Comparative analysis of different methodological approaches to the in vitro study of tumour cells chemosensitivity

Drug sensitivity assay was performed using two human tumour celi lines: HeLa and Hep-2 cultivated in two-dimensional monolayer celi cultures and three-dimensional cultures on gelatine sponge SpongostanK. Two cytostatics with different mechanisms of anti-tumour action were used: cisplatin and etoposide. Chemosensitivity of tumour cells was assessed by counting the number of viable and nonviable cells (cytostatic and cytotoxic activity of drugs), by counting the number of apoptotic cells and by clonogenic assay of viable cells. We found that the clonogenic assay was morę sensitive than the other tests used, especially after long-term (7 days) treatment of tumour cells with cytostatics. A short (24h) treatment with cytostatics gave false results which were not confirmed after prolonged treatment with cytostatics. We suppose that short treatment tests should not be used for examination of the chemosensitivity of tumour cells isolated from patients. Tumour cells growing on SpongostanH were viable for a longer time than in monolayer cultures and exhibited chemosensitivity comparable to monolayer celi cultures despite of their multilayer growth on gelatine sponge.Do badań wrażliwości na cytostatyki użyto dwie ludzkie linie nowotworowe: HeLa i Hep-2, hodowane w formie murawy dwuwymiarowej (płaskiej) oraz w formie przestrzennej, trójwymiarowej, na gąbce żelatynowej Spongostan". W badaniach użyto cytostatyki posiadające różny mechanizm działania przeciwnowotworowego, a mianowicie cisplatynę i etopozyd. Wrażliwość komórek nowotworowych na chemioterapeutyki określano ilością komórek przeżywających i martwych (cytostatyczne i cytotoksyczne właściwości leków), ilością komórek apoptotycznych oraz oceniano klonogenne właściwości komórek przeżywających. Stwierdzono, że ocena klonogennych właściwości komórek jest metodą bardziej czułą w porównaniu z innymi testami, szczególnie po długim (7-dniowym) kontakcie komórek z cytostatykami. Ocena krótkotrwałego (24-godz.) kontaktu komórek nowotworowych z cytostatykami dawała fałszywe wyniki, nie potwierdzone po długotrwałej inkubacji komórek z cytostatykami. Uważamy, że testy polegające na ocenie efektu krótkotrwałej inkubacji komórek z lekami nie powinny być stosowane do oceny wrażliwości na chemioterapeutyki komórek nowotworowych izolowanych od pacjenta. Komórki nowotworowe rosnące na Spongostanie” były żywe dłużej niż rosnące w hodowlach płaskich i pomimo wielowarstwowego wzrostu na gąbce żelatynowej wykazywały wrażliwość na leki porównywalną z hodowlami płaskimi

Antiproliferative Activity of Melanoidins Isolated from Heated Potato Fiber (Potex) in Glioma Cell Culture Model

Potex constitutes a potato fiber preparation widely used as an ingredient to meat and bakery products which thermal treatment results in creation of new compounds. Melanoidins are high molecular weight brown end products of Maillard reaction, and few data presenting tumor cell growth inhibiting activity of melanoidins have been reported. Thus, in present study we utilized water extract of Potex roasted (180 degrees C for 2 h), whose chemical characterization revealed the presence of melanoidin complexes. Heated Potex extract inhibited C6 glioma cell proliferation in a dose-dependent manner measured by MTT method. High molecular weight components present in initial extract were responsible for stronger antiproliferative effect compared with low molecular weight fraction. Impaired MAPK (mitogen-activated protein kinase) and Akt signaling was found in cells treated with the extract. Moreover, flow cytometry analyses revealed the extract to induce G(1)/S arrest in glioma cells. Simultaneously, Western blot analysis showed elevated levels of p21 protein with concomitant decrease of cyclin D1. In conclusion, observed antiproliferative activity of melanoidins present in heated Potex was linked to disregulated MAPK and Akt signaling pathways, as well as to cell cycle cessation. These results suggest potential application of Potex preparation as a functional food ingredient and chemopreventive agent

Melanoidins isolated from heated potato fiber (Potex) affect human colon cancer cells growth via modulation of cell cycle and proliferation regulatory proteins

Melanoidins are brown, nitrogen containing, high molecular weight end products of Maillard reaction with poorly established activity towards tumor cells. The goal of present study was to verify whether both heated potato fiber Potex extract (180 degrees C for 2 h) and melanoidins isolated from the extract exerts growth-inhibiting activity in human colon cancer cells in vitro. The cells of LS180 colon cancer cell line were tested upon treatment with roasted potato fiber extract (AM4) as well as with high (HMW) and low (LMW) molecular weight fractions isolated from the extract, since both may be regarded as/or contain melanoidins. The tested compounds at concentration of 1000 mu g/ml reduced cell growth down to 45%, 69% and 54%, respectively. Furthermore, deregulated ERK1/2 signaling was revealed upon treatment. Moreover, multiple alternations in cell cycle regulators activity were found (i.e. cyclinD1, cyclin-dependent kinase 4 and 6, p21, p27, p53, pRb) leading to cell cycle cessation in GO phase. Importantly, LMW compounds revealed markedly stronger potential to alter specific molecular targets comparing to HMW compounds. Summarizing, the results emphasize that both high and low molecular weight melanoidins contribute to antiproliferative activity of heated potato fiber in LS180 colon cancer cells in vitro. (C) 2013 Elsevier Ltd. All rights reserved