The production of three cytokines, interferon gamma (IFN-y), interleukin 10 (1L-10) and interleukin 12 (IL-12), was measured after intraperitoneal infection of immunocompetent Balb/c mice and immunodeficient SCID mice with the microsporidian, Encephalitozoon cuniculi Levaditi, Nicolau ct Schoen, 1923. High levels of IFN-y were detected in ex vivo cultures of peritoneal exudate cells (PEC) of Balb/c mice, a lower, but earlier IFN-y response was observed in PEC from SCID mice. The early 1L-10 response was detected in ex vivo cultures of splenocytes from Balb/c but not from SCID mice, explaining a delay in the IFN-y response in Balb/c mice. IL-12 was detected in PEC cultures from SCID mice, indicating an alternative pathway of IFN-y production by NK. cells stimulated by IL-12 derived from macrophages.
The survival of Encephalitozoon cuniculi Lcvaditi, Nicolau et Schoen, 1923 spores suspended in distilled water and exposed at defined temperatures was investigated. Infectivity of E. cuniculi spores was tested by inoculation of SCID mice. There was no marked loss of infectivity of spores stored at 4°C for two years or frozen at -12°C and -24°C for 1, 8, and 24 h. Although there was a remarkable loss of infectivity, spores remained infective after freezing at -70°C for 1 and 8 h. Heating at 60°C and 70°C for 5 min and 1 min, respectively, rendered the microsporidia non-infective. These findings demonstrate that E. cuniculi spores suspended in water can survive freezing temperatures but lost infectivity in water that reached a temperature of 60°C at 5 min.
An experimental infection with the microsporidian Encephalitozoon cuniculi Levaditi, Nicolau et Schoen, 1923 was studied using a model of immunocompetent BALB/c mice and immunodeficient SCID mice. The course of infection after intraperitoneal inoculation of E. cuniculi spores was evaluated using the presence of spores in peritoneal macrophages as a criterion. First significant decrease in the proportion of infected cells was recorded on day 9 post infection (p.i.) in BALB/c mice. From day 14 p.i. no spores were observed in macrophages from BALB/c mice, while the number of infected macrophages from SCID mice increased until the death of the mice. The natural killer (NK) cell activity of mouse splenocytes was compared with the production of interferon gamma (IFN-γ) by these cells. While in BALB/c mice NK activity peaked on days 9 and 14 p.i., in SCID mice the marked increase of NK activity was recorded close before death of mice, on day 21 p.i. in correlation with the production of IFN-γ. Production of specific antibodies was demonstrated from day 9 p.i. in sera from BALB/c mice. It is concluded that intraperitoneal infection of SCID mice with spores of E. cuniculi results in the marked increase in the number of peritoneal exudate cells and in the percentage of infected cells close before death of mice. Neither high activity of NK cells nor increased production of IFN-γ are sufficient for the recovery of SCID mice from an E. cuniculi infection.
Cancer development is a highly complicated process in which tumour growth depends on the development of its vascularization system. To support their own growth, tumour cells significantly modify their microenvironment. One of such modifications inflicted by tumours is stimulation of endothelial cell migration and proliferation. There is accumulating evidence that extracellular vesicles (EVs) secreted by tumour cells (tumour-derived EVs, TEVs) may be regarded as “messengers” with the potential for affecting the biological activities of target cells. Interaction of TEVs with different cell types occurs in an auto- and paracrine manner and may lead to changes in the function of the latter, e.g., promoting motility, proliferation, etc. This study analysed the proangiogenic activity of EVs derived from human pancreatic adenocarcinoma cell line (HPC-4, TEVHPC) in vitro and their effect in vivo on Matrigel matrix vascularization in severe combined immunodeficient (SCID) mice. TEVHPC enhanced proliferation of HPC-4 cells and induced their motility. Moreover, TEVHPC stimulated human umbilical vein endothelial cell (HUVEC) proliferation and migration in vitro. Additionally, TEVHPC influenced secretion of proangiogenic factors (IL-8, VEGF) by HUVEC cells and supported Matrigel matrix haemoglobinization in vivo. These data show that TEVs may support tumour propagation in an autocrine manner and may support vascularization of the tumour. The presented data are in line with the theory that tumour cells themselves are able to modulate the microenvironment via TEVs to maximize their growth potential.
Susceptibility of three strains of immunodeficient mice to two related microsporidian species Encephalitozoon cuniculi Levaditi, Nicolau et Schoen, 1923 and Encephalitozoon intestinalis (Cali, Kotler et Orenstein, 1993) was compared. While both, severe combined immunodeficient (SCID) and interferon-gamma knock-out (IFN-γ KO) mice, succumbed to either intraperitoneal (i.p.) or peroral (p.o.) (natural) infection with both parasites, only i.p. infection with E. cuniculi killed interleukin-12 knock-out (IL-12 KO) mice. IFN-γ KO mice died earlier than SCID mice. Adoptive transfer of naive splenocytes from IFN-γ KO mice did not protect the SCID mice from a lethal infection with either of the Encephalitozoon species. However, reconstituted mice survived significantly longer (P<0.05), thus indicating the role of IFN-γ produced by host NK cells in the development of mechanisms of anti-microsporidial protective immunity. Non-lethal outcome of the infection always correlated with the increase in CD8+ T lymphocyte subpopulation. Both E. intestinalis-infected IFN-γ KO and IL-12 KO mice produced comparable levels of specific antibodies, suggesting that antibodies did not protect IFN-γ KO mice from lethal infection.