MMP-2 and TIMP-2 Expression Correlates with Poor Prognosis in Cervical Carcinoma—A Clinicopathologic Study Using Immunohistochemistry and mRNA in Situ Hybridization

Objective. The spread of malignant neoplasms is closely associated with matrix and basement membrane degradation, mediated by various classes of proteolytic enzymes. Matrix metalloproteinases (MMP) appear to have a key role in the sequence of events that lead to local invasion and metastasis. The present study evaluated the role of matrix metalloproteinase-2 (MMP-2), tissue inhibitor of metalloproteinases-2 (TIMP-2), and membrane-type metalloproteinase (MT1-MMP) in cervical neoplasia.Methods. We have analyzed 49 uterine cervical squamous cell carcinomas, 10 cases of high-grade cervical intraepithelial neoplasia (CIN II–III), and 10 control cervices for the presence of MMP-2, TIMP-2, and MT1-MMP using in situ hybridization. MMP-2 protein expression was evaluated using immunohistochemistry. Results were analyzed for possible correlation with disease outcome.Results. MMP-2, TIMP-2, and MT1-MMP mRNA were localized to both stromal and tumor cells. However, an intense signal for MMP-2 was detected almost exclusively in tumor cells and was uniformly absent from CIN lesions and control cervices. Conversely, intense signals for TIMP-2 and MT1-MMP were detected in both stromal and tumor cells of invasive carcinomas, more often for the former. As with MMP-2, they were absent from CIN lesions. MMP-2 protein expression was enhanced in tumor cells compared to CIN cases and controls, significantly compared to the latter (P = 0.01). The presence of both MMP-2 and TIMP-2 mRNA in tumor cells correlated with advanced stage (P = 0.003 for MMP-2, P = 0.002 for TIMP-2) and with poor survival (P = 0.003 for MMP-2, P = 0.002 for TIMP-2) in univariate analysis. In addition, their presence in tumor cells intercorrelated (P = 0.002). In multivariate survival analysis, MMP-2 presence retained its association with survival (P = 0.004), in addition to patient age (P = 0.027) and advanced stage (P = 0.0002).Conclusions. Both MMP-2 and TIMP-2 have a key role in extracellular matrix invasion in cervical carcinoma, largely through their elaboration by tumor cells. The presence of mRNA for both proteins is interrelated and is associated with poor survival.     

The immediate and short‐term effects of bilateral intrahippocampal depth electrodes on verbal memory

In contrast to previous studies, Ljung et al. provide evidence of permanent cognitive consequences of bilateral intrahippocampal depth electrodes for verbal memory in patients who were not operated or operated in the right temporal lobe. Stimulated by this, we provide historical confirmatory and supplementary evidence of the detrimental effect of bilateral depth electrodes implanted along the longitudinal axis of the hippocampus on verbal learning and especially on delayed verbal memory and recognition performance. This is demonstrated in 31 patients with memory assessments before implantation, after explantation, and 3 months later after left/right temporal lobe surgery. After surgery, significant recovery from postimplantation impairment is found in right temporal patients. Left temporal resection patients stay on the level seen after implantation and do not recover. Surgery, however, has its own effects in addition to the implantation. Intracranial electrodes for electroencephalographic monitoring or electrical stimulation are commonly and increasingly used for diagnosis or treatment in pharmacoresistant epilepsies. Thus, the monitoring of invasive stereotactic approaches is recommended to find safe procedures for the patients. In response to the findings, we restricted indications and used different implantation schemes, different trajectories, and targets to minimize the risk of additional damage.     

New records of Culicoides species (Diptera: Ceratopogonidae) for Bolivia

Culicoides debilipalpis, C insignis, C. lahillei, and C venezuelensis are reported for the first time for Bolivia. The geographical distribution of C. paraensis extends to the Tarija Department.     

