Influence of Lactobacillus bacteria on apoptosis of human epithelial cells in vitro
bacteria ; epithelium ; Lactobacilli ; cancer ; apoptosis
Apoptosis, called programmed cell death, is one of the main physiological proceses that maintain tissue homeostasis. Apoptosis dysfunction can be observed in many pathological conditions like inflammation, cancer, etc. Cytokines, reactive oxygen species, hypoxia can initiate programmed cell death. Pathogen and commensal bacteria are in close contact with epithelial tissue and they can influence apoptotic pathways. The Lactobacillus species that are potential probiotic strains can also be important in maintaining tissue balance. They can secrete several different substances that can act on programmed cell death. The aim of this doctoral dissertation was the effect of different Lactobacillus strains on epithelial cells apoptosis. Apoptosis of cell line HT-29, CaCo-2, A431 and mouse primary colonocytes was tested. Potentially probiotic and hydrogen peroxide producing Lactobacillus strains and their supernatants have been chosen for these experiments. The level of activated caspases and cell membrane damage has been estimated by flow cytometry. Fluorescent FAM-VAD-FMK caspase inhibitor and propidium iodide have been used to distinguish apoptosis from necrosis. Apoptotic pathway was confirmed by DNA ladder electrophoresis. Changes of the actin filaments structure have been observed after Lactobacilli incubation have been performed. Among many Lactobacillus strains chosen three diffe ; rent groups have isolated. Lactobacilli that produced and secreted hydrogen peroxide had proapoptotic influence on cancer cell lines and primary colonocytes. Apoptotic fate of the some cells has been observed after incubation with L. fermentum and L. reuteri, but in most cases Lactobacilli that did not secrete hydrogen peroxide did not have significant influence on caspases activation in vitro. Some of the strains can even protect from the apoptotic process initiated by pathogen and proapoptotic substances. Protective effect of these H2O2 non-producing Lactobacilli can also be seen in case of actin filaments cytoskeleton. Addition of hydrogen peroxide producing Lactobacilli or their supernatant resulted in destruction of actin filaments and the loss of cell shape and contact. The purpose of this work was to better understand the role of Lactobacillus as commensal bacteria and their role in tissue homeostasis. Better understanding the role of Lactobacillus strains can result in improved prophylactic effect of probiotics.