RESUMEN
Acute pancreatitis (AP) is a common disease with no targeted therapy and has varied outcomes ranging from spontaneous resolution to being lethal. Although typically painful, AP can also be painless. Various agents, including opioids, are used for pain control in AP; the risks and benefits of which are often debated. As experimental AP in mice is used to study the efficacy of potential therapies, we studied the effect of a commonly used opioid, buprenorphine, on the initiation and progression of AP. For this, we administered extended-release buprenorphine subcutaneously before inducing the previously established severe AP model that uses interleukins 12 and 18 (IL12,18) in genetically obese (ob/ob) mice and compared this to mice with AP but without the drug. Mice were monitored over 3 days, and parameters of AP induction and progression were compared. Buprenorphine significantly reduced serum amylase, lipase, pancreatic necrosis, and AP-associated fat necrosis, which is ubiquitous in obese mice and humans. Buprenorphine delayed the AP-associated reduction of carotid artery pulse distention and the development of hypothermia, hastened renal injury, and muted the early increase in respiratory rate versus IL12,18 alone. The site of buprenorphine injection appeared erythematous, inflamed, and microscopically showed thinning, loss of epidermal layers that had increased apoptosis. In summary, subcutaneous extended-release buprenorphine interfered with the induction of AP by reducing serum amylase, lipase, pancreatic and fat necrosis, the worsening of AP by delaying hypotension, hypothermia, while hastening renal injury, respiratory depression, and causing cutaneous injury at the site of injection.NEW & NOTEWORTHY Extended-release buprenorphine interferes with the initiation and progression of acute pancreatitis at multiple levels.
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Buprenorfina , Pancreatitis , Animales , Buprenorfina/farmacología , Pancreatitis/inducido químicamente , Pancreatitis/patología , Ratones , Analgésicos Opioides/farmacología , Modelos Animales de Enfermedad , Masculino , Interleucina-12/metabolismo , Interleucina-18/metabolismo , Interleucina-18/sangre , Ratones Obesos , Enfermedad Aguda , Páncreas/patología , Páncreas/efectos de los fármacos , Ratones Endogámicos C57BLRESUMEN
BACKGROUND & AIMS: Although transient bacteremia is common during dental and endoscopic procedures, infections developing during sterile diseases like acute pancreatitis (AP) can have grave consequences. We examined how impaired bacterial clearance may cause this transition. METHODS: Blood samples from patients with AP, normal controls, and rodents with pancreatitis or those administered different nonesterified fatty acids (NEFAs) were analyzed for albumin-unbound NEFAs, microbiome, and inflammatory cell injury. Macrophage uptake of unbound NEFAs using a novel coumarin tracer were done and the downstream effects-NEFA-membrane phospholipid (phosphatidylcholine) interactions-were studied on isothermal titration calorimetry. RESULTS: Patients with infected AP had higher circulating unsaturated NEFAs; unbound NEFAs, including linoleic acid (LA) and oleic acid (OA); higher bacterial 16S DNA; mitochondrial DNA; altered ß-diversity; enrichment in Pseudomonadales; and increased annexin V-positive myeloid (CD14) and CD3-positive T cells on admission. These, and increased circulating dead inflammatory cells, were also noted in rodents with unbound, unsaturated NEFAs. Isothermal titration calorimetry showed progressively stronger unbound LA interactions with aqueous media, phosphatidylcholine, cardiolipin, and albumin. Unbound NEFAs were taken into protein-free membranes, cells, and mitochondria, inducing voltage-dependent anion channel oligomerization, reducing ATP, and impairing phagocytosis. These were reversed by albumin. In vivo, unbound LA and OA increased bacterial loads and impaired phagocytosis, causing infection. LA and OA were more potent for these amphipathic interactions than the hydrophobic palmitic acid. CONCLUSIONS: Release of stored LA and OA can increase their circulating unbound levels and cause amphipathic liponecrosis of immune cells via uptake by membrane phospholipids. This impairs bacterial clearance and causes infection during sterile inflammation.
