Impact of inflammatory preconditioning on murine microglial proteome response induced by focal ischemic brain injury.

Preconditioning with lipopolysaccharide (LPS) induces neuroprotection against subsequent cerebral ischemic injury, mainly involving innate immune pathways. Microglia are resident immune cells of the central nervous system (CNS) that respond early to danger signals through memory-like differential reprogramming. However, the cell-specific molecular mechanisms underlying preconditioning are not fully understood. To elucidate the distinct molecular mechanisms of preconditioning on microglia, we compared these cell-specific proteomic profiles in response to LPS preconditioning and without preconditioning and subsequent transient focal brain ischemia and reperfusion, - using an established mouse model of transient focal brain ischemia and reperfusion. A proteomic workflow, based on isolated microglia obtained from mouse brains by cell sorting and coupled to mass spectrometry for identification and quantification, was applied. Our data confirm that LPS preconditioning induces marked neuroprotection, as indicated by a significant reduction in brain infarct volume. The established brain cell separation method was suitable for obtaining an enriched microglial cell fraction for valid proteomic analysis. The results show a significant impact of LPS preconditioning on microglial proteome patterns by type I interferons, presumably driven by the interferon cluster regulator proteins signal transducer and activator of transcription1/2 (STAT1/2).

Impact of perinatal administration of probiotics on immune cell composition in neonatal mice.

BACKGROUND: Newborns and especially preterm infants are much more susceptible to infections than adults. The pathogens causing infections in newborns are often detectable in the intestinal flora of affected children even before disease onset. Therefore, it seems reasonable to prevent dysbiosis in newborns and preterm infants. An approach followed in many neonatal intensive care units (NICUs) is to prevent infections in preterm infants with probiotics however their mechanisms of action of probiotics are incompletely understood. Here, we investigated the effect of perinatal probiotic exposure on immune cells in newborn mice. METHODS: Pregnant mice were orally treated with a combination of Lactobacillus acidophilus and Bifidobacterium bifidum (Infloran®) from mid-pregnancy until the offspring were harvested. Immune cell composition in organs of the offspring were analyzed by flow cytometry. RESULTS: Perinatal probiotic exposure had profound effects on immune cell composition in the intestine, liver and lungs of newborn mice with reduction of myeloid and B cells and induction of T cells in the probiotic treated animals' organs at weaning. Furthermore, probiotic exposure had an effect on T cell development in the thymus. CONCLUSION: Our results contribute to a better understanding of the interaction of probiotics with the developing immune system. IMPACT: probiotics have profound effects on immune cell composition in intestines, livers and lungs of newborn mice. probiotics modulate T cell development in thymus of newborn mice. effects of probiotics on neonatal immune cells are particularly relevant in transition phases of the microbiome. our results contribute to a better understanding of the mechanisms of action of probiotics in newborns.

Depletion of Ly6G-Expressing Neutrophilic Cells Leads to Altered Peripheral T-Cell Homeostasis and Thymic Development in Neonatal Mice.

Newborns and especially preterm infants are much more susceptible to infections than adults. Due to immature adaptive immunity, especially innate immune cells play an important role in a newborn's infection defense. Neonatal neutrophils exhibit profound differences in their functionality compared to neutrophils of adults. In particular, neonates possess a relevant population of suppressive neutrophils, which not only inhibit but also specifically modulate the function of T-cells. In this study, we investigated whether neonatal neutrophils are already involved in T-cell development in the thymus. For this purpose, we used a newly developed model of antibody-mediated immune cell depletion in which we administered a depleting antibody to pregnant and then lactating dams. Using this method, we were able to sufficiently deplete Ly6G-positive neutrophils in offspring. We demonstrated that the depletion of neutrophils in newborn mice resulted in altered peripheral T-cell homeostasis with a decreased CD4+/CD8+ T-cell ratio and decreased expression of CD62L. Neutrophil depletion even affected T-cell development in the thymus, with increased double positive thymocytes and a decreased CD4+/CD8+ single positive thymocyte ratio. Altogether, we demonstrated a previously unknown mechanism mediating neutrophils' immunomodulatory effects in newborns.

