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1.
Am J Respir Crit Care Med ; 200(9): 1126-1133, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31260638

RESUMO

Rationale: Infections caused by multidrug-resistant bacteria are a major clinical challenge. Phage therapy is a promising alternative antibacterial strategy.Objectives: To evaluate the efficacy of intravenous phage therapy for the treatment of ventilator-associated pneumonia due to methicillin-resistant Staphylococcus aureus in rats.Methods: In a randomized, blinded, controlled experimental study, we compared intravenous teicoplanin (3 mg/kg, n = 12), a cocktail of four phages (2-3 × 109 plaque-forming units/ml of 2003, 2002, 3A, and K; n = 12), and a combination of both (n = 11) given 2, 12, and 24 hours after induction of pneumonia, and then once daily for 4 days. The primary outcome was survival at Day 4. Secondary outcomes were bacterial and phage densities in lungs and spleen, histopathological scoring of infection within the lungs, and inflammatory biomarkers in blood.Measurements and Main Results: Treatment with either phages or teicoplanin increased survival from 0% to 58% and 50%, respectively (P < 0.005). The combination of phages and antibiotics did not further improve outcomes (45% survival). Animal survival correlated with reduced bacterial burdens in the lung (1.2 × 106 cfu/g of tissue for survivors vs. 1.2 × 109 cfu/g for nonsurviving animals; P < 0.0001), as well as improved histopathological outcomes. Phage multiplication within the lung occurred during treatment. IL-1ß increased in all treatment groups over the course of therapy.Conclusions: Phage therapy was as effective as teicoplanin in improving survival and decreasing bacterial load within the lungs of rats infected with methicillin-resistant S. aureus. Combining antibiotics with phage therapy did not further improve outcomes.

2.
J Neuroinflammation ; 16(1): 156, 2019 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-31351490

RESUMO

BACKGROUND: Pneumococcal meningitis is associated with high risk of neurological sequelae such as cognitive impairment and hearing loss. These sequelae are due to parenchymal brain and inner ear damage primarily induced by the excessive inflammatory reaction in response to bacterial brain invasion. Metformin-a biguanide drug to treat diabetes mellitus type 2-was recently found to suppress neuroinflammation and induce neuroregeneration. This study evaluated the effect of metformin adjunctive to antibiotics on neuroinflammation, brain and inner ear damage, and neurofunctional outcome in experimental pediatric pneumococcal meningitis. METHODS: Eleven-day-old Wistar rats were infected intracisternally with 5.22 ± 1.27 × 103 CFU Streptococcus pneumoniae and randomized for treatment with metformin (50 mg/kg, i.p., once daily for 3 weeks) plus ceftriaxone (100 mg/kg, i.p., bid, n = 61) or ceftriaxone monotherapy (n = 79). Cortical damage and hippocampal apoptosis were evaluated histomorphometrically 42 h post infection. Cerebrospinal fluid cytokine levels were analyzed during acute infection. Five weeks post infection, auditory brainstem responses were measured to determine hearing thresholds. Spiral ganglion neuron density and abundance of recently proliferated and integrated hippocampal granule neurons were assessed histologically. Additionally, the anti-inflammatory effect of metformin was studied in primary rat astroglial cells in vitro. RESULTS: Upon pneumococcal infection, metformin treatment significantly reduced levels of inflammatory cytokines and nitric oxide production in cerebrospinal fluid and in astroglial cell cultures in vitro (p < 0.05). Compared to animals receiving ceftriaxone monotherapy, adjunctive metformin significantly reduced cortical necrosis (p < 0.02) during acute infection and improved median click-induced hearing thresholds (60 dB vs. 100 dB, p < 0.002) 5 weeks after infection. Adjuvant metformin significantly improved pure tone hearing thresholds at all assessed frequencies compared to ceftriaxone monotherapy (p < 0.05) and protected from PM-induced spiral ganglion neuron loss in the inner ear (p < 0.05). CONCLUSION: Adjuvant metformin reduces brain injury during pneumococcal meningitis by decreasing the excessive neuroinflammatory response. Furthermore, it protects spiral ganglion neurons in the inner ear and improves hearing impairments after experimental pneumococcal meningitis. These results identify adjuvant metformin as a promising therapeutic option to improve the outcome after pediatric pneumococcal meningitis.

