Benzyl isothiocyanate promotes apoptosis of oral cancer cells via an acute redox stress-mediated DNA damage response.

Benzyl isothiocyanate (BITC) is a cruciferous vegetable-derived compound with anticancer properties in human cancer cells. However, its anticancer potential and underlying mechanisms remain absent in human oral cancer cells. Results indicate that BITC inhibits growth, promotes G2/M phase arrest and triggers apoptosis of OC2 cells with a minimal toxicity to normal cells. BITC-induced cell death was completely prevented by pretreatment with thiol-containing redox compounds including N-acetyl-l-cysteine (NAC), glutathione (GSH), dithiothreitol, and 2-mercaptoethanol, but not free radical scavengers mito-TEMPO, catalase, apocynin, l-NAME and mannitol. BITC rapidly produced reactive oxygen species and nitric oxide, triggered oxidative DNA damage. BITC effectively decreased the intracellular GSH and GSH/GSSG ratio and redox balance recovery by thiol-containing redox compounds, but not by free radical scavengers. Accordingly, redox stresses-DNA damage response (DDR) activated ATM, Chk2, p53, and p21 and subsequently resulted in G2/M phase arrest by inhibiting Cdc2 and cyclin B1. Notably, BITC-induced apoptosis was associated with reduced Mcl-1 and Bcl-2 expression, diminished mitochondrial membrane potential (ΔΨm), and increased PARP cleavage. These BITC-induced redox stress-mediated DDR and apoptosis could be blocked by NAC and GSH. Therefore, BITC can be a rational drug candidate for oral cancer and acted via a redox-dependent pathway.

Attenuating heat-induced cellular autophagy, apoptosis and damage in H9c2 cardiomyocytes by pre-inducing HSP70 with heat shock preconditioning.

PURPOSE: We sought to assess whether heat-induced autophagy, apoptosis and cell damage in H9c2 cells can be affected by pre-inducing HSP70 (heat shock protein 70). MATERIALS AND METHODS: Cell viability was determined using 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide staining and a lactate dehydrogenase assay. Apoptosis was evidenced using both flow cytometry and counting caspase-3 positive cells, whereas autophagy was evidenced by the increased LC3-II expression and lysosomal activity. RESULTS: The viability of H9c2 cells was temperature-dependently (40-44 °C) and time-dependently (90-180 min) significantly (p < 0.05) reduced by severe heat, which caused cell damage, apoptosis and autophagy. Heat-induced cell injury could be attenuated by pretreatment with 3-methylademine (an autophagy inhibitor) or Z-DEVD-FMK (a caspase-3 inhibitor). Neither apoptosis nor autophagy over the levels found in normothermic controls was induced in heat-shock preconditioned controls (no subsequent heat injury). The beneficial effects of mild heat preconditioning (preventing heat-induced cell damage, apoptosis and autophagy) were significantly attenuated by inhibiting HSP70 overexpression with triptolide (Tripterygium wilfordii) pretreatment. CONCLUSION: We conclude that pre-inducing HSP70 attenuates heat-stimulated cell autophagy, apoptosis and damage in the heart. However, this requires in vivo confirmation.

Tetramethylpyrazine decreases hypothalamic glutamate, hydroxyl radicals and prostaglandin-E2 and has antipyretic effects.

OBJECTIVE: We studied the effects of tetramethylpyrazine (TMP) on the fever, increased plasma levels of tumor necrosis factor-α (TNF-α) and increased hypothalamic levels of glutamate, hydroxyl radicals and prostaglandin-E2 (PGE2) induced by lipopolysaccharide (LPS). MATERIALS AND METHODS: The microdialysis probes were stereotaxically and chronically implanted into the hypothalamus of rabbit brain for determining extracellular levels of glutamate, hydroxyl radials, and PGE2. In addition, both the body core temperature and plasma levels of TNF-α were measured. RESULTS: All the body core temperature, plasma levels of TNF-α, and hypothalamic levels of glutamate, hydroxyl radicals, and PGE2 were up-graded by an intravenous dose of LPS (2 µg/kg). Pretreatment with intravenous TMP (10-40 mg/kg) or intracerebroventricular TMP (130 µg in 20 µl per animal) 1 h before LPS administration significantly attenuated the LPS-induced fever as well as the increased hypothalamic levels of glutamate, hydroxyl radicals, and PGE2. LPS-induced fever could also be attenuated by intravenous or intracerebroventricular TMP 1 h after LPS injection. CONCLUSION: TMP preconditioning may cause its antipyretic action by reducing plasma levels of TNF-α as well as hypothalamic levels of glutamate, hydroxyl radicals, and PGE2 in rabbits.