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Pancreatitis , Humanos , Enfermedad Aguda , Ácidos Grasos no Esterificados , Ácido Oléico , Inflamación , Albúminas , FosfatidilcolinasRESUMEN
BACKGROUND: Neuroinflammation causing disruption of the blood-brain barrier and immune cell extravasation into the brain parenchyma may cause delirium; however, knowledge of the exact pathophysiologic mechanism remains incomplete. The purpose of our study was to determine whether cytokine profiles differ depending on whether delirium occurs in the setting of sepsis, coronavirus disease 2019 (COVID-19), or recent surgery. METHODS: This prospective observational cohort study involved 119 critically ill patients admitted to a multidisciplinary intensive care unit (ICU) during 2019 and 2020. Delirium was identified using the validated confusion assessment method for the ICU. Multiple delirium risk factors were collected daily including clinical characteristics, hospital course, lab values, vital signs, surgical exposure, drug exposure, and COVID-19 characteristics. Serums samples were collected within 12 hours of ICU admission and cytokine levels were measured. RESULTS: The following proinflammatory cytokines were elevated in our delirium population: tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-18, C-C motif ligand (CCL) 2, CCL3, C-X-C motif chemokine ligand (CXCL)1, CXCL10, IL-8, IL-1 receptor antagonist, and IL-10. Analysis of relative cytokine levels in those patients that developed delirium in the setting of sepsis, COVID-19, and recent surgery showed elevations of CCL2, CXCL10, and TNF-α in both the sepsis and COVID-19 group in comparison to the postsurgical population. In the postsurgical group, granulocyte colony-stimulating factor was elevated and CXCL10 was decreased relative to the opposing groups. CONCLUSIONS: We identify several cytokines and precipitating factors known to be associated with delirium. However, our study suggests that the cytokine profile associated with delirium is variable and contingent upon delirium precipitating factors.
RESUMEN
OBJECTIVES: The pathophysiology of delirium is complex and incompletely understood. Inflammation is hypothesized to be integral to its development due to effects on blood brain barrier integrity, facilitation of leukocyte extravasation into brain parenchyma, and propagation of neuroinflammation. Septic shock is the prototypical condition associated with ICU delirium; however, the relative contribution of resultant hypotension and systemic inflammation to the development of delirium is unknown. DESIGN: This was a prospective exploratory study. SETTING: A multidisciplinary ICU at an academic medical center in Phoenix, AZ. PATIENTS: Critically ill patients older than or equal to 18 years old admitted to the ICU. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Screening for delirium was performed using the Confusion Assessment Method for the ICU tool. The levels of C-C motif ligand 2 (CCL2), C-C motif ligand 3, C-X-C motif chemokine ligand 1, C-X-C motif chemokine ligand 10, and interleukin-8 were measured in serum samples obtained within 12 hours of ICU admission. Univariate and multivariate analyses were performed to assess the association of delirium with patient data pertaining to hospital course, laboratory values, vital signs, medication administration, and levels of the aforementioned chemokines. Forty-one of 119 patients (34.5%) in the study cohort developed ICU delirium. Each chemokine studied was associated with delirium on univariate analyses; however, CCL2 was the only chemokine found to be independently associated with the development of delirium on multivariable analysis. The association of increased CCL2 levels with delirium remained robust in various models controlling for age, presence of shock, Sequential Organ Failure Assessment score, Acute Physiology and Chronic Health Evaluation IV score, mean arterial pressure at presentation, lowest mean arterial pressure, and total opioid, midazolam, propofol, and dexmedetomidine exposure. CONCLUSIONS: The demonstrated relationship between CCL2 and delirium suggests this chemokine may play a role in the development of delirium and warrants further investigation.