3.
Cell Mol Life Sci ; 76(16): 3097-3116, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31172218

RESUMO

Metalloproteinases-such as matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinases (ADAMs)-are involved in various diseases of the nervous system but also contribute to nervous system development, synaptic plasticity and neuroregeneration upon injury. MMPs and ADAMs proteolytically cleave many substrates including extracellular matrix components but also signaling molecules and receptors. During neuroinfectious disease with associated neuroinflammation, MMPs and ADAMs regulate blood-brain barrier breakdown, bacterial invasion, neutrophil infiltration and cytokine signaling. Specific and broad-spectrum inhibitors for MMPs and ADAMs have experimentally been shown to decrease neuroinflammation and brain damage in diseases with excessive neuroinflammation as a common denominator, such as pneumococcal meningitis and multiple sclerosis, thereby improving the disease outcome. Timing of metalloproteinase inhibition appears to be critical to effectively target the cascade of pathophysiological processes leading to brain damage without inhibiting the neuroregenerative effects of metalloproteinases. As the critical role of metalloproteinases in neuronal repair mechanisms and regeneration was only lately recognized, the original idea of chronic MMP inhibition needs to be conceptually revised. Recently accumulated research urges for a second chance of metalloproteinase inhibitors, which-when correctly applied and dosed-harbor the potential to improve the outcome of different neuroinflammatory diseases.


Assuntos
Proteínas ADAM/metabolismo , Metaloproteinases da Matriz/metabolismo , Esclerose Múltipla/patologia , Doenças do Sistema Nervoso/patologia , Barreira Hematoencefálica/metabolismo , Citocinas/metabolismo , Humanos , Esclerose Múltipla/metabolismo , Doenças do Sistema Nervoso/metabolismo , Neurônios/metabolismo , Neutrófilos/citologia , Neutrófilos/imunologia , Neutrófilos/metabolismo , Transdução de Sinais
4.
J Neuroinflammation ; 16(1): 105, 2019 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-31103037

RESUMO

BACKGROUND: Streptococcus pneumoniae is the cause of a highly lethal form of meningitis in humans. Microglial cells in the brain represent the first line of defense against pathogens, and they participate in the inflammatory response. The cholesterol-dependent cytolysin pneumolysin and the bacterial capsule are key pathogenic factors, known to exacerbate the course of pneumococcal meningitis. METHODS: We utilized live imaging and immunostaining of glial cells in dissociated and acute brain slice cultures to study the effect of pneumococcal factors, including the cholesterol-dependent cytolysin pneumolysin and the pneumococcal capsule, on microglial motility and taxis. RESULTS: In brain tissue, primary microglia cells showed an enhanced response towards lysates from bacteria lacking capsules and pneumolysin as they moved rapidly to areas with an abundance of bacterial factors. The presence of bacterial capsules and pneumolysin cumulatively inhibited microglial taxis. In mixed cultures of astrocytes and microglia, the motility of microglia was inhibited by capsular components within minutes after exposure. The reduced motility was partially reversed by mannan, a mannose receptor inhibitor. The effects on microglia were not mediated by astrocytes because pure microglial cells responded to various pneumococcal lysates similarly with distinct cell shape changes as seen in mixed cultures. CONCLUSIONS: Our data indicate that microglia possess the capacity for a very agile response towards bacterial pathogens, but key pathogenic factors, such as pneumococcal capsules and pneumolysin, inhibited this response shortly after a bacterial challenge. Furthermore, we demonstrate for the first time that the bacterial capsule affects cellular behaviors such as motility and taxis.

5.
Artigo em Inglês | MEDLINE | ID: mdl-31061158

RESUMO

Despite appropriate antibiotic therapy, pneumococcal meningitis (PM) is associated with a case fatality rate of up to 30% in high-income countries. Survivors often suffer from severe lifelong disabilities. An excessive inflammatory reaction drives the pathophysiology, leading to brain damage and neurologic sequelae. We aimed to improve the outcome of experimental PM by simultaneously targeting different pathophysiological mechanisms with combined adjunctive therapies previously shown to be neuroprotective. In vitro, the anti-inflammatory effects of doxycycline and daptomycin were evaluated on primary rat astroglial cells stimulated with Streptococcus pneumoniae Eleven-day-old infant Wistar rats were infected intracisternally with S. pneumoniae and randomized for treatment with ceftriaxone or combination adjuvant therapy consisting of ceftriaxone, daptomycin, and doxycycline. During acute PM, combined-adjuvant therapy with ceftriaxone, daptomycin, and doxycycline increased the survival rate from 64.1% to 85.8% (P < 0.01) and alleviated weight loss compared to ceftriaxone monotherapy (P < 0.01). Levels of inflammatory cytokines were significantly reduced by combined-adjuvant therapy in vitro (P < 0.0001) and in cerebrospinal fluid in vivo (P < 0.05). In infected animals treated with combined adjunctive therapy, cortical damage was significantly reduced (P < 0.05), and animals showed a trend toward better hearing capacity 3 weeks after the infection (P = 0.089), an effect which was significant in mildly infected animals (48 decibels [dB] versus 67.22 dB; P < 0.05). These mildly infected animals showed significantly reduced cochlear fibrous occlusion (P < 0.01). By combining nonbacteriolytic daptomycin and anti-inflammatory doxycycline with ceftriaxone, the previously reported beneficial effects of the drugs were cumulated and identified the triple-antibiotic therapy as a promising therapeutic option for pediatric PM.