HIF-1α targeted deletion in myeloid cells decreases MDSC accumulation and alters microbiome in neonatal mice.

The newborn's immune system is faced with the challenge of having to learn quickly to fight off infectious agents, but tolerating the colonization of the body surfaces with commensals without reacting with an excessive inflammatory response. Myeloid-derived suppressor cells (MDSC) are innate immune cells with suppressive activity on other immune cells that regulate fetal-maternal tolerance during pregnancy and control intestinal inflammation in neonates. Until now, nothing is known about the role of MDSC in microbiome establishment. One of the transcription factors regulating MDSC homeostasis is the hypoxia-inducible factor 1α (HIF-1α). We investigated the impact of HIF-1α on MDSC accumulation and microbiome establishment during the neonatal period in a mouse model with targeted deletion of HIF-1α in myeloid cells (Hif1a loxP/loxP LysMCre+). We show that in contrast to wildtype mice, where an extensive expansion of MDSC was observed, MDSC expansion in neonatal Hif1a loxP/loxP LysMCre+ mice was dramatically reduced both systemically and locally in the intestine. This was accompanied by an altered microbiome composition and intestinal T-cell homeostasis. Our results point toward a role of MDSC in inflammation regulation in the context of microbiome establishment and thus reveal a new aspect of the biological role of MDSC during the neonatal period.

Training vs. Tolerance: The Yin/Yang of the Innate Immune System.

For almost nearly a century, memory functions have been attributed only to acquired immune cells. Lately, this paradigm has been challenged by an increasing number of studies revealing that innate immune cells are capable of exhibiting memory-like features resulting in increased responsiveness to subsequent challenges, a process known as trained immunity (known also as innate memory). In contrast, the refractory state of endotoxin tolerance has been defined as an immunosuppressive state of myeloid cells portrayed by a significant reduction in the inflammatory capacity. Both training as well tolerance as adaptive features are reported to be accompanied by epigenetic and metabolic alterations occurring in cells. While training conveys proper protection against secondary infections, the induction of endotoxin tolerance promotes repairing mechanisms in the cells. Consequently, the inappropriate induction of these adaptive cues may trigger maladaptive effects, promoting an increased susceptibility to secondary infections-tolerance, or contribute to the progression of the inflammatory disorder-trained immunity. This review aims at the discussion of these opposing manners of innate immune and non-immune cells, describing the molecular, metabolic and epigenetic mechanisms involved and interpreting the clinical implications in various inflammatory pathologies.

Decreased expression of hypoxia-inducible factor 1α (HIF-1α) in cord blood monocytes under anoxia.

BACKGROUND: Infections are a major cause for morbidity and mortality in neonates; however, the underling mechanisms for increased infection susceptibility are incompletely understood. Hypoxia, which is present in inflamed tissues, has been identified as an important activation signal for innate immune cells in adults and is mainly mediated by hypoxia-inducible factor 1α (HIF-1α). Fetal tissue pO2 physiologically is low but rises immediately after birth. METHODS: In this study, the effect of low oxygen partial pressure (pO2) on HIF-1α expression and its targets phagocytosis, reactive oxygen species (ROS) production and vascular endothelial growth factor (VEGF) secretion was compared in vitro between immune cells from adult peripheral blood and cord blood using anoxia, HIF-1α stabilizer desferroxamin (DFO) and E. coli as stimuli. RESULTS: We show that anoxia-induced HIF-1α protein accumulation, phagocytosis, ROS-production and VEGF-expression were greatly diminished in cord blood compared to adult cells. E. coli led to HIF-1α gene expression in adult and cord blood immune cells; however, cord blood cells failed to accumulate HIF-1α protein and VEGF upon E. coli stimulation. CONCLUSIONS: Taken together, our results show a diminished activation of cord blood immune cells by low pO2, which might contribute to impaired reactivity in the context of infection. IMPACT: Neonatal immune cells do not accumulate HIF-1α under low oxygen partial pressure leading to decreased phagocytosis and decreased ROS production. We demonstrate a previously unknown mechanism of reduced activation of neonatal immune cells in the context of an inflammatory response. This could contribute to the increased susceptibility of newborns and preterm infants to infection.