Oxidative stress and pyrogenic fever pathogenesis.

The causative/regulatory connections between changes in tissue redox state and fever induction were investigated herein. Wherefore, LPS, the primary element of bacterial cell wall, in addition to inducing pro-inflammatory cytokines, activated macrophages and other leukocytes to secrete hydroxyl radical (OH), nitric oxide metabolites (NO(x)(-)), superoxide (O(2)) and other reactive oxygen/nitrogen species. Furthermore, inflammation response-associated hypoxia stimulated glutamate release, which caused excitotoxicity of cells by increasing extracellular Ca(2+). Cytokines and glutamate in turn also triggered the release of large amounts of NO(x)(-), OH, O(2), and other radicals. Those reactive nitrogen species in turn caused cellular injury via the peroxidation of membrane lipids and oxidative damage of proteins and DNA. Glutamate, NO(x)(-), OH and antioxidants participated in the pathogenesis and regulation of LPS- or cytokines-induced fever. In particular, to highlight the role of glutamate, prostaglandin E(2), NO(x)(-) and OH generated in the hypothalamus during pyrogenic fever was attempted hereby. To find the link among the signaling with the glutamate, NO(x)(-) and OH/prostaglandin E(2) in the hypothalamus during pyrogenic fever will be challenging and could now clinically suppress pyrogenic fever.

Central interleukin-10 attenuated lipopolysaccharide-induced changes in core temperature and hypothalamic glutamate, hydroxyl radicals and prostaglandin-E(2).

It has been documented that intravenous lipopolysaccharide (LPS) in rabbits causes fever accompanied by increased levels of extracellular glutamate, hydroxyl radicals, and prostaglandin E(2) (PGE(2)) in the hypothalamus and circulating tumor necrosis factor-alpha (TNF-α). This investigation was to determine whether central interleukin-10 (IL-10) exerted its antipyresis by reducing changes in circulating TNF-α and extracellular glutamate, hydroxyl radicals and PGE(2) in the hypothalamus. The microdialysis probes were stereotaxically and chronically implanted into the preoptic anterior hypothalamus of rabbit brain for determinating extracellular glutamate, hydroxyl radicals, and PGE(2) in situ. It was found that systemically injected LPS (2µg/kg, intravenously) increased the levels of core temperature, and extracellular glutamate, hydroxyl radicals, and PGE(2) in the hypothalamus accompanied by increased plasma levels of TNF-α. Pretreatment with IL-10 (10-100ng, intracerebroventricularly) 1h before intravenous LPS significantly reduced the LPS-induced changes in extracellular glutamate, hydroxyl radicals, and PGE(2) in the hypothalamus and fever, but not the increased levels of TNF-α in rabbits. These findings suggested that directly injected IL-10 into the lateral cerebral ventricle 1h before intravenous LPS exerted its antipyresis by inhibiting the changes in extracellular glutamate, hydroxyl radicals and PGE(2) in the hypothalamus during LPS fever in rabbits.

Interleukin-1 receptor antagonist inhibits the release of glutamate, hydroxyl radicals, and prostaglandin E(2) in the hypothalamus during pyrogen-induced fever in rabbits.

The present study was attempted to determine whether interleukin-1 receptor antagonist (IL-1ra) pretreatment exerts its antipyresis by reducing organum vasculosum laminae terminalis (OVLT) release of glutamate, hydroxyl radicals and prostaglandin E(2) in rabbits. It was found that systemic administration of lipopolysaccharide induced increased levels of both core temperature and OVLT levels of glutamate, hydroxyl radicals, and prostaglandin E(2). The rise in both the core temperature and OVLT glutamate, hydroxyl radicals and prostaglandin E(2) could also be induced by intracerebroventricular injection of interleukin-1beta. Pretreatment with an intracerebroventricular dose of IL-1ra significantly prevented the lipopolysaccharide or IL-1beta-induced overproduction of glutamate, hydroxyl radicals, and prostaglandin E(2) in OVLT of rabbit's brain. The febrile response caused by systemic administration of lipopolysaccharide or central injection of interleukin-1beta could also be IL-1ra pretreatment-ameliorated. These results indicate that IL-1ra pretreatment may exert its antipyresis by inhibiting the glutamate-hydroxyl radicals-prostaglandin E(2) pathways in the OVLT of rabbit's brain during lipopolysaccharide fever.

