Eosinophils in the cerebrospinal fluid of mice infected with Angiostrongylus cantonensis are resistant to apoptosis

Interleukin-5 (IL-5) transgenic mice were used to assess the immunological features of CSF eosinophils from mice infected with Angiostrongylus cantonensis. CSF eosinophils were hypodense by day 14 post infection (p.i.). CSF eosinophils survived longer in vitro than peritoneal eosinophils collected from cadmium sulphate (CdSO₄) -treated normal IL-5 transgenic mice. Apoptosis was measured by Annexin V binding and the presence of a distinct laddering pattern of DNA fragmentation on agarose electrophoresis. Regardless of the presence or absence of Actinomycin D, CSF eosinophils collected from IL-5 transgenic mice from days 15–36 p.i. exhibited less apoptosis than peritoneal eosinophils collected from uninfected IL-5 transgenic mice. CSF eosinophils collected from A. cantonensis infected C57BL/6 mice at days 15–34 p.i. showed elongation of survival time and less apoptosis during in vitro cultivation. Reduced apoptosis was noted only in CSF eosinophils, but not in peritoneal eosinophils recovered from the same infected IL-5 transgenic mice. CPP32/Caspase 3 activity of cultured peritoneal eosinophils from both infected and uninfected IL-5 transgenic mice was higher than that of cultured CSF eosinophils. Stimulation with A23187 readily induced apoptosis of peritoneal eosinophils, but not CSF eosinophils or peritoneal eosinophils cultured with mouse recombinant IL-5. The latter cells were morphologically identical to hypodense eosinophils. RT-PCR analysis indicated that bcl-2 and bcl-x_L mRNA expression was higher in CSF eosinophils compared with peritoneal eosinophils and this expression in the latter cells was upregulated after culture with mouse recombinant IL-5. These results suggest that CSF eosinophils, shifting to hypodense status through an accumulation from peripheral blood, are resistant to apoptosis. These changes may explain the long-lasting, helminthotoxic and neurotoxic actions of CSF eosinophils in A. cantonensis infection.     

Matrix Metalloproteinase-9 Leads to Claudin-5 Degradation via the NF-κB Pathway in BALB/c Mice with Eosinophilic Meningoencephalitis Caused by Angiostrongylus cantonensis

The epithelial barrier regulates the movement of ions, macromolecules, immune cells and pathogens. The objective of this study was to investigate the role of the matrix metalloproteinase (MMP)-9 in the degradation of tight junction protein during infection with rat nematode lungworm Angiostrongylus cantonensis. The results showed that phosphorylation of IκB and NF-κB was increased in mice with eosinophilic meningoencephalitis. Treatment with MG132 reduced the phosphorylation of NF-κB and the activity of MMP-9, indicating upregulation of MMP-9 through the NF-κB signaling pathway. Claudin-5 was reduced in the brain but elevated in the cerebrospinal fluid (CSF), implying that A. cantonensis infection caused tight junction breakdown and led to claudin-5 release into the CSF. Degradation of claudin-5 coincided with alteration of the blood-CSF barrier permeability and treatment with the MMP inhibitor GM6001 attenuated the degradation of claudin-5. These results suggested that degradation of claudin-5 was caused by MMP-9 in angiostrongyliasis meningoencephalitis. Claudin-5 could be used for the pathophysiologic evaluation of the blood-CSF barrier breakdown and tight junction disruption after infection with A. cantonensis.     

Compartmentalization of TNF and IL-6 in meningitis and septic shock

We examined the compartmentalization of bioactive tumour necrosis factor (TNF) and interleukin 6 (IL-6) to the subarachnoid space and systemic circulation in patients with meningococcal meningitis and septic shock/bacteraemia. In patients with meningitis, median levels of TNF in 31 paired samples of cerebrospinal fluid (CSF) and serum were respectively 783 pg/ml and below detection limit (p < 0.001) and median levels of IL-6 were 150 ng/ml and 0.3 ng/ml (p < 0.0001). In patients with septic shock without meningitis, median levels in paired samples of CSF and serum were respectively below detection limit and 65 pg/ml (not significant, (ns)) (TNF, eleven patients) and 1.3 ng/ml-3 ng/ml (ns) (IL-6, nine patients). The data show that TNF and IL-6 are localized to the subarachnoid space in patients with meningitis although the blood-brain barrier is penetrable to serum proteins. On the other hand, patients with septic shock tend to have cytokines in both serum and CSF.     

