Suzanne R. Broussard

Cytokine-induced sickness behavior

The behavioral repertoire of humans and animals changes dramatically following infection. Sick individuals have little motivation to eat, are listless, complain of fatigue and malaise, loose interest in social activities and have significant changes in sleep patterns. They display an inability to experience pleasure, have exaggerated responses to pain and fail to concentrate. Proinflammatory cytokines acting in the brain cause sickness behaviors. These nearly universal behavioral changes are a manifestation of a central motivational state that is designed to promote recovery. Exaggerated symptoms of sickness in cancer patients, such as cachexia, can be life-threatening. However, quality of life is often drastically impaired before the cancer becomes totally debilitating. Although basic studies in psychoneuroimmunology have defined proinflammatory cytokines as the central mediators of sickness behavior, a much better understanding of how cytokine and neurotransmitter receptors communicate with each other is needed. Advances that have been made during the past decade should now be extended to clinical studies in an attempt to alleviate sickness symptoms and improve quality of life for cancer patients.

IL-1β impairs insulin-like growth factor I-induced differentiation and downstream activation signals of the insulin-like growth factor I receptor in myoblasts

Proinflammatory cytokines are elevated in disorders characterized by muscle wasting and weakness, such as inflammatory myopathies and AIDS wasting. We recently demonstrated that TNF-α impairs the ability of insulin-like growth factor (IGF)-I to promote protein synthesis in muscle precursor cells. In this study we extend these findings by showing that low concentrations of IL-1β impair IGF-I-dependent differentiation of myoblasts, as assessed by expression of the muscle specific protein, myosin heavy chain. In the absence of exogenous IGF-I, IL-1β (1 ng/ml) did not impair muscle cell development. However, in the presence of IGF-I, 100-fold lower concentrations of IL-1β (0.01 ng/ml) significantly suppressed myoblast differentiation, protein synthesis, and myogenin expression. Increasing IL-1β to 1 ng/ml completely blocked the anabolic actions of IGF-I in murine C2C12 myoblasts. Similarly, IL-1β inhibited IGF-I-stimulated protein synthesis in primary porcine myoblasts. IL-1β impaired the actions of IGF-I at a point distal to the IGF receptor, and this was not due to IL-1β-induced cell death. Instead, IL-1β inhibited the ability of IGF-I to phosphorylate tyrosine residues on both of its downstream docking proteins, insulin receptor substrate 1 and insulin receptor substrate 2. These data establish that physiological concentrations of IL-1β block the ability of IGF-I to promote protein synthesis, leading to reduced expression of the myogenic transcription factor, myogenin, and the subsequent development of more mature differentiated cells that express myosin heavy chain. Collectively, the results are consistent with the notion that very low concentrations of IL-1β significantly impair myogenesis, but they are unable to do so in the absence of the growth factor IGF-I.

IL-10 Inhibits Apoptosis of Promyeloid Cells by Activating Insulin Receptor Substrate-2 and Phosphatidylinositol 3′-Kinase

IL-10 is well known to be a potent inhibitor of the synthesis of proinflammatory cytokines, but noninflammatory hemopoietic cells also express IL-10Rs. Here we show that IL-10 directly affects progenitor myeloid cells by protecting them from death following the removal of growth factors. Murine factor-dependent cell progenitors cultured in the absence of growth factors were 43 ± 1% apoptotic after 12 h. Addition of IL-10 at a concentration as low as 100 pg/ml significantly reduced the apoptotic population to 32 ± 3%. At 10 ng/ml, IL-10 caused a 4-fold reduction in the apoptotic population (11 ± 1%). The anti-apoptotic activity of IL-10 was significantly inhibited with a neutralizing IL-10R Ab. Factor-dependent cell progenitor promyeloid cells expressed functional IL-10Rs, as assessed by precipitation of a 110-kDa protein with an Ab to the IL-10R and by the ability of IL-10 to activate Jak1 and Tyk2 and to phosphorylate tyrosine 705 on Stat-3. IL-10 increased tyrosyl phosphorylation of insulin receptor substrate-2 and stimulated the enzymatic activity of both phosphatidylinositol 3′-kinase and Akt. The anti-apoptotic activity of IL-10 was blocked by inhibition of phosphatidylinositol 3′-kinase. Wortmannin and LY294002 also totally inhibited activation of extracellular signal-related kinase (ERK)1/2 by IL-10. Direct inhibition of ERK1/2 with the mitogen-activated protein kinase/ERK kinase inhibitor PD98059 partially, but significantly, impaired the anti-apoptotic activity of IL-10. These data establish that activation of the IL-10R promotes survival of progenitor myeloid cells. This survival-promoting activity is totally due to IL-10 stimulating the insulin receptor substrate-2/PI 3-kinase/Akt pathway, which increases the anti-apoptotic activity of ERK1/2.

