TNF receptor I mediates chemokine production and neutrophil accumulation in the lung following systemic lipopolysaccharide.

BACKGROUND: Tumor necrosis factor (TNF)-alpha is a critical effector of lipopolysaccharide (LPS)-induced acute lung injury, and its effects are mediated by two structurally related receptors, RI and RII. Cellular adhesion molecules and C-X-C chemokines (Keratinocyte chemoattractant (KC) and macrophage inflammatory protein [MIP]-2) regulate tissue neutrophil polymorphonuclear neutrophil (PMN) accumulation in a multitude of inflammatory states. We hypothesized that TNFRI signaling dictates PMN accumulation in the lung via regulation of chemokine molecule production. Therefore, the purposes of this study were to (1) delineate LPS-induced lung TNF-alpha production and (2) characterize the contribution of both TNF receptors to lung chemokine production and neutrophil influx following systemic LPS. METHODS: Wild-type or TNFRI and TNFRII knockout (KO) mice were injected with vehicle (saline) or LPS (Escherichia coli 0.5 mg/kg intraperitoneally). After 2, 4, 6, or 24 h, lungs were analyzed for TNF-alpha and chemokine (KC and MIP-2) protein expression (enzyme-linked immunosorbent assay) and PMN accumulation (myeloperoxidase assay). RESULTS: There was an increase in total lung TNF-alpha (vehicle, 5.0 +/- 1.2 pg/mg total protein vs LPS, 950 +/- 318; P < 0.05) after LPS. Lung chemokine production and PMN accumulation were also increased compared to vehicle-injected mice. Lung chemokine production and PMN accumulation were significantly lower in TNFRI KO, but not TNFRII KO, mice, despite no difference in TNF-alpha production (TNFRI KO, 925 +/- 301 vs TNFRII KO, 837 +/- 267, P = 0.82). CONCLUSIONS: Acute lung injury following systemic LPS administration is characterized by increased lung (1) TNF-alpha production, (2) C-X-C chemokine production, and (3) neutrophil accumulation. The maximal effect of LPS-induced lung neutrophil accumulation appears to be dependent upon the TNFRI receptor but not the TNFRII receptor. .

Suppression of tumor necrosis factor alpha production by cAMP in human monocytes: dissociation with mRNA level and independent of interleukin-10.

BACKGROUND: Elevation of cellular cAMP inhibits lipopolysaccharide(LPS)-stimulated tumor necrosis factor alpha (TNF-alpha) production and increases the expression of interleukin (IL)-10 in mononuclear cells. TNF-alpha gene expression obligates activation of the transcription factor nuclear factor kappaB (NF-kappaB). Exogenous IL-10 inhibits NF-kappaB in monocytes and thus attenuates TNF-alpha production. We examined the role of endogenous IL-10 in the regulation of NF-kappaB activation and TNF-alpha production in human monocytes by cAMP. METHODS: Human monocytes were stimulated with Escherichia coli LPS (100 ng/ml) with and without forskolin (FSK, 50 microM) or dibutyryl cyclic AMP (dbcAMP, 100 microM). Cytokine (TNF-alpha and IL-10) release was measured by immunoassay. TNF-alpha mRNA was measured by reverse transcription polymerase chain reaction, and NF-kappaB DNA binding activity was assessed by gel mobility shift assay. RESULTS: cAMP-elevating agents inhibited LPS-stimulated TNF-alpha release (0.77 +/- 0.13 ng/10(6) cells in LPS + dbcAMP and 0.68 +/- 0.19 ng/10(6) cells in LPS + FSK, both P < 0.05 vs 1.61 +/- 0.34 ng/10(6) cells in LPS alone). Conversely, cAMP enhanced LPS-stimulated IL-10 release (100 +/- 21.5 pg/10(6) cells in LPS + dbcAMP and 110 +/- 25.2 pg/10(6) cells in LPS + FSK, both P < 0.05 vs 53.3 +/- 12.8 pg/10(6) cells in LPS alone). Neither TNF-alpha mRNA expression nor NF-kappaB activation stimulated by LPS was inhibited by the cAMP-elevating agents. Neutralization of IL-10 with a specific antibody did not attenuate the effect of cAMP-elevating agents on TNF-alpha production. CONCLUSION: The results indicate that cAMP inhibits LPS-stimulated TNF-alpha production through a posttranscriptional mechanism that is independent of endogenous IL-10.

