SYS Feng, VYH Yu, AM Walker

Abstract

Endotoxin induces complex changes in the newborn cerebral circulation, including vasoconstriction by down-regulating endothelial nitric oxide synthase and stimulating the production of endothelin and cyclooxygenase-2 derived vasoconstrictors including thromboxane A₂, while also promoting vasodilatation by increasing the expression of inducible nitric oxide synthase and cyclooxygenase-2 derived vasodilators. Together, these changes represent a complex dysregulation of the newborn cerebral circulation. This review considers the effects of neonatal endotoxaemia on cerebral circulatory changes and endothelium-derived factors, and their implications for cerebral circulatory control.

Introduction

Due to deficiencies in the host defense systems, neonates are particularly vulnerable to bacterial infections.¹ Neonatal infection has been reported to affect 6.6 per 1000 live births in Australia.² It has become an increasing problem in neonatal intensive care units with an overall rate of 21.3 infected infants or 29.9 infections per 100 ventilated admissions since the late 1990s.³ Endotoxaemia, a systemic inflammatory response to endotoxin is one of the major causes of morbidity and mortality in the newborn.

Endotoxin is the lipopolysaccharide (LPS) outer coat of all Gram-negative bacteria.⁴ It consists of three main parts: an outer branched chain polysaccharide portion (the O antigen), a mid portion R antigen polysaccharide core; and an inner toxic lipid A portion which accounts for the majority of the toxicity of endotoxin.⁵ Normally, only small amounts of naturally occurring endotoxin are released systemically, and these play an important role in the development of host immune responses. Excessive amounts of LPS are released when the bacterial cell wall breaks down in conditions such as infection, immune disturbances resulting in the 'overgrowth' of pathogenic bacteria, and when bacteria are killed by bactericidal antibiotics.⁵

Among endotoxin-induced haemodynamic changes, blood pressure instability is a common feature observed in septic neonates in neonatal intensive care units.⁶ A reduced responsiveness to vasoconstrictor stimuli associated with decreased systemic vascular resistance and mean blood pressure had been observed in humans following Gram-negative bacteraemia.^7,8 Furthermore, endotoxin-induced septic shock might not be associated with a positive blood culture.⁹ Endotoxin can cause not only systemic hypotension, but also multiple organ dysfunction in the newborn due to dysregulation of the systemic circulation, such as myocardial, respiratory and renal failure and coagulopathy.

Effect of Endotoxins on the Systemic Endothelium and Circulation

The endothelium plays a critical role in blood pressure and blood flow regulation by releasing vasodilators such as nitric oxide synthase (NOS) and cyclooxygenase-2 (COX-2) derived vasodilator factors, as well as vasoconstrictors, such as endothelin (ET) and COX-2 derived vasoconstrictors including thromboxane A₂ (TXA₂).^10,11 During infection, endotoxin stimulates widespread endothelial damage by interacting with soluble CD14, a surface receptor of mononuclear cells.¹² The LPS-binding protein complex together with CD14 promote mononuclear phagocytes to produce reactive oxygen molecules, cytokines (such as tumour necrosis factor-a and interleukins-1) and prostaglandin metabolites. The complex also induces endothelial cell apoptosis.¹³ Following LPS administration, histological examinations show significant endothelial cell disruption, such as areas of denuded endothelium, cytoplasmic swelling and fragmentation. These anatomical characteristics of endothelial cell dysfunction are in keeping with functional impairment which have been demonstrated in patients with sepsis, and in animal models of endotoxin-induced septic shock.¹⁴ Functional impairments such as endothelium-dependent vasodilator responses to LPS vary in different organs. For example, they are unchanged in rat renal arteries¹⁵ but increased in rat aorta.¹⁶ In the brain, vascular resistance is increased by a combination of vasoconstriction as well as vasodilatation (see below).

