Montelukast, Leukotriene Inhibitor, Reduces LPS-Induced Acute Lung Inflammation and Human Neutrophil Activation / El montelukast, un inhibidor de leucotrienos, reduce la inflamación pulmonar aguda inducida por LPS y la activación de neutrófilos humanos

Objectives: Some pro-inflammatory lipids derived from 1 lipooxygenase enzyme are potent neutrophil chemoattractant, a cell centrally involved in acute respiratory distress syndrome (ARDS); a syndrome lacking effective treatment. Considering the beneficial effects of the leukotriene receptor inhibitor, montelukast, on other lung diseases, whether montelukast attenuates inflammation in a mouse model of ARDS, and whether it reduces LPS stimulated activation of human neutrophils was investigated. Methods: Thirty-five C57Bl/6 mice were distributed into control (PBS) + 24h, LPS + 24h (10 μg/mouse), control + 48 h, LPS+48 h, and LPS 48 h+Montelukast (10 mg/kg). In addition, human neutrophils were incubated with LPS ( 1μg/mL) and treated with montelukast (10 μM). Results: Oral-tracheal administration of montelukast significantly attenuated total cells (P < .05), macrophages (P < .05), neutrophils (P < .01), lymphocytes (P < .001) and total protein levels in BAL (P < .05), as well as IL-6 (P < .05), CXCL1/KC (P < .05), IL-17 (P < .05) and TNF-alfa (P < .05). Furthermore, montelukast reduced neutrophils (P < .001), lymphocytes (P < .01) and macrophages (P < .01) in the lung parenchyma. In addition, montelukast restored BAL VEGF levels (P < .05). LTB4 receptor expression (P < .001) as well as NF-κB (P <. 001), a downstream target of LPS, were also reduced in lung parenchymal leukocytes. Furthermore, montelukast reduced IL-8 (P < .001) production by LPS-treated human neutrophils. Conclusion: In conclusion, montelukast efficiently attenuated both LPS-induced lung inflammation in a mouse model of ARDS and in LPS challenged human neutrophils

Objetivos: Algunos lípidos proinflamatorios derivados de la enzima lipooxigenasa 1 son potentes quimioatrayentes de neutrófilos, un tipo celular con una implicación principal en el síndrome de distrés respiratorio agudo (SDRA), para el que no hay tratamiento efectivo. Considerando los efectos beneficiosos del inhibidor de los receptores de leucotrienos montelukast en otras enfermedades pulmonares, se investigó si este fármaco era capaz de atenuar la inflamación en un modelo de ratón de SDRA y de reducir la activación de los neutrófilos humanos inducida por LPS. Métodos: Se utilizaron 35 ratones C57BL/6 distribuidos en los siguientes grupos: control (PBS) + 24 h, LPS+(24 h [10 μg/ratón]), control + 48 h y LPS 48 h + montelukast (10 mg/kg). Por otro lado, se incubaron neutrófilos humanos con LPS (1 μg/ml) y se trataron con montelukast (10 μM). Resultados: La administración orotraqueal de montelukast redujo el número total de células (p < 0,05), de macrófagos (p < 0,05), de neutrófilos (p < 0,01), de linfocitos (p < 0,001) y los niveles totales de proteína en el lavado broncoalveolar (p < 0,05), así como de IL-6 (p < 0,05), CXCL1/KC (p < 0,05), IL-17 (p < 0,05) y TNF-alfa (p < 0,05). Además, el montelukast redujo los neutrófilos (p < 0,001), los linfocitos (p < 0,01) y los macrófagos (p < 0,01) en el parénquima pulmonar. Asimismo, restauró los niveles de VEGF en el lavado broncoalveolar (p < 0,05) y disminuyó la expresión del receptor LTB4 (p < 0,001) y de NF-κB (p < 0,001), una diana downstream del LPS, en los leucocitos del parénquima pulmonar. Por último, redujo la producción de IL-8 por parte de los neutrófilos humanos tratados con LPS. Conclusión: En conclusión, el montelukast atenuó de manera eficaz tanto la inflamación pulmonar inducida por LPS en un modelo de ratón de SDRA como en neutrófilos humanos estimulados con LPS

Clinical and hematological changes in sheep induced by Escherichia coli lipopolysaccharide / alterações clínicas e hematológicas induzidas pela administração de lipopolisscarídeo de Escherichia coli em ovelhas / Alteraciones clínicas y hematológicas inducidas por la administración de lipopoliscárido de Escherichia coli en ovejas