IL-10 promotes survival of microglia without activating Akt

IL-10 is an anti-inflammatory cytokine that has recently been shown to promote survival of neurons and glia. Here we establish that IL-10 induces phosphorylation of Stat3 on Tyr(705) and serves as a survival factor for N13 microglial cells. Recombinant IL-10 (10 ng/ml) decreases growth factor withdrawal-induced apoptosis by 50%, as assessed by TUNEL. In contrast to IL-10, IGF-I increases enzymatic activity of PI 3-kinase and causes phosphorylation on serine(473) of Akt but does not prevent microglial apoptosis. These data establish that IL-10 activates Stat3 and inhibits the mitochondrial pathway of cell death without activating the Akt cell survival pathway.

Exacerbated fatigue and motor deficits in interleukin-10-deficient mice after peripheral immune stimulation

The anti-inflammatory cytokine interleukin (IL)-10 is important for regulating inflammation in the periphery and brain, but whether it protects against infection- or age-related psychomotor disturbances and fatigue is unknown. Therefore, the present study evaluated motor coordination, time to fatigue, and several central and peripheral proinflammatory cytokines in male young adult (3-mo-old) and middle-aged (12-mo-old) wild-type (IL-10(+/+)) and IL-10-deficient (IL-10(-/-)) mice after intraperitoneal injection of lipopolysaccharide (LPS) or saline. No age-related differences were observed; therefore, data from the two ages were pooled and analyzed to determine effects of genotype and treatment. LPS treatment increased IL-1beta, IL-6, and TNFalpha mRNA in all brain areas examined in IL-10(+/+) and IL-10(-/-) mice, but to a greater extent and for a longer time in IL-10(-/-) mice. Plasma IL-1beta and IL-6 were increased similarly in IL-10(+/+) and IL-10(-/-) mice 4 h after LPS but remained elevated longer in IL-10(-/-) mice, whereas TNFalpha was higher in IL-10(-/-) mice throughout after LPS treatment. Motor performance and motor learning in IL-10(+/+) mice were not affected by LPS treatment; however, both were reduced in IL-10(-/-) mice treated with LPS compared with those treated with saline. Furthermore, although LPS reduced the time to fatigue in IL-10(+/+) and IL-10(-/-) mice, the effects were exacerbated in IL-10(-/-) mice. Thus the increased brain and peripheral inflammation induced by LPS in IL-10(-/-) mice was associated with increased coordination deficits and fatigue. These data suggest that IL-10 may inhibit motor deficits and fatigue associated with peripheral infections via its anti-inflammatory effects.

Insulin-like growth factor-I enhances the biological activity of brain-derived neurotrophic factor on cerebrocortical neurons

Insulin-like growth factor (IGF)-I and brain-derived neurotrophic factor (BDNF) act within the brain to enhance neuronal survival and plasticity. We extend these findings by showing that the presence of both neurotrophins is required to depress the rise in intracellular Ca2+ caused by glutamate in primary cultures of cerebrocortical neurons. IGF-I enhanced expression of BDNF receptors (Trk-B) and increased the ability of BDNF to induce ERK1/2 phosphorylation. This IGF-I-induced increase in BDNF responsiveness describes a new interaction between these peptides in the brain.

Tumor Necrosis Factor α Inhibits Cyclin A Expression and Retinoblastoma Hyperphosphorylation Triggered by Insulin-like Growth Factor-I Induction of New E2F-1 Synthesis

Cyclin A is required for cell cycle S phase entry, and its overexpression contributes to tumorigenesis. Release of pre-existing E2Fs from inactive complexes of E2F and hypophosphorylated retinoblastoma (RB) is the prevailing dogma for E2F transcriptional activation of target genes such as cyclin A. Here we explored the hypothesis that new synthesis of E2F-1 is required for insulin-like growth factor-I (IGF-I) to induce cyclin A accumulation and RB hyperphosphorylation, events that are targeted by tumor necrosis factor α (TNFα) to arrest cell cycle progression. We first established that IGF-I increases expression of cyclin A, causes hyperphosphorylation of RB, and augments the mass of E2F-1 in a time-dependent manner. As expected, E2F-1 small interfering RNA blocks the ability of IGF-I to increase synthesis of E2F-1. Most important, this E2F-1 small interfering RNA also blocks the ability of IGF-I to increase cyclin A accumulation and to hyperphosphorylate RB. We next established that TNFα dose-dependently inhibits IGF-I-induced phosphorylation of both RB and histone H1 by cyclin A-dependent cyclin-dependent kinases. Cyclin-dependent kinase 2 (Cdk2) mediates this suppression because co-immunoprecipitation experiments revealed that TNFα reduces the amount of IGF-I-induced cyclin A that binds Cdk2, leading to a reduction in Cdk2 enzymatic activity. TNFα antagonizes the ability of IGF-I to increase mass of both E2F-1 and cyclin A but not cyclin E or D1. The cytostatic property of TNFα is also shown by its ability to block IGF-I-stimulated luciferase activity of a cyclin A promoter reporter. Deletion of an E2F recognition site from this reporter eliminates the regulatory effects of both IGF-I and TNFα on cyclin A transcription, indicating the essential role of E2F-1 in mediating their cross-talk. Collectively, these results establish that TNFα targets IGF-I-induced E2F-1 synthesis, leading to inhibition of the subsequent accumulation in cyclin A, formation of cyclin A-Cdk2 complexes, hyperphosphorylation of RB, and cell cycle arrest.

