Diese Arbeit wurde vom NIH gewährt 5R01CA100750-07, 5R01CA125541-04, 3R01CA125541-03S1, 5R01CA129501-03, 3R01CA129501-02S1 zu RS unterstützt

<ol>
	<li>Adamski, J., Finnegan, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+Perspectives+on+Microsoft+Office+Access.">New Perspectives on Microsoft Office Access.</a> , (2007).</li><li>Giltnane, J., Rimm, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Technology+Insight:+Identification+of+biomarkers+with+tissue+microarray+technology.">Technology Insight: Identification of biomarkers with tissue microarray technology.</a> Nat Clin Pract Oncol. 1, 104-111 (2004).</li></ol>

<table border="1"><tbody><tr><td> Material Name </td><td> Firma </td><td> Katalog-Nummer </td></tr><tr><td> Zentrifuge </td><td> Eppendorf </td><td> </td></tr><tr><td> Konische Zentrifugenröhrchen </td><td> Falke </td><td> 518-PG </td></tr><tr><td> Minimum Essential Medium Eagle (MEM) </td><td> Sigma </td><td> M4655-500ML </td></tr><tr><td> Fetal Calf Serum </td><td> CellGro FBS HALLO </td><td> MTT35011CV </td></tr><tr><td> Dimethylsulfoxid (DMSO) </td><td> Amerikanischen Bioanalytische </td><td> AB03091 </td></tr><tr><td> BD Vacutainer Serumröhrchen </td><td> Fischer Scientific </td><td> 367815 </td></tr></tbody></table>

generation of comprehensive thoracic oncology database - tool for translational research

Das Thoracic Oncology Program Database Project wurde geschaffen, um ein umfassendes, verifiziert und zugänglich Repository für gut kommentierten Krebs Proben und klinischen Daten zur Verfügung zu stehen Forschern innerhalb der Thoracic Oncology Research Program dienen. Diese Datenbank erfasst auch eine große Menge von Genom-und Proteom-Daten aus verschiedenen Tumorgewebe Studien erhalten. Ein Team von klinischen und wissenschaftlichen Forschern, ein Biostatistiker, und ein Bioinformatik-Experte wurde einberufen, um die Datenbank-Design. Variablen von Interesse waren klar definiert und ihre Beschreibungen wurden in einem Standard-Betriebsanleitung geschrieben, um die Konsistenz der Daten-Annotation zu gewährleisten. Mit einem Protokoll für angehende Gewebe Banking und ein anderes Protokoll für die retrospektive Banking, Tumor-und Normalgewebe von Patienten zugestimmt, diese Protokolle gesammelt wurden. Klinische Daten wie Demographie, Krebs Charakterisierung und Behandlung Pläne für diese Patienten wurden abstrahiert und in eine Access-Datenbank. Proteomik und Genomik Daten in die Datenbank aufgenommen und haben zu klinischen Informationen für Patienten in der Datenbank beschrieben in Verbindung gebracht worden. Die Daten aus den einzelnen Tabellen verknüpft wurden unter Verwendung der Beziehungen Funktion in Microsoft Access, damit die Datenbank-Manager zu klinischen Informationen während einer Abfrage verbinden. Die abgefragten Daten können dann für statistische Analysen und Hypothesen Generation exportiert werden.

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Generation of Comprehensive Thoracic Oncology Database - Tool for Translational Research

Eine Brust-Onkologie-Datenbank wurde entwickelt, um als eine umfassende Quelle für klinische Daten und Laborwerte für die Zwecke der translationalen Forschung dienen. Die Datenbank wird translationalen Krebsforschung in der Thoracic Oncology Research Program dienen. Diese Datenbank ist anpassungsfähig zu anderen Krebs-Modelle, sowie andere Erkrankungen des Menschen.

Generation umfassender Thoracic Oncology Database - Tool für Translational Research

The Thoracic Oncology Program Database Project was created to serve as a comprehensive, verified, and accessible repository for well-annotated cancer ...

