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Home » Archive » 2015

TDK conference 2015

3D tumor cell culture for testing chemicals and treatments: a promising method for future in-vitro studies
Oster-Weinberg Jake - year 4
SzIU, Faculty of Veterinary Science, Department of Pharmacology and Toxicology
Supervisor: Csaba Kővágó

Abstract:

Multilayer or 3D cell culture systems have been used in research and biotechnology for several years, for mimicking solid tissues much better than a monolayer cell-cultures. Extracellular matrix scaffold using the protein extract of Engelbreth–Holm–Swarm tumor, also known as Matrigel, is possibly the most common for growing 3D tumor cultures.

Modulated elector-hyperthermia (mEHT) is a special treatment method of tumors, which utilizes electromagnetic energy to interfere with the neoplastic tissues. The source of heating is impedance coupled electromagnetic energy. The mEHT effect is complex acting through cellular stress pathways in tumors.

Our goal was to compare the effects of mEHT on 3D cell culture to the results gained from in vivo experiments. We used Matrigel as matrix scaffold for maintaining a colon carcinoma cell line, C26. We formed four groups: HT group; Non-cont. mEHT group., mEHT group; Cont. mEHT group and a C group of untreated control kept at 37oC. All groups consisted of 3 samples.

The cultures were micrographed before treatment and 24h after treatment. Hematoxilin-eosin (HE) stained sections were examined for basic cell morphology, cell connections, immune-histochemistry (IHC) stainings were performed to identify the proteins HSP60, HSP70, and BAX. TUNEL assay was also performed to identify the apoptotic cells

In cultures examined right before formalin fixation, the typical cellular structure of the tumor were seen. The examination of the HE stained slides resulted that in the treated slides were more pycnotic nuclei or fragmented. apoptotic bodies. As for ICH stainings, HSP70 positivity emerged in the treated samples, especially in case of "ont. mEHT group, where strong local activation of HSP70 was seen. TUNEL assay and BAX showed apoptotic nuclei in the treated samples.

Our results showing high similarity to previous in-vivo experiments, thus we can conclude that our 3D cell culture model works in very close way to the living animal models. We are convinced that with developing the system, we can earn an effective however in-vitro model for mEHT research and anticancer drug candidate screening. However, we should mention that this kind of in-vitro model still can not entirely substituate the living animal experiments.



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