Tumor necrosis factorα and insulin-like growth factor-I in the brain: Is the whole greater than the sum of its parts?

Abstract The cytokine tumor necrosis factorα (TNFα) and the hormone insulin-like growth factor-I (IGF-I) have both been shown to regulate inflammatory events in the central nervous system (CNS). This review summarizes the seemingly independent roles of TNFα and IGF-I in promoting and inhibiting neurodegenerative diseases. We then offer evidence that the combined effects of IGF-I and TNFα on neuronal survival can be vastly different when both receptors are stimulated simultaneously, as is likely to occur in vivo. We propose the framework of a molecular model of hormone-cytokine receptor cross talk in which disparate cell surface receptors share intracellular substrates that regulate neuronal survival.

IL-1β suppresses prolonged Akt activation and expression of E2F-1 and cyclin a in breast cancer cells

Cell cycle aberrations occurring at the G1/S checkpoint often lead to uncontrolled cell proliferation and tumor growth. We recently demonstrated that IL-1β inhibits insulin-like growth factor (IGF)-I-induced cell proliferation by preventing cells from entering the S phase of the cell cycle, leading to G0/G1 arrest. Notably, IL-1β suppresses the ability of the IGF-I receptor tyrosine kinase to phosphorylate its major docking protein, insulin receptor substrate-1, in MCF-7 breast carcinoma cells. In this study, we extend this juxtamembrane cross-talk between cytokine and growth factor receptors to downstream cell cycle machinery. IL-1β reduces the ability of IGF-I to activate Cdk2 and to induce E2F-1, cyclin A, and cyclin A-dependent phosphorylation of a retinoblastoma tumor suppressor substrate. Long-term activation of the phosphatidylinositol 3-kinase/Akt signaling pathway, but not the mammalian target of rapamycin or mitogen-activated protein kinase pathways, is required for IGF-I to hyperphosphorylate retinoblastoma and to cause accumulation of E2F-1 and cyclin A. In the absence of IGF-I to induce Akt activation and cell cycle progression, IL-1β has no effect. IL-1β induces p21Cip1/Waf1, which may contribute to its inhibition of IGF-I-activated Cdk2. Collectively, these data establish a novel mechanism by which prolonged Akt phosphorylation serves as a convergent target for both IGF-I and IL-1β; stimulation by growth factors such as IGF-I promotes G1-S phase progression, whereas IL-1β antagonizes IGF-I-induced Akt phosphorylation to induce cytostasis. In this manner, Akt serves as a critical bridge that links proximal receptor signaling events to more distal cell cycle machinery.

Insulin-like growth factor-I and the cytokines IL-3 and IL-4 promote survival of progenitor myeloid cells by different mechanisms

Hormones, such as insulin-like growth factor-I (IGF-I), and cytokines, like IL-3 and IL-4, promote survival of progenitor myeloid cells. Here we demonstrate that IGF-I, IL-3 and IL-4 all significantly block activation of caspase-3 in promyeloid cells following growth factor deprivation. However, only IL-3 and IGF-I increase enzymatic activity and phosphorylation of the survival-promoting kinase Akt. IGF-I fails to reduce caspase-3 activity and cell death in the presence of the PI 3-kinase inhibitors, wortmannin and LY294002, whereas these blockers do not affect the ability of IL-3 to maintain cell survival. IL-4 inhibits caspase-3 activity and promotes promyeloid cell survival by a substrate for PI 3-kinase that is not Akt. These data establish that IGF-I inhibits activation of caspase-3 and promotes promyeloid cell survival through a PI 3-kinase-dependent pathway, whereas IL-3 does not. It therefore appears that signal transduction pathways for all three receptors converge upstream of caspase-3 to prevent apoptosis of progenitor myeloid cells, but their receptors differ in the intracellular substrates that are used to promote cell survival.