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We compared three hospitalized patient cohorts and conducted mechanistic studies to determine if lipotoxicity worsens COVID-19. Cohort-1 (n = 30) compared COVID-19 patients dismissed home to those requiring intensive-care unit (ICU) transfer. Cohort-2 (n = 116) compared critically ill ICU patients with and without COVID-19. Cohort-3 (n = 3969) studied hypoalbuminemia and hypocalcemia's impact on COVID-19 mortality. Patients requiring ICU transfer had higher serum albumin unbound linoleic acid (LA). Unbound fatty acids and LA were elevated in ICU transfers, COVID-19 ICU patients and ICU non-survivors. COVID-19 ICU patients (cohort-2) had greater serum lipase, damage-associated molecular patterns (DAMPs), cytokines, hypocalcemia, hypoalbuminemia, organ failure and thrombotic events. Hypocalcemia and hypoalbuminemia independently associated with COVID-19 mortality in cohort-3. Experimentally, LA reacted with albumin, calcium and induced hypocalcemia, hypoalbuminemia in mice. Endothelial cells took up unbound LA, which depolarized their mitochondria. In mice, unbound LA increased DAMPs, cytokines, causing endothelial injury, organ failure and thrombosis. Therefore, excessive unbound LA in the circulation may worsen COVID-19 outcomes.
RESUMEN
BACKGROUND & AIMS: The role of fatty acid ethyl esters (FAEEs) during human alcoholic pancreatitis is unknown. We compared FAEEs levels with their nonesterified fatty acids (NEFAs) precursors during alcohol intoxication and clinical alcoholic pancreatitis. The pathophysiology underlying FAEEs increase and their role as diagnostic biomarkers for alcoholic pancreatitis was investigated. METHODS: A prospective blinded study compared FAEEs, NEFAs, and ethanol blood levels on hospitalization for alcoholic pancreatitis (n = 31), alcohol intoxication (n = 25), and in normal controls (n = 43). Serum FAEEs were measured at admission for nonalcoholic pancreatitis (n = 75). Mechanistic cell and animal studies were done. RESULTS: Median FAEEs were similarly elevated during alcohol intoxication (205 nmol/L; 95% confidence interval [CI], 71.8-515 nmol/L, P < .001) and alcoholic pancreatitis (103.1 nmol/L; 95% CI, 53-689 nmol/L, P < .001) vs controls (1.7 nmol/L; 95% CI, 0.02-4.3 nmol/L) or nonalcoholic pancreatitis (8 nmol/L; 95% CI, 1.1-11.5 nmol/L). Alcoholic pancreatitis increased serum NEFAs (1024 ± 710 µmol/L vs 307 ± 185 µmol/L in controls, P < .05). FAEEs comprised 0.1% to 2% of the parent NEFA concentrations. FAEES correlated strongly with NEFAs independent of ethanol levels in alcoholic pancreatitis but not during alcohol intoxication. On receiver operating characteristic curve analysis for diagnosing alcoholic pancreatitis, the area under the curve for serum FAEEs was 0.87 (95% CI, 0.78-0.95, P < .001). In mice and cells, alcohol administration transiently increased all FAEEs. Oleic acid ethyl ester was the only FAEE with a sustained increase up to 24 hours after intraperitoneal oleic acid plus ethanol administration. CONCLUSIONS: The sustained, alcohol-independent, large (20- to 50-fold) increase in circulating FAEEs during alcoholic pancreatitis results from their visceral release and mirrors the 2- to 4-fold increase in parent NEFA. The large areas under the curve of FAEEs on receiver operating characteristic curve analysis supports their role as alcoholic pancreatitis biomarkers.