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15.9K Aufrufe.  University of Chicago. Eine Brust-Onkologie-Datenbank wurde entwickelt, um als eine umfassende Quelle für klinische Daten und Laborwerte für die Zwecke der translationalen Forschung dienen. Die Datenbank wird translationalen Krebsforschung in der Thoracic Oncology Research Program dienen. Diese Datenbank ist anpassungsfähig zu anderen Krebs-Modelle, sowie andere Erkrankungen des Menschen.

Serie: Generation of Comprehensive Thoracic Oncology Database - Tool for Translational Research

Time-lapse Imaging of Primary Preneoplastic Mammary Epithelial Cells Derived from Genetically Engineered Mouse Models of Breast Cancer

Time-lapse imaging can be used to compare behavior of cultured primary preneoplastic mammary epithelial cells derived from different genetically engineered mouse models of breast cancer. For example, time between cell divisions (cell lifetimes), apoptotic cell numbers, evolution of morphological changes, and mechanism of colony formation can be quantified and compared in cells carrying specific genetic lesions. Primary mammary epithelial cell cultures are generated from mammary glands without palpable tumor. Glands are carefully resected with clear separation from adjacent muscle, lymph nodes are removed, and single-cell suspensions of enriched mammary epithelial cells are generated by mincing mammary tissue followed by enzymatic dissociation and filtration. Single-cell suspensions are plated and placed directly under a microscope within an incubator chamber for live-cell imaging. Sixteen 650 &mu;m x 700 &mu;m fields in a 4x4 configuration from each well of a 6-well plate are imaged every 15 min for 5 days. Time-lapse images are examined directly to measure cellular behaviors that can include mechanism and frequency of cell colony formation within the first 24 hr of plating the cells (aggregation versus cell proliferation), incidence of apoptosis, and phasing of morphological changes. Single-cell tracking is used to generate cell fate maps for measurement of individual cell lifetimes and investigation of cell division patterns. Quantitative data are statistically analyzed to assess for significant differences in behavior correlated with specific genetic lesions.

Time-lapse imaging is used to assess behavior of primary preneoplastic mammary epithelial cells derived from genetically engineered mouse models of breast cancer risk to determine if there are correlations between specific behavioral parameters and distinct genetic lesions.

Zeitraffer-Imaging of Primary Präkanzeröse Brustepithelzellen aus gentechnisch veränderten Mausmodellen der Breast Cancer Abgeleitet

Time-lapse  imaging can be used to compare behavior of cultured primary preneoplastic  mammary epithelial cells derived from different genetically ...

Forschung

Medizin

A Modified Heterotopic Swine Hind Limb Transplant Model for Translational Vascularized Composite Allotransplantation (VCA) Research

Vascularized Composite Allotransplantation (VCA) such as hand and face transplants represent a viable treatment option for complex musculoskeletal trauma and devastating tissue loss. Despite favorable and highly encouraging early and intermediate functional outcomes, rejection of the highly immunogenic skin component of a VCA and potential adverse effects of chronic multi-drug immunosuppression continue to hamper widespread clinical application of VCA. Therefore, research in this novel field needs to focus on translational studies related to unique immunologic features of VCA and to develop novel immunomodulatory strategies for immunomodulation and tolerance induction following VCA without the need for long term immunosuppression.
This article describes a reliable and reproducible translational large animal model of VCA that is comprised of an osteomyocutaneous flap in a MHC-defined swine heterotopic hind limb allotransplantation. Briefly, a well-vascularized skin paddle is identified in the anteromedial thigh region using near infrared laser angiography. The underlying muscles, knee joint, distal femur, and proximal tibia are harvested on a femoral vascular pedicle. This allograft can be considered both a VCA and a vascularized bone marrow transplant with its unique immune privileged features. The graft is transplanted to a subcutaneous abdominal pocket in the recipient animal with a skin component exteriorized to the dorsolateral region for immune monitoring.
Three surgical teams work simultaneously in a well-coordinated manner to reduce anesthesia and ischemia times, thereby improving efficiency of this model and reducing potential confounders in experimental protocols. This model serves as the groundwork for future therapeutic strategies aimed at reducing and potentially eliminating the need for chronic multi-drug immunosuppression in VCA.