6.
J Neurointerv Surg ; 11(10): 1050-1054, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30852526

RESUMO

BACKGROUND: Advances in stent-assisted coiling have incrementally expanded endovascular treatment options for complex cerebral aneurysms. After successful coil consolidation and aneurysm occlusion, endovascular scaffolds are no longer needed. Thus, bioresorbable stents that disappear after aneurysm healing could avoid future risks of in-stent thrombosis and the need for lifelong antiplatelet therapy. OBJECTIVE: To assess the applicability and compatibility of a bioresorbable magnesium- alloy stent (brMAS) for assisted coiling. METHODS: Saccular sidewall aneurysms were created in 84 male Wistar rats and treated with brMAS alone, brMAS + aspirin, or brMAS + coils + aspirin. Control groups included no treatment (natural course), solely aspirin treatment, or conventional cobalt-chromium stent + coils + aspirin treatment. After 1 and 4 weeks, aneurysm specimens were harvested and macroscopically, histologically, and molecularly examined for healing, parent artery perfusion status, and inflammatory reactions. Stent degradation was monitored for up to 6 months with micro-computed and optical coherence tomography. RESULTS: Aneurysms treated with brMAS showed advanced healing, neointima formation, and subsequent stent degradation. Additional administration of aspirin sustained aneurysm healing while reducing stent-induced intraluminal and periadventitial inflammatory responses. No negative interaction was detected between platinum coils and brMAS. Progressive brMAS degradation was confirmed. CONCLUSIONS: brMAS induced appropriate healing in this sidewall aneurysm model. The concept of using bioresorbable materials to promote complete aneurysm healing and subsequent stent degradation seems promising. These results should encourage further device refinements and clinical evaluation of this treatment strategy for cerebrovascular aneurysms.

7.
Infect Immun ; 87(4)2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30718288

RESUMO

In serogroup C Neisseria meningitidis, the cssA (siaA) gene codes for an UDP-N-acetylglucosamine 2-epimerase that catalyzes the conversion of UDP-N-acetyl-α-d-glucosamine into N-acetyl-d-mannosamine and UDP in the first step in sialic acid biosynthesis. This enzyme is required for the biosynthesis of the (α2→9)-linked polysialic acid capsule and for lipooligosaccharide (LOS) sialylation. In this study, we have used a reference serogroup C meningococcal strain and an isogenic cssA knockout mutant to investigate the pathogenetic role of surface-exposed sialic acids in a model of meningitis based on intracisternal inoculation of BALB/c mice. Results confirmed the key role of surface-exposed sialic acids in meningococcal pathogenesis. The 50% lethal dose (LD50) of the wild-type strain 93/4286 was about four orders of magnitude lower than that of the cssA mutant. Compared to the wild-type strain, the ability of this mutant to replicate in brain and spread systemically was severely impaired. Evaluation of brain damage evidenced a significant reduction in cerebral hemorrhages in mice infected with the mutant in comparison with the levels in those challenged with the wild-type strain. Histological analysis showed the typical features of bacterial meningitis, including inflammatory cells in the subarachnoid, perivascular, and ventricular spaces especially in animals infected with the wild type. Noticeably, 80% of mice infected with the wild-type strain presented with massive bacterial localization and accompanying inflammatory infiltrate in the corpus callosum, indicating high tropism of meningococci exposing sialic acids toward this brain structure and a specific involvement of the corpus callosum in the mouse model of meningococcal meningitis.


Assuntos
Proteínas de Bactérias/genética , Meningite Meningocócica/microbiologia , Ácido N-Acetilneuramínico/metabolismo , Neisseria meningitidis Sorogrupo C/patogenicidade , Animais , Proteínas de Bactérias/metabolismo , Encéfalo/microbiologia , Encéfalo/patologia , Carboidratos Epimerases/genética , Carboidratos Epimerases/metabolismo , Modelos Animais de Doenças , Feminino , Técnicas de Inativação de Genes , Humanos , Meningite Meningocócica/mortalidade , Meningite Meningocócica/patologia , Camundongos , Camundongos Endogâmicos BALB C , Neisseria meningitidis Sorogrupo C/genética , Neisseria meningitidis Sorogrupo C/metabolismo , Virulência
8.
PLoS One ; 13(10): e0205294, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30300398

RESUMO

Neurotropic tick borne encephalitis virus (TBEV) causes life-threatening disease, and accounts for most cases of tick-transmitted viral infections in Central and Eastern Europe and Russia. No specific treatment for TBEV infections exists, and vaccination is recommended for people at risk. So far, various nucleoside analogues have been investigated in vitro as potential candidates for treatment of TBEV infections. However, in vitro experiments with more complex cell culture systems, such as organotypic culture slices which model the sophisticated architecture of the target tissue are lacking. Using TBEV as a model, we investigated the suitability of rat organotypic cerebellum slices (OCS) to study the effectiveness of nucleoside analogues with a well-known anti-TBEV activity. In these OCS, 50 µM of the nucleoside analogues 2'-C-methyladenosine (2'-CMA) and especially 7-deaza-2'-C-methyladenosine (7-deaza-2'-CMA) exhibited strong inhibitory effects on TBEV replication, reducing viral titers to an average of 103-fold and TBEV RNA content 60-90-fold. In contrast, the influence of 2'-C-methylcytidine (2'-CMC) on TBEV replication was very weak, reducing virus titers by 10-fold and TBEV RNA content by 3-fold. In agreement with other studies, there was no noticeable difference in TBEV titers between OCS treated with 50 µM of Ribavirin and the DMSO treated controls. All tested nucleoside analogues exhibited excellent cytotoxicity profiles at concentrations of 50 µM. Our findings in OCS were highly comparable to data obtained in cell line culture systems. Therefore, OCS represent an ideal in vitro approach to study antivirals against TBEV and possibly other neurotropic viruses.

