common and early event in the metastatic process of head and neck squamous cell carcinoma (HNSCC) and is the most powerful prognostic factor for survival of HNSCC patients. 3-O-acetyloleanolic acid (3AOA), a pentacyclic triterpenoid compound isolated from seeds of Vigna sinensis K., has been reported to have potent anti-angiogenesis and anti-tumor activities. However, its effects on tumor-related lymphangiogenesis and lymph node metastasis are not yet understood.MethodsThe in vitro inhibitory effects of 3AOA on VEGF-A-induced lymphangiogenesis were investigated via in vitro experiments using mouse oral squamous cell carcinoma (SCCVII) cells and human lymphatic microvascular endothelial cells (HLMECs). The in vivo inhibitory effects of 3AOA on VEGF-A-induced lymphangiogenesis and sentinel lymph node metastasis were investigated in an oral cancer sentinel lymph node (OCSLN) animal model.Results3AOA inhibited tumor-induced lymphangiogenesis and sentinel lymph node metastasis in an OCSLN animal model, and reduced expression of VEGF-A, a lymphangiogenic factor in hypoxia mimetic agent CoCl2-treated SCCVII cells. 3AOA inhibited proliferation, tube formation, and migration of VEGF-A-treated HLMECs. The lymphatic vessel formation that was stimulated in vivo in a by VEGF-A Matrigel plug was reduced by 3AOA. 3AOA suppressed phosphorylation of vascular endothelial growth factor (VEGFR) -1 and - 2 receptors that was stimulated by VEGF-A. In addition, 3AOA suppressed phosphorylation of the lymphangiogenesis-related downstream signaling factors PI3K, FAK, AKT, and ERK1/2. 3AOA inhibited tumor growth, tumor-induced lymphangiogenesis, and sentinel lymph node metastasis in a VEGF-A-induced OCSLN animal model that was established using VEGF-A overexpressing SCCVII cells.Conclusion3AOA inhibits VEGF-A-induced lymphangiogenesis and sentinel lymph node metastasis both in vitro and in vivo. The anti-lymphangiogenic effects of 3AOA are probably mediated via suppression of VEGF-A/VEGFR-1 and VEGFR-2 signaling in HLMECs, and can be a useful anti-tumor agent to restrict the metastatic spread of oral cancer.

Project description:Purpose: Sentinel lymph node biopsy (SLNB) has emerged as the preferred standard procedure in patients with breast cancer, melanoma and other types of cancer. Herein, we developed a method to intra-operatively map SLNs and differentiate tumor metastases within SLNs at the same time, with the aim to provide more accurate and real-time intraoperative guidance. Experimental Design: Hyaluronic acid (HA), a ligand of lymphatic vessel endothelial hyaluronan receptor (LYVE)-1, is employed as a SLN mapping agent after being conjugated with a near-infrared fluorophore (Cy5.5). Different sized HAs (5, 10 and 20K) were tested in normal mice and mice with localized inflammation to optimize LN retention time and signal to background ratio. Cetuximab, an antibody against epidermal growth factor receptor (EGFR), and trastuzumab, an antibody against human epidermal growth factor receptor 2 (HER2), were labeled with near-infrared fluorophore (IRDye800) for detecting metastatic tumors. LN metastasis model was developed by hock injection of firefly luciferase engineered human head neck squamous carcinoma cancer UM-SCC-22B cells or human ovarian cancer SKOV-3 cells. The metastases within LNs were confirmed by bioluminescence imaging (BLI). IRDye800-Antibodies were intravenously administered 24 h before local administration of Cy5.5-HA. Optical imaging was then performed to identify nodal metastases. Results: Binding of HA with LYVE-1 was confirmed by ELISA and fluorescence staining. HA with a size of 10K was chosen based on the favorable migration and retention profile. After sequential administration of IRDye800-antibodies intravenously and Cy5.5-HA locally to a mouse model with LN metastases and fluorescence optical imaging, partially metastasized LNs were successfully distinguished from un-metastasized LNs and fully tumor occupied LNs, based on the different signal patterns. Conclusions: Fluorophore conjugated HA is a potential lymphatic mapping agent for SLNB. Dual-tracer imaging with the combination of lymphatic mapping agents and tumor targeting agents can identify tumor metastases within SLNs, thus may provide accurate and real-time intra-operative guidance to spare the time spent waiting for a biopsy result.