Endothelial NO Synthase (eNOS)

eNOS is stimulated via the shear stress in the endothelial cells produced by blood flow and by various receptor agonists.¹⁷ Under normal basal conditions, eNOS synthesises endothelial NO, which plays important roles in endogenous vasodilatation counterbalancing the vasoconstriction produced by the sympathetic nervous system and the renin-angiotensin system,^18,19 and in anti-thrombosis by inhibiting platelet aggregation and adhesion to the vascular wall.²⁰ During sepsis, the constitutive isoforms of NOS including eNOS and neuronal NO synthase (nNOS) are downregulated or even inactivated.^21,22

Inducible NO Synthase (iNOS)

iNOS has the ability to synthesise NO, but it is not present in significant amounts under normal conditions. iNOS is induced by a wide range of factors, including cellular products of endotoxin, Gram-positive bacteria and proinflammatory cytokines²¹ in a variety of cell types including macrophages,²³ vascular smooth muscle cells,²⁴ astrocytes,²⁵ and neurons.²⁶ Induction of iNOS reduces NO-mediated vasodilatation initiated by the other two major NOS, i.e. eNOS and nNOS,²⁷ and NOS inhibitors were able to reverse LPS-induced hypotension in animals,²⁸ so that over-production of NO by iNOS for prolonged period of time appears primarily responsible for endotoxin-induced hypotension.²⁹

Endothelin (ET)

ET is the most potent, long-acting vasoconstrictor secreted from endothelial cells upon mechanical stimuli.³⁰ There are three isopeptides of ET: endothelin-1 (ET-1), endothelin-2 (ET-2), and endothelin-3 (ET-3).³¹ ET-1 is normally produced by endothelium.³⁰ Because of ET-1's affinity for endothelin-A (ET_A) receptors, its vasoconstriction is eight times stronger than ET-2, and 1000 times stronger than ET-3. ET-1 is believed to play a key role in vasoconstriction during sepsis,^32,33 increasing at 4-12 hours and returning to normal at 24-48 hours.³⁴ ET-1 may be elevated as early as 15 minutes after peripheral injection of tumour necrosis factor-a (TNF-a).³⁵ The significantly increased plasma ET-1 levels in sepsis may explain the increased vascular resistance and decreased organ blood flow under such conditions.^36,37 On the other hand, the activation of ET_A receptors by increasing ET-1 concentration also enhances vasodilatation via stimulated iNOS expression and NO production in systemic vessels, contributing to LPS-induced hypotension in both human and animal studies.^38,39 Furthermore, elevated plasma ET-1 is related to the severity of clinical manifestations, especially oliguria, acidosis and systemic hypotension in neonatal sepsis.³⁹

Cyclooxygenase-2 (COX-2) and Prostaglandins

Cyclooxygenase is an enzyme involved in the endothelial production of prostaglandins, which is present in constitutive cyclooxygenase-1 and inducible COX-2 isoforms. COX-2 though present in resting human endothelial cells, it does not appear to regulate basal systemic vascular tone.⁴⁰ Following an inflammatory response induced by LPS, reactive oxygen species, some cytokines, and growth factors, COX-2 expression is up-regulated.^41,42 Functionally, infusion of COX-2 altered regional blood flow.⁴³ The production of prostaglandins increases after COX-2 is expressed.⁴⁴ The main prostaglandins produced during inflammation are the vasodilators PGE₂,⁴⁵ PGI₂ and PGD₂.⁴ These are potent dilators⁴⁶ whereas other prostaglandins, such as PGF_2α and TXA₂ act as vasoconstrictors.⁴⁷ TXA₂ can also cause platelet aggregation.

Apart from systemic effects, LPS can also induce subsequent hypoxic ischaemic damage in the neonatal rat brain.⁴⁸ The exact mechanisms regarding the changes in cerebral blood flow (CBF) and cerebral vascular resistance (CVR) remain unclear.