Identification of a matrix-degrading phenotype in human tuberculosis in vitro and in vivo

Tuberculous meningitis is characterized by cerebral tissue destruction. Monocytes, pivotal in immune responses to Mycobacterium tuberculosis, secrete matrix metalloproteinase-9 (MMP-9), which facilitates leukocyte migration across the blood-brain barrier, but may cause cerebral injury. In vitro, human monocytic (THP-1) cells infected by live, virulent M. tuberculosis secreted MMP-9 in a dose-dependent manner. At 24 h, MMP-9 concentrations increased 10-fold to 239 +/- 75 ng/ml (p = 0.001 vs controls). MMP-9 mRNA became detectable at 24--48 h. In contrast, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) gene expression and secretion were similar to constitutive levels from controls at 24 h and increased just 5-fold by 48 h. In vivo investigation revealed MMP-9 concentration per leukocyte in cerebrospinal fluid (CSF) from tuberculous meningitis patients (n = 23; median (range), 3.19 (0.19--31.00) ng/ml/cell) to be higher than that in bacterial (n = 12; 0.23 (0.01--18.37) ng/ml/cell) or viral meningitis (n = 20; 0.20 (0.04--31.00) ng/ml/cell; p < 0.01). TIMP-1, which was constitutively secreted into CSF, was not elevated in tuberculous compared with bacterial meningitis or controls. Thus, a phenotype in which MMP-9 activity is relatively unrestricted by TIMP-1 developed both in vitro and in vivo. This is functionally significant, since MMP-9 concentrations per CSF leukocyte (but not TIMP-1 concentrations) were elevated in fatal tuberculous meningitis and in patients with signs of cerebral tissue damage (unconsciousness, confusion, or neurological deficit; p < 0.05). However, MMP-9 activity was unrelated to the severity of systemic illness. In summary, M. tuberculosis-infected monocytic cells develop a matrix-degrading phenotype, which was observed in vivo and relates to clinical signs reflecting cerebral injury in tuberculous meningitis.     

Oxidative stress in mice infected with Angiostrongylus cantonensis coincides with enhanced glutathione-dependent enzymes activity

This study aimed to estimate reactive oxygen species (ROS) production, antioxidants activity, and biomarkers level of oxidative damage to protein and DNA in the cerebrospinal fluid (CSF) of C57BL/6 mice infected with Angiostrongylus cantonensis. The mean ROS concentration in the CSF of infected mice increased gradually, and the increase in ROS in CSF became statistical significance at days 12-30 post-infection compared to that before infection (P < 0.001), and then ROS returned to normal level at day 45 after infection. In parallel with the increase in ROS in the CSF, infected mice showed similar of changes in reduced glutathione (GSH), glutathione reductase (GR), glutathione peroxidase (GPx), and glutathione S-transferase (GST) as that in ROS in the CSF. GSH, GR, GPx, and GST in the CSF of infected mice were all significantly higher than they were before infection during days 12-30 post-infection. However, protein carbonyl content and 8-hydroxy-2′-deoxyguanosine, biomarkers of oxidative damage to protein and DNA, respectively, were also significantly higher in the CSF of infected mice during this period. These results suggest that oxidative stress occur in the cells of central nervous system of mice infected with A. cantonensis during days 12-30 after infection due to ROS overproduction in CSF despite the increase in antioxidants during this period.     

