No abstract

Although in vitro endoplasmic reticulum (ER) stress studies have been carried out using Tunicamycin in human trophoblast cell lines in recent years, the effect of calcium homeostasis impaired by the effect of Thapsigargin on cell survival - death pathways have not been clearly demonstrated. Here, the effects of ER stress and impaired calcium homeostasis on cell death pathways such as apoptosis and autophagy in 2-dimensional and 3-dimensional cell cultures were investigated using the HTR8 / SVneo cell line representing human trophoectoderm cells and the ER stressor Thapsigargin. By using Real Time PCR, gene and immunofluorescence analyzes were studied at the protein level. In this study, it has been established that the Thapsigargin creates ER stress by increasing the level of GRP78 gene and protein in 2 and 3 dimensions of human trophoectoderm cells and that cells show different characterization properties in 2 and 3 dimensions. It has been determined that while it moves in the direction of EIF2A and IRE1A mechanisms in 2 dimensions, it proceeds in the direction of EIF2A and ATF6 mechanisms in 3 dimensions and creates different responses in survival and programmed cell death mechanisms such as apoptosis and autophagy. With forthcoming studies, it is thought that the effects of Thapsigargin on the intrinsic pathway of apoptosis and the linkage of the autophagy mechanism, the examination of the survival-death pathways in the co-culture model with endometrial cells, therapeutic target molecules that will contribute to the elucidation of intracellular cell dynamics may increase the success of implantation.

The current cancer studies focus on the molecular-targeting diagnostics and their interaction with surrounding microenvironment, however, there are some missing points on the characterization of the cells with their topological differences and elemental composition. Glioblastoma multiforme (GBM) which is an astrocytic aggressive brain tumour with short survival time. GBM and astrocyte cells may differ at molecular level and the elemental and topological evaluation of these cells are vital for a definition of new potential targets for cancer research. Here, we report the topology and chemistry of cancer (GBM) and healthy (astrocyte) cells by atomic force microscopy (AFM), scanning electron microscopy (SEM) supported with energy dispersive X-ray spectroscopy (EDS) and X-ray photoelectron spectroscopy (XPS), for the first time. Additionally, F-actin Immunofluorescence staining and Real-Time Polymerase chain reaction (RT-PCR) techniques were performed. Actin related genes were similar in level of gene expression; however, F-actin protein intensities were different. The gene expressions related to the invasion were elevated in GBM cells. Morphologically, GBM cells were found to be longer and narrower while astrocytes were shorter and more disseminated based on AFM. Furthermore, roughness values of these cells were relatively close to each other. SEM-EDS analysis demonstrated that elongated GBM cells exhibited several filopodial protrusions whereas the astrocyte surfaces were rougher in lamellipodial area. Our investigation provides considerable further insight into rapid cancer cell characterization field in terms of its combinatorial spectroscopic and microscopic approach.