L-arginine attenuates lipopolysaccharide-induced lung chemokine production.

Chemokines stimulate the influx of leukocytes into tissues. Their production is regulated by nuclear factor-kappaB (NF-kappaB), an inducible transcription factor under the control of inhibitory factor kappaB-alpha (IkappaB-alpha). We have previously demonstrated that L-arginine (L-Arg) attenuates neutrophil accumulation and pulmonary vascular injury after administration of lipopolysaccharide (LPS). We hypothesized that L-Arg would attenuate the production of lung chemokines by stabilizing IkappaB-alpha and preventing NF-kappaB DNA binding. We examined the effect of L-Arg on chemokine production, IkappaB-alpha degradation, and NF-kappaB DNA binding in the lung after systemic LPS. To block nitric oxide (NO) production, a NO synthase inhibitor was given before L-Arg. LPS induced the production of chemokine protein and mRNA. L-Arg attenuated the production of chemokine protein and mRNA, prevented the decrease in IkappaB-alpha levels, and inhibited NF-kappaB DNA binding. NO synthase inhibition abolished the effects of L-Arg on all measured parameters. Our results suggest that L-Arg abrogates chemokine protein and mRNA production in rat lung after LPS. This effect is dependent on NO and is mediated by stabilization of IkappaB-alpha levels and inhibition of NF-kappaB DNA binding.

Laparoscopic approach to adrenal and endocrine pancreatic tumors.

Laparoscopic adrenalectomy quickly has become the procedure of choice for benign adrenal lesions because it results in less pain, shorter hospital stay, comparable safety, and more patient satisfaction overall. The laparoscopic approach requires advanced laparoscopic surgical skills. Surgeons should be familiar with these techniques and the open approaches before attempting this procedure. When first learning the technique, small left-sided lesions are likely the easiest, and a more experienced surgeon should be present for the initial few cases; however, at this point, the laparoscopic approach to pancreatic endocrine tumors does not have a clear benefit, and it should be considered primarily investigational without clearly established benefits.

Neutralization of IL-18 reduces neutrophil tissue accumulation and protects mice against lethal Escherichia coli and Salmonella typhimurium endotoxemia.

In addition to stimulating IFN-gamma synthesis, IL-18 also possesses inflammatory effects by inducing synthesis of the proinflammatory cytokines TNF and IL-1beta and the chemokines IL-8 and macrophage inflammatory protein-1alpha. We hypothesized that neutralization of IL-18 would have a beneficial effect in lethal endotoxemia in mice. IL-1beta converting enzyme (ICE)-deficient mice, lacking the ability to process mature IL-18 and IL-1beta, were completely resistant to lethal endotoxemia induced by LPS derived from either Escherichia coli or Salmonella typhimurium. In contrast, both wild-type and IL-1beta-/- mice were equally susceptible to the lethal effects of LPS, implicating that absence of mature IL-18 or IFN-gamma but not IL-1beta in ICE-/- mice is responsible for this resistance. However, IFN-gamma-deficient mice were not resistant to S. typhimurium LPS, suggesting an IFN-gamma-independent role for IL-18. Anti-IL-18 Abs protected mice against a lethal injection of either LPS. Anti-IL-18 treatment also reduced neutrophil accumulation in liver and lungs. The increased survival was accompanied by decreased levels of IFN-gamma and macrophage inflammatory protein-2 in anti-IL-18-treated animals challenged with E. coli LPS, whereas IFN-gamma and TNF concentrations were decreased in treated mice challenged with S. typhimurium. In conclusion, neutralization of IL-18 during lethal endotoxemia protects mice against lethal effects of LPS. This protection is partly mediated through inhibition of IFN-gamma production, but mechanisms involving decreased neutrophil-mediated tissue damage due to the reduction of either chemokines (E. coli LPS) or TNF (S. typhimurium LPS) synthesis by anti-IL-18 treatment may also be involved.