A Modified Heterotopic Swine Hind Limb Transplant Model for Translational Vascularized Composite Allotransplantation VCA Research

Vascularized Composite Allotransplantations (VCA) have become a clinical reality. However, broad clinical application of VCA is limited by chronic multi-drug immunosuppression. The authors present a reliable and reproducible large animal model to translate novel immunomodulatory strategies that can minimize or potentially eliminate the need of immunosuppression in VCA.

Eine modifizierte Heterotope Schweine Hind Gliedmaßen-Transplantation Modell für Translationale Vaskularisierte Composite-Allotransplantation (VCA) Forschung

Vascularized Composite Allotransplantation (VCA) such as hand and face transplants represent a viable treatment option for complex musculoskeletal trauma ...

Three Dimensional Cultures: A Tool To Study Normal Acinar Architecture vs. Malignant Transformation Of Breast Cells

Invasive breast carcinomas are a group of malignant epithelial tumors characterized by the invasion of adjacent tissues and propensity to metastasize. The interplay of signals between cancer cells and their microenvironment exerts a powerful influence on breast cancer growth and biological behavior1. However, most of these signals from the extracellular matrix are lost or their relevance is understudied when cells are grown in two dimensional culture (2D) as a monolayer. In recent years, three dimensional (3D) culture on a reconstituted basement membrane has emerged as a method of choice to recapitulate the tissue architecture of benign and malignant breast cells. Cells grown in 3D retain the important cues from the extracellular matrix and provide a physiologically relevant ex&#160;vivo system2,3. Of note, there is growing evidence suggesting that cells behave differently when grown in 3D as compared to 2D4. 3D culture can be effectively used as a means to differentiate the malignant phenotype from the benign breast phenotype and for underpinning the cellular and molecular signaling involved3. One of the distinguishing characteristics of benign epithelial cells is that they are polarized so that the apical cytoplasm is towards the lumen and the basal cytoplasm rests on the basement membrane. This apico-basal polarity is lost in invasive breast carcinomas, which are characterized by cellular disorganization and formation of anastomosing and branching tubules that haphazardly infiltrates the surrounding stroma. These histopathological differences between benign gland and invasive carcinoma can be reproduced in 3D6,7. Using the appropriate read-outs like the quantitation of single round acinar structures, or differential expression of validated molecular markers for cell proliferation, polarity and apoptosis in combination with other molecular and cell biology techniques, 3D culture can provide an important tool to better understand the cellular changes during malignant transformation and for delineating the responsible signaling.

Three dimensional culture of mammary epithelial cells on a reconstituted basement membrane is a useful method to recapitulate the in vivo architecture of the benign breast, and to differentiate the malignant phenotype from the benign breast phenotype. Importantly, this system can be applied to study invasive carcinomas in other tissues.

Dreidimensional Kulturen: Ein Werkzeug, um Architektur zu studieren Normale acinar gegen maligne Transformation von Brustzellen

Invasive breast carcinomas are a group of malignant epithelial tumors characterized by the invasion of adjacent tissues and propensity to metastasize. The ...

Working with Human Tissues for Translational Cancer Research

Medical research for human benefit is greatly impeded by the necessity for human tissues and subjects. However, upon obtaining consent for human specimens, precious samples must be handled with the greatest care in order to ensure integrity of organs, tissues, and cells to the highest degree. Unfortunately, tissue processing by definition requires extraction of tissues from the host, a change which can cause great cellular stress and have major repercussions on subsequent analyses. These stresses could result in the specimen being no longer representative of the site from which it was retrieved. Therefore, a strict protocol must be adhered to while processing these specimens to ensure representativeness. The desired assay(s) must also be taken into consideration in order to ensure that an optimal technique is used for sample processing. Outlined here is a protocol for tissue retrieval, processing and various analyses which may be performed on processed tissue in order to maximize downstream production from limited tissue samples.