Gram-positive Staphylococcus aureus LTA promotes distinct memory-like effects in murine bone marrow neutrophils.

Neutrophils primarily act as first responders in acute infection and directly maintain inflammatory responses. However, a growing body of evidence suggests that neutrophils also bear the potential to mediate chronic inflammation by exhibiting memory-like features. We now asked whether bone marrow-derived murine neutrophils can be primed by lipoteichoic acid (LTA) from gram-positive S. aureus. We found that low-dose (1 ng/mL) LTA-priming promoted increased production of pro-inflammatory mediators (TNF-α, IL-6, ROS), whereas high-dose (10 µg/mL) priming resulted in opposing reactions marked by increased IL-10 and suppressed pro-inflammatory mediators upon a second stimulus. A similar pattern of pro-inflammatory activation (trained sensitivity) and anti-inflammatory properties (tolerance) was recapitulated in cellular functional in vitro assays (transmigration and phagocytosis). Priming by LTA correlated with TLR2/MyD88-mediated regulation of NFκB-p65 through intermediate PI3Ks/MAPK. Collectively, our data suggest a previously unknown capacity of neutrophils to be differentially primed by varying doses of LTA, endorsing memory-like features in neutrophils.

Gut Microbiota-Derived Small Extracellular Vesicles Endorse Memory-like Inflammatory Responses in Murine Neutrophils.

Neutrophils are classically characterized as merely reactive innate effector cells. However, the microbiome is known to shape the education and maturation process of neutrophils, improving their function and immune-plasticity. Recent reports demonstrate that murine neutrophils possess the ability to exert adaptive responses after exposure to bacterial components such as LPS (Gram-negative bacteria) or LTA (Gram-positive bacteria). We now ask whether small extracellular vesicles (EVs) from the gut may directly mediate adaptive responses in neutrophils in vitro. Murine bone marrow-derived neutrophils were primed in vitro by small EVs of high purity collected from colon stool samples, followed by a second hit with LPS. We found that low-dose priming with gut microbiota-derived small EVs enhanced pro-inflammatory sensitivity as indicated by elevated levels of TNF-α, IL-6, ROS and MCP-1 and increased migratory and phagocytic activity. In contrast, high-dose priming resulted in a tolerant phenotype, marked by increased IL-10 and decreased transmigration and phagocytosis. Alterations in TLR2/MyD88 as well as TLR4/MyD88 signaling were correlated with the induction of adaptive cues in neutrophils in vitro. Taken together, our study shows that small EVs from stools can drive adaptive responses in neutrophils in vitro and may represent a missing link in the gut-immune axis.

PI3Kγ Mediates Microglial Proliferation and Cell Viability via ROS.

(1) Background: Rapid microglial proliferation contributes to the complex responses of the innate immune system in the brain to various neuroinflammatory stimuli. Here, we investigated the regulatory function of phosphoinositide 3-kinase γ (PI3Kγ) and reactive oxygen species (ROS) for rapid proliferation of murine microglia induced by LPS and ATP. (2) Methods: PI3Kγ knockout mice (PI3Kγ KO), mice expressing catalytically inactive PI3Kγ (PI3Kγ KD) and wild-type mice were assessed for microglial proliferation using an in vivo wound healing assay. Additionally, primary microglia derived from newborn wild-type, PI3Kγ KO and PI3Kγ KD mice were used to analyze PI3Kγ effects on proliferation and cell viability, senescence and cellular and mitochondrial ROS production; the consequences of ROS production for proliferation and cell viability after LPS or ATP stimulation were studied using genetic and pharmacologic approaches. (3) Results: Mice with a loss of lipid kinase activity showed impaired proliferation of microglia. The prerequisite of induced microglial proliferation and cell viability appeared to be PI3Kγ-mediated induction of ROS production. (4) Conclusions: The lipid kinase activity of PI3Kγ plays a crucial role for microglial proliferation and cell viability after acute inflammatory activation.