Effects of Endotoxins on the Cerebral Endothelium and Circulation

Brain infection has the potential to substantially modify the expression of genes that have crucial roles in cerebral vascular regulation. By injecting group B streptococcus (GBS), a Gram-positive bacteria, into the cerebral ventricle of newborn piglets, the upper limit of cerebral vascular autoregulation is impaired within the first 2 hours post-injection. This autoregulatory change correlates well with the induction of NOS gene expression.⁴⁹ Both eNOS and COX-2 were down-regulated post-GBS treatment in the brain of newborn piglets.⁵⁰

Systemic injected LPS does not cross the capillary endothelium of the central nervous system, that is, the blood-brain barrier,⁵¹ but it increases blood-brain barrier permeation by binding to LPS receptors such as toll-like receptors (TLR) TLR-4 and TLR-2, as well as CD14 in the brain-endothelial cells.⁵² This activates a cascade of events involving proinflammatory genes such as IL-1β and iNOS genes, which, in turn, lead to inflammation⁵¹ and microglia activation in the brain.⁵³ It also causes a capillary leak and diffusion of low molecular weight proteins from cerebral vessels into the cerebral spinal fluid, leading to cerebral oedema and increased intracranial pressure.⁵¹

The endotoxin-induced alterations of the cerebral circulation vary in different studies, perhaps reflecting species differences. LPS dilates cerebral arterioles in rabbit,⁵⁴ but vasoconstricts large arteries, such as the middle cerebral artery (MCA) in rats.⁴⁷ CBF responses to LPS are complex. There have been reports of no change in sepsis-induced normotensive rats⁵⁵ and sheep.⁵⁶ In contrast, an increased regional CBF was recorded by laser-Doppler flowmetry without a fall in blood pressure within 4 hours of intracerebro-ventricular injection of LPS (1 mg/kg) in rats anaesthetised by halothane inhalation.⁴² Increased CBF has also been reported in ovine models of sepsis.⁵⁷ On the other hand, by using an autoradiographic technique employing ¹⁴C-iodoantipyrine, Young et al showed endotoxin-induced hypotension with reduced rCBF in periventricular and occipital white matter in newborn dogs.⁵⁸ By applying an electromagnetic probe around the intact superior sagittal sinus or measuring directly from the cannulated confluence of the sagittal, straight and lateral sinuses, a reduction in CBF with increased CVR during the first hour of endotoxin shock were detected in anaesthetised dogs.⁵⁹

At Monash University, we have studied the cerebral haemodynamics and the neutrophil and platelet counts in conscious newborn lambs at 1-4 weeks of age following a single LPS infusion (2 mg/kg). Endotoxaemia in the lambs resulted in a marked reduction of neutrophils and platelet counts by 73% and 19% from baseline respectively soon after LPS infusion (Figure 1). These results in lambs are compatible with the findings in LPS-treated neonatal calves.⁶⁰

CVR = cerebral vascular resistance, CBF = cerebral blood flow, Pca = mean arterial blood pressure

Figure 1 Cerebral haemodynamics, neutrophils and platelet count changes in LPS-treated newborn lambs.

We used a transonic flow probe placed over the sagittal sinus to measure CBF. This method not only allows continuous recording of the blood flow, but also provides a simple quantitative measurement of CBF that is linearly related to arterial inflow.⁶¹ The technique measures blood flow of the entire frontal lobe and the superior portion of the anterior parietal lobe, which represents 35% of the total brain mass of the lamb.⁶¹ We demonstrated that CBF was reduced by 33% and CVR was increased by 129% despite no reduction in blood pressure during the first 2-4 hour post-LPS (Figure 1).

Cerebral Vasoconstriction

The possible mechanisms of endothelial-mediated, LPS-induced cerebral circulatory impairment are an alteration of vasoconstrictor and vasodilator factors:

1. eNOS down-regulation. LPS-induced cerebral endothelium impairment⁶² may lead to disruption of eNOS, impairment of cerebral vasodilatation, and reduction in CBF. Furthermore, induction of iNOS by LPS reduces NO-mediated vasodilatation initiated by eNOS in cerebral arteries⁶³ and enhances the effect on cerebral vasoconstriction.