9.
J Neuroinflammation ; 15(1): 233, 2018 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-30131074

RESUMO

BACKGROUND: Pneumococcal meningitis is associated with high mortality and morbidity rates. Up to 50% of survivors show neurologic sequelae including hearing loss, cognitive impairments and learning disabilities, being particularly detrimental in affected infants and children where adjuvant therapy with dexamethasone has no proven beneficial effect. We evaluated the effect of concomitantly targeting specific pathophysiological mechanisms responsible for brain damage-i.e. matrix-metalloproteinase (MMP) activity and the exacerbated cerebral inflammation provoked through antibiotic-induced bacterial lysis. Here, we combined adjunctive therapies previously shown to be neuroprotective when used as single adjuvant therapies. METHODS: Eleven-day-old Wistar rats were infected intracisternally with 6.44 ± 2.17 × 103 CFU Streptococcus pneumoniae and randomised for treatment with ceftriaxone combined with (a) single adjuvant therapy with daptomycin (n = 24), (b) single adjuvant therapy with Trocade (n = 24), (c) combined adjuvant therapy (n = 66) consisting of daptomycin and Trocade, or (d) ceftriaxone monotherapy (n = 42). Clinical parameters and inflammatory CSF cytokine levels were determined during acute meningitis. Cortical damage and hippocampal apoptosis were assessed 42 h after infection. Morris water maze and auditory brainstem responses were used to assess neurofunctional outcome 3 weeks after infection. RESULTS: We found significantly reduced apoptosis in the hippocampal subgranular zone in infant rats receiving adjuvant Trocade (p < 0.01) or combined adjuvant therapy (p < 0.001). Cortical necrosis was significantly reduced in rats treated with adjuvant daptomycin (p < 0.05) or combined adjuvant therapy (p < 0.05) compared to ceftriaxone monotherapy. Six hours after treatment initiation, CSF cytokine levels were significantly reduced for TNF-α (p < 0.01), IL-1ß (p < 0.01), IL-6 (p < 0.001) and IL-10 (p < 0.01) in animals receiving combined adjuvant intervention compared to ceftriaxone monotherapy. Importantly, combined adjuvant therapy significantly improved learning and memory performance in infected animals and reduced hearing loss (77.14 dB vs 60.92 dB, p < 0.05) by preserving low frequency hearing capacity, compared to ceftriaxone monotherapy. CONCLUSION: Combined adjuvant therapy with the non-bacteriolytic antibiotic daptomycin and the MMP inhibitor Trocade integrates the neuroprotective effects of both single adjuvants in experimental paediatric pneumococcal meningitis by reducing neuroinflammation and brain damage, thereby improving neurofunctional outcome. This strategy represents a promising therapeutic option to improve the outcome of paediatric patients suffering from pneumococcal meningitis.


Assuntos
Antibacterianos/uso terapêutico , Lesões Encefálicas/prevenção & controle , Transtornos da Audição/prevenção & controle , Transtornos de Aprendizagem/tratamento farmacológico , Inibidores de Metaloproteinases de Matriz/uso terapêutico , Transtornos da Memória/tratamento farmacológico , Animais , Animais Recém-Nascidos , Apoptose/efeitos dos fármacos , Lesões Encefálicas/etiologia , Ceftriaxona/uso terapêutico , Córtex Cerebral/patologia , Citocinas/líquido cefalorraquidiano , Daptomicina/uso terapêutico , Modelos Animais de Doenças , Quimioterapia Combinada , Potenciais Evocados Auditivos do Tronco Encefálico , Transtornos da Audição/etiologia , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Transtornos de Aprendizagem/etiologia , Metaloproteinases da Matriz/metabolismo , Aprendizagem em Labirinto/efeitos dos fármacos , Transtornos da Memória/etiologia , Meningite Pneumocócica/complicações , Ratos , Streptococcus pneumoniae/patogenicidade
10.
Front Microbiol ; 9: 919, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29867830