LPS Induces Opposing Memory-like Inflammatory Responses in Mouse Bone Marrow Neutrophils.

A growing body of evidence suggests that innate immune cells can respond in a memory-like (adaptive) fashion, which is referred to as trained immunity. Only few in vivo studies have shown training effects in neutrophils; however, no in vitro setup has been established to study the induction of trained immunity or tolerance in neutrophils by microbial agents. In light of their short lifespan (up to 48 h), we suggest to use the term trained sensitivity for neutrophils in an in vitro setting. Here, we firstly describe a feasible two-hit model, using different doses of lipopolysaccharide (LPS) in bone marrow neutrophils. We found that low doses (10 pg/mL) induce pro-inflammatory activation (trained sensitivity), whereas priming with high doses (100 ng/mL) leads to suppression of pro-inflammatory mediators such as TNF-α or IL-6 (tolerance) (p < 0.05). On a functional level, trained neutrophils displayed increased phagocytic activity and LFA-1 expression as well as migrational capacity and CD11a expression, whereas tolerant neutrophils show contrasting effects in vitro. Mechanistically, TLR4/MyD88/PI3Ks regulate the activation of p65, which controls memory-like responses in mouse bone marrow neutrophils (p < 0.05). Our results open a new window for further in vitro studies on memory-like inflammatory responses of short-lived innate immune cells such as neutrophils.

The Role of the Pathogen Dose and PI3Kγ in Immunometabolic Reprogramming of Microglia for Innate Immune Memory.

Microglia, the innate immune cells of the CNS, exhibit long-term response changes indicative of innate immune memory (IIM). Our previous studies revealed IIM patterns of microglia with opposing immune phenotypes: trained immunity after a low dose and immune tolerance after a high dose challenge with pathogen-associated molecular patterns (PAMP). Compelling evidence shows that innate immune cells adopt features of IIM via immunometabolic control. However, immunometabolic reprogramming involved in the regulation of IIM in microglia has not been fully addressed. Here, we evaluated the impact of dose-dependent microglial priming with ultra-low (ULP, 1 fg/mL) and high (HP, 100 ng/mL) lipopolysaccharide (LPS) doses on immunometabolic rewiring. Furthermore, we addressed the role of PI3Kγ on immunometabolic control using naïve primary microglia derived from newborn wild-type mice, PI3Kγ-deficient mice and mice carrying a targeted mutation causing loss of lipid kinase activity. We found that ULP-induced IIM triggered an enhancement of oxygen consumption and ATP production. In contrast, HP was followed by suppressed oxygen consumption and glycolytic activity indicative of immune tolerance. PI3Kγ inhibited glycolysis due to modulation of cAMP-dependent pathways. However, no impact of specific PI3Kγ signaling on immunometabolic rewiring due to dose-dependent LPS priming was detected. In conclusion, immunometabolic reprogramming of microglia is involved in IIM in a dose-dependent manner via the glycolytic pathway, oxygen consumption and ATP production: ULP (ultra-low-dose priming) increases it, while HP reduces it.

The Role of Microbiota in Neutrophil Regulation and Adaptation in Newborns.