Synthesis and Ex Vivo Autoradiographic Evaluation of Ethyl-��-d-galactopyranosyl-(1,4��)-2��-deoxy-2��-[18F]fluoro-��-d-glucopyranoside��A Novel Radioligand for Lactose-Binding Protein: Implications for Early Detection of Pancreatic Carcinomas with PET

Introduction

Previous studies demonstrated that the lactose-binding protein (hepatocellular carcinoma?Cintestine?Cpancreas and pancreatitis-associated proteins (HIP/PAP)) is upregulated >130 times in peritumoral pancreatic tissue as compared to normal pancreatic tissue. Therefore, we developed a new radiolabeled ligand of HIP/PAP, the ethyl-??-d-galactopyranosyl-(1,4??)-2??-deoxy-2??-[¹⁸F]fluoro-??-d-glucopyranoside (Et-[¹⁸F]FDL) for noninvasive imaging of pancreatic carcinoma using positron emission tomography and computerized tomography (PET/CT).

Methods

The novel precursor and radiolabeling methods for synthesis of Et-[¹⁸F]FDL produced no isomers; the average decay-corrected radiochemical yield was 68%, radiochemical purity >99%, and specific activity >74 GBq/µmol. The radioligand properties of Et-[¹⁸F]FDL were evaluated using an ex vivo autoradiography and immunohistochemistry in pancreatic tissue sections obtained from mice-bearing orthotopic pancreatic tumor xenografts.

Results and Discussion

Et-[¹⁸F]FDL binding to peritumoral pancreatic tissue sections strongly correlated with HIP/PAP expression (r?=?0.81) and could be completely blocked by treatment with 1 mM lactose.

Conclusion

These results suggest that Et-[¹⁸F]FDL is a promising agent which should be evaluated for detection of early pancreatic carcinomas by PET/CT imaging.     

Pharmacokinetics and magnetic resonance imaging of biodegradable macromolecular blood‐pool contrast agent PG–Gd in non‐human primates: a pilot study

The purpose of this study was to evaluate poly(L ‐glutamic acid)‐benzyl‐DTPA–Gd (PG–Gd), a new biodegradable macromolecular magnetic resonance imaging contrast agent, for its pharmacokinetics and MRI enhancement in nonhuman primates. Studies were performed in rhesus monkeys at intravenous doses of 0.01, 0.02 and 0.08 mmol Gd/kg. T₁‐weighted MR images were acquired at 1.5 T using fast spoiled gradient recalled echo and fast spin echo imaging protocols. The small‐molecule contrast agent Magnevist was used as a control. PG–Gd in the monkey showed a bi‐exponential disposition. The initial blood concentrations within 2 h of PG–Gd administration were much higher than those for Magnevist. The high blood concentration of PG–Gd was consistent with the MR imaging data, which showed prolonged circulation of PG–Gd in the blood pool. Enhancement of blood vessels and organs with a high blood perfusion (heart, liver, and kidney) was clearly visualized at 2 h after contrast injection at the three doses used. A greater than proportional increase of the area under the blood concentration–time curve was observed when the administered single dose was increased from 0.01 to 0.08 mmol/kg. By 2 days after PG–Gd injection, the contrast agent was mostly cleared from all major organs, including kidney. The mean residence time was 15 h at the 0.08 mmol/kg dose. A similar pharmacokinetic profile was observed in mice, with a mean residence time of 5.4 h and a volume of distribution at steady‐state of 85.5 ml/kg, indicating that the drug was mainly distributed in the blood compartment. Based on this pilot study, further investigations on the potential systemic toxicity of PG–Gd in both rodents and large animals are warranted before testing this agent in humans. Copyright © 2011 John Wiley & Sons, Ltd.     

A novel method for analyzing images of live nerve cells

Analysis of images from live-cell experiments is a central activity to studying the effects of stimulation on neuronal behavior. Image analysis techniques currently used to study these effects rely for the most part on the salience of the neuronal structures within the image. In both fluorescent and electron microscopy, neuronal structures are enhanced and therefore easy to distinguish in an image. Unlike images obtained via fluorescent or electron microscopy, however, images produced via transmission microscopy (e.g., bright field, phase contrast, DIC) are significantly more difficult to analyze because there is little contrast between the object-of-interest and the image background. This difficulty is amplified when a time-dependent sequence of images are to be analyzed, because of the corresponding large data sets. To address this problem, we introduce a novel approach to the analysis of images of live cells captured via transmission microscopy that takes advantage of commercially available software and the Fourier transform. Specifically, our approach utilizes several morphological functions in MATLAB to enhance the contrast of the cells with respect to the background, which is followed by 2-D Fourier analysis to generate a spectrum from which the orientation and alignment of cells and their processes can be measured. We show that this method can be used to simplify the interpretation of complex structure in images of live neurons obtained via transmission microscopy and consequently, discover trends in neurite development following different types of stimulation. This approach provides a consistent and reliable tool for analyzing changes in cell structure that occurs during live-cell experiments.     