RESUMO

The rate of invasive group B Streptococcus (GBS) infections is steadily increasing, particularly in older persons and in adults with diabetes and other comorbidities. This population includes persons with a foreign body (e.g., who have undergone arthroplasty). In a rat tissue cage model, we evaluated the efficacy of adjunctive gentamicin (GEN) administered systemically (5 mg/kg body weight) every 24 h, or locally (12.5 mg/L tissue cage concentration) every 24 or 72 h, in combination with penicillin (PEN) administered systemically (250,000 IU/kg body weight three times per day). The efficacy was evaluated on two different sessile forms of GBS: transition (i.e., in between planktonic and biofilm) and biofilm. After 3 days of treatment, the mean bacterial load reduction of transition-form GBS was greater in all PEN-GEN combination groups than in the PEN monotherapy group (P ≤ 0.03). The 6-day regimen decreased the bacterial load significantly in comparison to the 3-day regimen, irrespective of growth form and adjunctive GEN (P < 0.01). After 6 days of treatment, the mean reduction in transition-form GBS was greater with PEN plus GEN administered locally every 24 h than with PEN monotherapy (P = 0.03). These results were not confirmed with biofilm GBS. The difference in mean bacterial load reduction between all PEN-GEN and PEN monotherapy groups was <100 CFU/mL. Hence, synergy criteria were not fulfilled. Adjunctive systemic GEN consists of potential side effects and showed poor efficacy in this study. Combining systemic PEN and local GEN has a potential application in the treatment of streptococcal implant-associated infections.

11.
BMC Microbiol ; 18(1): 23, 2018 03 27.
Artigo em Inglês | MEDLINE | ID: mdl-29580217

RESUMO

BACKGROUND: Nonencapsulated Streptococcus pneumoniae bacteria are successful colonizers of the human nasopharynx and often possess genes aliB-like ORF 1 and 2 in place of capsule genes. AliB-like ORF 2 binds peptide FPPQSV, found in Prevotella species, resulting in enhanced colonization. How this response is mediated is so far unknown. RESULTS: Here we show that the peptide increases expression of genes involved in release of host carbohydrates, carbohydrate uptake and carbohydrate metabolism. In particular, the peptide increased expression of 1,5-anhydro-D-fructose reductase, a metabolic enzyme of an alternative starch and glycogen degrading pathway found in many organisms, in both transcriptomic and proteomic data. The peptide enhanced pneumococcal growth giving a competitive advantage to a strain with aliB-like ORF 2, over its mutant lacking the gene. Possession of aliB-like ORF 2 did not affect release of inflammatory cytokine CXCL8 from epithelial cells in culture and the nonencapsulated wild type strain was not able to establish disease or inflammation in an infant rat model of meningitis. CONCLUSIONS: We propose that AliB-like ORF 2 confers an advantage in colonization by enhancing carbohydrate metabolism resulting in a boost in growth. This may explain the widespread presence of aliB-like ORF 2 in the nonencapsulated pneumococcal population in the human nasopharynx.

12.
Shock ; 49(2): 229-234, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28562478

RESUMO

BACKGROUND: The small molecule pifithrin-µ reversibility inhibits the mitochondrial pathway of apoptosis. The neuronal effects of pifithrin-µ applied after cardiac arrest are unknown. We hypothesized that pifithrin-µ reduces neuronal damage in the most vulnerable brain region, the hippocampus, after cardiac arrest. METHODS: In two randomized controlled series we administered pifithrin-µ or control in 109 rats resuscitated after 8 or 10 min of cardiac arrest. Neuronal damage was blindly assessed with histology (Fluoro Jade B: FJB, cresyl violet: CV) in the most vulnerable brain region (CA1 segment of hippocampus) and with a series of neurobehavioral tests (Open Field Task, Tape-Removal Test, Morris Water Maze test). Mixed ANOVA was used to combine both series, simple comparisons were done with t tests or Mann-Whitney U test. RESULTS: Pifithrin-µ reduced the number of degenerating, FJB-positive neurons by 25% (mixed ANOVA p group = 0.014). This was more prominent after 8 min cardiac arrest (8 min arrest pifithrin-µ 94 ±â€Š47 vs control 128 ±â€Š37; n = 11 each; 10 min arrest pifithrin-µ 78 ±â€Š44, n = 15 vs control 101 ±â€Š31, n = 18; p group* arrest length interaction = 0.622). The reduction of ischemic CV-positive neurons in pifithrin-µ animals was not significant (ANOVA p group = 0.063). No significant group differences were found in neurobehavioral testing. CONCLUSION: Temporarily inhibition of apoptosis with pifithrin-µ after cardiac arrest decreases the number of injured neurons in the CA1 segment of hippocampus in a cardiac arrest rat model, without clinical correlate. Further studies should elucidate the role of this neuroprotective agent in different settings and with longer cardiac arrest.