Interleukin-11 attenuates pulmonary inflammation and vasomotor dysfunction in endotoxin-induced lung injury.

Interleukin (IL)-11, like other members of the gp130 receptor class, possesses anti-inflammatory properties. We hypothesized that IL-11 pretreatment would attenuate endotoxin [lipopolysaccharide (LPS)]-induced lung inflammation and diminish injury to endothelium-dependent and -independent mechanisms of pulmonary vasorelaxation that require cGMP in Sprague-Dawley rats. LPS (20 mg/kg ip) increased lung tumor necrosis factor (TNF)-alpha compared with the saline control (0.7 +/- 0.15 ng/g lung wet wt for control vs. 3.5 +/- 0.09 ng/g lung wet wt for LPS; P < 0.05). IL-11 (200 mg/kg ip) injected 10 min before LPS administration attenuated the LPS-induced lung TNF-alpha levels (1.6 +/- 0.91 ng/g lung wet wt; P < 0.05 vs. LPS). IL-11 also diminished LPS-induced lung neutrophil sequestration as assessed by myeloperoxidase units (2.1 +/- 0.25 U/g lung wet wt for saline and 15.6 +/- 2.02 U/g lung wet wt for LPS vs. 7.07 +/- 1.65 U/g lung wet wt for LPS plus IL-11; P < 0.05). Similarly, TNF-alpha binding protein (175 mg/kg) attenuated LPS-induced myeloperoxidase activity (6.04 +/- 0.14 U/g lung wet wt; P < 0.05). Both IL-11 and TNF-alpha binding protein similarly attenuated LPS-induced endothelium-dependent vasomotor dysfunction with improved relaxation responses to 10(-7) and 10(-6) M acetylcholine and A-23187 in phenylephrine-preconstricted isolated pulmonary artery rings (P < 0.05 vs. LPS). Endothelium-independent relaxation responses to sodium nitroprusside were also improved after LPS at 10(-6) M (P < 0.05 vs. LPS). Moreover, IL-11 decreased endotoxin-induced mortality in CF1 mice from 90 to 50% (P

Inhibition of PARS attenuates endotoxin-induced dysfunction of pulmonary vasorelaxation.

Endotoxin (Etx) causes excessive activation of the nuclear repair enzyme poly(ADP-ribose) synthase (PARS), which depletes cellular energy stores and leads to vascular dysfunction. We hypothesized that PARS inhibition would attenuate injury to mechanisms of pulmonary vasorelaxation in acute lung injury. The purpose of this study was to determine the effect of in vivo PARS inhibition on Etx-induced dysfunction of pulmonary vasorelaxation. Rats received intraperitoneal saline or Etx (Salmonella typhimurium; 20 mg/kg) and one of the PARS inhibitors, 3-aminobenzamide (3-AB; 10 mg/kg) or nicotinamide (Nic; 200 mg/kg), 90 min later. After 6 h, concentration-response curves were determined in isolated pulmonary arterial rings. Etx impaired endothelium-dependent (response to ACh and calcium ionophore) and -independent (sodium nitroprusside) cGMP-mediated vasorelaxation. 3-AB and Nic attenuated Etx-induced impairment of endothelium-dependent and -independent pulmonary vasorelaxation. 3-AB and Nic had no effect on Etx-induced increases in lung myeloperoxidase activity and edema. Lung ATP decreased after Etx but was maintained by 3-AB and Nic. Pulmonary arterial PARS activity increased fivefold after Etx, which 3-AB and Nic prevented. The beneficial effects were not observed with benzoic acid, a structural analog of 3-AB that does not inhibit PARS. Our results suggest that PARS inhibition with 3-AB or Nic improves pulmonary vasorelaxation and preserves lung ATP levels in acute lung injury.