Asunto(s)
Intoxicación Alcohólica/sangre , Ácidos Grasos/sangre , Pancreatitis Alcohólica/sangre , Adulto , Intoxicación Alcohólica/diagnóstico , Intoxicación Alcohólica/fisiopatología , Biomarcadores/sangre , Nivel de Alcohol en Sangre , Estudios de Casos y Controles , Ácidos Grasos no Esterificados/sangre , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pancreatitis Alcohólica/diagnóstico , Pancreatitis Alcohólica/fisiopatología , Valor Predictivo de las Pruebas , Estudios Prospectivos , Regulación hacia ArribaRESUMEN
Obesity sometimes seems protective in disease. This obesity paradox is predominantly described in reports from the Western Hemisphere during acute illnesses. Since adipose triglyceride composition corresponds to long-term dietary patterns, we performed a meta-analysis modeling the effect of obesity on severity of acute pancreatitis, in the context of dietary patterns of the countries from which the studies originated. Increased severity was noted in leaner populations with a higher proportion of unsaturated fat intake. In mice, greater hydrolysis of unsaturated visceral triglyceride caused worse organ failure during pancreatitis, even when the mice were leaner than those having saturated triglyceride. Saturation interfered with triglyceride's interaction and lipolysis by pancreatic triglyceride lipase, which mediates organ failure. Unsaturation increased fatty acid monomers in vivo and aqueous media, resulting in greater lipotoxic cellular responses and organ failure. Therefore, visceral triglyceride saturation reduces the ensuing lipotoxicity despite higher adiposity, thus explaining the obesity paradox.
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Pancreatitis , Enfermedad Aguda , Tejido Adiposo , Animales , Inflamación , Ratones , Obesidad/complicaciones , Pancreatitis/etiología , TriglicéridosAsunto(s)
COVID-19/epidemiología , Hipoalbuminemia/epidemiología , Hipocalcemia/epidemiología , Albúminas/uso terapéutico , COVID-19/sangre , COVID-19/terapia , Calcio/sangre , Calcio/uso terapéutico , Humanos , Hipoalbuminemia/sangre , Hipoalbuminemia/terapia , Hipocalcemia/sangre , Hipocalcemia/terapia , Lipólisis , SARS-CoV-2 , Albúmina SéricaAsunto(s)
Infecciones por Coronavirus , Hipocalcemia , Pandemias , Neumonía Viral , Betacoronavirus , COVID-19 , Calcio , Hospitalización , Humanos , Hipocalcemia/epidemiología , Hipocalcemia/etiología , SARS-CoV-2RESUMEN
Bile acids (BA), with their large hydrophobic steroid nucleus and polar groups are amphipathic molecules. In bile, these exist as micelles above their critical micellar concentration (CMC). In blood at low concentrations, these exist as monomers, initiating cellular signals. This micellar to monomer transition may involve complex thermodynamic interactions between bile salts alone or with phospholipids, i.e. mixed micelles and the aqueous environment. We therefore went on to test if therapeutically relevant changes in temperature could influence micellar behavior of bile salts, and in turn whether this affected the biological responses in cells, and in vivo. Sodium taurocholate (STC) belongs to a major class of bile salts. STC has a CMC in the 5-8 mM range and its infusion into the pancreatic duct is commonly used to study pancreatitis. We thus studied micellar breakdown of STC using isothermal titration calorimetry (ITC), dynamic light scattering and cryogenic transmission electron microscopy. Under conditions relevant to the in vivo environment (pH 7.4, Na 0.15 M), ITC showed STC to have a U shaped reduction in micellar breakdown between 37 °C and 15 °C with a nadir at 25 °C approaching ≈90% inhibition. This temperature dependence paralleled pancreatic acinar injury induced by monomeric STC. Mixed micelles of STC and 1-palmitoyl, 2-oleyl phosphatidylcholine, a phospholipid present in high proportions in bile, behaved similarly, with ≈75% reduction in micellar breakdown at 25 °C compared to 37 °C. In vivo pancreatic cooling to 25 °C reduced the increase in circulating BAs after infusion of 120 mM (5%) STC into the pancreatic duct, and duct ligation. Lower BA levels were associated with improved cardiac function, reduced myocardial damage, shock, lung injury and improved survival independent of pancreatic injury. Thus micellar breakdown of bile salts is essential for their entry into the systemic circulation, and thermodynamic interference with this may reduce their systemic entry and consequent injury during cholestasis, such as from biliary pancreatitis.