13.
Curr Opin Infect Dis ; 31(1): 85-92, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29095719

RESUMO

PURPOSE OF REVIEW: Pneumococcal meningitis is the most frequent form of bacterial meningitis in Europe and the United States. Although early antimicrobial and adjuvant therapy with dexamethasone have helped to improve disease outcome in adults, mortality and morbidity rates remain unsatisfactorily high, emphasizing the need for additional treatment options. Promising targets for adjuvant therapy have been identified recently and will be the focus of this review. RECENT FINDINGS: Brain disease in pneumococcal meningitis is caused by direct bacterial toxicity and excessive meningeal inflammation. Accordingly, promising targets for adjuvant therapy comprise limiting the release of toxic bacterial products and suppressing inflammation in a way that maximally protects against tissue injury without hampering pathogen eradication by antibiotics. Among the agents tested so far in experimental models, complement inhibitors, matrix-metalloproteinase inhibitors, and nonbacteriolytic antibiotics or a combination of the above have the potential to more efficiently protect the brain either alone (e.g., in children and outside the high-income settings) or in addition to adjuvant dexamethasone. Additionally, new protein-based pneumococcal vaccines are being developed that promise to improve disease prevention, namely by addressing the increasing problem of serotype replacement seen with pneumococcal conjugate vaccines. SUMMARY: Pneumococcal meningitis remains a life-threatening disease requiring early antibiotic and targeted anti-inflammatory therapy. New adjuvant therapies showed promising results in animal models but need systematic clinical testing.

14.
Hear Res ; 350: 100-109, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28460251

RESUMO

Hearing loss remains the most common long-term complication of pneumococcal meningitis (PM) reported in up to 30% of survivors. Streptococcus pneumoniae have been shown to possess different ototoxic properties. Here we present a novel ex vivo experimental setup to examine in detail the pattern of hair cell loss upon exposure to different S. pneumoniae strains, therefore recapitulating pathogen derived aspects of PM-induced hearing loss. Our results show a higher susceptibility towards S. pneumoniae-induced cochlear damage for outer hair cells (OHC) compared to inner hair cells (IHC), which is consistent with in vivo data. S. pneumoniae-induced hair cell loss was both time and dose-dependent. Moreover, we have found significant differences in the level of cell damage between tissue from the basal and the apical turns. This shows that the higher vulnerability of hair cells located at high frequency regions observed in vivo cannot be explained solely by the spatial organisation and bacterial infiltration from the basal portion of the cochlea. Using a wild type D39 strain and a mutant defective for the pneumolysin (PLY) gene, we also have shown that the toxin PLY is an important factor involved in ototoxic damages. The obtained results indicate that PLY can cause both IHC and OHC loss. Finally, we are reporting here for the first time a higher vulnerability of HC located at the basal and middle cochlear region to pneumolysin-induced damage. The detailed description of the susceptibility of hair cells to Streptococcus pneumoniae provided in this report can in the future determine the choice and the development of novel otoprotective therapies during pneumococcal meningitis.


Assuntos
Perda Auditiva/microbiologia , Meningite Pneumocócica/microbiologia , Órgão Espiral/microbiologia , Streptococcus pneumoniae/patogenicidade , Animais , Animais Recém-Nascidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sobrevivência Celular , Genótipo , Células Ciliadas Auditivas Internas/microbiologia , Células Ciliadas Auditivas Internas/patologia , Células Ciliadas Auditivas Externas/microbiologia , Células Ciliadas Auditivas Externas/patologia , Perda Auditiva/patologia , Meningite Pneumocócica/patologia , Mutação , Órgão Espiral/patologia , Ratos Wistar , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/metabolismo , Estreptolisinas/genética , Estreptolisinas/metabolismo , Técnicas de Cultura de Tecidos , Virulência
15.
Front Neurol ; 8: 119, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28421031

RESUMO

Tick-borne encephalitis is a viral disease affecting the central nervous system. It is endemic in Switzerland with 200-250 notified cases annually. Active immunization is effective for persons in all age groups. Vaccine failure is rare, in particular after a completed vaccination course. Here, we describe the case of 67-year-old man with a fatal outcome despite vaccination. The diagnosis was confirmed by extensive postmortem analyses. The diagnostic challenges of vaccine failure in tick-borne encephalitis and the dynamics of the immune response in vaccination breakthrough are discussed.

16.
Front Pharmacol ; 8: 73, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28280466

RESUMO

Clostridium botulinum neurotoxins (BoNTs) are the most poisonous naturally occurring protein toxins known to mankind and are the causative agents of the severe and potentially life-threatening disease botulism. They are also known for their application as cosmetics and as unique bio-pharmaceuticals to treat an increasing number of neurological and non-neurological disorders. Currently, the potency of biologically active BoNT for therapeutic use is mainly monitored by the murine LD50-assay, an ethically disputable test causing suffering and death of a considerable number of mice. The aim of this study was to establish an in vitro assay as an alternative to the widely used in vivo mouse bioassay. We report a novel BoNT detection assay using mouse embryonic stem cell-derived neurons (mESN) cultured on multi-electrode arrays (MEAs). After 21 days in culture, the mESN formed a neuronal network showing spontaneous bursting activity based on functional synapses and express the necessary target proteins for BoNTs. Treating cultures for 6 h with 16.6 pM of BoNT serotype A and incubation with 1.66 pM BoNT/A or 33 Units/ml of Botox® for 24 h lead to a significant reduction of both spontaneous network bursts and average spike rate. This data suggests that mESN cultured on MEAs pose a novel, biologically relevant model that can be used to detect and quantify functional BoNT effects, thus accelerating BoNT research while decreasing animal use.