Resveratrol,a multitargeted agent,can enhance antitumor activity of gemcitabine in vitro and in orthotopic mouse model of human pancreatic cancer

Gemcitabine, while a standard treatment of advanced pancreatic cancer (PaCa), alone is not very effective. New agents that are safe and effective are highly needed. Resveratrol is one such agent which is safe and multitargeted; and has been linked with suppression of survival, proliferation, invasion and angiogenesis of cancer. Whether resveratrol can sensitize PaCa to gemcitabine in vitro and in vivo was investigated. We established PaCa xenografts in nude mice, randomized into 4 groups, and treated with vehicle, gemcitabine, resveratrol and with combination. Modulation of NF‐κB and markers of proliferation, angiogenesis and invasion were ascertained using electrophoretic mobility shift assay (EMSA), immunohistochemistry and western blot analysis. Resveratrol inhibited the proliferation of 4 different human PaCa cell lines, synergized the apoptotic effects of gemcitabine, inhibited the constitutive activation of NF‐κB and expression of bcl‐2, bcl‐xL, COX‐2, cyclin D1 MMP‐9 and VEGF. In an orthotopic model of human PaCa, we found that resveratrol significantly suppressed the growth of the tumor (p < 0.001) and this effect was further enhanced by gemcitabine (p < 0.001). Both the markers of proliferation index Ki‐67 and the micro vessel density CD31 were significantly downregulated in tumor tissue by the combination of gemcitabine and resveratrol (p < 0.001 vs. control; p < 0.01 vs. gemcitabine). As compared to vehicle control, resveratrol also suppressed the NF‐κB activation and expression of cyclin D1, COX‐2, ICAM‐1, MMP‐9 and survivin. Overall our results demonstrate that resveratrol can potentiate the effects of gemcitabine through suppression of markers of proliferation, invasion, angiogenesis and metastasis.     

Blockade of transforming growth factor‐β signaling enhances oncolytic herpes simplex virus efficacy in patient‐derived recurrent glioblastoma models

Despite the current standard of multimodal management, glioblastoma (GBM) inevitably recurs and effective therapy is not available for recurrent disease. A subset of tumor cells with stem‐like properties, termed GBM stem‐like cells (GSCs), are considered to play a role in tumor relapse. Although oncolytic herpes simplex virus (oHSV) is a promising therapeutic for GBM, its efficacy against recurrent GBM is incompletely characterized. Transforming growth factor beta (TGF‐β) plays vital roles in maintaining GSC stemness and GBM pathogenesis. We hypothesized that oHSV and TGF‐β inhibitors would synergistically exert antitumor effects for recurrent GBM. Here we established a panel of patient‐derived recurrent tumor models from GBMs that relapsed after postsurgical radiation and chemotherapy, based on GSC‐enriched tumor sphere cultures. These GSCs are resistant to the standard‐of‐care temozolomide but susceptible to oHSVs G47Δ and MG18L. Inhibition of TGF‐β receptor kinase with selective targeted small molecules reduced clonogenic sphere formation in all tested recurrent GSCs. The combination of oHSV and TGF‐βR inhibitor was synergistic in killing recurrent GSCs through, in part, an inhibitor‐induced JNK‐MAPK blockade and increase in oHSV replication. In vivo, systemic treatment with TGF‐βR inhibitor greatly enhanced the antitumor effects of single intratumoral oHSV injections, resulting in cures in 60% of mice bearing orthotopic recurrent GBM. These results reveal a novel synergistic interaction of oHSV therapy and TGF‐β signaling blockade, and warrant further investigations aimed at clinical translation of this combination strategy for GBM patients.