MicroRNA miR-155-5p knockdown attenuates Angiostrongylus cantonensis-induced eosinophilic meningitis by downregulating MMP9 and TSLP proteins

Angiostrongylus cantonensis infection is a major cause of eosinophilic meningitis (EM). Severe cases or cases that involve infants and children present poor prognoses. MicroRNAs (miRNAs), which are important regulators of gene expression in many biological processes, were recently found to be regulators of the host response to infection by parasites; however, their roles in brain inflammation caused by A. cantonensis are still unclear. The current study confirmed that miR-155-5p peaked at 21 days after A. cantonensis infection, and its expression was positively correlated with the concentration of excretory and secretory products (ESPs). We found that miR-155-5p knockdown lentivirus successfully ameliorated brain injury and downregulated the expression of major basic protein (MBP) in vivo, and the number of eosinophils in CSF (and the percentage of eosinophils in peripheral blood were also decreased in the miR-155-5p knockdown group. Moreover, the expression of several eosinophilic inflammation cytokines such as CCL6/C10, ICAM-1, and MMP9, declined after the miR-155-5p knockdown. SOCS1 protein, which is an important negative regulator of inflammation activation, was identified as a direct miR-155-5p target. We further detected the effect of miR-155-5p knockdown on phosphorylated-STAT3 and phosphorylated-p65 proteins, which were found to be negatively regulated by SOCS1 and play an important role in regulating the inflammatory response. We found that miR-155-5p knockdown decreased the activity of p-STAT3 and p-p65, thereby leading to lower expression of MMP9 and TSLP proteins, which were closely related to the chemotaxis and infiltration of eosinophils. Interestingly, the inhibition of p-STAT3 or p-p65 was found to induce the downregulation of miR-155-5p in an opposite manner. These observations suggest that a positive feedback loop was formed between miR-155-5p, STAT3, and NF-κB in A. cantonensis infection and that miR-155-5p inhibition might provide a novel strategy to attenuate eosinophilic meningitis.     

Angiostrongylus cantonensis: identification and characterization of microRNAs in male and female adults

Angiostrongylus cantonensis causes eosinophilic meningitis and eosinophilic pleocytosis in humans and is of significant socio-economic importance globally. microRNAs (miRNAs) are endogenous small non-coding RNAs that play crucial roles in gene expression regulation, cellular function and defense, homeostasis and pathogenesis. They have been identified in a diverse range of organisms. The objective of this study was to determine and characterize miRNAs of female and male adults of A. cantonensis by Solexa deep sequencing. A total of 8861,260 and 10,957,957 high quality reads with 20 and 23 conserved miRNAs were obtained in females and males, respectively. No new miRNA sequence was found. Nucleotide bias analysis showed that uracil was the prominent nucleotide, particularly at positions of 1, 10, 14, 17 and 22, approximately at the beginning, middle and the end of the conserved miRNAs. To our knowledge, this is the first report of miRNA profiles in A. cantonensis, which may represent a new platform for studying regulation of genes and their networks in A. cantonensis.     

Circulation of marker substances in the cerebrospinal fluid of an amphibian,Rana pipiens

Summary Solutions of fluorescein-labelled dextran or Evans blue-albumin were infused into the lateral cerebral ventricle of Rana pipiens. The subsequent distribution in the cerebrospinal fluid (CSF) was investigated between 2 and 24 h after infusion by freezing and examination of the cut blocks of the head and vertebral column of the stage of a freezing microtome. These marker substances move out of the ventricles into the subarachnoid space at the caudal end of the fourth ventricle and spread rapidly along the subarachnoid space of the spinal cord. The spreading of marker substances is slower into the brain subarachnoid space. When the marker is infused into the subarachnoid space of the forebrain, it becomes distributed throughout the subarachnoid space of the brain and spinal cord but not in the ventricles.Partial clearance of markers from the ventricles takes place within 5 h and total clearance within 8 h. Clearance from the brain and cord subarachnoid space is somewhat slower and can only be detected in experiments lasting 10 h or more. Absorption of the markers from the CSF occurs via the intervertebral foramina of the spinal cord. Fluorescence microscopy of sections of the cord show that the fluorescence leaves the subarachnoid space at the point where the spinal nerves traverse the arachnoid membrane.     