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Ácidos y Sales Biliares/metabolismo , Colestasis/complicaciones , Inflamación/prevención & control , Lesión Pulmonar/prevención & control , Micelas , Contusiones Miocárdicas/prevención & control , Choque/prevención & control , Animales , Humanos , Inflamación/etiología , Inflamación/metabolismo , Inflamación/patología , Lesión Pulmonar/etiología , Lesión Pulmonar/metabolismo , Lesión Pulmonar/patología , Masculino , Ratones , Contusiones Miocárdicas/etiología , Contusiones Miocárdicas/metabolismo , Contusiones Miocárdicas/patología , Choque/etiología , Choque/metabolismo , Choque/patología , Temperatura , TermodinámicaAsunto(s)
Albúminas/administración & dosificación , Calcio de la Dieta/administración & dosificación , Infecciones por Coronavirus/mortalidad , Suplementos Dietéticos , Grasas Insaturadas/análisis , Neumonía Viral/mortalidad , Adulto , Anciano , Betacoronavirus , COVID-19 , Calcio de la Dieta/sangre , Infecciones por Coronavirus/sangre , Infecciones por Coronavirus/terapia , Femenino , Humanos , Hipoalbuminemia/mortalidad , Hipoalbuminemia/terapia , Hipoalbuminemia/virología , Hipocalcemia/mortalidad , Hipocalcemia/terapia , Hipocalcemia/virología , Masculino , Persona de Mediana Edad , Pandemias , Neumonía Viral/sangre , Neumonía Viral/terapia , SARS-CoV-2 , Albúmina Sérica/análisisRESUMEN
Pseudomonas aeruginosa (PA) is commonly associated with nosocomial and chronic infections of lungs. We have earlier demonstrated that an acidic sugar, sialic acid, is present in PA which is recognized and bound by sialic acid binding immunoglobulin type lectins (siglecs) expressed on neutrophils. Here, we have tried to gain a detailed insight into the immunosuppressive role of sialic acid-siglec interactions in macrophage-mediated clearance of sialylated PA (PA+Sia). We have demonstrated that PA+Sia shows enhanced binding (~1.5-fold) to macrophages due to additional interactions between sialic acids and siglec-E and exhibited more phagocytosis. However, internalization of PA+Sia is associated with a reduction in respiratory burst and increase in anti-inflammatory cytokines secretion which is reversed upon desialylation of the bacteria. Phagocytosis of PA+Sia is also associated with reduced intracellular calcium ion concentrations and altered calcium-dependent signaling which negatively affects phagosome maturation. Consequently, although more PA+Sia was localized in early phagosomes (Rab5 compartment), only fewer bacteria reach into the late phagosomal compartment (Rab7). Possibly, this leads to reduced phagosome lysosome fusion where reduced numbers of PA+Sia are trafficked into lysosomes, compared to PA-Sia. Thus, internalized PA+Sia remain viable and replicates intracellularly in macrophages. We have also demonstrated that such siglec-E-sialic acid interaction recruited SHP-1/SHP-2 phosphatases which modulate MAPK and NF-κB signaling pathways. Disrupting sialic acid-siglec-E interaction by silencing siglec-E in macrophages results in improved bactericidal response against PA+Sia characterized by robust respiratory burst, enhanced intracellular calcium levels and nuclear translocation of p65 component of NF-κB complex leading to increased pro-inflammatory cytokine secretion. Taken together, we have identified that sialic acid-siglec-E interactions is another pathway utilized by PA in order to suppress macrophage antimicrobial responses and inhibit phagosome maturation, thereby persisting as an intracellular pathogen in macrophages.