2. ET-1 upregulation. ET-1 mRNA can be detected in cerebral endothelium.⁶⁴ Its production is increased by TNF-α,⁶⁵ and vascular smooth muscle cells.⁶⁶ It produces potent and long-lasting contraction of cerebral vessels, which is mediated by ETA receptors both in vivo and in vitro.^67,68 ET-1 can also reduce cerebral artery sensitivity to NO by a protein kinase C-independent pathway, and therefore enhances cerebral vasoconstriction.⁶⁹

3. COX-2 induction. Intravenous LPS or proinflammatory cytokines increases expression of COX-2 not only in brain neuron including hippocampus, cerebral cortex, amygdale and hypothalamus,⁷⁰ but also in the cerebral endothelium.⁷¹ Injection of NS-398, a COX-2 inhibitor, can inhibit vascular contraction of the MCA from the first hour in LPS-treated rats, suggesting vasoconstrictors derived from COX-2 are involved in early endotoxin-induced cerebral vasoconstriction.⁴⁷ As these changes occur within one hour of LPS injection, they have the potential to explain the cerebral vasoconstriction that is illustrated in Figure 1.

4. Proinflammatory cytokines. Inhibition of endothelium-dependent relaxation following treatment with proinflammatory cytokines has been shown in peripheral arteries.⁷² Proinflammatory cytokines such as TNF-α was found to be increased in septic neonates.⁷³ It was also found in post-mortem brain tissue from patients suffering from bacterial meningitis.⁷⁴ Moreover, TNF-α has been found to reduce cerebral blood volume by 15-30%, and causes cerebral vasoconstriction via an endothelin- and TNF-α-type-2 receptor dependent pathway.⁷⁵

Cerebral Vasodilatation

1. iNOS. iNOS expression increases after endotoxin exposure in the MCA in rats around 5 hours post-LPS,^42,47 and may be responsible for subsequent reduction in vascular resistance and systemic hypotension. However, in our own observations, CVR increased but hypotension did not develop. Therefore, iNOS induction was not a mitigating factor in the early vasoconstriction responses.

2. COX-2 derived vasodilators. LPS not only induces COX-2 expression causing vasodilatation in systemic circulation,⁴³ but also increases COX-2 expression in brain tissue. LPS application induces PGE₂-dependent dilatation of cerebral arterioles, with the maximum dilatation occurring at 4 hours.⁵⁴ It is possible that LPS induces cyclooxygenase which in turn may also produce reactive oxygen species which are vasodilators in the brain.⁴⁶

Anaesthetic Agents

These should be taken into account when comparing experimental results between anaesthetised and conscious animals. For example, halothane produces a dose-dependent vasodilatation of intraparenchymal cerebral microvessels, which is similar to those of the potent vasodilator sodium nitroprusside, an endothelium dependent NO donor.⁷⁶

Clinical Implications

Our experimental data demonstrating that a reduced CBF and increased cerebral vasoconstriction occur in normotensive lambs post-LPS administration, suggest that impaired cerebral perfusion is possible despite normal blood pressure during the early stages of endotoxaemia in the newborn. Thus, disturbances in cerebral perfusion should be considered in managing the septic newborn, even when the blood pressure remains in the normal range.

Potential treatments including anti-LPS antibody,⁷⁷ NOS inhibitors,^29,78 endothelin-1 (ET-1) antagonists,⁷⁹ and proinflammatory cytokine blockers⁸⁰ are still under experimental study, but they are thought to be an important potential additions to future haemodynamic therapy in septic shock.

In conclusion, endotoxin-induced cerebral circulatory changes in the newborn may be altered by a series of factors following proinflammatory cytokines mediation. By disrupting the balance of vasoconstrictor and vasodilator factors, cerebral vasoconstriction and flow impairment become the net results in the early stage of neonatal endotoxaemia.

References

1. Ford HR RM. Sepsis and related considerations. O'Neil JAJ RM, Grosfeld JL, Fonkalsrud EW, Coran AG, editor. St Louis: Mosby; 1998.

2. Isaacs D, Barfield CP, Grimwood K, McPhee AJ, Minutillo C, Tudehope DI. Systemic bacterial and fungal infections in infants in Australian neonatal units. Australian Study Group for Neonatal Infections. Med J Aust 1995;162:198-201.

3. Mehr SS, Sadowsky JL, Doyle LW, Carr J. Sepsis in neonatal intensive care in the late 1990s. J Paediatr Child Health 2002;38:246-51.