Transgenic expression of matrix metalloproteinase-9 causes adult-onset emphysema in mice associated with the loss of alveolar elastin

Matrix metalloproteinase (MMP)-9 has been consistently identified in the lungs of patients with chronic obstructive pulmonary disease (COPD). However, its role in the development of the disease remains undefined. Mice that specifically express human MMP-9 in their macrophages were generated, and morphometric, biochemical, and histological analyses were conducted on the transgenic and littermate control mice over 1 yr to determine the effect of macrophage MMP-9 expression on emphysema formation and lung matrix content. Lung morphometry was normal in transgenic mice at 2 mo of age (mean linear intercept = 50+/-3 littermate mice vs. 51+/-2 transgenic mice). However, after 12 mo of age, the MMP-9 transgenic mice developed significant air space enlargement (mean linear intercept = 53+/-3 littermate mice vs. 61+/-2 MMP-9 transgenic mice; P<0.04). Lung hydroxyproline content was not significantly different between wild-type and transgenic mice, but MMP-9 did significantly decrease alveolar wall elastin at 1 yr of age (4.9+/-0.3% area of alveolar wall in the littermate mice vs. 3.3+/-0.3% area of alveolar wall in the MMP-9 mice; P<0.004). Thus these results establish a central role for MMP-9 in the pathogenesis of this disease by demonstrating that expression of this protease in macrophages can alter the extracellular matrix and induce progressive air space enlargement in mice.     

Intrathecal levels of matrix metalloproteinases in systemic lupus erythematosus with central nervous system engagement

Symptoms originating from the central nervous system (CNS) occur frequently in patients with systemic lupus erythematosus (SLE), and CNS involvement in lupus is associated with increased morbidity and mortality. We recently showed that neurones and astrocytes are continuously damaged during the course of CNS lupus. The matrix metalloproteinases (MMPs) are a group of tissue degrading enzymes that may be involved in this ongoing brain destruction. The aim of this study was to examine endogenous levels of free, enzymatically active MMP-2 and MMP-9 in cerebrospinal fluid from patients with SLE. A total of 123 patients with SLE were evaluated clinically, with magnetic resonance imaging of brain and cerebrospinal fluid (CSF) analyses. Levels of free MMP-2 and MMP-9 were determined in CSF using an enzymatic activity assay. CSF samples from another 22 cerebrally healthy individuals were used as a control. Intrathecal MMP-9 levels were significantly increased in patients with neuropsychiatric SLE as compared with SLE patients without CNS involvement (P < 0.05) and healthy control individuals (P = 0.0012). Interestingly, significant correlations between MMP-9 and intrathecal levels of neuronal and glial degradation products were noted, indicating ongoing intrathecal degeneration in the brains of lupus patients expressing MMP-9. In addition, intrathecal levels of IL-6 and IL-8 – two cytokines that are known to upregulate MMP-9 – both exhibited significant correlation with MMP-9 levels in CSF (P < 0.0001), suggesting a potential MMP-9 activation pathway. Our findings suggest that proinflammatory cytokine induced MMP-9 production leads to brain damage in patients with CNS lupus.     

Elevation of plasminogen activators in cerebrospinal fluid of mice with eosinophilic meningitis caused by Angiostrongylus cantonensis

A hallmark of parasitic meningitis is the infiltration of eosinophils into the subarachnoid space. Infection with Angiostrongylus cantonensis in mice induced proteinase activity in parallel with the pathological changes of eosinophilic meningitis. Zymogram analysis demonstrated that 70 and 55 kDa proteinases from cerebrospinal fluid (CSF) were active against the casein/plasminogen substrate. The proteinase activities were clearly inhibited by phenylmethanesulphonyl fluoride but not by ethylenediamine tetraacetic acid, 1,10-phenanthroline or leupeptin. Western blotting confirmed these enzymes to be tissue-type plasminogen activator and urokinase-type plasminogen activator, respectively. High activities of tissue-type plasminogen activator and urokinase-type plasminogen activator were detected in the CSF of mice with eosinophilic meningitis, and correlated positively with CSF eosinophil numbers and total protein, respectively. Immunohistochemistry demonstrated that tissue-type plasminogen activator and urokinase-type plasminogen activator localised in the endothelial cells of blood vessels, in blood clots and in infiltrated leukocytes. These results suggest that tissue-type plasminogen activator and urokinase-type plasminogen activator may be play a role in the pathogenesis of eosinophilic meningitis of angiostrongyliasis.     

Effect of naloxone administration upon the diurnal concentrations of oxytocin in the cerebrospinal fluid of rhesus and cynomolgus monkeys.