17.
J Neurosci ; 36(29): 7740-9, 2016 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-27445150

RESUMO

UNLABELLED: Hearing loss is an important sequela of pneumococcal meningitis (PM), occurring in up to 30% of survivors. The role of the severity of infection on hearing function and pathomorphological consequences in the cochlea secondary to PM have not been investigated to date. Using a well-established model of PM, we systematically investigated the functional hearing outcome and the long-term fate of neurosensory cells in the cochlea, i.e., hair cells and spiral ganglion neurons (SGNs), with a focus on their tonotopic distribution. Intracisternal infection of infant rats with increasing inocula of Streptococcus pneumoniae resulted in a dose-dependent increase in CSF levels of interleukin-1ß, interleukin-6, tumor necrosis factor α, interleukin-10, and interferon-γ in acute disease. The severity of long-term hearing loss at 3 weeks after infection, measured by auditory brainstem response recordings, correlated to the initial inoculum dose and to the levels of proinflammatory cytokines determined in the acute phase of PM. Quantitative cochlear histomorphology revealed a significant loss of SGNs and outer hair cells that strongly correlated to the level of infection, with the most severe damage occurring in the basal part of the cochlea. Inner hair cells (IHCs) were not significantly affected throughout the entire cochlea. However, surviving IHCs lost synaptic connectivity to remaining SGNs in all cochlear regions. These findings provide evidence that the inoculum concentration, i.e., severity of infection, is the major determinant of long-term morphological cell pathologies in the cochlea and functional hearing loss. SIGNIFICANCE STATEMENT: Hearing loss is a neurofunctional deficit occurring in up to 30% of patients surviving pneumococcal meningitis (PM). Here, we analyze the correlation between the severity of infection and the inflammatory response in the CSF, the tonotopic distribution of neurosensory pathologies in the cochlea, and the long-term hearing function in a rat model of pneumococcal meningitis. Our study identifies the severity of infection as the key determinant of long-term hearing loss, underlining the importance of the prompt institution of antibiotic therapy in patients suffering from PM. Furthermore, our findings reveal in detail the spatial loss of cochlear neurosensory cells, providing new insights into the pathogenesis of meningitis-associated hearing loss that reveal new starting points for the development of otoprotective therapies.


Assuntos
Citocinas/líquido cefalorraquidiano , Perda Auditiva Neurossensorial/etiologia , Perda Auditiva Neurossensorial/patologia , Meningite Pneumocócica/complicações , Neurônios/metabolismo , Estimulação Acústica , Animais , Animais Recém-Nascidos , Cóclea/patologia , Citocinas/genética , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Potenciais Evocados Auditivos do Tronco Encefálico/fisiologia , Feminino , Células Ciliadas Auditivas/metabolismo , Humanos , Técnicas In Vitro , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurônios/patologia , Fosfoproteínas/metabolismo , Ratos , Ratos Wistar , Gânglio Espiral da Cóclea/patologia , Streptococcus pneumoniae/fisiologia
18.
BMC Genomics ; 17: 491, 2016 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-27411385

RESUMO

BACKGROUND: Biocides and antibiotics are used to eradicate or prevent the growth of microbial species on surfaces (occasionally on catheters), or infected sites, either in combination or sequentially, raising concerns about the development of co-resistance to both antimicrobial types. The effect of such compounds on Salmonella enterica, a major food-borne and zoonotic pathogen, has been analysed in different studies, but only few works evaluated its biological cost, and the overall effects at the genomic and transcriptomic levels associated with diverse phenotypes resulting from biocide exposure, which was the aim of this work. RESULTS: Exposure to triclosan, clorhexidine, benzalkonium, (but not to hypochlorite) resulted in mutants with different phenotypes to a wide range of antimicrobials even unrelated to the selective agent. Most biocide-resistant mutants showed increased susceptibility to compounds acting on the cell wall (ß-lactams) or the cell membranes (poly-L-lysine, polymyxin B, colistin or toxic anions). Mutations (SNPs) were found in three intergenic regions and nine genes, which have a role in energy production, amino acids, carbohydrates or lipids metabolism, some of them involved in membrane transport and pathogenicity. Comparative transcriptomics of biocide-resistant mutants showed over-expression of genes encoding efflux pumps (sugE), ribosomal and transcription-related proteins, cold-shock response (cpeE) and enzymes of microaerobic metabolism including those of the phosphotransferase system. Mainly ribosomal, metabolic and pathogenicity-related genes had affected expression in both in vitro-selected biocide mutants and field Salmonella isolates with reduced biocide susceptibility. CONCLUSIONS: Multiple pathways can be involved in the adaptation of Salmonella to biocides, mainly related with global stress, or involving metabolic and membrane alterations, and eventually causing "collateral sensitivity" to other antimicrobials. These changes might impact the bacterial-environment interaction, imposing significant bacterial fitness costs which may reduce the chances of fixation and spread of biocide resistant mutants.


