Reliability and spatial specificity of rat brain sensorimotor functional connectivity networks are superior under sedation compared with general anesthesia

Functional connectivity networks derived from resting‐state functional MRI (rsfMRI) have received increasing interest to further our understanding of brain function. The anesthesia in rodent models may influence the interpretation and comparison of results from functional connectivity MRI (fcMRI). More research is required on this aspect. In this study, we investigated rat brain connectivity networks under 1.5% isoflurane anesthesia in comparison with medetomidine sedation. rsfMRI data were acquired under both anesthesia conditions within one imaging session. Male Wistar rats (n = 17) were scanned at 11.7 T with focus on the sensorimotor system. The data underwent a per‐subject independent component analysis (ICA), after which individual components were grouped using hierarchical clustering. Consistent and reliable networks were identified under medetomidine in sensorimotor cortex (three networks) and striatum (two networks). The incidence of these networks was drastically reduced under isoflurane. Seed correlation analysis confirmed these results and revealed globally elevated correlations with low topical specificity under isoflurane, stemming from low‐frequency global signal fluctuations. Global signal removal thus enhanced slightly regional specificity under isoflurane and showed anti‐correlations of cortico‐striatal connections in both anesthesia regimes. Functional connectivity networks are thus reliably detected in medetomidine‐sedated animals on an individual basis using ICA. Their occurrence, however, is heavily compromised under isoflurane as a result of global signal fluctuations potentially stemming from burst‐suppression‐like neural activity. Anesthesia and pharmacologically induced modulations may provide insight into network mechanisms in the future. As an agent for fcMRI in brain disease studies, light sedation using medetomidine preserves connectivity networks in a greater level of detail, and may therefore be considered superior to standard isoflurane anesthesia. Copyright © 2013 John Wiley & Sons, Ltd.     

Adjuvant treatment of pancreatic carcinoma in a clinically adapted mouse resection model.

BACKGROUND: The high rate of local recurrence after radical resection of pancreatic adenocarcinoma fosters intensive efforts to develop new approaches for adjuvant treatment. The established animal models show significant limitations in simulating an adjuvant therapeutic setting. For optimal approximation to the clinical situation we therefore improved a murine orthotopic human xenotransplantation model. METHODS: Subtotal pancreatectomy in mice was performed after orthotopic inoculation of human pancreatic cancer cells and manifestation of solid tumours. The natural course of disease, tumour growth and metastases were analysed. Gemcitabine as a cytotoxic drug was tested in vitro on the cell line used in this model and the effect of adjuvant treatment with gemcitabine in vivo was investigated. RESULTS: All tumour-resected animals showed local recurrence. Organ metastases occurred in 67% in resected compared to 25% of non-resected animals. Gemcitabine in vitro was ineffective, but as adjuvant monotherapy resulted in a highly significant reduction of tumour weight and metastatic events. CONCLUSION: Subtotal pancreatectomy for xenotransplanted pancreatic cancer in SCID beige mice is feasible. Due to high rates of local recurrence and increased organ metastases, this model offers a relevant option for preclinical adjuvant testing, especially as in vitro and in vivo effects of cytotoxic drugs differ enormously.     

TRAIL-R2 promotes skeletal metastasis in a breast cancer xenograft mouse model

Despite improvements in detection, surgical approaches and systemic therapies, breast cancer remains typically incurable once distant metastases occur. High expression of TRAIL-R2 was found to be associated with poor prognostic parameters in breast cancer patients, suggesting an oncogenic function of this receptor. In the present study, we aimed to determine the impact of TRAIL-R2 on breast cancer metastasis. Using an osteotropic variant of MDA-MB-231 breast cancer cells, we examine the effects of TRAIL-R2 knockdown in vitro and in vivo. Strikingly, in addition to the reduced levels of the proliferation-promoting factor HMGA2 and corresponding inhibition of cell proliferation, knockdown of TRAIL-R2 increased the levels of E-Cadherin and decreased migration. In vivo, these cells were strongly impaired in their ability to form bone metastases after intracardiac injection. Evaluating possible underlying mechanisms revealed a strong downregulation of CXCR4, the receptor for the chemokine SDF-1 important for homing of cancers cells to the bone. In accordance, cell migration towards SDF-1 was significantly impaired by TRAIL-R2 knockdown. Conversely, overexpression of TRAIL-R2 upregulated CXCR4 levels and enhanced SDF-1-directed migration. We therefore postulate that inhibition of TRAIL-R2 expression could represent a promising therapeutic strategy leading to an effective impairment of breast cancer cell capability to form skeletal metastases.     

