Jeffrey D. Hasday

Inflammatory markers in intrauterine and fetal blood and cerebrospinal fluid compartments are associated with adverse pulmonary and neurologic outcomes in preterm infants.

Recent evidence strongly implicates the inflammatory response to intrauterine infection in the pathogenesis of neonatal brain and lung injury. We hypothesized that lung and brain injury in preterm infants occurs during a common developmental window of vulnerability as the result of an inflammatory response in different compartments. To determine whether inflammatory markers in these compartments are associated with bronchopulmonary dysplasia (BPD) or cranial ultrasound (CUS) abnormalities in infants <33 wk gestation age (GA) and <1501 g birth weight, we analyzed placental pathology and serum and cerebrospinal fluid (CSF) IL-6, IL-1β, and tumor necrosis factor-α (TNF-α) concentrations in 276 infants. Logistic regressions were performed stratified by GA. Histologic chorioamnionitis was significantly associated with BPD in infants ≤28 wk GA (OR 3.6, p = 0.027). Maternal stage of chorioamnionitis significantly correlated with severity of BPD. Presence of a fetal inflammatory response indicated by fetal vasculitis or elevated cytokines was not associated with the development of BPD. Serum IL-6 ≥17 pg/mL was associated with an abnormal CUS in infants >28 wk GA (OR 3.36, p = 0.023) but not ≤28 wk GA. CSF concentrations of IL-6 ≥6.5 pg/mL and TNF-α ≥3 pg/mL were associated with abnormal CUS in infants ≤28 wk GA (IL-6 OR 3.0; TNF-α OR 3.5; p < 0.05 each case) but not ≥28 wk GA. These data suggest that in infants ≤28 wks GA, BPD may be initiated by inflammatory mediators in amniotic fluid, but brain injury may involve variations in the systemic inflammatory response.

Elevated levels of proinflammatory cytokines in the semen of patients with chronic prostatitis/chronic pelvic pain syndrome ☆

OBJECTIVES Chronic prostatitis/chronic pelvic pain syndrome is a common diagnosis, but the disease is poorly understood. The diagnosis is based only on symptoms, and no measurable parameter can help in defining the presence of the disease, its severity, or its cause. Cytokines are soluble proteins secreted by cells of the immune system that principally regulate inflammatory and immune responses. To provide an objective measure of inflammation in the genital tract, we measured levels of the proinflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) in the semen of men with chronic prostatitis/chronic pelvic pain syndrome and compared these with levels in normal men. METHODS We obtained semen samples from 18 men with chronic prostatitis/chronic pelvic pain syndrome and from 8 normal male volunteers. Cytokine levels were measured in the seminal plasma by two-antibody enzyme-linked immunosorbent assay. RESULTS Men with prostatitis had higher mean levels of IL-1 beta and TNF-alpha in seminal plasma (mean +/-SEM) than normal men: TNF-alpha 98+/-39 versus 17+/-8; IL-1 beta 246+/-63 versus 27+/-10, respectively; P <0.05. There was a strong correlation between the levels of TNF-alpha and IL-1 beta in the semen of men with chronic prostatitis/chronic pelvic pain syndrome. There was no correlation between either TNF-alpha or IL-1 beta levels and the number of leukocytes per high power field in expressed prostatic secretions in patients. CONCLUSIONS Some men with chronic prostatitis/chronic pelvic pain syndrome have elevated levels of TNF-alpha and IL-1 beta in the semen. This suggests that inflammation of the genital tract is a feature of this disease, irrespective of the presence or absence of leukocytes in the expressed prostatic secretions. Seminal cytokine levels may provide an objective measure of disease in these patients and suggest specific therapeutic strategies to treat chronic prostatitis/chronic pelvic pain syndrome in such patients.

Febrile Core Temperature Is Essential for Optimal Host Defense in Bacterial Peritonitis

ABSTRACT Fever, a nonspecific acute-phase response, has been associated with improved survival and shortened disease duration in infections, but the mechanisms of these beneficial responses are poorly understood. We previously reported that increasing core temperature of bacterial endotoxin (LPS)-challenged mice to the normal febrile range modified expression of tumor necrosis factor alpha (TNF-α), interleukin 1β (IL-1β), and IL-6, three cytokines critical to mounting an initial defense against microbial pathogens, but survival was not improved in the warmer animals. We speculated that our inability to show a survival benefit of optimized cytokine expression in the warmer animals reflected our use of LPS, a nonreplicating agonist, rather than an infection with viable pathogens. The objective of this study was to determine if increasing murine core temperature altered cytokine expression and improved survival in an experimental bacterial peritonitis model. We showed that housing mice at 35.5°C rather than 23°C increased core temperature from 36.5 to 37.5°C to 39.2 to 39.7°C, suppressed plasma TNF-α expression for the initial 48 h, delayed gamma interferon expression, improved survival, and reduced the bacterial load in mice infected with Klebsiella pneumoniae peritonitis. We showed that the reduced bacterial load was not caused by a direct effect on bacterial proliferation and probably reflected enhanced host defense. These data suggest that the increase in core temperature that occurs during bacterial infections is essential for optimal antimicrobial host defense.

