Réactions solide-gaz suivies par microscopie électronique en atmosphère controlée (M.E.A.C.). - I. Réalisation d'un microréacteur fonctionnant à haute température et en M.E.A.C.

A high temperature reaction specimen chamber for an electron microscope has been realized. The specimen under electron microscope observation can be heated at about 1300 K in a gas at a pressure of about 200 Torr. The specimen is placed on a film covering the minute hole (0.5 mm) in the tantalum ribbon and heated by an electric current sent through the ribbon. The minute hole in the ribbon and molybdenum apertures on either side of the ribbon are set to enable the passage of electron beam. The displacement of the heater is controlled by an accessory operating lever. The overflow gas from the specimen chamber is pumped out separately from the evacuation system of main column. The working parameters: temperature, gas pressure and cross-sections of scattered electrons through gas layers, have been determined for the best use of the microreactor.Une chambre de réaction en présence de gaz fonctionnant à haute température a été réalisée. Eéchantillon observé au microscope électronique peut être chauffé à 1300 K sous une pression de gaz de 200 Torr. Léchantillon est placé sur un film couvrant un trou (0.5 mm) du ruban chauffant en tantale. Le ruban est compris entre deux diaphragmes en molybdène de très faible diamètre pour le passage du faisceau électronique. Une commande accessoire permet le déplacement du four entre les deux diaphragmes du microréacteur. Un pompage auxiliaire assure l'évacuation du gaz du microréacteur vers l'extérieur de la colonne du microscope. Les paramètres de fonctionnement : température, pression du gaz, sections efficaces de la diffusion des électrons à travers les couches gazeuses, ont été déterminés pour les meilleures conditions d'utilisation du microréacteur

New chondrosarcoma cell lines and mouse models to study the link between chondrogenesis and chemoresistance

Chondrosarcomas are cartilage-forming, poorly vascularized tumors. They represent the second malignant primary bone tumor of adults after osteosarcoma, but in contrast to osteosarcoma they are resistant to chemotherapy and radiotherapy, surgical excision remaining the only therapeutic option. Few cell lines and animal models are available, and the mechanisms behind their chemoresistance remain largely unknown. Our goal was to establish new cell lines and animal cancer models from human chondrosarcoma biopsies to study their chemoresistance. Between 2007 and 2012, 10 chondrosarcoma biopsies were collected and used for cell culture and transplantation into nude mice. Only one transplanted biopsy and one injected cell line has engrafted successfully leading to conventional central high-grade chondrosarcoma similar to the original biopsies. In culture, two new stable cell lines were obtained, one from a dedifferentiated and one from a grade III conventional central chondrosarcoma biopsy. Their genetic characterization revealed triploid karyotypes, mutations in IDH1, IDH2, and TP53, deletion in CDKN2A and/or MDM2 amplification. These cell lines expressed mesenchymal membrane markers (CD44, 73, 90, 105) and were able to produce a hyaline cartilaginous matrix when cultured in chondrogenic three-dimensional (3D) pellets. Using a high-throughput quantitative RT-PCR approach, we observed that cell lines cultured in monolayer had lost expression of several genes implicated in cartilage development (COL2A1, COMP, ACAN) but restored their expression in 3D cultures. Chondrosarcoma cells in monolayer were sensitive to several conventional chemotherapeutic agents but became resistant to low doses of mafosfamide or doxorubicin when cultured in 3D pellets, in parallel with an altered nucleic accumulation of the drug. Our results indicate that the cartilaginous matrix produced by chondrosarcoma cells may impair diffusion of several drugs and thus contribute to chemoresistance. Therefore, 3D chondrogenic cell pellets constitute a more relevant model to study chondrosarcoma chemoresistance and may be a valuable alternative to animal experimentations