Organic response to infection is characterized by a systemic reaction known as acute phase response (APR). In order to know the effect of the administration of Escherichia coli lipopolysaccharide (LPS) on physiological, hematological and biochemical variables, 10 sheep weighing 45 ± 5 kg were divided in two groups: Experimental group treated with 3 doses of 1 µg.kg-1 LPS and control group treated with saline solution (SS) at the same frequency as experimental group. Body temperature (BT°), heart rate (HR) and respiratory rate (RR) were monitored. Blood samples for hemogram and enzyme activity for aspartate amino transferase (AST) and gamma glutamyl transferase (GGT) were collected between 1 and 24 h post-LPS. LPS-treated sheep presented mean values of BT (41.2 ± 0.4°C), HR (132 ± 12.3 beats/min) and RR (107.2 ± 25 cycles/min) higher than those observed in control sheep (39.8 ± 0.2°C, 88.8 ± 8.7 beats/min and 53.6 ± 17.1 cycles/min respectively). Between 4 and 8 hours post-injection (hpi) of LPS the leukocyte count was associated with lymphopenia, followed by leukocytosis at 24 hours. No changes were observed in the activity of AST and GGT enzymes. The results characterize APR induced by LPS in sheep, representing a useful model to study cardiovascular, hematological and biochemical responses to infection. (AU)

A resposta orgânica à infecção é caracterizada por uma reação sistêmica conhecida como resposta de fase aguda (RFA). Para conhecer o efeito da administração de lipopolissacárideo (LPS) de Escherichia coli sobre as variáveis fisiológicas, hematológicas e bioquímicas, 10 carneiros pesando 45 ± 5 kg foram divididos em dois grupos: o grupo experimental tratado com três doses de 1 µg.kg -1 de LPS e grupo controle tratado com solução salina (SS) na mesma frequência que o grupo experimental. Temperatura corporal (T°C), frequência cardíaca (FC) e frequência respiratória (FR) foram monitoradas. As amostras de sangue foram tomadas para hemograma e atividade da enzima aspartato aminotransferase (AST) e gama- -glutamiltransferase (GGT) entre uma e 24 h pós-LPS. Ovelhas tratadas com LPS apresentaram valores médios de T°C (41,2 ± 0,4°C), FC (132 ± 12,3 batimentos/min) e FR (107,2 ± 25 ciclos/min) acima dos observados em ovelhas do tratamento controle (39,8 ± 0,2°C, 88,8 ± 8,7 batimentos/min e 53,6 ± 17,1 ciclos/min, respectivamente). Entre 4 e 8 horas após a injeção de LPS, a contagem de leucocitos foi asociado com linfopenia, seguida de leucocitose as 24 horas. Nenhuma mudança na atividade das enzimas AST e GGT foi observada. Os resultados caracterizam uma resposta de fase aguda induzida por LPS em ovelhas, o que representa um modelo útil para estudar os sistemas cardiovascular, hematológico e bioquímico em resposta à infecção.(AU)

La respuesta orgánica a la infección se caracteriza por una reacción sistémica conocida como respuesta de fase aguda (RFA). Para conocer el efecto de la administración de lipopolisacárido (LPS) de Escherichia coli sobre las variables fisiológicas, hematológicas y bioquímicas, 10 ovejas con un peso de 45 ± 5 kg se dividieron en dos grupos: grupo experimental tratado con 3 dosis de 1 µg.kg-1de LPS y grupo control tratado con solución salina (SS) en la misma frecuencia que el grupo experimental. Temperatura corporal (T°C), frecuencia cardíaca (FC) y frecuencia respiratoria (FR) fueron monitoreadas. Se tomaron muestras de sangre para hemograma y actividad enzimática para el aspartato amino transferasa (AST) y gamma glutamil transferasa (GGT) entre 1 y 24 h post-LPS. Las ovejas tratadas con LPS presentaron valores medios de T°C (41.2 ± 0.4°C), FC (132 ± 12.3 latidos / min) y FR (107.2 ± 25 ciclos/min) por encima de los observados en ovinos controles (39.8 ± 0.2°C, 88.8 ± 8.7 latidos/min y 53.6 ± 17.1 ciclos/min respectivamente). Entre las 4 y 8 horas después de la inyección de LPS, el recuento de leucocitos se asoció a linfopenia, seguida de leucocitosis a las 24 horas. No se observaron cambios en la actividad de las enzimas AST y GGT. Los resultados caracterizan una respuesta de fase aguda inducida por LPS en ovinos, representando un modelo útil para estudiar los sistemas cardiovascular, hematológico y bioquímico en respuesta a la infección.(AU)

Alpha-lactalbumin in human milk alters the proteolytic degradation of soluble CD14 by forming a complex.