Newborns are highly susceptible to infections and mainly rely on innate immune functions. Reduced reactivity, delayed activation and subsequent failure to resolve inflammation however makes the neonatal immune system a very volatile line of defense. Perinatal microbiota, nutrition and different extra-uterine factors are critical elements that define long-term outcomes and shape the immune system during the neonatal period. Neutrophils are first responders and represent a vital component of the immune system in newborns. They have long been regarded as merely executive immune cells, however this notion is beginning to shift. Neutrophils are shaped by their surrounding and adaptive elements have been described. The role of "innate immune memory" and the main triangle connection microbiome-neutrophil-adaptation will be discussed in this review.

Memory-Like Responses of Brain Microglia Are Controlled by Developmental State and Pathogen Dose.

Microglia, the innate immune cells of the central nervous system, feature adaptive immune memory with implications for brain homeostasis and pathologies. However, factors involved in the emergence and regulation of these opposing responses in microglia have not been fully addressed. Recently, we showed that microglia from the newborn brain display features of trained immunity and immune tolerance after repeated contact with pathogens in a dose-dependent manner. Here, we evaluate the impact of developmental stage on adaptive immune responses of brain microglia after repeated challenge with ultra-low (1 fg/ml) and high (100 ng/ml) doses of the endotoxin LPS in vitro. We find that priming of naïve microglia derived from newborn but not mature and aged murine brain with ultra-low LPS significantly increased levels of pro-inflammatory mediators TNF-α, IL-6, IL-1ß, MMP-9, and iNOS as well as neurotrophic factors indicating induction of trained immunity (p < 0.05). In contrast, stimulation with high doses of LPS led to a robust downregulation of pro-inflammatory cytokines and iNOS independent of the developmental state, indicating induced immune tolerance. Furthermore, high-dose priming with LPS upregulated anti-inflammatory mediators IL-10, Arg-1, TGF- ß, MSR1, and IL-4 in newborn microglia (p < 0.05). Our data indicate pronounced plasticity of the immune response of neonate microglia compared with microglia derived from mature and aged mouse brain. Induced trained immunity after priming with ultra-low LPS doses may be responsible for enhanced neuro-inflammatory susceptibility of immature brain. In contrast, the immunosuppressed phenotype following high-dose LPS priming might be prone to attenuate excessive damage after recurrent systemic inflammation.

Impact of ambient temperature on inflammation-induced encephalopathy in endotoxemic mice-role of phosphoinositide 3-kinase gamma.

BACKGROUND: Sepsis-associated encephalopathy (SAE) is an early and frequent event of infection-induced systemic inflammatory response syndrome. Phosphoinositide 3-kinase γ (PI3Kγ) is linked to neuroinflammation and inflammation-related microglial activity. In homeotherms, variations in ambient temperature (Ta) outside the thermoneutral zone lead to thermoregulatory responses, mainly driven by a gradually increasing sympathetic activity, and may affect disease severity. We hypothesized that thermoregulatory response to hypothermia (reduced Ta) aggravates SAE in PI3Kγ-dependent manner. METHODS: Experiments were performed in wild-type, PI3Kγ knockout, and PI3Kγ kinase-dead mice, which were kept at neutral (30 ± 0.5 °C) or moderately lowered (26 ± 0.5 °C) Ta. Mice were exposed to lipopolysaccharide (LPS, 10 µg/g, from Escherichia coli serotype 055:B5, single intraperitoneal injection)-evoked systemic inflammatory response (SIR) and monitored 24 h for thermoregulatory response and blood-brain barrier integrity. Primary microglial cells and brain tissue derived from treated mice were analyzed for inflammatory responses and related cell functions. Comparisons between groups were made with one-way or two-way analysis of variance, as appropriate. Post hoc comparisons were made with the Holm-Sidak test or t tests with Bonferroni's correction for adjustments of multiple comparisons. Data not following normal distribution was tested with Kruskal-Wallis test followed by Dunn's multiple comparisons test. RESULTS: We show that a moderate reduction of ambient temperature triggers enhanced hypothermia of mice undergoing LPS-induced systemic inflammation by aggravated SAE. PI3Kγ deficiency enhances blood-brain barrier injury and upregulation of matrix metalloproteinases (MMPs) as well as an impaired microglial phagocytic activity. CONCLUSIONS: Thermoregulatory adaptation in response to ambient temperatures below the thermoneutral range exacerbates LPS-induced blood-brain barrier injury and neuroinflammation. PI3Kγ serves a protective role in suppressing release of MMPs, maintaining microglial motility and reinforcing phagocytosis leading to improved brain tissue integrity. Thus, preclinical research targeting severe brain inflammation responses is seriously biased when basic physiological prerequisites of mammal species such as preferred ambient temperature are ignored.