The NFkappaB inhibitory peptide, IkappaBalpha, prevents human vascular smooth muscle proliferation.

BACKGROUND: Vessel injury results in an inflammatory response characterized by the elaboration of cytokines and growth factors, which ultimately influence vascular smooth muscle cell (VSMC) growth and contribute to atherogenesis. Nuclear factor-kappa B (NFkappaB) is a central transcription factor important in mediating stress and inflammatory-induced signals. We hypothesized that strategies aimed at inhibiting NFkappaB would abrogate mitogen-induced human VSMC proliferation. METHODS: Human aortic VSMC were stimulated with basic fibroblast growth factor (FGF) and tumor necrosis factor-alpha (TNF), and proliferation was quantified by a colormetric assay. The influence of NFkappaB on VSMC proliferation was examined by both nonspecific NFkappaB blockade with calpain inhibitor-1 (CI-1) and dexamethasone (Dex) and specific NFkappaB blockade with liposomal delivery of the NFkappaB inhibitory peptide, IkappaBalpha. RESULTS: FGF and TNF induced concentration-dependent VSMC proliferation (p < 0.002). Neither CI-1, Dex, nor liposomal IkappaBalpha influenced proliferation of unstimulated VSMC. However, both FGF- and TNF-stimulated VSMC proliferation was inhibited to the level of control with CI-1, Dex, and liposomal IkappaBalpha (p < 0.001). CONCLUSION: The mitogenic effect of FGF and TNF on human arterial VSMC may be prevented by inhibiting NFkappaB. Furthermore, liposomal delivery of endogenous inhibitory proteins such as IkappaBalpha may represent a novel, therapeutically accessible method for selective transcriptional suppression in the response to vascular injury.

Cardiotrophin-1 attenuates endotoxin-induced acute lung injury.

Cardiotrophin-1 (CT-1) is a recently discovered member of the gp130 cytokine family, which includes IL-6, IL-11, leukemia inhibitory factor, ciliary neurotrophic factor, and oncostatin M. Recent evidence suggests that, like other members of this family, CT-1 may possess anti-inflammatory properties. We hypothesized that in vivo CT-1 administration would attenuate endotoxin (ETX)-induced acute lung injury. We studied the effects of CT-1 (100 microgram/kg ip, 10 min prior to ETX) in a rat model of ETX-induced acute lung injury (Salmonella typhimurium lipopolysaccharide, 20 mg/kg ip). Six hours after ETX, lungs were harvested for determination of neutrophil accumulation (myeloperoxidase, MPO, assay) and lung edema (wet-to-dry weight ratio). Mechanisms of pulmonary vasorelaxation were examined in isolated pulmonary artery rings at 6 h by interrogating endothelium-dependent (response to acetylcholine) and endothelium-independent (response to sodium nitroprusside) relaxation following alpha-adrenergic (phenylephrine)-stimulated preconstriction. CT-1 abrogated the endotoxin-induced lung neutrophil accumulation: 2.3 +/- 0.2 units MPO/g wet lung (gwl) vs 6. 3 +/- 0.3 units MPO/gwl in the ETX group (P < 0.05 vs ETX, P > 0.05 vs control). Similarly, CT-1 prevented ETX-induced lung edema: wet-to-dry-weight ratio, 4.473 +/- 0.039 vs 4.747 +/- 0.039 in the ETX group (P < 0.05 vs ETX, P > 0.05 vs control). Endotoxin caused significant impairment of both endothelium-dependent and -independent pulmonary vasorelaxation, and CT-1 attenuated this injury. Thus, cardiotrophin-1 possesses significant anti-inflammatory properties in a model of endotoxin-induced acute lung injury.