A diurnal pattern in oxytocin concentrations is present in cerebrospinal fluid (CSF) removed from the spinal subarachnoid space of monkeys, with elevated levels occurring in the early light hours. In order to investigate the possible role of endogenous opioid peptides in the generation of this oxytocin rhythm, we administered naloxone (0.4 mg/kg/h x 48 h) to rhesus and cynomolgus monkeys and examined the effects on the diurnal pattern of oxytocin in CSF collected from the lumbar subarachnoid spinal space. Monkeys maintained on jacket/tether/swivel systems and in a 12 h light: 12 h dark cycle (lights on 07.00-19.00 h) were implanted with temporary spinal subarachnoid catheters. CSF was continuously collected from the lumbar subarachnoid space and assayed for oxytocin. Oxytocin concentrations in CSF showed a diurnal variation with peak and nadir concentrations during light and dark hours, respectively. The lumbar CSF concentrations of oxytocin were not significantly different during naloxone vs. saline infusion. Plasma oxytocin concentrations, measured in the same animals, displayed no diurnal variation and were not significantly different during naloxone vs. saline infusion. We conclude that naloxone administration for 48 h does not perturb the diurnal variation in oxytocin concentrations in the CSF of monkeys. Mu opioid receptors are unlikely to be involved in modulating the diurnal rhythm of oxytocin in the CSF of monkeys.     

Leukocyte Attraction by CCL20 and Its Receptor CCR6 in Humans and Mice with Pneumococcal Meningitis

We previously identified CCL20 as an early chemokine in the cerebrospinal fluid (CSF) of patients with pneumococcal meningitis but its functional relevance was unknown. Here we studied the role of CCL20 and its receptor CCR6 in pneumococcal meningitis. In a prospective nationwide study, CCL20 levels were significantly elevated in the CSF of patients with pneumococcal meningitis and correlated with CSF leukocyte counts. CCR6-deficient mice with pneumococcal meningitis and WT mice with pneumococcal meningitis treated with anti-CCL20 antibodies both had reduced CSF white blood cell counts. The reduction in CSF pleocytosis was also accompanied by an increase in brain bacterial titers. Additional in vitro experiments showed direct chemoattractant activity of CCL20 for granulocytes. In summary, our results identify the CCL20-CCR6 axis as an essential component of the innate immune defense against pneumococcal meningitis, controlling granulocyte recruitment.     

Meningitis patients with Angiostrongylus cantonensis may present without eosinophilia in the cerebrospinal fluid in northern Vietnam

BackgroundEosinophilic meningitis (EM) is a rare clinical syndrome caused by both infectious and noninfectious diseases. In tropical pacific countries, Angiostrongylus cantonensis is the most common cause. However, the EM definition varies in the literature, and its relation to parasitic meningitis (PM) remains unclear.Methodology/Principal findingsAdult and adolescent patients of 13 years old or above with suspected central nervous system (CNS) infections with abnormal CSF findings were prospectively enrolled at a tertiary referral hospital in Hanoi, Vietnam from June 2012 to May 2014. Patients with EM or suspected PM (EM/PM) were defined by the presence of either ≥10% eosinophils or an absolute eosinophil cell counts of ≥10/mm³ in the CSF or blood eosinophilia (>16% of WBCs) without CSF eosinophils. In total 679 patients were enrolled: 7 (1.03%) had ≥10% CSF eosinophilia, 20 (2.95%) had ≥10/mm³ CSF eosinophilia, and 7 (1.03%) had >16% blood eosinophilia. The patients with ≥10% CSF eosinophilia were significantly younger (p = 0.017), had a lower body temperature (p = 0.036) than patients with ≥10/mm³ CSF eosinophilia among whom bacterial pathogens were detected in 72.2% (13/18) of those who were tested by culture and/or PCR. In contrast, the characteristics of the patients with >16% blood eosinophilia resembled those of patients with ≥10% CSF eosinophilia. We further conducted serological tests and real-time PCR to identify A. cantonensis. Serology or real-time PCR was positive in 3 (42.8%) patients with ≥10% CSF eosinophilia and 6 (85.7%) patients with >16% blood eosinophilia without CSF eosinophils but none of patients with ≥10/mm³ CSF eosinophilia.ConclusionsThe etiology of PM in northern Vietnam is A. cantonensis. The eosinophil percentage is a more reliable predictor of parasitic EM than absolute eosinophil count in the CSF. Patients with PM may present with a high percentage of eosinophils in the peripheral blood but not in the CSF.     