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Pulmón/inmunología , Macrófagos/inmunología , Neutrófilos/inmunología , Fagosomas/metabolismo , Infecciones por Pseudomonas/inmunología , Pseudomonas aeruginosa/fisiología , Infecciones del Sistema Respiratorio/inmunología , Animales , Antígenos CD/metabolismo , Antígenos de Diferenciación de Linfocitos B/metabolismo , Calcio/metabolismo , Señalización del Calcio , Humanos , Evasión Inmune , Tolerancia Inmunológica , Pulmón/microbiología , Ratones , Ácido N-Acetilneuramínico/genética , Ácido N-Acetilneuramínico/metabolismo , FN-kappa B/metabolismo , Oxidación-Reducción , Fagocitosis , Unión Proteica , ARN Interferente Pequeño/genética , Células THP-1RESUMEN
Visceral adipose tissue plays a critical role in numerous diseases. Although imaging studies often show adipose involvement in abdominal diseases, their outcomes may vary from being a mild self-limited illness to one with systemic inflammation and organ failure. We therefore compared the pattern of visceral adipose injury during acute pancreatitis and acute diverticulitis to determine its role in organ failure. Acute pancreatitis-associated adipose tissue had ongoing lipolysis in the absence of adipocyte triglyceride lipase (ATGL). Pancreatic lipase injected into mouse visceral adipose tissue hydrolyzed adipose triglyceride and generated excess nonesterified fatty acids (NEFAs), which caused organ failure in the absence of acute pancreatitis. Pancreatic triglyceride lipase (PNLIP) increased in adipose tissue during pancreatitis and entered adipocytes by multiple mechanisms, hydrolyzing adipose triglyceride and generating excess NEFAs. During pancreatitis, obese PNLIP-knockout mice, unlike obese adipocyte-specific ATGL knockouts, had lower visceral adipose tissue lipolysis, milder inflammation, less severe organ failure, and improved survival. PNLIP-knockout mice, unlike ATGL knockouts, were protected from adipocyte-induced pancreatic acinar injury without affecting NEFA signaling or acute pancreatitis induction. Therefore, during pancreatitis, unlike diverticulitis, PNLIP leaking into visceral adipose tissue can cause excessive visceral adipose tissue lipolysis independently of adipocyte-autonomous ATGL, and thereby worsen organ failure.
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Adipocitos/enzimología , Grasa Intraabdominal/enzimología , Lipasa/metabolismo , Pancreatitis/enzimología , Transducción de Señal , Enfermedad Aguda , Adipocitos/patología , Animales , Ácidos Grasos no Esterificados/genética , Ácidos Grasos no Esterificados/metabolismo , Femenino , Humanos , Inflamación/enzimología , Inflamación/genética , Inflamación/patología , Grasa Intraabdominal/patología , Lipasa/genética , Masculino , Ratones , Ratones Noqueados , Pancreatitis/genética , Pancreatitis/patologíaRESUMEN
OBJECTIVE: Ringer's lactate may improve early systemic inflammation during critical illnesses like severe acute pancreatitis, which are associated with hypocalcemia. Ringer's lactate is buffered and contains lactate and calcium. We, thus analyzed extracellular calcium or lactate's effects on the mechanisms, intermediary markers, and organ failure in models mimicking human disease with nonesterified fatty acid (NEFA) elevation. METHODS: Meta-analyses and experimental studies were performed. Experimentally, extracellular calcium and lactate were compared in their interaction with linoleic acid (LA; a NEFA increased in human severe pancreatitis), and its subsequent effects on mitochondrial depolarization and cytosolic calcium signaling resulting in cell injury. In vivo, the effect of LA was studied on organ failure, along with the effect of calcium or lactate (pH 7.4) on severe acute pancreatitis-associated organ failure. A meta-analysis of human randomized control trials comparing Ringer's lactate to normal saline was done, focusing on necrosis and organ failure. RESULTS: Calcium reacted ionically with LA and reduced lipotoxic necrosis. In vivo, LA induced organ failure and hypocalcemia. During severe pancreatitis, calcium supplementation in saline pH 7.4, unlike lactate, prevented hypocalcemia, increased NEFA saponification, reduced circulating NEFA and C-reactive protein , reduced pancreatic necrosis adjacent to fat necrosis, and normalized shock (carotid pulse distension) and blood urea nitrogen elevation on day 1. This, however, did not prevent the later increase in serum NEFA which caused delayed organ failure. Meta-analysis showed Ringer's lactate reduced necrosis, but not organ failure, compared with normal saline. CONCLUSION: Hypocalcemia occurs due to excess NEFA binding calcium during a critical illness. Ringer's lactate's early benefits in systemic inflammation are by the calcium it provides reacting ionically with NEFA. This, however, does not prevent later organ failure from sustained NEFA generation. Future studies comparing calcium supplemented saline resuscitation to Ringer's lactate may provide insights to this pathophysiology.