Translational cancer research is dependent on extraction of human tissues. Much work has gone into optimizing extraction methods for ex vivo analysis. Here, we describe tissue processing methods allowing for maximal data output from limited samples.

Arbeiten mit menschlichen Geweben für Translationale Krebsforschung

Medical research for human benefit is greatly impeded by the necessity for human tissues and subjects. However, upon obtaining consent for human ...

Exosomal miRNA Analysis in Non-small Cell Lung Cancer (NSCLC) Patients' Plasma Through qPCR: A Feasible Liquid Biopsy Tool

The discovery of alterations in the EGFR and ALK genes, amongst others, in NSCLC has driven the development of targeted-drug therapy using selective tyrosine kinase inhibitors (TKIs). To optimize the use of these TKIs, the discovery of new biomarkers for early detection and disease progression is mandatory. These plasma-isolated exosomes can be used as a non-invasive and repeatable way for the detection and follow-up of these biomarkers. One ml of plasma from 12 NSCLC patients, with different mutations and treatments (and 6 healthy donors as controls), were used as exosome sources. After RNAse treatment, in order to degrade circulating miRNAs, the exosomes were isolated with a commercial kit and resuspended in specific buffers for further analysis. The exosomes were characterized by western blotting for ALIX and TSG101 and by transmission electron microscopy (TEM) analysis, the standard techniques to obtain biochemical and dimensional data of these nanovesicles. Total RNA extraction was performed with a high yield commercial kit. Due to the limited miRNA-content in exosomes, we decided to perform retro-transcription PCR using an individual assay for each selected miRNA. A panel of miRNAs (30b, 30c, 103, 122, 195, 203, 221, 222), all correlated with NSCLC disease, were analyzed taking advantage of the remarkable sensitivity and specificity of Real-Time PCR analysis; mir-1228-3p was used as endogenous control and data were processed according to the formula 2-&#916;&#916;ct 13. Control values were used as baseline and results are shown in logarithmic scale.

Exosomal miRNA Analysis in Non-small Cell Lung Cancer NSCLC Patients&#039; Plasma Through qPCR: A Feasible Liquid Biopsy Tool

This protocol describes the feasibility to perform miRNA profiling in exosomes, released in plasma of NSCLC patients, through a commercial exosome isolation kit with Proteinase K and RNAse treatments, in order to avoid circulating miRNAs contamination and evaluate their biomarker features in NSCLC.

Exosomalen miRNA-Analyse in nicht-kleinzelligem Lungenkrebs (NSCLC) der Patienten Plasma durch qPCR: ein gangbarer Flüssiges Biopsy-Tool

The discovery of alterations in the EGFR and ALK genes, amongst others, in NSCLC has driven the development of targeted-drug therapy using selective ...

Development of a Direct Pulp-capping Model for the Evaluation of Pulpal Wound Healing and Reparative Dentin Formation in Mice

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is often capped with biocompatible materials in order to expedite pulpal wound healing. The ultimate goal is to regenerate reparative dentin, a physical barrier that functions as a "biological seal" and protects the underlying pulp tissue. Although this direct pulp-capping procedure has long been used in dentistry, the underlying molecular mechanism of pulpal wound healing and reparative dentin formation is still poorly understood. To induce reparative dentin, pulp capping has been performed experimentally in large animals, but less so in mice, presumably due to their small sizes and the ensuing technical difficulties. Here, we present a detailed, step-by-step method of performing a pulp-capping procedure in mice, including the preparation of a Class-I-like cavity, the placement of pulp-capping materials, and the restoration procedure using dental composite. Our pulp-capping mouse model will be instrumental in investigating the fundamental molecular mechanisms of pulpal wound healing in the context of reparative dentin in vivo by enabling the use of transgenic or knockout mice that are widely available in the research community.