Hyperbaric oxygen causes both antiinflammation and antipyresis in rabbits.

Current study was attempted to assess whether hyperbaric oxygen pretreatment or treatment exerts its antipyresis by reducing circulating interleukin-6 and hypothalamic glutamate, hydroxyl radicals and prostaglandin-E(2) in rabbits. It was found that systemic administration of lipopolysaccharide induced increased levels of both core temperature and hypothalamic levels of glutamate, hydroxyl radicals, and prostaglandin E(2) accompanied by increased plasma levels of interleukin-6. The rise in both the core temperature and hypothalamic glutamate, hydroxyl radicals and prostaglandin-E(2) could also be induced by intracerebroventricular injection of interleukin-6. Pretreatment or treatment with hyperbaric oxygen significantly reduced the lipopolysaccharide-induced overproduction of circulating interleukin-6, and hypothalamic glutamate, prostaglandin E(2), and hydroxyl radicals. The febrile response caused by central administration of interleukin-6 could also be suppressed by hyperbaric oxygen pretreatment or treatment. Simultaneous administration of an anti-oxidant (e.g., N-acetyl-L-cysteine) significantly enhanced the antipyretic effects exerted by hyperbaric oxygen treatment. These results indicate that hyperbaric oxygen pretreatment or treatment may exert its antipyresis by inhibiting the glutamate-hydroxyl radicals-prostaglandin-E(2) pathways in the hypothalamus and circulating interleukin-6 accumulation during lipopolysaccharide-fever.

Curcumin inhibits the increase of glutamate, hydroxyl radicals and PGE2 in the hypothalamus and reduces fever during LPS-induced systemic inflammation in rabbits.

Evidence has accumulated to suggest that systemic administration of lipopolysaccharide (LPS), in addition to elevating tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and interleukin-6 (IL-6) as well as fever, induces overproduction of glutamate, hydroxyl radicals and prostaglandin E(2) (PGE(2)) in the rabbit's hypothalamus. Current study was attempted to assess whether Curcumin exerts its antipyresis by reducing circulating pro-inflammatory cytokines and hypothalamic glutamate, hydroxyl radicals and PGE(2) in rabbits. The microdialysis probes were stereotaxically and chronically implanted into the preoptic anterior hypothalamus of rabbit brain for determination of glutamate, hydroxyl radicals, and PGE(2) in situ. It was found that systemic administration of LPS (2 microg/kg) induced increased levels of both core temperature and hypothalamic levels of both glutamate and hydroxyl radicals accompanied by increased plasma levels of TNF-alpha, IL-1beta, and IL-6. The rise in both the core temperature and hypothalamic glutamate and hydroxyl radicals could also be induced by direct injection of TNF-alpha, IL-1beta, or IL-6 into the lateral ventricle of rabbit brain. Pretreatment with Curcumin (5-40 mg/kg, i.p.) 1 h before an i.v. dose of LPS significantly reduced the LPS-induced overproduction of circulating TNF-alpha, IL-1beta, and IL-6, and brain glutamate, PGE(2), and hydroxyl radicals. Both the febrile response and overproduction of both glutamate and hydroxyl radicals in the hypothalamus caused by central administration of TNF-alpha, IL-1beta, or IL-6 could be suppressed by Curcumin. These results indicate that systemic injection of Curcumin may exert its antipyresis by inhibiting the glutamate-hydroxyl radicals-PGE(2) pathways in the hypothalamus and circulating TNF-alpha, IL-1beta, and IL-6 accumulation during LPS fever.

Lipopolysaccharide- and glutamate-induced hypothalamic hydroxyl radical elevation and fever can be suppressed by N-methyl-D-aspartate-receptor antagonists.