LPS-Induced NF-kappaB activation and TNF-alpha release in human monocytes are protein tyrosine kinase dependent and protein kinase C independent.

BACKGROUND: Tumor necrosis factor alpha (TNF-alpha) is an important mediator of septic shock. Endotoxin (LPS) signal transduction in human monocytes leads to activation of nuclear factor-kappa B (NF-kappaB) and TNF-alpha release. Previous studies have implicated activation of both protein kinase C (PKC) and protein tyrosine kinases (PTK) in LPS-induced NF-kappaB activation and TNF-alpha production. We hypothesized that inhibition of either PKC or PTK would decrease LPS-induced NF-kappaB DNA binding and TNF-alpha release in human monocytes. MATERIALS AND METHODS: Human monocytes were stimulated with PMA (50 ng/ml) alone or LPS (100 ng/ml) with and without a nonspecific serine/threonine protein kinase inhibitor staurosporine (Stauro), a specific pan-PKC inhibitor bisindolylmaleimide (Bis), or an inhibitor of PTK genistein (Gen). TNF-alpha release in culture supernatants was measured by an ELISA. NF-kappaB DNA binding was evaluated by electrophoretic mobility shift assay. RESULTS: LPS increased NF-kappaB DNA binding and TNF-alpha release in human monocytes. Nonspecific protein kinase inhibition inhibited NF-kappaB activation and TNF-alpha release, while specific PKC inhibition with Bis had no effect on LPS-induced NF-kappaB DNA binding or TNF-alpha release. PTK inhibition with Gen attenuated both LPS-induced NF-kappaB DNA binding and TNF-alpha production in human monocytes. Direct activation of PKC with PMA induced both NF-kappaB activation and TNF-alpha production by human monocytes. CONCLUSIONS: These results suggest that LPS-induced NF-kappaB activation and TNF-alpha release in human monocytes are independent of PKC activity. Furthermore, our results provide evidence that PTK plays a role in LPS-induced NF-kappaB activation and TNF-alpha release in human monocytes and thus could be a potential therapeutic target in inflammatory states.

L-arginine attenuates endothelial dysfunction in endotoxin-induced lung injury.

BACKGROUND: Pulmonary vasorelaxation to endothelium-dependent and independent agonists is dysfunctional in endotoxin-induced acute lung injury. L-arginine is the precursor to endothelial production of nitric oxide (NO), suggesting that arginine and NO are intimately linked. We hypothesized that L-arginine would attenuate endotoxin-induced dysfunction of guanosine 3',5'-cyclic monophosphate-mediated pulmonary vasorelaxation. METHODS: Concentration-response curves were generated for acetylcholine, calcium ionophore A23187, and sodium nitroprusside (SNP) in isolated phenylepherine-preconstricted pulmonary artery rings (10(-9) to 10(-6) mol/L) 4 hours after endotoxin (500 mg/kg intraperitoneal) or saline injection. The effect of L-arginine in vitro was determined with L- or D-arginine (50 mmol/L) 30 minutes before dose response. RESULTS: Endothelium-dependent pulmonary vasorelaxation was dysfunctional after endotoxin injection as demonstrated by impaired responses to acetylcholine and A23187 (P < .05 vs control). Endotoxin-induced dysfunction of these endothelium-dependent responses was attenuated by L-arginine (P < .05 vs endotoxin). Endothelium-independent vasorelaxation (SNP) was also dysfunctional after endotoxin treatment (P < .05 vs control). L-arginine failed to attenuate the endotoxin-induced dysfunction of the response to SNP. The concentration responses for endothelium-dependent and independent vasorelaxing agonists in endotoxin-treated rats were not influenced by D-arginine. CONCLUSION: L-arginine supplementation attenuates endotoxin-induced dysfunction of endothelium-dependent pulmonary vasorelaxation.