L-Arginine Modulates Neonatal Leukocyte Recruitment in a Gestational Age-Dependent Manner.

(1) Background: L-arginine is a complex modulator of immune functions, and its levels are known to decrease under septic conditions. L-arginine may suppress leukocyte recruitment in vivo; however, little is known about the gestational age-specific effects of L-arginine on leukocyte recruitment in preterm infants. We now asked whether L-arginine alters leukocyte recruitment in preterm and term neonates. (2) Methods: Leukocytes were isolated from preterm (28 + 0 to 32 + 6 weeks of gestation) and term (>37 weeks of gestation) newborns as well as from healthy adults. After incubation with 10 µg/mL L-arginine, we assessed leukocyte rolling and adhesion in dynamic microflow chamber experiments and leukocyte transmigration in fluorescence assays. In addition, we measured the expression of inducible nitric oxide synthase (iNOS) and Arginase 1 (Arg-1) in neutrophils by flow cytometry. (3) Results: Leukocyte rolling, adhesion, and transmigration increased with gestational age. Leukocyte rolling, adhesion, and transmigration were decreased by L-arginine in term-born infants and adults. Preterm leukocytes showed no change in recruitment upon L-arginine exposure. Leukocyte adhesion after L-arginine exposure reached similar levels among all groups. In line, the expression of iNOS and Arg-1 was similar in all three age groups. (4) Conclusion: L-arginine dampens the ex vivo recruitment capacity of leukocytes from term-born infants, whereas no effect was seen in premature infants. As levels of iNOS and Arg-1 in neutrophils remain ontogenetically unchanged, the anti-inflammatory effect of L-arginine on the leukocyte recruitment cascade needs further investigation. These results add to the controversial debate of L-arginine supplementation in premature infants in sepsis.

Leukocyte Infiltration of Cremaster Muscle in Mice Assessed by Intravital Microscopy.

Intravital microscopy (IVM) is widely used to monitor physiological and pathophysiological processes within the leukocyte recruitment cascade in vivo. The current protocol represents a practical and reproducible method to visualize the leukocyte endothelium interaction leading to leukocyte recruitment in skeletal muscle derived tissue within the intact organism of the mouse. The model is applicable to all fields of research that focus on granulocyte activation and their role in disease. We provide a step by step protocol to guide through the method and to highlight potential pitfalls and technical difficulties. The protocol covers the following aspects: experimental settings and required material, anesthesia of the mouse, dissection of the cremaster muscle as well as tracheal and carotid cannulation, IVM recordings and offline analysis. Data formats like adherent leukocytes, rolling flux (RF) and rolling flux fraction (RFF) are explained in detail and appropriate applications are discussed. Representative results from dystrophin deficient mdx mice are provided in the results section. IVM is a powerful tool to assess leukocyte recruitment in an in vivo setting; however, delineating for example endothelial and leukocyte function may require a combination with ex vivo setups like flow chamber experiments. Furthermore, the genetic background of animals of interest may greatly influence baseline recruitment, requiring individual fine tuning of the protocol provided. Despite its limitations, IVM may serve as a platform to readily translate in vitro findings into a living vertebrate organism.

Memory-Like Inflammatory Responses of Microglia to Rising Doses of LPS: Key Role of PI3Kγ.