尿激酶型纤溶酶原激活因子及抑制剂表达与人大肠癌细胞转移能力的关系

目的　探讨尿激酶型纤溶酶原激活因子 ( u PA)、尿激酶型纤溶酶原激活因子受体 ( u PAR)和纤溶酶原抑制剂 1( PAI- 1)的表达与人大肠癌细胞系转移能力的关系。方法　用 EL ISA方法测定 3个人大肠癌细胞系培养上清液中 u PA、u PAR和 PAI- 1含量 ;用免疫组化 ABC方法检测 u PA、u PAR和 PAI- 1在细胞中的表达 ;分析其表达与大肠癌细胞转移能力的关系。结果　在培养上清液中具有高转移能力的 HT- 2 9d细胞的 u PA、u PAR及 PAI- 1含量明显高于低转移的 HT- 2 9和不转移的 Wi Dr细胞 ,而 HT- 2 9细胞的 u PA、u PAR及 PAI- 1含量则高于 Wi Dr细胞。免疫组化显示 u PA和 PAI- 1在 HT- 2 9d细胞中的表达高于 HT- 2 9和 Wi Dr细胞。结论　 u PA、u PAR和 PAI-1的表达与大肠癌细胞的转移能力密切相关。     

Influence of tumor-associated E-cadherin mutations on tumorigenicity and metastasis

In this study, we investigated whether tumor-associated E-cadherin mutations impair the tumor-suppressive function of the cell adhesion molecule and influence metastasis formation in a severe combined immunodeficiency mouse model. The investigated E-cadherin mutations were in frame deletions of exons 8 (del 8) or 9 (del 9) and a point mutation in exon 8 (p8). Transfected human MDA-MB-435S carcinoma cells stably expressing wild-type (wt) or mutant E-cadherin were injected into the mouse mammary fat pad. Mice transplanted with wt E-cadherin transfectants developed significantly smaller tumors than animals transplanted with the E-cadherin-negative parental cell line. Animals transplanted with del 9 or p8 E-cadherin transfectants produced medium size tumors, indicating that these mutations impair the tumor-suppressive function of E-cadherin. In contrast, mice transplanted with del 8 E-cadherin transfectants developed tumors of approximately the same sizes as animals transplanted with wt E-cadherin expressing cells. Lung metastases were induced by all cell lines without significant differences. Immunohistochemical analysis of E-cadherin expression in the tumors revealed a heterogeneous staining pattern, indicating loss or down-regulation of E-cadherin in some tumor cells. Metastases were completely negative for E-cadherin. Our data suggest that the type of mutation determines whether the tumor-suppressive function of E-cadherin is impaired.     

Combretastatin A-4 prodrug: a potent inhibitor of malignant hemangioendothelioma cell proliferation