Endotoxin differentially impairs receptor-mediated relaxation in rat isolated pulmonary and thoracic aortic vessels.

PURPOSE: The purpose of this study was to determine the effect of endotoxin on vasorelaxation in the pulmonary and systemic circulations in response to the following agonists that require generation of cyclic adenosine monophosphate: (1) beta-adrenergic receptor stimulation with isoproterenol; (2) H2 receptor stimulation with dimaprit; and (3) adenylate cyclase stimulation with forskolin. METHODS: Male Sprague-Dawley rats weighing 250 to 350 g were injected with endotoxin (20 mg/kg intraperitoneal) or saline. Six hours later, the cumulative dose response to beta-adrenergic receptor stimulation (isoproterenol), H2 receptor stimulation (dimaprit), and adenylate cyclase stimulation (forskolin) was determined in isolated rat pulmonary artery and thoracic aortic rings preconstricted with phenylephrine. RESULTS: Endotoxin caused significant impairment of relaxation to isoproterenol in the pulmonary artery, but the response in the aorta was not different from the control response. In the pulmonary circulation, endotoxin converted the response to dimaprit from vasorelaxation to vasoconstriction. On the other hand, dimaprit resulted in vasorelaxation in the thoracic aorta after endotoxin; however, the response was impaired compared with the control response. Endotoxin did not affect the dose response to forskolin in either the pulmonary artery or the thoracic aorta. CONCLUSION: From these data, we conclude that endotoxin causes regional specific changes in vascular reactivity. These changes in vascular reactivity result in preserved vasorelaxation in the systemic circulation and impairment of vasorelaxation in the pulmonary circulation in response to endotoxin.

Nitric oxide downregulates lung macrophage inflammatory cytokine production.

BACKGROUND: Inflammatory cytokine production contributes to lung injury after lung ischemia reperfusion and during lung transplant rejection. Although nitric oxide has been demonstrated to reduce lung injury associated with the adult respiratory distress syndrome, it remains unknown whether the mechanism of nitric oxide's beneficial effects involves reducing lung macrophage inflammatory cytokine production. The purpose of this study was to determine whether nitric oxide downregulates lung macrophage inflammatory cytokine production. METHODS: Lung macrophages were harvested by bronchoalveolar lavage (10(6) macrophage per milliliter from normal Sprague-Dawley rats, 6 animals per group) and treated under ex vivo tissue culture conditions with the nitric oxide releasing compound S-nitoso-N-acetyl-D, L-penicillamine (0, 10(-5) 10(-4), 10(-3), 10(-2) mol/L) before induction of inflammatory cytokines with endotoxin, (50 ng/mL for 24 hours). Supernatants were assayed for inflammatory cytokine production (tumor necrosis factor alpha, interleukin-1beta) by enzyme-linked immunosorbent assay. RESULTS: Continuous nitric oxide release by S-nitoso-N-acetyl-D, L-penicillamine decreased lung macrophage tumor necrosis factor-alpha and interleukin-1beta production in a dose-dependent fashion (6 rats per group; data were analyzed for significance [p < 0.05] using two-way analysis of variance with Tukey's post-hoc correction). CONCLUSIONS: Nitric oxide decreases inflammatory cytokine production by lung macrophage. The mechanism of nitric oxide's beneficial effects may be partially attributable to decreased production of inflammatory cytokines. Nitric oxide may serve an expanded role for reducing inflammatory cytokine production during acute lung injury, ischemia-reperfusion-induced inflammation, or lung transplant rejection.

Class II cytokine receptor ligands inhibit human vascular smooth muscle proliferation.