4. Worthley LI. Shock: a review of pathophysiology and management. Part II. Crit Care Resusc 2000;2:66-84.

5. Shenep JL, Barton RP, Mogan KA. Role of antibiotic class in the rate of liberation of endotoxin during therapy for experimental gram-negative bacterial sepsis. J Infect Dis 1985;151:1012-8.

6. Lee SY, Ng DK, Fung GP, et al. Chorioamnionitis with or without funisitis increases the risk of hypotension in very low birthweight infants on the first postnatal day but not later. Arch Dis Child Fetal Neonatal Ed 2006;91:F346-8.

7. Sessler CN, Shepherd W. New concepts in sepsis. Curr Opin Crit Care 2002;8:465-72.

8. Miki S, Takeyama N, Tanaka T, Nakatani T. Immune dysfunction in endotoxicosis: role of nitric oxide produced by inducible nitric oxide synthase. Crit Care Med 2005;33:716-20.

9. Danner RL, Elin RJ, Hosseini JM, Wesley RA, Reilly JM, Parillo JE. Endotoxemia in human septic shock. Chest 1991;99:169-75.

10. Cines DB, Pollak ES, Buck CA, et al. Endothelial cells in physiology and in the pathophysiology of vascular disorders. Blood 1998;91:3527-61.

11. Levin ER. Endothelins as cardiovascular peptides. Am J Nephrol 1996;16:246-51.

12. Landmann R, Zimmerli W, Sansano S, et al. Increased circulating soluble CD14 is associated with high mortality in gram-negative septic shock. J Infect Dis 1995;171:639-44.

13. Bannerman DD, Goldblum SE. Mechanisms of bacterial lipopolysaccharide-induced endothelial apoptosis. Am J Physiol Lung Cell Mol Physiol 2003;284:L899-914.

14. Leclerc J, Pu Q, Corseaux D, et al. A single endotoxin injection in the rabbit causes prolonged blood vessel dysfunction and a procoagulant state. Crit Care Med 2000;28:3672-8.

15. Piepot HA, Boer C, Groeneveld AB, Van Lambalgen AA, Sipkema P. Lipopolysaccharide impairs endothelial nitric oxide synthesis in rat renal arteries. Kidney Int 2000;57:2502-10.

16. Pu Q, Bordet R, Robin E, Puisieux F, Vallet B, Dupuis B. Low dose of lipopolysaccharide induces a delayed enhanced nitric oxide-mediated relaxation in rat aorta. Eur J Pharmacol 1999;377(2-3):209-14.

17. Li H, Forstermann U. Nitric oxide in the pathogenesis of vascular disease. J Pathol 2000;190:244-54.

18. Halbrugge T, Lutsch K, Thyen A, Graefe KH. Vasodilatation by endothelium-derived nitric oxide as a major determinant of noradrenaline release. J Neural Transm Suppl 1991;34:113-9.

19. Klabunde RE, Ritger RC. NG-monomethyl-l-arginine (NMA) restores arterial blood pressure but reduces cardiac output in a canine model of endotoxic shock. Biochem Biophys Res Commun 1991;178:1135-40.

20. Alheid U, Frolich JC, Forstermann U. Endothelium-derived relaxing factor from cultured human endothelial cells inhibits aggregation of human platelets. Thromb Res 1987;47:561-71.

21. Szabo C. Alterations in nitric oxide production in various forms of circulatory shock. New Horiz 1995;3:2-32.

22. Traber DL. Presence and absence of nitric oxide synthase in sepsis. Crit Care Med 1996;24:1102-3.

23. Lorsbach RB, Murphy WJ, Lowenstein CJ, Snyder SH, Russell SW. Expression of the nitric oxide synthase gene in mouse macrophages activated for tumor cell killing. Molecular basis for the synergy between interferon-gamma and lipopolysaccharide. J Biol Chem 1993;268:1908-13.

24. Gonzalez C, Fernandez A, Martin C, Moncada S, Estrada C. Nitric oxide from endothelium and smooth muscle modulates responses to sympathetic nerve stimulation: implications for endotoxin shock. Biochem Biophys Res Commun 1992;186:150-6.