We describe a step-by-step method of performing direct pulp capping on mice teeth for the evaluation of pulpal wound healing and reparative dentin formation in vivo.

Entwicklung eines Direkt Pulp-Kappungsmodell für die Beurteilung von Pulpal Wundheilung und Reparative Dentinbildung bei Mäusen

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is ...

Second Harmonic Generation Signals in Rabbit Sclera As a Tool for Evaluation of Therapeutic Tissue Cross-linking (TXL) for Myopia

Methods to strengthen tissue by introducing chemical bonds (non-enzymatic cross-linking) into structural proteins (fibrillar collagens) for therapy include photochemical cross-linking and tissue cross-linking (TXL) methods. Such methods for inducing mechanical tissue property changes are being employed to the cornea in corneal thinning (mechanically weakened) disorders such as keratoconus as well as the sclera in progressive myopia, where thinning and weakening of the posterior sclera occurs and likely contributes to axial elongation. The primary target proteins for such tissue strengthening are fibrillar collagens which constitute the great majority of dry weight proteins in the cornea and sclera. Fortuitously, fibrillar collagens are the main source of second harmonic generation signals in the tissue extracellular space. Therefore, modifications of the collagen proteins, such as those induced through cross-linking therapies, could potentially be detected and quantitated through the use of second harmonic generation microscopy (SHGM). Monitoring SHGM signals through the use of a laser scanning microscopy system coupled with an infrared excitation light source is an exciting modern imaging method that is enjoying widespread usage in the biomedical sciences. Thus, the present study was undertaken in order to evaluate the use of SHGM microscopy as a means to measure induced cross-linking effects in ex vivo rabbit sclera, following an injection of a chemical cross-linking agent into the sub-Tenon&#39;s space (sT), an injection approach that is standard practice for causing ocular anesthesia during ophthalmologic clinical procedures. The chemical cross-linking agent, sodium hydroxymethylglycinate (SMG), is from a class of cosmetic preservatives known as formaldehyde releasing agents (FARs). Scleral changes following reaction with SMG resulted in increases in SHG signals and correlated with shifts in thermal denaturation temperature, a standard method for evaluating induced tissue cross-linking effects.

Second Harmonic Generation Signals in Rabbit Sclera As a Tool for Evaluation of Therapeutic Tissue Cross-linking TXL for Myopia

This protocol describes techniques for evaluating chemical cross-linking of the rabbit sclera using&#160;second harmonic generation imaging and differential scanning calorimetry.

Zweite harmonische Generation Signale in Kaninchen Sklera als Instrument für die Bewertung des therapeutischen Gewebes Vernetzung (TXL) für Kurzsichtigkeit

Methods to strengthen tissue by introducing chemical bonds (non-enzymatic cross-linking) into structural proteins (fibrillar collagens) for therapy ...

Generation of Human 3D Lung Tissue Cultures (3D-LTCs) for Disease Modeling

Translation of novel discoveries to human disease is limited by the availability of human tissue-based models of disease. Precision-cut lung slices (PCLS) used as 3D lung tissue cultures (3D-LTCs) represent an elegant and biologically highly relevant 3D cell culture model, which highly resemble in situ tissue due to their complexity, biomechanics and molecular composition. Tissue slicing is widely applied in various animal models. 3D-LTCs derived from human PCLS can be used to analyze responses to novel drugs, which might further help to better understand the mechanisms and functional effects of drugs in human tissue. The preparation of PCLS from surgically resected lung tissue samples of patients, who experienced lung lobectomy, increases the accessibility of diseased and peritumoral tissue. Here, we describe a detailed protocol for the generation of human PCLS from surgically resected soft-elastic patient lung tissue. Agarose was introduced into the bronchoalveolar space of the resectates, thus preserving lung structure and increasing the tissue&#39;s stiffness, which is crucial for subsequent slicing. 500 &#181;m thick slices were prepared from the tissue block with a vibratome. Biopsy punches taken from PCLS ensure comparable tissue sample sizes and further increase the amount of tissue samples. The generated lung tissue cultures can be applied in a variety of studies in human lung biology, including the pathophysiology and mechanisms of different diseases, such as fibrotic processes at its best at (sub-)cellular levels. The highest benefit of the 3D-LTC ex vivo model is its close representation of the in situ human lung in respect of 3D tissue architecture, cell type diversity and lung anatomy as well as the potential for assessment of tissue from individual patients, which is relevant to further develop novel strategies for precision medicine.