BACKGROUND: Vessel injury provokes the release of proinflammatory cytokines and growth factors that influence vascular smooth muscle cell (VSMC) proliferation and migration. Produced by T lymphocytes, interleukin-10 (IL-10) and interferon-gamma (IFN-gamma) both have immunoregulatory functions and act on similar receptors, designated class II cytokine receptors. We hypothesized that the class II cytokine receptor participates in vascular remodeling by inhibiting VSMC proliferation. The purposes of this study were to determine the influence of class II cytokine receptor stimulation on (1) unstimulated, (2) cytokine-stimulated, and (3) growth factor-stimulated VSMC proliferation. METHODS: Human aortic VSMCs were isolated and cultured. VSMCs were treated with IL-10 or IFN with or without tumor necrosis factor-alpha (TNF-alpha) or basic fibroblast growth factor (FGF). Proliferation was quantified by colormetric assay. RESULTS: Compared to control, both TNF and FGF stimulated concentration-dependent VSMC proliferation (P < .005). IL-10 and IFN alone had no effect on unstimulated cell growth. With TNF or FGF stimulation, both IL-10, at a dose as low as 10 fg/ml, and IFN, at a dose as low as 1.0 U/ml, inhibited cell growth (P < .001). CONCLUSIONS: The class II cytokine receptor ligands, IL-10 and IFN, inhibit cytokine-(TNF) and growth factor-(FGF) induced VSMC proliferation. The class II cytokine receptor may provide a novel therapeutic target in regulating vessel wall remodeling after vascular injury.

Tissue-specific protein kinase C isoforms differentially mediate macrophage TNFalpha and IL-1beta production.

UNLABELLED: Macrophage subpopulations are differentially activated during sepsis, shock, or trauma; however, it is unknown whether inherent mechanistic and phenotypic differences exist between macrophage subpopulations that may account for region-specific inflammation. We hypothesized that macrophage expression/function of protein kinase C (PKC) isoforms is tissue specific (alveolar versus peritoneal). Rat alveolar and peritoneal macrophages were each probed for the expression of PKC isoforms alpha, beta1, beta2, gamma, delta, epsilon, zeta, and theta by immunoblot. PKC isoforms alpha, beta1, beta2, and zeta were detected in both populations; however, isoforms epsilon, gamma, and eta were found in alveolar macrophages only. To investigate the functional role of the Ca2+-dependent PKC (cPKC) versus Ca2+-independent PKC (nPKC) isoforms, pan-PKC isoform inhibition (cPKC and nPKC), or cPKC isoform selective inhibition (alpha, beta1, beta2, gamma) was performed before endotoxin (lipopolysaccharide, Salmonella minnesota, 100 ng/mL) stimulation in vitro. Pan-PKC isoform inhibition attenuated TNFalpha and IL-1beta production by each population; however, selective cPKC (alpha, beta1, beta2, gamma) inhibition decreased peritoneal, but not alveolar, macrophage TNFalpha production. IL-1beta production was not affected by cPKC inhibition in either population. CONCLUSIONS: 1) alveolar and peritoneal macrophages constitutively express different PKC isoforms; 2) alveolar macrophages uniquely express isoforms epsilon, gamma, eta; 3) TNFalpha production is regulated by cPKCs in peritoneal macrophages, but by nPKCs in alveolar macrophages; 4) nPKCs regulate IL-1beta production in both populations. These results suggest that tissue-specific PKC isoforms differentially mediate macrophage function, which may have important regulatory implications in the compartmentalization of immune function. Further understanding may allow region-specific manipulation of inflammation.

Interleukin-10 inhibits human vascular smooth muscle proliferation.