25. Galea E, Reis DJ, Feinstein DL. Cloning and expression of inducible nitric oxide synthase from rat astrocytes. J Neurosci Res 1994;37:406-14.

26. Minc-Golomb D, Tsarfaty I, Schwartz JP. Expression of inducible nitric oxide synthase by neurones following exposure to endotoxin and cytokine. Br J Pharmacol 1994;112:720-2.

27. Buga GM, Griscavage JM, Rogers NE, Ignarro LJ. Negative feedback regulation of endothelial cell function by nitric oxide. Circ Res 1993;73:808-12.

28. Kilbourn RG, Jubran A, Gross SS, et al. Reversal of endotoxin-mediated shock by NG-methyl-L-arginine, an inhibitor of nitric oxide synthesis. Biochem Biophys Res Commun 1990;172:1132-8.

29. Nava E, Palmer RM, Moncada S. Inhibition of nitric oxide synthesis in septic shock: how much is beneficial? Lancet 1991;338(8782-8783):1555-7.

30. Yanagisawa M, Kurihara H, Kimura S, et al. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature 1988;332:411-5.

31. Inoue A, Yanagisawa M, Kimura S, et al. The human endothelin family: three structurally and pharmacologically distinct isopeptides predicted by three separate genes. Proc Natl Acad Sci U S A 1989;86:2863-7.

32. Hemsen A. Biochemical and functional characterization of endothelin peptides with special reference to vascular effects. Acta Physiol Scand Suppl 1991;602:1-61.

33. Takayanagi R, Ohnaka K, Takasaki C, Ohashi M, Nawata H. Multiple subtypes of endothelin receptors in human and porcine tissues: characterization by ligand binding, affinity labeling, and regional distribution. J Cardiovasc Pharmacol 1991;17 Suppl 7:S127-30.

34. Sharma AC, Motew SJ, Farias S, et al. Sepsis alters myocardial and plasma concentrations of endothelin and nitric oxide in rats. J Mol Cell Cardiol 1997;29:1469-77.

35. Klemm P, Warner TD, Hohlfeld T, Corder R, Vane JR. Endothelin 1 mediates ex vivo coronary vasoconstriction caused by exogenous and endogenous cytokines. Proc Natl Acad Sci U S A 1995;92:2691-5.

36. Szalay L, Kaszaki J, Nagy S, Boros M. The role of endothelin-1 in circulatory changes during hypodynamic sepsis in the rat. Shock 1998;10:123-8.

37. Forni M, Mazzola S, Ribeiro LA, et al. Expression of endothelin-1 system in a pig model of endotoxic shock. Regul Pept 2005;131:89-96.

38. Fujii Y, Magder S, Cernacek P, Goldberg P, Guo Y, Hussain SN. Endothelin receptor blockade attenuates lipopolysaccharide-induced pulmonary nitric oxide production. Am J Respir Crit Care Med 2000;161(3 Pt 1):982-9.

39. Figueras-Aloy J, Gomez-Lopez L, Rodriguez-Miguelez JM, et al. Plasma endothelin-1 and clinical manifestations of neonatal sepsis. J Perinat Med 2004;32:522-6.

40. FitzGerald GA, Pedersen AK, Patrono C. Analysis of prostacyclin and thromboxane biosynthesis in cardiovascular disease. Circulation 1983;67:1174-7.

41. Akarasereenont P, Bakhle YS, Thiemermann C, Vane JR. Cytokine-mediated induction of cyclo-oxygenase-2 by activation of tyrosine kinase in bovine endothelial cells stimulated by bacterial lipopolysaccharide. Br J Pharmacol 1995;115:401-8.

42. Okamoto H, Ito O, Roman RJ, Hudetz AG. Role of inducible nitric oxide synthase and cyclooxygenase-2 in endotoxin-induced cerebral hyperemia. Stroke 1998;29:1209-18.

43. Campbell WB HP. Eicosanoids and platelet-activating factor. In: Hardman JG LL, Molinoff PM, Ruddon RW, Gilman AG, editors. New York: McGraw-Hill; 1996, p601.