The purpose of the current study was to explore the effects of N-methyl-D-aspartate (NMDA)-receptor antagonists (MK-801 and LY235959) administered intracerebroventricularly on the changes of both core temperature and hypothalamic levels of 2,3-dihydroxybenzoic acid (2,3-DHBA) induced by intracerebroventricular injection of glutamate (100 - 400 microg at 10 microl/rabbit) or intravenous administration of lipopolysaccharide (LPS) (2 microg/kg) in rabbits. The measurements of 2,3-DHBA were used as an index of the intrahypothalamic levels of hydroxyl radicals. The rise in both the core temperature and hypothalamic 2,3-DHBA could be induced by intracerebroventricular injection of glutamate or intravenous administration of LPS. The glutamate- or LPS-induced fever and increased hypothalamic levels of 2,3-DHBA were significantly antagonized by pretreatment with injection of MK-801 or LY235959 1 h before glutamate or LPS injection. The increased levels of prostaglandin E2 in the hypothalamus induced by glutamate or LPS could be suppressed by MK-801 or LY235959. The data demonstrate that prior antagonism of NMDA receptors in the brain, in addition to reducing prostaglandin E2 production in the hypothalamus, suppresses both the glutamate- and LPS-induced fever and increased hypothalamic hydroxyl radicals.

Aspirin may exert its antipyresis by inhibiting the N-methyl-D-aspartate receptor-dependent hydroxyl radical pathways in the hypothalamus.

Recent findings have suggested that the N-methyl-D-aspartate (NMDA) receptor-dependent hydroxyl radical pathway in the hypothalamus of rabbit brain may mediate the fever induced by lipopolysaccharide (LPS). The aim of this study was to investigate whether aspirin exerts its antipyresis by suppressing hypothalamic glutamate and hydroxyl radicals in rabbits. The microdialysis probes were stereotaxically and chronically implanted into the preoptic anterior hypothalamus of rabbit brain for determination of both glutamate and hydroxyl radicals in situ. It was found that intravenous (i.v.) injection of LPS, in addition to inducing fever, caused increased levels of both glutamate and hydroxyl radicals in the hypothalamus. Pretreatment with aspirin (10 - 60 mg/kg, i.v.) one hour before an i.v. dose of LPS significantly reduced the febrile response and attenuated the LPS-induced increased levels of both glutamate and hydroxyl radicals in the hypothalamus. The increased levels of prostaglandin E(2) (PGE(2)) in the hypothalamus induced by LPS could be suppressed by aspirin pretreatment. The data indicate that systemic administration of aspirin, in addition to suppressing PGE(2) production, may exert its antipyresis by inhibiting the NMDA receptor-dependent hydroxyl radical pathways in the hypothalamus during LPS fever.

The flavonoid baicalin protects against cerebrovascular dysfunction and brain inflammation in experimental heatstroke.

The present study was performed to assess the prophylactic effect of baicalin, a flavonoid compound, in an animal model of heatstroke. Anesthetized rats, immediately before the start of heat stress, were divided into two major groups and given the following: vehicle solution (1mL per kg body weight) or baicalin (10-40mg per kg body weight) intravenously. They were exposed to ambient temperature of 43 degrees C to induce heatstroke. Another group of rats was exposed to room temperature (24 degrees C) and used as normothermic controls. Their physiologic and biochemical parameters were continuously monitored. When the vehicle-pretreated rats underwent heat stress, their survival time values were found to be 20-28min. Pretreatment with intravenous doses of baicalin significantly improved survival during heatstroke (65-248min). As compared to those of normothermic controls, all vehicle-pretreated heatstroke animals displayed higher levels of core temperature, intracranial pressure, and nitric oxide metabolite (NO(2)(-)), glutamate, glycerol, lactate/pyruvate ratio, and dihydroxybenzoic acid (DHBA) in hypothalamus. In addition, both serum and hypothalamic levels of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) as well as plasma levels of creatinine, serum urea nitrogen, glutamic oxaloacetic transaminase, glutamic pyruvic transaminase and alkaline phosphatase were elevated after heatstroke onset. In contrast, all vehicle-pretreated heatstroke animals had lower levels of mean arterial pressure, cerebral perfusion pressure, cerebral blood flow, and brain PO(2). Administration of baicalin before the start of heat exposure significantly reduced the hyperthermia, intracranial hypertension, and the increased levels of NO(2)(-), glutamate, glycerol, lactate/pyruvate ratio, and DHBA in the hypothalamus that occurred during heatstroke. The heatstroke-induced increased levels of IL-1beta and TNF-alpha in both the serum and hypothalamus, and renal and hepatic dysfunction were suppressed by baicalin pretreatment. In contrast, both the serum and hypothalamic levels of IL-10 were significantly elevated by baicalin during heatstroke. We successfully demonstrated that baicalin can be used as a prophylactic agent for heatstroke. In particular, baicalin may protect against cerebrovascular dysfunction and brain inflammation in heatstroke.