Arterial injury results in the elaboration of pro-inflammatory substances including cytokines and peptide growth factors which act to modify vascular smooth muscle (VSMC) proliferation and migration with resultant vessel stenosis. Produced by T-lymphocytes and macrophages, interleukin-10 (IL-10) is an anti-inflammatory cytokine in several cell lines. We hypothesized that IL-10 may participate in vascular remodeling by inhibiting VSMC proliferation. Human aortic VSMCs were isolated and cultured. Proliferation assays were performed to determine the effect of the effect of IL-10 on (1) unstimulated, (2) cytokine (tumor necrosis factor-alpha: TNF alpha)-stimulated, and (3) growth factor (basic fibroblast growth factor: bFGF)-stimulated VSMC proliferation. Compared to control, both TNF alpha and bFGF-stimulated VSMC proliferation (P < 0.002). IL-10 alone had no effect on cell growth. However, with TNF alpha or bFGF-stimulation, physiologic doses of IL-10 inhibited both VSMC DNA synthesis and VSMC growth (P < 0.001). Furthermore, IL-10 was effective in inhibiting TNF alpha-induced proliferation at a dose as low as 10 fg/ml (P < 0.001) and bFGF-induced proliferation at a dose as low as 1 pg/ml (P < 0.001). In conclusion, TNF alpha and bFGF stimulate human VSMC growth. IL-10 potently abrogates the proliferative response to these atherogenic mitogens. IL-10 might represent an endogenous source of immune-mediated atherprotection and when given exogenously, may prove to be a novel therapeutic agent in regulating vessel wall remodeling following vascular injury.

Interleukin-10 stabilizes inhibitory kappaB-alpha in human monocytes.

Interleukin-10 (IL-10) protects animals from lethal endotoxemia. This beneficial effect is mediated, in part, by inhibition of inflammatory cytokine production, including tumor necrosis factor-alpha (TNF-alpha). Evidence suggests that IL-10 may inhibit activation of the transcription factor nuclear factor-kappaB (NF-kappaB) through an unknown mechanism. NF-kappaB activation in response to inflammatory signals is dependent upon degradation of its associated inhibitory peptide, inhibitory kappaB-alpha (IkappaB-alpha). We hypothesized that IL-10 prevents human monocyte NF-kappaB activation and resultant TNF-alpha production by stabilization of IkappaB-alpha. The purpose of this study was to determine the effect of IL-10 on lipopolysaccharide (LPS)-induced human monocyte TNF-alpha production, NF-kappaB activation, and IkappaB-alpha degradation. Monocytes were isolated from human donors. Cells were stimulated with endotoxin (LPS, 100 ng/mL) with and without human IL-10 (10 ng/mL). Following stimulation, TNF-alpha was measured in cell supernatants by ELISA, NF-kappaB activity by electrophoretic mobility shift assay, and IkappaB-alpha levels by Western blot. We observed that after LPS stimulation of human monocytes, TNF-alpha increased to 798+/-67 pg/mL (p < .001 versus control). IL-10 attenuated LPS-stimulated TNF-alpha production (297+/-54; p < .001 versus LPS alone). After LPS stimulation in human monocytes, IkappaB-alpha protein levels decreased, and NF-kappaB DNA binding increased. IL-10 pretreatment prevented LPS-induced decreases in IkappaB-alpha protein levels and attenuated NF-kappaB DNA binding. IL-10 appears to prevent activation of NF-kappaB by preserving IkappaB-alpha protein levels, leading to a reduction in TNF-alpha release.

Cost-effective management of gynecomastia.

BACKGROUND: Routine endocrine screening of idiopathic gynecomastia has been advocated, but may not be cost effective. We carried out a cost-benefit analysis of this approach. METHODS: A retrospective study (1992 to 1997) of 87 adult males with symptomatic gynecomastia was performed. RESULTS: Thirty-four (39%) patients had extrinsic causes; 53 (61%) were considered idiopathic. Forty-five idiopathic cases underwent endocrine testing: beta human chorionic gonadotropin alone, 16; and beta human chorionic gonadotropin, LH, estradiol, testosterone+/-testicular ultrasound, 29. One (2%) occult Leydig cell testicular tumor was detected. Forty-four patients had normal studies and remain well after local excision. CONCLUSION: Routine endocrine evaluation of idiopathic gynecomastia is rarely productive; such testing is best done selectively.