IL-4 and IL-13 Induce Protection of Porcine Endothelial Cells from Killing by Human Complement and from Apoptosis through Activation of a Phosphatidylinositide 3-Kinase/Akt Pathway

Abstract

Vascular endothelial cells (EC) perform critical functions that require a balance of cell survival and cell death. EC death by apoptosis and EC activation and injury by the membrane attack complex of complement are important mechanisms in atherosclerosis and organ graft rejection. Although the effects of various cytokines on EC apoptosis have been studied, little is known about their effects on complement-mediated EC injury. Therefore, we studied the abilities of various cytokines to induce protection of porcine aortic EC against apoptosis and killing by human complement, a model of pig-to-human xenotransplantation. We found that porcine EC incubated with IL-4 or IL-13, but not with IL-10 or IL-11, became protected from killing by complement and apoptosis induced by TNF-α plus cycloheximide. Maximal protection required 10 ng/ml IL-4 or IL-13, developed progressively from 12 to 72 h of incubation, and lasted 48–72 h after cytokine removal. Protection from complement was not associated with reduced complement activation, C9 binding, or changes in CD59 expression. Inhibition of PI3K prevented development of protection; however, inhibition of p38 MAPK or p42/44 MAPK had no effect. IL-4 and IL-13 induced rapid phosphorylation of Akt. Although protection was inhibited by an Akt inhibitor and a dominant negative Akt mutant transduced into EC, it was induced by transduction of EC with the constitutively active Akt variant, myristylated Akt. We conclude that IL-4 and IL-13 can induce protection of porcine EC against killing by apoptosis and human complement through activation of the PI3K/Akt signaling pathway.

STAT6 Mediates Apoptosis of Human Coronary Arterial Endothelial Cells by Interleukin-13

“…There are conflicting reports on the effects of IL-13 and IL-4 on EC apoptosis (12,13,22,23). Our results support the findings of Volpert et al (13), who reported that IL-13 and IL-4 inhibited angiogenesis in the rat cornea, and in human and bovine microvascular ECs.…”

“…In addition, induction of the IL-4 gene significantly suppressed angiogenesis in a rat model of adjuvant-induced arthritis (24). On the other hand, there are reports (22,23) that apoptosis induced by TNF-α in combination with cycloheximide was inhibited by IL-4 and IL-13. The discrepancy between the previous reports and the present report might be due to the difference in experimental conditions.…”

“…We presume that IL-13 has dual effects on apoptosis; i.e., it shows proapoptotic activity in the presence of growth factors, and it shows anti-apoptotic activity in the presence of proapoptotic factors and/or in the absence of factors supporting survival. It is also likely that considerable numbers of ECs were already dead of apoptosis under the conditions of the previous study (22,23), because the cells were pretreated with IL-4 and IL-13 for 72 h before the treatment with TNF-α in combination with cycloheximide. Nonetheless, the precise causes of these discrepant findings need to be clarified in a later study.…”

IL‐4 induces protection of vascular endothelial cells against killing by complement and melittin through lipid biosynthesis

“…However, the small but significant decreased permeability in IL-4-treated cells may preserve viability and capacity for ATP synthesis, as it occurs in transiently permeabilized nucleated cells following exposure to low MAC concentrations, allowing for membrane repair [5,36]. IL-4 does not appear to significantly change MAC removal, since we found only a slight, not statistically significant, reduction in bound MAC at the end of the complement treatment (2 h) and no differences at an earlier time (30 min) [8,11].Our finding that protected EC had minimal reduction in MACmediated loss of permeability control suggested to us that IL-4 would induce, through activation of Akt/SREBP-1/lipid synthesis, strong cellular mechanisms that assure cell viability until the membrane lesions resealed. Therefore, we asked whether activation of SREBP-1 and subsequent lipid synthesis might impart protection by endowing the mitochondria with resistance against complement injury.…”

“…This observation is likely explained through a mechanism in which control EC lose both cytoplasmic and mitochondrial ATP, whereas IL-4-treated EC lose cytoplasmic ATP but little or no mitochondrial ATP. These experiments demonstrated that, despite showing only minimal reduction in MAC-induced loss of permeability control, pretreatment of EC with IL-4 induces protection from MAC-mediated killing, as shown by their ability to take up a vital dye and to retain a significant amount of cellular ATP.We previously reported that IL-4-induced protection from the MAC is not associated with reduced binding of MAC to the cell membrane when MAC was measured after 30 min of incubation with complement [8,11]. We now extended the study by measuring bound MAC at the end of the incubation with complement for 2 h, the incubation time used in most experiments.…”