Cellular Interactions in Autologous Culture of Human Leukocytes: Mitochondrial Dynamics, Cellular Activation and Extracellular Traps

In this work of scientific research of the Mitochondrial Dynamics (MD) in lymphocytes in autologous cultures of human total leukocytes from peripheral blood stimulated with the Lypopolisaccharide (LPS) endotoxin and the Ovalbumin (OVA) heterologous antigen was carried out. In addition, the activated phenotype (effector cells and memory cells) of T lymphocytes was marked for CD45RO and MD was studied in lymphocytes in relation to the presence of B7 costimulatory molecules (CD80 and CD86) in antigen presenting cells (APCs) in autologous cultures. Extracellular traps (ETs) generated in cultures stimulated with LPSwere visualized with fluorescence microscopy and were observed like diffuse staining or fibrillar appearance with DAPI. In paired control samples without stimulation, the occurrence of ETs was in average value: 3.4% of the cells. A higher percentage of cells that release ETs was observed in cultures stimulated with LPS versus the paired control group (p <0.0001). In ultrastructural observations differences were registered in the size of the lymphocyte cytoplasm. On the other hand, MD in stimulated lymphocytes shows changes with respect to the paired controlled samples: in the size (area) mitochondrial and the position of the mitochondria: polarization. No statistically significant differences were observed in number of mitochondria per cell at 30 minutes of culture. In multiple trials stimulated with LPS, lymphocytes with conventional mitochondria and others with morphologically altered mitochondria with electrolucent ridges were observed. After stimulation, in paired samples cell activation in lymphocytes was selected for CD45RO positivity. The changes in MD in stimulated lymphocytes correspond to the state of cellular activation in parallel detected by immunofluorescence (IF) with anti-CD45RO. The increase in the expression of B7 molecules in the APCs given by the stimuli contributed to the state of lymphocyte activation.

Keywords: human leukocytes; mitochondrial dynamics; cellular activation; extracellular traps.