Liberation of desmosine and isodesmosine as amino acids from insoluble elastin by elastolytic proteases

The development of atherosclerotic lesions and abdominal aortic aneurysms involves degradation and loss of extracellular matrix components, such as collagen and elastin. Releases of the elastin cross-links desmosine (DES) and isodesmosine (IDE) may reflect elastin degradation in cardiovascular diseases. This study investigated the production of soluble elastin cross-linking structures by proteinases implicated in arterial diseases. Recombinant MMP-12 and neutrophil elastase liberated DES and IDE as amino acids from insoluble elastin. DES and IDE were also released from insoluble elastin exposed to monocyte/macrophage cell lines or human primary macrophages derived from peripheral blood monocytes. Elastin oxidized by reactive oxygen species (ROS) liberated more unconjugated DES and IDE than did non-oxidized elastin when incubated with MMP-12 or neutrophil elastase. These results support the exploration of free DES and IDE as biomarkers of elastin degradation.     

Analysis of cytokine levels in cerebrospinal fluid in mumps meningitis: comparison with echovirus type 30 meningitis

BACKGROUND: It is unclear whether or not the CSF cytokine profiles in viral meningitis differ with the kind of causative virus. METHODS: We measured the concentrations of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2), IL-4, IL-6, and IL-10 in CSF during the acute stage in 15 children with mumps meningitis (MM), and 34 with echovirus type 30 meningitis (EM). RESULTS: The CSF IFN-gamma, IL-2, IL-6, and IL-10 levels were elevated in MM, and the CSF IFN-gamma, IL-2, and IL-6 levels were elevated in EM. The CSF IFN-gamma, IL-2, and IL-10 levels in MM were significantly higher than those in EM (p<0.0001, p<0.0001, and p<0.0001, respectively). The CSF IL-6 levels in EM were significantly higher than those in MM (p=0.0255). The CSF TNF-alpha and IL-4 levels were not elevated in MM or EM. In MM, the IL-6 level was correlated with the IL-2 and IL-10 levels in CSF (p=0.0347 and p=0.0120, respectively). In EM, the IFN-gamma level was correlated with the IL-10 level in CSF (p=0.0002). CONCLUSION: CSF cytokine profiles in MM were different from those in EM. Therefore, it is likely that the pathogenesis of MM is different from that of EM.     

Apoptosis in meningoencephalitis of Angiostrongylus cantonensis-infected mice

A hallmark of eosinophilic meningoencephalitis is infiltration of leukocytes into brain parenchyma and subarachnoid space infected by Angiostrongylus cantonensis. Apoptosis, a process that eliminates useless cells and counterbalances tissue homeostasis, is important for homeostasis of the immune system. In this study, we investigated the characteristics of cell death induced in BABL/c mice infected with A. cantonensis. We observed increased expression of the apoptotic proteins, caspase-3, caspase-8, caspase-9, and cytochrome c, and decreased expression of anti-apoptotic proteins, B-cell leukemia 2 and inhibitor of apoptosis protein 1. On immunohistochemistry, apoptotic proteins were localized within the leukocytes infiltrate. A terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick-end labeling assay to detect DNA fragmentation confirmed these observations. The infiltration of leukocytes present in the brain parenchyma and subarachnoid space in vivo may also express these apoptotic regulatory molecules, which demonstrates the capacity of these cells to undergo apoptosis.     