44. Hempel SL, Monick MM, Hunninghake GW. Lipopolysaccharide induces prostaglandin H synthase-2 protein and mRNA in human alveolar macrophages and blood monocytes. J Clin Invest 1994;93:391-6.

45. Appleton I, Tomlinson A, Willoughby DA. Induction of cyclo-oxygenase and nitric oxide synthase in inflammation. Adv Pharmacol 1996;35:27-78.

46. Kontos HA, Wei EP, Kukreja RC, Ellis EF, Hess ML. Differences in endothelium-dependent cerebral dilation by bradykinin and acetylcholine. Am J Physiol 1990;258(5 Pt 2):H1261-6.

47. Hernanz R, Alonso MJ, Briones AM, Vila E, Simonsen U, Salaices M. Mechanisms involved in the early increase of serotonin contraction evoked by endotoxin in rat middle cerebral arteries. Br J Pharmacol 2003;140:671-80.

48. Coumans AB, Middelanis JS, Garnier Y, et al. Intracisternal application of endotoxin enhances the susceptibility to subsequent hypoxic-ischemic brain damage in neonatal rats. Pediatr Res 2003;53:770-5.

49. Mertineit C, Samlalsingh-Parker J, Glibetic M, Ricard G, Noya FJ, Aranda JV. Nitric oxide, prostaglandins, and impaired cerebral blood flow autoregulation in group B streptococcal neonatal meningitis. Can J Physiol Pharmacol 2000;78:217-27.

50. Hauck W, Samlalsingh-Parker J, Glibetic M, et al. Deregulation of cyclooxygenase and nitric oxide synthase gene expression in the inflammatory cascade triggered by experimental group B streptococcal meningitis in the newborn brain and cerebral microvessels. Semin Perinatol 1999;23:250-60.

51. Wubbel L, McCracken GH Jr. Management of bacterial meningitis: 1998. Pediatr Rev 1998;19:78-84.

52. Singh AK, Jiang Y. How does peripheral lipopolysaccharide induce gene expression in the brain of rats? Toxicology 2004;201(1-3):197-207.

53. Mayer AM. Therapeutic implications of microglia activation by lipopolysaccharide and reactive oxygen species generation in septic shock and central nervous system pathologies: a review. Medicina (B Aires) 1998;58:377-85.

54. Brian JE Jr, Heistad DD, Faraci FM. Dilatation of cerebral arterioles in response to lipopolysaccharide in vivo. Stroke 1995;26:277-81.

55. Martin CM, Sibbald WJ. Modulation of hemodynamics and organ blood flow by nitric oxide synthase inhibition is not altered in normotensive, septic rats. Am J Respir Crit Care Med 1994;150(6 Pt 1):1539-44.

56. Raper RF, Sibbald WJ, Hobson J, Rutledge FS. Effect of PGE1 on altered distribution of regional blood flows in hyperdynamic sepsis. Chest 1991;100:1703-11.

57. Lingnau W, McGuire R, Dehring DJ, et al. Changes in regional hemodynamics after nitric oxide inhibition during ovine bacteremia. Am J Physiol 1996;270(1 Pt 2):R207-16.

58. Young RS, Hernandez MJ, Yagel SK. Selective reduction of blood flow to white matter during hypotension in newborn dogs: a possible mechanism of periventricular leukomalacia. Ann Neurol 1982;12:445-8.

59. Ekstrom-Jodal B, Elfverson J, Larsson LE. Early effects of E. coli endotoxin on superior sagittal sinus blood flow. An experimental study in dogs. Acta Anaesthesiol Scand 1982;26:171-4.

60. Gerros TC, Semrad SD, Proctor RA. Alterations in clinical, hematological and metabolic variables in bovine neonatal endotoxemia. Can J Vet Res 1995;59:34-9.

61. Grant DA, Franzini C, Wild J, Walker AM. Continuous measurement of blood flow in the superior sagittal sinus of the lamb. Am J Physiol 1995;269(2 Pt 2):R274-9.