Anti-vascular treatment by targeting proliferating endothelial cells has become a promising option in anti-neoplastic therapy. Combretastatin A-4 prodrug (CA-4PD) has been identified as a selective inhibitor of endothelial cell proliferation, acting by the interruption of microtubule assembly. In this study, the effect of CA-4PD on proliferating endothelial cells derived from a primary tumor of the vascular endothelium was investigated in vitro and in vivo. In vitro, CA-4PD was an effective inhibitor of endothelial cell proliferation in a time- and dose-dependent manner, displaying a certain selectivity toward endothelial cells in comparison to proliferating fibroblasts. Analysis of DNA profiles by FACS revealed an increasing proportion of cells arrested in the G(2) cell-cycle phase with correlation to the duration of drug exposure. A decrease in cell viability correlated with duration of drug exposure, whereas FACS analysis, DNA fragmentation assay, and DNA gel electrophoresis failed to demonstrate that DNA fragmentation was indicative of apoptosis up to 48 hr of continued drug exposure. In vivo, CA-4PD induced excessive regressive alterations in experimentally allotransplanted hemangioendotheliomas within 24 hr after singular i. p. injection of 100 mg CA-4PD/kg body weight. This represented less than one-tenth of the maximum tolerated dose. In conclusion, our findings characterize CA-4PD as a potent inhibitor of malignant endothelial cell proliferation in vitro, effecting arrest of proliferating cells in the G(2) cell-cycle phase with subsequent cell death on a pathway different from apoptosis. in vivo, CA-4PD induces extensive intratumoral cell loss within 24 hr following systemic administration, suggesting a synergistic effect of direct cell killing and the induction of vascular shutdown.     

TLR-3 polymorphism is an independent prognostic marker for stage II colorectal cancer

Background

Clinicopathologic stage is still the main parameter to evaluate the prognosis of newly diagnosed colorectal cancer (CRC) patients. Although molecular markers have been suggested for follow up of treated CRC patients, their complete clinical application is still under evaluation.

Materials and methods

To evaluate the association of immune-related genes with CRC prognosis and survival, a total of 19 single nucleotide polymorphisms (SNPs) were genotyped in 614 German patients within the Kiel cohort (POPGEN).

Results

A promoter variant (rs1800872) in the Interleukin-10 (IL-10) gene was associated with an increased lymph node metastasis involvement [odds ratio (OR) = 2.1, 95% confidence interval (CI) = 1.03-4.2, for carriers of the TT genotype]. More importantly, among 582 followed up patients the SNP rs3775291 in the toll-like receptor 3 (TLR-3) gene was associated with CRC specific survival (150 events). Patients carrying the TT genotype had a 93% increased risk of death compared with the CC carriers [hazard ratio (HR) = 1.93, 95% CI 1.14-3.28]. The observed effect of the TLR-3 variant was restricted to stage II patients (HR = 4.14, 95% CI 1.24-13.84) and to patients who did not receive adjuvant therapy (HR = 3.2, 95% CI 1.4-7.7).

Conclusions

Our results may provide additional candidates for risk assessment in stage II CRC patients for treatment decision. Further validation of the presented findings is warranted.     

Epicatechin gallate and catechin gallate are superior to epigallocatechin gallate in growth suppression and anti-inflammatory activities in pancreatic tumor cells

Green tea catechins are considered as possible cancer preventive agents for several cancer types but little is known regarding their effects on pancreatic cancer cells. The best studied catechin and the major polyphenol present in green tea is epigallocatechin gallate (EGCG). In the present study, we investigated the in vitro anti-tumoral properties of EGCG on human pancreatic ductal adenocarcinoma (PDAC) cells PancTu-I, Panc1, Panc89 and BxPC3 in comparison with the effects of two minor components of green tea catechins, catechin gallate (CG) and epicatechin gallate (ECG). We found that all three catechins inhibited proliferation of PDAC cells in a dose- and time-dependent manner. Interestingly, CG and ECG exerted much stronger anti-proliferative effects than EGCG. Western blot analyses performed with PancTu-I cells revealed catechin-mediated modulation of cell cycle regulatory proteins (cyclins, cyclin-dependent kinases [CDK], CDK inhibitors). Again, these effects were clearly more pronounced in CG or ECG than in EGCG-treated cells. Importantly, catechins, in particular ECG, inhibited TNFα-induced activation of NF-κB and consequently secretion of pro-inflammatory and invasion promoting proteins like IL-8 and uPA. Overall, our data show that green tea catechins ECG and CG exhibit potent and much stronger anti-proliferative and anti-inflammatory activities on PDAC cells than the most studied catechin EGCG.