Matrix metalloproteinase inhibitors attenuate endotoxemia induced cardiac dysfunction: A potential role for MMP-9

Enhanced cardiac generation of peroxynitrite contributes to septic cardiomyopathy. Since matrix metalloproteinases (MMPs) are activated in vitro by peroxynitrite, we hypothezised that MMPs may contribute to cardiac mechanical dysfunction in sepsis. Rats were injected (i.p.) with either lipopolysaccharide (LPS, 4mg/kg) or vehicle. MMP inhibitors, either Ro 31-9790 (20 mg/kg), doxycycline (4mg/kg), or vehicle were administered i.p. 30 min after LPS. At 6 h, when the symptoms of endotoxemia peak, hearts were excised and perfused as working hearts with Krebs-Henseleit buffer at 37°C. Cardiac work (cardiac output x peak systolic pressure product) was measured. Perfusate and ventricle samples were analyzed by gelatin zymography to quantify MMP activity.Cardiac function was significantly depressed in LPS-treated rats compared to control rats (control: 55 ± 4, LPS: 26 ± 6 mmHg*mL*min^–1). LPS also caused a loss of 72 kDa MMP-2 activity in the ventricles and the perfusate. Although MMP-9 activity was not detected in the ventricles, LPS resulted in an increase in perfusate 92 kDa MMP-9 activity. The MMP inhibitors significantly improved cardiac function of LPS-treated rats (Ro31-9790: 38 ± 3, doxycycline: 51 ± 3 mmHg*mL*min^–1), had no effect on the loss of MMP-2 activity, and significantly reduced the MMP-9 activity in the perfusate. These results demonstrate, for the first time, that LPS induced cardiac dysfunction is associated with a loss in ventricular MMP-2 activity and the release of MMP-9 from the heart. MMP inhibitors can significantly preserve cardiac mechanical function during septic shock.     

Expression of matrix metalloproteinases and their tissue inhibitor during viral encephalitis

Matrix metalloproteinases (MMPs) participate in remodeling the extracellular matrix and facilitate entry of inflammatory cells into tissues. Infection of the murine central nervous system (CNS) with a neurotropic coronavirus induces encephalitis associated with increased levels of mRNA encoding MMP-3 and MMP-12. Whereas virus-induced MMP-3 expression was restricted to CNS resident astrocytes, MMP-12 mRNA was expressed by both inflammatory cells and CNS resident cells. Immunosuppression increased both MMP-3 and MMP-12 mRNA levels in CNS resident cells, suggesting that the presence of virus rather than inflammation induced protease up-regulation. MMP activity is partially regulated by a small family of genes encoding tissue inhibitors of matrix metalloproteinases (TIMPs); among the TIMPs, only TIMP-1 mRNA expression increased in the CNS following coronavirus infection. During inflammation TIMP-1 mRNA was most prominently expressed by infiltrating cells. By contrast, in the immunosuppressed host TIMP-1 mRNA was expressed by CNS resident cells. Analysis of cytokine and chemokine mRNA induction within the infected CNS of healthy and immunocompromised mice suggested a possible correlation between increased viral replication and increased levels of beta interferon, MMP-3, MMP-12, and TIMP-1 mRNA. CD4+ T cells which localize to the perivascular and subarachnoid spaces were identified as the primary source of TIMP-1 protein. By contrast, protein expression was undetectable in astrocytes or CD8+ T cells, the primary antiviral effectors that localize to the CNS parenchyma in response to infection. These data suggest that in contrast to the results seen with MMPs, inhibition of protease activity via TIMP-1 expression correlates with the differential tissue distribution of T-cell subsets during acute coronavirus-induced encephalitis.     

Unusual presentations of inflammatory conditions in cerebrospinal fluid

Unusual inflammatory reactions in cerebrospinal fluid (CSF) in five patients were explicable by the type of intracranial injury or surgical intervention that they had received or by their basic disease process. Lumbar puncture fluid from a 64-year-old man with multiple facial fractures contained neutrophils, bacteria, Candida sp. and ciliated columnar cells, findings consistent with a basilar skull fracture allowing paranasal sinus contents to enter the subarachnoid space. A 59-year-old man with angioimmunoblastic lymphadenopathy developed meningitis and suffered a respiratory arrest; a ventricular fluid contained acute inflammatory cells as well as numerous corpora amylacea. Lumbar CSF obtained during surgery from a 26-year-old man with a pontine glioma contained numerous histiocytes clustered around polarizable filaments, probably strands of gauze introduced during surgery. A specimen of CSF obtained intraoperatively from a 54-year-old man with an acoustic neuroma undergoing a second craniotomy contained multinucleated giant cells bearing suture material. A 19-year-old girl with systemic sarcoidosis had noncaseating granulomas in the right temporal lobe and multinucleated giant cells in her CSF.