62. Veszelka S, Pasztoi M, Farkas AE, Krizbai I, Dung NT, Niwa M, Abraham CS, Deli MA. Pentosan polysulfate protects brain endothelial cells against bacterial lipopolysaccharide-induced damages. Neurochem Int 2007;50:219-28.

63. Mathewson AM, Wadsworth RM. Induction of iNOS restricts functional activity of both eNOS and nNOS in pig cerebral artery. Nitric Oxide 2004;11:331-9.

64. Yoshimoto S, Ishizaki Y, Kurihara H, et al. Cerebral microvessel endothelium is producing endothelin. Brain Res 1990;508:283-5.

65. Skopal J, Turbucz P, Vastag M, et al. Regulation of endothelin release from human brain microvessel endothelial cells. J Cardiovasc Pharmacol 1998;31 Suppl 1:S370-2.

66. Kedzierski RM, Yanagisawa M. Endothelin system: the double-edged sword in health and disease. Annu Rev Pharmacol Toxicol 2001;41:851-76.

67. Adner M, Jansen I, Edvinsson L. Endothelin-A receptors mediate contraction in human cerebral, meningeal and temporal arteries. J Auton Nerv Syst 1994;49 Suppl:S117-21.

68. Patel TR, Galbraith S, McAuley MA, McCulloch J. Endothelin-mediated vascular tone following focal cerebral ischaemia in the cat. J Cereb Blood Flow Metab 1996;16:679-87.

69. Gilbert P, Tremblay J, Thorin E. Endothelium-derived endothelin-1 reduces cerebral artery sensitivity to nitric oxide by a protein kinase C-independent pathway. Stroke 2001;32:2351-5.

70. Vidensky S, Zhang Y, hand T, et al. Neuronal overexpression of COX-2 results in dominant production of PGE2 and altered fever response. Neuromolecular Med 2003;3:15-28.

71. Pittet JF, Morel DR, Hemsen A, et al. Elevated plasma endothelin-1 concentrations are associated with the severity of illness in patients with sepsis. Ann Surg 1991;213:261-4.

72. Kessler P, Bauersachs J, Busse R, Schini-Kerth VB. Inhibition of inducible nitric oxide synthase restores endothelium-dependent relaxations in proinflammatory mediator-induced blood vessels. Arterioscler Thromb Vasc Biol 1997;17:1746-55.

73. Atici A, Satar M, Cetiner S, Yaman A. Serum tumor necrosis factor-alpha in neonatal sepsis. Am J Perinatol 1997;14:401-4.

74. Waage A, Halstensen A, Shalaby R, Brandtzaeg P, Kierulf P, Espevik T. Local production of tumor necrosis factor alpha, interleukin 1, and interleukin 6 in meningococcal meningitis. Relation to the inflammatory response. J Exp Med 1989;170:1859-67.

75. Sibson NR, Blamire AM, Perry VH, Gauldie J, Styles P, Anthony DC. TNF-alpha reduces cerebral blood volume and disrupts tissue homeostasis via an endothelin- and TNFR2-dependent pathway. Brain 2002;125(Pt 11):2446-59.

76. Harkin CP, Hudetz AG, Schmeling WT, Kampine JP, Farber NE. Halothane-induced dilatation of intraparenchymal arterioles in rat brain slices: a comparison to sodium nitroprusside. Anesthesiology 1997;86:885-94.

77. Ziegler EJ, McCutchan JA, Fierer J, et al. Treatment of gram-negative bacteremia and shock with human antiserum to a mutant Escherichia coli. N Engl J Med 1982;307:1225-30.

78. Petros A, Bennett D, Vallance P. Effect of nitric oxide synthase inhibitors on hypotension in patients with septic shock. Lancet 1991;338(8782-8783):1557-8.

79. Benigni A, Remuzzi G. Endothelin antagonists. Lancet 1999;353:133-8.

80. Fisher CJ Jr, Slotman GJ, Opal SM, et al. Initial evaluation of human recombinant interleukin-1 receptor antagonist in the treatment of sepsis syndrome: a randomized, open-label, placebo-controlled multicenter trial. Crit Care Med 1994;22:12-21.