Trained immunity and immune tolerance have been identified as long-term response patterns of the innate immune system. The causes of these opposing reactions remain elusive. Here, we report about differential inflammatory responses of microglial cells derived from neonatal mouse brain to increasing doses of the endotoxin LPS. Prolonged priming with ultra-low LPS doses provokes trained immunity, i.e., increased production of pro-inflammatory mediators in comparison to the unprimed control. In contrast, priming with high doses of LPS induces immune tolerance, implying decreased production of inflammatory mediators and pronounced release of anti-inflammatory cytokines. Investigation of the signaling processes and cell functions involved in these memory-like immune responses reveals the essential role of phosphoinositide 3-kinase γ (PI3Kγ), one of the phosphoinositide 3-kinase species highly expressed in innate immune cells. Together, our data suggest profound influence of preceding contacts with pathogens on the immune response of microglia. The impact of these interactions-trained immunity or immune tolerance-appears to be shaped by pathogen dose.

Reduced ambient temperature exacerbates SIRS-induced cardiac autonomic dysregulation and myocardial dysfunction in mice.

Sepsis-induced myocardial depression (SIMD) is an early and frequent consequence of the infection-induced systemic inflammatory response syndrome. In homiotherms, variations in ambient temperature (Ta) outside the thermoneutral zone induce thermoregulatory responses mainly driven by a gradually increased sympathetic activity, which may affect disease severity. We hypothesized that thermoregulatory responses upon reduced Ta exposition aggravate SIMD in mice. Mice were kept at neutral Ta (30 ± 0.5 °C), moderately lowered Ta (26 ± 0.5 °C) or markedly lowered Ta (22 ± 0.5 °C), exposed to lipopolysaccharide- (LPS, 10 µg/g, from Escherichia coli serotype 055:B5, single intraperitoneal injection) evoked shock and monitored for survival, cardiac autonomic nervous system function and left ventricular performance. Primary adult cardiomyocytes and heart tissue derived from treated mice were analyzed for inflammatory responses and signaling pathways of myocardial contractility. We show that a moderate reduction of Ta to 26 °C led to a 40% increased mortality of LPS-treated mice when compared to control mice and that a marked reduction of Ta to 22 °C resulted in an early mortality of all mice. Mice kept at 26 °C exhibited increased heart rate and altered indices of heart rate variability (HRV), indicating sympathovagal imbalance along with aggravated LPS-induced SIMD. This SIMD was associated with reduced myocardial ß-adrenergic receptor expression and suppressed adrenergic signaling, as well as with increased myocardial iNOS expression, nitrotyrosine formation and leukocyte invasion as well as enhanced apoptosis and appearance of contraction band necrosis in heart tissue. While ineffective separately, combined treatment with the ß2-adrenergic receptor (AR) antagonist ICI 118551 (10 ng/gbw) and the inducible nitric oxide synthase (iNOS) inhibitor 1400 W (5 µg/gbw) reversed the increase in LPS-induced mortality and aggravation of SIMD at reduced Ta. Thus, consequences of thermoregulatory adaptation in response to ambient temperatures below the thermoneutral range increase the mortality from LPS-evoked shock and markedly prolong impaired myocardial function. These changes are mitigated by combined ß2-AR and iNOS inhibition.

Toxin-induced hormesis may restrain aging.

Mild environmental stress might have beneficial effects in aging by activating maintenance and repair processes in cells and organs. These beneficial stress effects fit to the concept of hormesis. Prominent stressors acting in a hormetic way are physical exercises, fasting, cold and heat. This review will introduce some toxins, which have been found to induce hormetic responses in animal models of aging research. To highlight the molecular signature of these hormetic effects we will depict signaling pathways affected by low doses of toxins on cellular and organismic level. As prominent examples for signaling pathways involved in both aging processes as well as toxin responses, PI3K/Akt/mTOR- and AMPK-signal transduction will be described in more detail. Due to the striking overlap of signaling pathways mediating toxin induced responses and aging processes we propose considering the ability of low doses of toxins to slow down the rate of aging.