The antipyretic effects of baicalin in lipopolysaccharide-evoked fever in rabbits.

Evidence has accumulated to indicate that systemic administration of lipopolysaccharide (LPS), in addition to elevating tumor necrosis factor-alpha (TNF-alpha) as well as fever, induces overproduction of both glutamate and hydroxyl radicals in the rabbit's hypothalamus. Current investigation was attempted to determine whether baicalin exerts its antipyresis by suppressing overproduction of circulating TNF-alpha and hypothalamic glutamate and hydroxyl radicals in rabbits. The microdialysis probes were stereotaxically and chronically implanted into the preoptic anterior hypothalamus of rabbit brain for determination of both glutamate and hydroxyl radicals in situ. It was found that systemic administration of LPS (0.5-10 microg/kg) induced dose-related increased levels of both core temperature and hypothalamic levels of both glutamate and hydroxyl radicals accompanied by increased plasma levels of TNF-alpha. The rise in both the core temperature and hypothalamic glutamate and hydroxyl radicals could also be induced by direct injection of TNF-alpha (1-20 ng) into the lateral ventricle of rabbit brain. Pretreatment with baicalin (2-20 mg/kg, i.v.) one hour before an i.v. dose of LPS significantly reduced the LPS-induced overproduction of circulating TNF-alpha and brain glutamate and hydroxyl radicals. Both the febrile response and overproduction of both glutamate and hydroxyl radicals in the hypothalamus caused by central administration of TNF-alpha could be suppressed by baicalin. These findings suggest that systemic administration of baicalin may exert its antipyresis by inhibiting the N-methyl-D-aspartate receptor-dependent hydroxyl radicals pathways in the hypothalamus and circulating TNF-alpha accumulation during LPS-fever.

An NMDA receptor-dependent hydroxyl radical pathway in the rabbit hypothalamus may mediate lipopolysaccharide fever.

The aim of this study was to investigate the effects of antioxidants (e.g. alpha-lipoic acid and N-acetyl-L-cysteine) as well as N-methyl-D-aspartate (NMDA) receptor antagonists (e.g. MK-801 and LY235959) on the changes of both core temperature and hypothalamic levels of 2,3-dihydroxybenzoic acid (2,3-DHBA) induced by systemic administration of lipopolysaccharide (LPS) in rabbits. The measurements of 2,3-DHBA were used as an index of the intrahypothalamic levels of hydroxyl radicals. Intravenous administration of LPS (2-10 microg/kg) elicited a biphasic febrile response, with the core temperature maxima at 80 and 200 min post-injection. Each core temperature rise was accompanied by a distinct wave of cellular concentrations of 2,3-DHBA in the hypothalamus. The rise in both the core temperature and hypothalamic 2,3-DHBA could be induced by direct injection of glutamate (100-400 microg in 10 microl/rabbit) into the cerebroventricular fluid system. Either the early or the late phase of fever rise and increased hypothalamic levels of 2,3-DHBA following systemic injection of LPS were significantly antagonized by pretreatment with injection of alpha-lipoic acid (5-60 mg/kg, i.v.), N-acetyl-L-cysteine (2-20 mg/kg, i.v.), MK-801 (0.1-1 mg/kg, i.m.), or LY235959 (0.1-1 mg/kg, i.v.) 1 h before LPS injection. The increased levels of prostaglandin E(2) in the hypothalamus induced by LPS could be suppressed by alpha-lipoic acid or N-acetyl-L-cysteine pretreatment. These findings suggest that an NMDA receptor-dependent hydroxyl radical pathway in the hypothalamus of rabbit brain may mediate both the early and late phases of the fever induced by LPS.

Inhibition of nuclear factor-kappa B prevents staphylococcal enterotoxin A-induced fever.