Generation of Human 3D Lung Tissue Cultures 3D-LTCs for Disease Modeling

Here, we present a protocol for the preparation of agarose-filled human precision-cut lung slices from resected patient tissue that are suitable for generating 3D lung tissue cultures to model human lung diseases in biological and biomedical studies.

Generation von menschlichen 3D Lungengewebe Kulturen (3D-LTCs) für Krankheit Modellierung

Translation of novel discoveries to human disease is limited by the availability of human tissue-based models of disease. Precision-cut lung slices (PCLS) ...

Modified Heterotopic Hindlimb Osteomyocutaneous Flap Model in the Rat for Translational Vascularized Composite Allotransplantation Research

Vascularized composite allotransplantation (VCA) is a relatively new field in the reconstructive surgery. Clinical achievements in human VCA include hand and face transplants and, more recently, abdominal wall, uterus, and urogenital transplants. Functional outcomes have exceeded initial expectations, and most recipients enjoy an improved quality of life. However, as clinical experience accumulates, chronic rejection and complications from the immunosuppression must be addressed. In many cases where grafts have failed, the causative pathology has been ischemic vasculopathy. <span style="font-size: 11pt; line-height: 115%; font-family: Arial, sans-serif; color: rgb(41, 43, 49); background-image: initial; background-position: initial; background-size: initial; background-repeat: initial; background-attachment: initial; background-origin: initial; background-clip: initial;">The biological mechanisms of the acute and chronic rejection associated with VCA, especially ischemic vasculopathy, are important areas of research. However, due to the very small number of VCA patients, the evaluation of proposed mechanisms is better addressed in an experimental model. Multiple groups have used animal models to address some of the relevant unsolved questions in VCA rejection and vasculopathy. Several model designs involving a variety of species are described in the literature. Here we present a reproducible model of VCA heterotopic hindlimb osteomyocutaneous flap in the rat that can be utilized for translational VCA research. This model allows for the serial evaluation of the graft, including biopsies and different imaging modalities, while maintaining a low level of morbidity.

Vascularized composite allograft offers life-altering benefits to transplant recipients, but the biological causes of graft rejection and vasculopathy remain poorly understood. The rodent surgical model presented here offers a reproducible, clinically relevant model of transplantation, allowing researchers to evaluate rejection events and potential therapeutic strategies to prevent their occurrence.

Modifiziertes Heterotopisches Hindlimb Osteomyoskan Flap Model im Rat für translational Vasculariisierte Composite Allotransplantation Research

Vascularized composite allotransplantation (VCA) is a relatively new field in the reconstructive surgery. Clinical achievements in human VCA include hand ...

Transaxillary First Rib Resection for Treatment of the Thoracic Outlet Syndrome

Thoracic outlet syndrome (TOS) is a common disorder that causes a significant loss of productivity. The transaxillary first rib resection (TFRR) protocol has been used for the decompression of trapped neurovascular structures in the TOS. Among the other surgical procedures, the advantage of the TFRR is that it has the smallest rate of recurrence and better cosmetic outcomes. The disadvantage of TFRR is that it provides a narrow, and deep working corridor that makes obtaining vascular control challenging.

Here, we present a protocol of the transaxillary resection of the first rib for treatment of thoracic outlet syndrome caused by compression of the brachial plexus, subclavian vein and artery.

Transaxilläre erste Rippenresektion zur Behandlung des Thoracic-Outlet-Syndroms

Thoracic outlet syndrome (TOS) is a common disorder that causes a significant loss of productivity. The transaxillary first rib resection (TFRR) protocol ...