Project description:Background The rate of breast-conserving surgery is very low in China, compared with that in developed countries; most breast cancer patients receive mastectomy. It is great important to explore the possibility of omitting axillary lymph node dissection (ALND) in early-stage breast cancer patients with 1 or 2 positive sentinel lymph nodes (SLNs) in China. The aim of this study was to develop a nomogram based on elastography for the prediction of the risk of non-SLN (NSLN) metastasis in early-stage breast cancer patients with 1 or 2 positive SLNs. Methods A total of 601 breast cancer patients were initially recruited. According to the inclusion and exclusion criteria, 118 early-stage breast cancer patients with 1 or 2 positive SLNs were finally enrolled and were assigned to the training cohort (n=82) and the validation cohort (n=36), respectively. In the training cohort, the independent predictors were screened by logistic regression analysis and then were used to conducted the nomogram for the prediction of NSLN metastasis in early-stage breast cancer patients with 1 or 2 positive SLNs. The calibration curves, concordance index (C-index), the area under the receiver operating characteristic (ROC) curve (AUC), and Decision curve analysis (DCA) were used to verified the performance of the nomogram. Results The multivariable analysis showed that the enrolled patients with positive HER2 expression (OR=6.179, P=0.013), Ki67≥14% (OR=8.976, P=0.015), larger lesion size (OR=1.038, P=0.045), and higher Emean (OR=2.237, P=0.006) were observed to be the independent factors of NSLN metastasis. Based on the above four independent predictors, a nomogram was conducted to predict the risk of the NSLN metastasis in early-stage breast cancer patients with 1 or 2 positive SLNs. The nomogram showed good discrimination in the prediction of NSLN metastasis, with bias-corrected C-index of 0.855 (95% CI, 0.754-0.956) and 0.853 (95% CI, 0.724-0.983) in the training and validation cohorts, respectively. Furthermore, the AUC was 0.877 (95%CI: 0.776- 0.978) and 0.861 (95%CI: 0.732-0.991), respectively, indicating a good performance of the nomogram. The calibration curve suggested a satisfactory agreement between the predictive and actual risk in both the training (χ2 = 11.484, P=0.176, HL test) and validation (χ2 = 6.247, p = 0.620, HL test) cohorts, and the obvious clinical nets were revealed by DCA. Conclusions We conducted a satisfactory nomogram model to evaluate the risk of NSLN metastasis in early-stage breast cancer patients with 1 or 2 SLN metastases. This model could be considered as an ancillary tool to help such patients to be selectively exempted from ALND.

Project description:BackgroundIn recent years, sentinel lymph node excision and ultrastaging have been performed in endometrial carcinomas to obtain information about lymph node status, avoiding unnecessary complete pelvic and paraaortic lymphadenectomy. The purpose of this retrospective study was to provide a comprehensive evaluation of the pathological features of endometrial carcinomas and their significance in association with sentinel lymph node involvement.MethodsPatients with endometrial carcinomas, preceded by sentinel lymph node mapping, were classified into Group-I and Group-II with negative and positive involvement, respectively. The pathological features, associated with sentinel lymph node involvement, were statistically analyzed, including determination of test performance parameters.ResultsAmong 70 patients who had undergone hysterectomy and sentinel lymph node excision, 61 had carcinoma and 9 had atypical hyperplasia. There were 50 patients in Group-I and 10 in Group-II. In Group-II, the significant pathological features were: 1) lower uterine segment involvement (100%), 2) an average tumor size of ≥5 CM, 3) lymphovascular invasion (50%), 4) cervical stromal invasion (40%), and 5) depth of myometrial invasion of ≥50% (50%). The incidences of these pathological features were significantly less in Group-I. Statistical analyses singled out "lower uterine segment involvement" as the most important feature.ConclusionsWe have identified five pathological features which are associated with sentinel lymph node involvement. Since lower uterine segment involvement has occurred in all cases of the Group-II cohort, we recommend FIGO and other organizations that determine staging rules should consider whether tumors that involve the lower uterine segment should be staged as higher than "1a", if the findings in this small series are confirmed by other studies. The results of this study may guide pathologists and oncologists in the diagnostic and therapeutic approaches to management of endometrial carcinomas.

Dataset Information

Establishment of a model of sentinel lymph node metastasis using immunodeficient swine.

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Establishment of a model of sentinel lymph node metastasis using immunodeficient swine.

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