It has been shown that staphylococcal enterotoxin A (SEA) acts through human peripheral blood mononuclear cells (PBMC) to stimulate synthesis or release of pyrogenic cytokines. Nuclear factor-kappa B (NF-kappaB) is thought to play an important role in inflammatory responses through the regulation of genes encoding pro-inflammatory cytokines. The purpose of the present study was to determine whether the NF-kappaB mechanisms in human PBMC are involved in SEA-induced fever. Western blot evaluation revealed SEA was able to induce nuclear translocation of NF-kappaB from cytosol to nucleus in PBMC, which could be abolished by a NF-kappaB inhibitor such as pyrrolidine dithiocarbamate (PDTC), sodium pyrithione (Pyri), N-acetyl-L-cysteine (NAC), or curcumin (Cur). Electrophoretic mobility shift assay also showed that the NF-kappaB DNA-binding activity was increased in the SEA-treated PBMC. Again, the SEA-induced increased NF-kappaB binding activity was significantly attenuated by either PDTC, Pyri, NAC or Cur. The pyrogenic responses to supernatant fluids obtained from human PBMC stimulated with SEA were associated with increased levels of interleukin 1-beta (IL-1beta), IL-6, and tumor necrosis factor-alpha (TNF-alpha) in the supernatant fluids. Both the fever and the increased levels of IL-1beta, IL-6, and TNF-alpha in supernatant fluids obtained from the SEA-stimulated PBMC were decreased by incubating SEA-PBMC with either PDTC, Pyri, NAC, or Cur. Furthermore, the fever induced by systemic or central administration of SEA in rabbits were attenuated by pre-treatment with an systemic or central dose of either PDTC, Pyri, NAC, or Cur. The data indicate that inhibition of NF-kappaB prevents SEA-induced fever.

Cyclooxygenase inhibitors attenuate augmented glutamate release in organum vasculosum laminae terminalis and fever induced by staphylococcal enterotoxin A.

Both the hyperthermia and augmented glutamate release in the organum vasculosum laminae terminalis (OVLT) after an intravenous dose (30 ng/kg) of staphylococcal enterotoxin A (SEA) were significantly reduced by pretreatment with intravenous administration of cyclooxygenase inhibitors such as aspirin (1 - 10 mg/kg), sodium salicylate (1 - 10 mg/kg), or diclofenac (10 mg/kg). Intra-OVLT administration of 50 - 200 microg in 1.0 microl of either aspirin or sodium salicylate 60 min before or 120 min after an intra-OVLT dose (50 microg in 1.0 microl) of glutamate also significantly suppressed the glutamate-induced hyperthermia. These findings suggest that inhibition of cyclooxygenase receptor mechanisms suppresses SEA fever by inhibition of glutamate release in the OVLT of rabbit brain.

Antipyretic effect of acetaminophen by inhibition of glutamate release after staphylococcal enterotoxin A fever in rabbits.

The present study was designed to determine whether the inhibition of glutamate release in organum vasculosum laminae terminalis (OVLT) of rabbit brain by acetaminophen might be protective in a whole-animal model of staphylococcal enterotoxin A (SEA) fever. Unanesthetized rabbits were administered intravenously with SEA, and both colonic temperature (Tco) and glutamate release in OVLT were measured simultaneously. The glutamate release in OVLT was measured with a microdialysis probe previously implanted. Both the Tco and glutamate release in OVLT were simultaneously increased following intravenous administration of SEA. The SEA-induced rise in both the Tco and the levels of glutamate release in OVLT were suppressed by pretreatment with intravenous injection of acetaminophen (1, 5 or 10 mg/kg). Furthermore, treatment of OVLT with acetaminophen (50-150 microg) attenuated the fever-like hyperthermia induced by intra-OVLT injection of glutamate. Our results show acetaminophen may reduce glutamate release in OVLT and result in antipyresis.

Platonin, a cyanine photosensitizing dye, inhibits pyrogen release and results in antipyresis.