Increased Activity of Interleukin-6 but not Tumor Necrosis Factor-α in Lung Lavage of Premature Infants is Associated with the Development of Bronchopulmonaiy Dysplasia

ABSTRACT: Although considerable evidence suggests that bronchopulmonary dysplasia (BPD) is the result of prolonged inflammation and impaired healing of the immature lung, the mediators that regulate inflammation in neonatal lung injury have not been completely elucidated. We examined whether the cytokines IL-6 and tumor necrosis factor-α (TNF) interact to modulate a cascade of cell-cell signaling events involved in inflammation contributing to the development of BPD. To determine the relative activities of these cytokines in neonatal lung injury, lung lavage samples were serially obtained from 1 to 28 d from 11 infants with self-limited respiratory distress syndrome (RDS), 19 infants with evolving BPD, and 10 control infants ventilated for nonpulmonary reasons. On the first day of life, there were no differences in antigenic IL-6 concentrations in lavage fluids among the BPD, RDS, and control groups, but IL-6 activity determined by the 7TD1 proliferation assay was 15-fold and 6.6-fold higher in lung lavage of infants who developed BPD compared with activities in lavage from control and RDS infants, respectively (control, 49.4 ± 17.6; RDS, 117.3 ± 59.6; BPD, 779.5 ± 212.6 ± 103 hybridoma units/L, mean ± SEM, p = 0.02). This suggests that pathways for inactivating or inhibiting IL-6 that may be present in the lungs of RDS and control infants may be deficient in BPD infants. IL-6 activity remained elevated in lavage of BPD infants for the first 2 wk and declined to low levels by d 28. There were no differences among groups on the first day of life for TNF antigen concentration or TNF activity determined by the L929 bioassay. Detectable but low TNF activity was found in BPD samples, with peak activity found in d-14 samples. Differences in complex interactions among these and other cytokines with their receptors and inhibitors may predispose some infants with RDS to develop BPD.

The role of fever in the infected host

Sepsis is a highly lethal clinical syndrome characterized by a systemic inflammatory response to infection. Fever, a non-specific acute-phase response, has been associated with improved survival and shortened disease duration in non-life-threatening infections. However, the influence of fever and the effects of antipyresis in patients with sepsis has not been prospectively studied in humans. This paper reviews the state of our knowledge concerning the biological effects of fever in infected hosts and the influence of fever and antipyretic therapy on survival during sepsis in experimental models and in man.

A High Affinity HSF-1 Binding Site in the 5′-Untranslated Region of the Murine Tumor Necrosis Factor-α Gene Is a Transcriptional Repressor

Tumor necrosis factor-α (TNFα) is a pivotal early mediator of host defenses that is essential for survival in infections. We previously reported that exposing macrophages to febrile range temperatures (FRT) (38.5–40 °C) markedly attenuates TNFα expression by causing abrupt and premature cessation of transcription. We showed that this inhibitory effect of FRT is mediated by an alternatively activated repressor form of heat shock factor 1 (HSF-1) and that a fragment of the TNFα gene comprising a minimal 85-nucleotide (nt) proximal promoter and the 138-nt 5′-untranslated region (UTR) was sufficient for mediating this effect. In the present study we have used an electrophoretic mobility shift assay (EMSA) to identify a high affinity binding site for HSF-1 in the 5′-UTR of the TNFα gene and have used a chromosome immunoprecipitation assay to show that HSF-1 binds to this region of the endogenous TNFα gene. Mutational inactivation of this site blocks the inhibitory effect of overexpressed HSF-1 on activity of the minimal TNFα promoter (−85/+138) in Raw 264.7 murine macrophages, identifying this site as an HSF-1-dependent repressor. However, the same mutation fails to block repression of a full-length (−1080/+138) TNFα promoter construct by HSF-1 overexpression, and HSF-1 binds to upstream sequences in the regions −1080/−845, −533/−196, and −326/−39 nt in EMSA, suggesting that additional HSF-1-dependent repressor elements are present upstream of the minimal −85-nt promoter. Furthermore, although mutation of the HSF-1 binding site in the minimalTNFα promoter construct abrogates HSF-1-mediated repression, the same mutation fails to abrogate repression of this construct by high levels of HSF-1 overexpression or exposure to 39.5 °C. This suggests that HSF-1 might repress TNFα transcription through redundant mechanisms, some of which might not require high affinity binding of HSF-1.