Mother's milk represents a foundational step in the proper development of newborn immunity. This is achieved, in part, through the action of numerous regulatory proteins such as soluble cluster of differentiation 14 (sCD14) found in significant quantities in human milk (~25-50 µg/mL). In adults, CD14 stimulates cytokine production in response to lipopolysaccharide (LPS), the major lipid component found in the outer membrane of Gram-negative bacteria. However, the fate and function of sCD14 in the neonatal gastrointestinal (GI) tract are unknown and may function differently from adults. Therefore, we administered human sCD14 to experimental animals and observed that it persisted in the upper GI tract after feeding. In our search for potential proteolytic protectants, immunoprecipitation of sCD14 from human milk revealed a 15-kD novel protein that copurified with sCD14. Mass spectrometry analysis of the protein identified alpha-lactalbumin. CD14 was also identified by immunoblot after immunoprecipitation of alpha-lactalbumin from milk. In vitro digestion assays revealed that purified alpha-lactalbumin decreases the proteolytic degradation of human milk derived sCD14 in vitro, suggesting a mechanism by which this key LPS receptor may remain functional in the neonate gut.

Increased voluntary exercise in mice deficient for tumour necrosis factor-alpha and lymphotoxin-alpha.

BACKGROUND: The endogenous mediators playing a role in the sensing of fatigue and cessation of exercise are yet to be characterized. We hypothesized that proinflammatory cytokines, in particular tumour necrosis factor-alpha (TNFalpha) and lymphotoxin-alpha (LT) transmit signals leading to fatigue. MATERIALS AND METHODS: Mice were placed in a cage with a freely rotating exercise wheel and allowed to adapt for 24 h. The running distance was measured for two additional periods of 24 h. The effects of the administration of intravenous anti-TNF antibodies, intracerebral recombinant TNF, or intravenous lipopolysaccharide (LPS) were also determined. RESULTS: Compared to normal littermates, the voluntary daily running distance was 1.8-fold greater in mice with a disruption of the gene for TNFalpha, and 3-fold greater in mice with a gene disruption for both TNFalpha and LT. Intravenous administration of a monoclonal antibody against murine TNFalpha did not affect the running distance of wild-type mice, whereas administration of TNF intracerebrally reduced by 4-fold the voluntary running distance of the animals. This demonstrates that fatigue is mediated by TNFalpha expressed in the central nervous system (CNS) and not by increased peripheral TNFalpha concentrations. TNFalpha and LT are strong inducers of prostaglandins, but mice with disrupted prostaglandin or prostacyclin receptors exhibited running distances not significantly different from their wild-type littermates. Thus, signalling molecules other than prostaglandins mediate the effect of TNFalpha and LT on exercise capacity. CONCLUSIONS: Our finding that exercise capacity is controlled by TNFalpha is the first to define the endogenous mediators of fatigue, and may have important implications for diseases with impaired exercise tolerance.

Stimulation of macrophages and neutrophils by complexes of lipopolysaccharide and soluble CD14.

Sensitive responses of monocytes, macrophages, and neutrophils to bacterial LPS require membrane-bound CD14 (mCD14) and a plasma protein called LPS-binding protein (LBP). Cells lacking mCD14 respond to complexes of LPS and soluble CD14 (sCD14); these responses do not require LBP. To determine whether LBP is necessary for responses of mCD14-bearing cells to LPS, we measured responses of macrophages and neutrophils to complexes of LPS and sCD14 formed in the absence of LBP. We found that the amount of LPS needed to induce adhesive responses of neutrophils or cytokine production by macrophages was the same whether LPS was added with LBP or as LPS-sCD14 complexes, and was >100-fold less than when LPS was added alone. This result supports the view that LBP transfers LPS to CD14, but is not directly involved in responses of CD14-bearing cells to LPS. Responses of neutrophils to LPS-sCD14 complexes could be inhibited partially by blocking mCD14, suggesting that LPS may move rapidly from sCD14 to mCD14. Additionally, we found that responses of neutrophils to LBP and smooth LPS were made 30 to 100 times more sensitive when sCD14 was added. Our findings show that LBP is not necessary for the activation of CD14-bearing cells with LPS, and suggest that LPS-sCD14 complexes are an important intermediate in the inflammatory responses of leukocytes to LPS.