Intravenous injection of the supernatant fluids from human peripheral blood mononuclear cells (PBMC) incubated with lipopolysaccharide (LPS) caused fever in rabbits. The fever was in parallel with the levels of either interleukin-1 beta (IL-1 beta), IL-6, or tumor necrosis factor-alpha (TNF-alpha) in supernatant fluids. When incubating the platonin with the LPS-human PBMC, both the levels of IL-1 beta, IL-6, or TNF-alpha in supernatant fluids and the pyrogenicity of supernatant fluids were significantly suppressed. The febrile response to supernatant fluids from the LPS-stimulated PBMC was attenuated almost completely by adding anti-IL-1 beta, but not anti-IL-6 or anti-TNF-alpha, monoclonal antibody to supernatant fluids. In addition, both the fever and the increased levels of either IL-1 beta, IL-6, or TNF-alpha in rabbit serum following an intravenous administration of LPS were significantly attenuated by pretreatment with an intravenous dose of platonin. Furthermore, the fever induced by intravenous injection of IL-1 beta was reduced by pretreatment of rabbits with intravenous injection of platonin. The data indicate that platonin inhibits production of pyrogenic cytokines (in particular, IL-1 beta) from PBMC and results in antipyresis.

Pyrogens enhance beta-endorphin release in hypothalamus and trigger fever that can be attenuated by buprenorphine.

At first, we investigated whether both beta-endorphin release level in the hypothalamus and body temperature can be altered after intracerebroventricular (i.c.v.) injection of either lipopolysaccharide (LPS), interleukin-1beta (IL-1beta), or prostaglandin E(2) (PGE(2)) in rats. It was found that in the rat, i.c.v. administration of either LPS (0.5 microg in 10 microl), IL-1beta (10 ng in 10 microl), or PGE(2) (200 ng in 10 microl), in addition to producing fever, upregulated the immunoreactivity of beta-endorphin in the preoptic anterior hypothalamus of rat brain. Secondarily, we assessed whether the fever induced by either LPS, IL-1beta, or PGE(2) can be altered by pretreatment with buprenorphine (an opioid receptor antagonist). The results revealed that i.c.v. administration of buprenorphine (1 - 10 microg in 10 microl) alone had an insignificant effect on the body temperature. However, the fever induced by i.c.v. injection of either LPS, IL-1beta, or PGE(2) was significantly attenuated by pretreatment with i.c.v. injection of buprenorphine 1 h before the pyrogen injection in rats. The results suggest that pyrogens enhance beta-endorphin release in the hypothalamus and trigger fever which can be attenuated by buprenorphine, an opioid receptor antagonist.

Blocking NF-kappaB activation may be an effective strategy in the fever therapy.

Lipopolysaccharide (LPS) stimulates peripheral mononuclear cells (PBMC) to synthesize or release pyrogenic cytokines, including interleukin-1beta (IL-1beta), IL-6, and tumor necrosis factor-alpha (TNF-alpha). Nuclear factor-kappa B (NF-kappaB) influences inflammatory responses through the regulation of genes encoding cytokines. In the present study, experiments were carried out to determine whether an inhibition of NF-kappaB mechanisms causes an inhibition of pyrogenic cytokine synthesis or release from PBMC and results in antipyresis. Intravenous administration of the supernatant fluids obtained from the human PBMC incubated with LPS caused feverlike hyperthermia in rabbits. The febrile responses were in parallel with the levels of IL-1beta, IL-6, and TNF-alpha in supernatant fluids. Both the fever and the increased levels of these cytokines in supernatant fluids were decreased by incubating LPS-PBMC with NF-kappaB inhibitors, including pyrrolidine dithiocarbamate, sodium pyrithione, N-acetyl-cysteine, and curcumin. Moreover, an intravenous administration of LPS (0.5-2 microg/kg) produced dose-dependent fever in the rabbits. The fevers were in parallel with the levels of IL-1beta, IL-6, and TNF-alpha in rabbit serum. A pretreatment of rabbits with an intravenous injection of pyrrolidine dithiocarbamate, sodium pryithione, N-acetyl-cysteine, or curcumin 1 h before the intravenous administration of LPS significantly attenuated the LPS-induced fever and/or increased levels of these cytokines in the serum of rabbits. Furthermore, pretreatment with an intravenous dose of anti-IL-1beta, anti-IL-6, or anti-TNF-alpha monoclonal antibody significantly attenuated the fever induced by the intravenous injection of LPS in rabbits. The antipyretic effects exerted by anti-L-1beta monoclonal antibody were greater than those exerted by anti-L-6 or anti-NF-alpha monoclonal antibody. The data indicate that NF-kappaB activation correlates with an LPS-induced synthesis or a release of cytokines (in particular, IL-1beta) from PBMC and triggers fever. Blocking NF-kappaB mechanisms in the PBMC with NF-kappaB inhibitors may be an effective strategy in the fever therapy.