Assuntos
Adaptação Biológica , Antibacterianos/farmacologia , Desinfetantes/farmacologia , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/fisiologia , Análise por Conglomerados , Farmacorresistência Bacteriana , Perfilação da Expressão Gênica , Genes Bacterianos , Aptidão Genética , Genoma Bacteriano , Sequenciamento de Nucleotídeos em Larga Escala , Testes de Sensibilidade Microbiana , Mutação
19.
BMC Microbiol ; 16(1): 154, 2016 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-27430279

RESUMO

BACKGROUND: Streptococcus pneumoniae causes several human diseases, including pneumonia and meningitis, in which pathology is associated with an excessive inflammatory response. A major inducer of this response is the cholesterol dependent pneumococcal toxin, pneumolysin. Here, we measured the amount of inflammatory cytokine CXCL8 (interleukin (IL)-8) by ELISA released by human nasopharyngeal epithelial (Detroit 562) cells as inflammatory response to a 24 h exposure to different pneumococcal strains. RESULTS: We found pneumolysin to be the major factor influencing the CXCL8 response. Cholesterol and sphingomyelin-containing liposomes designed to sequester pneumolysin were highly effective at reducing CXCL8 levels from epithelial cells exposed to different clinical pneumococcal isolates. These liposomes also reduced CXCL8 response from epithelial cells exposed to pneumolysin knock-out mutants of S. pneumoniae indicating that they also reduce the CXCL8-inducing effect of an unidentified pneumococcal virulence factor, in addition to pneumolysin. CONCLUSION: The results indicate the potential of liposomes in attenuating excessive inflammation as a future adjunctive treatment of pneumococcal diseases.


Assuntos
Células Epiteliais/metabolismo , Interleucina-8/metabolismo , Lipossomos/farmacologia , Nasofaringe/metabolismo , Streptococcus pneumoniae/metabolismo , Cápsulas Bacterianas , Proteínas de Bactérias/genética , Proteínas de Bactérias/farmacologia , Linhagem Celular , Células Cultivadas , Colesterol/farmacologia , Citocinas/metabolismo , Células Epiteliais/efeitos dos fármacos , Humanos , Mutação , Nasofaringe/efeitos dos fármacos , Esfingomielinas/farmacologia , Streptococcus pneumoniae/isolamento & purificação , Streptococcus pneumoniae/patogenicidade , Estreptolisinas/genética , Estreptolisinas/farmacologia
20.
BMC Neurol ; 16: 43, 2016 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-27044425

RESUMO

BACKGROUND: The noble gas xenon is considered as a neuroprotective agent, but availability of the gas is limited. Studies on neuroprotection with the abundant noble gases helium and argon demonstrated mixed results, and data regarding neuroprotection after cardiac arrest are scant. We tested the hypothesis that administration of 50% helium or 50% argon for 24 h after resuscitation from cardiac arrest improves clinical and histological outcome in our 8 min rat cardiac arrest model. METHODS: Forty animals had cardiac arrest induced with intravenous potassium/esmolol and were randomized to post-resuscitation ventilation with either helium/oxygen, argon/oxygen or air/oxygen for 24 h. Eight additional animals without cardiac arrest served as reference, these animals were not randomized and not included into the statistical analysis. Primary outcome was assessment of neuronal damage in histology of the region I of hippocampus proper (CA1) from those animals surviving until day 5. Secondary outcome was evaluation of neurobehavior by daily testing of a Neurodeficit Score (NDS), the Tape Removal Test (TRT), a simple vertical pole test (VPT) and the Open Field Test (OFT). Because of the non-parametric distribution of the data, the histological assessments were compared with the Kruskal-Wallis test. Treatment effect in repeated measured assessments was estimated with a linear regression with clustered robust standard errors (SE), where normality is less important. RESULTS: Twenty-nine out of 40 rats survived until day 5 with significant initial deficits in neurobehavioral, but rapid improvement within all groups randomized to cardiac arrest. There were no statistical significant differences between groups neither in the histological nor in neurobehavioral assessment. CONCLUSIONS: The replacement of air with either helium or argon in a 50:50 air/oxygen mixture for 24 h did not improve histological or clinical outcome in rats subjected to 8 min of cardiac arrest.


Assuntos
Argônio/administração & dosagem , Parada Cardíaca/complicações , Hélio/administração & dosagem , Fármacos Neuroprotetores/farmacologia , Animais , Hipocampo/patologia , Masculino , Neuroproteção/efeitos dos fármacos , Gases Nobres/administração & dosagem , Oxigênio/administração & dosagem , Ratos , Ratos Wistar
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