Tobacco Smoking Inhibits Expression of Proinflammatory Cytokines and Activation of IL-1R-Associated Kinase, p38, and NF-κB in Alveolar Macrophages Stimulated with TLR2 and TLR4 Agonists

Tobacco smoking has been associated with impaired pulmonary functions and increased incidence of infections; however, mechanisms that underlie these phenomena are poorly understood. In this study, we examined whether smokers’ alveolar macrophages (AM) exhibit impaired sensing of bacterial components via TLR2 and TLR4 and determined the effect of smoking on expression levels of TLR2, TLR4 and coreceptors, and activation of signaling intermediates. Smokers’ AMs exhibited reduced gene expression and secretion of proinflammatory cytokines (TNF-α, IL-1β, IL-6) and chemokines (RANTES and IL-8) upon stimulation with TLR2 and TLR4 agonists, S-[2,3-bis(palmitoyloxy)-(2-RS)-propyl]-N-palmitoyl-(R)-Cys-(S)-Ser-Lys4-OH trihydrochloride (Pam3Cys), and LPS, whereas expression of anti-inflammatory cytokines (IL-10 and IL-1 receptor antagonist) was not affected. TLR3 activation with polyinosinic-polycytidylic acid led to comparable or even higher cytokine responses in smokers’ AMs, indicating that smoking-induced suppression does not affect all TLRs. Comparable expression of cytokines and chemokines was detected in PBMC and purified monocytes obtained from smokers and nonsmokers, demonstrating that the suppressive effect of smoking is restricted to the lung. TLR2/4-inducible IL-1R-associated kinase-1 (IRAK-1) and p38 phosphorylation and NF-κB activation was suppressed in smokers’ AMs, whereas TLR2, TLR4, CD14, MD-2 mRNA levels, and TLR4 protein expression were not altered. These data suggest that changes in expression and/or activities of signaling intermediates at the postreceptor level account for smoking-induced immunosuppression. Thus, exposure of AMs to tobacco smoke induces a hyporesponsive state similar to endotoxin tolerance as manifested by inhibited TLR2/4-induced expression of proinflammatory cytokines, chemokines, and impaired activation of IRAK-1, p38, and NF-κB, resulting in suppressed expression of proinflammatory mediators.

Fever and the heat shock response: distinct, partially overlapping processes.

Abstract The heat shock response is an ancient and highly conserved process that is essential for surviving environmental stresses, including extremes of temperature. Fever is a more recently evolved response, during which organisms temporarily subject themselves to thermal stress in the face of infections. We review studies showing that fever is beneficial in the infected host. We show that core temperatures achieved during fever can activate the heat shock response and discuss some of the biochemical consequences of such an effect. We present data suggesting 4 possible mechanisms by which fever might confer protection: (1) directly killing or inhibiting growth of pathogens; (2) inducing cytoprotective heat shock proteins (Hsps) in host cells; (3) inducing expression of pathogen Hsps, an activator of host defenses; and (4) modifying and orchestrating host defenses. Two of these mechanisms directly involve the heat shock response. We describe how heat shock factor-1, the predominant heat-induced transcriptional enhancer not only activates transcription of Hsps but also regulates expression of pivotal cytokines and early response genes. The relationship between fever and the heat shock response is an illuminating example of how a more recently evolved response might exploit preexisting biochemical pathways for a new function.

Inflammatory Cytokine mRNAs in Surgical Specimens of Necrotizing Enterocolitis and Normal Newborn Intestine

Coagulation necrosis, inflammation, and hemorrhage are pathologic hallmarks of necrotizing enterocolitis (NEC). Because cytokines are peptides that mediate inflammatory cell recruitment and amplify the immune response, several of the inflammatory cytokines have been implicated in NEC. We hypothesized that mRNA levels for the interrelated cytokines interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha), IL-6, and the neutrophil chemotactic factor IL-8 would be increased in NEC and would be associated with the presence of inflammation. In this study, we determined the relative levels and localization of mRNA for these cytokines in surgical pathology archival intestinal tissue from 29 premature infants with acute NEC and 15 control infants with congenital intestinal malformations using a novel quantitative in situ hybridization technique. Compared with controls, there were higher IL-1 beta mRNA levels in full-thickness sections and higher TNF-alpha mRNA levels in full-thickness and mucosa sections of acute NEC samples, suggesting a potential role for these cytokines in the pathogenesis of local inflammation in NEC. IL-6 and IL-8 mRNA levels were similar in samples of control and acute NEC cases. Analysis of covariance including all subjects showed that the presence of acute inflammation was associated with increased IL-1 beta mRNA levels in mucosa (P = .035) and increased IL-8 in full-thickness sections (P = .005) and mucosa (P = .01). In four of five NEC cases in which intestinal specimens were available from reanastomosis surgery, cytokine mRNA levels decreased to low or undetectable levels. These data suggest that the inflammatory cytokines are involved in neutrophil recruitment and augmentation of the inflammatory response in neonatal intestine.

Concepts of Fever: Recent Advances and Lingering Dogma

Fever has been a preoccupation of clinicians since medicines beginning. One might therefore expect that basic concepts relating to this physiological response would be well delineated and that such concepts would be widely known. In fact, only in the past several decades has the febrile response been subjected to scientific scrutiny. As a result of recent scientific investigation, modern concepts have evolved from a perception of fever as nothing more than a rise in core temperature to one in which fever is recognized as a complex physiological response characterized by a cytokine-mediated rise in temperature, as well as by generation of acute-phase reactants and activation of a panoply of physiological, endocrinologic, and immunologic systems. The average clinician appears to have little more than a regrettably rudimentary knowledge of these modern concepts of fever. This symposium summary considers many such concepts that have immediate relevance to the practice of medicine.