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Acute lipolysis of visceral fat or circulating triglycerides may worsen acute pancreatitis (AP)-associated local and systemic injury. The pancreas expresses pancreatic triacylglycerol lipase (PNLIP), pancreatic lipase-related protein 2 (PNLIPRP2), and carboxyl ester lipase (CEL), which may leak into the visceral fat or systemic circulation during pancreatitis. We, thus, aimed to determine the pancreatic lipase(s) regulating lipotoxicity during AP. For this AP, associated fat necrosis was analyzed using Western blot analysis. Bile acid (using liquid chromatography-tandem mass spectrometry) and fatty acid (using gas chromatography) concentrations were measured in human fat necrosis. The fat necrosis milieu was simulated in vitro using glyceryl trilinoleate because linoleic acid is increased in fat necrosis. Bile acid requirements to effectively hydrolyze glyceryl trilinoleate were studied using exogenous or overexpressed lipases. The renal cell line (HEK 293) was used to study lipotoxic injury. Because dual pancreatic lipase knockouts are lethal, exocrine parotid acini lacking lipases were used to verify the results. PNLIP, PNLIPRP2, and CEL were increased in fat necrosis. Although PNLIP and PNLIPRP2 were equipotent in inducing lipolysis and lipotoxic injury, CEL required bile acid concentrations higher than in human fat necrosis. The high bile acid requirements for effective lipolysis make CEL an unlikely mediator of lipotoxic injury in AP. It remains to be explored whether PNLIP or PNLIPRP2 worsens AP severity in vivo.
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Necrosis Grasa/enzimología , Grasa Intraabdominal/enzimología , Lipasa/metabolismo , Pancreatitis/enzimología , Animales , Necrosis Grasa/inducido químicamente , Necrosis Grasa/genética , Necrosis Grasa/patología , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Grasa Intraabdominal/patología , Ácido Linoleico/toxicidad , Lipasa/genética , Masculino , Ratones , Pancreatitis/inducido químicamente , Pancreatitis/genética , Pancreatitis/patologíaRESUMEN
BACKGROUND & AIMS: Acute pancreatitis (AP) of different etiologies is associated with the activation of different signaling pathways in pancreatic cells, posing challenges to the development of targeted therapies. We investigated whether local pancreatic hypothermia, without systemic hypothermia, could lessen the severity of AP induced by different methods in rats. METHODS: A urethane balloon with 2 polyurethane tubes was placed inside the stomach of rats. AP was induced in Wistar rats by the administration of cerulein or glyceryl tri-linoleate (GTL). Then, cold water was infused into the balloon to cool the pancreas. Pancreatic temperatures were selected based on those found to decrease acinar cell injury. An un-perfused balloon was used as a control. Pancreatic and rectal temperatures were monitored, and an infrared lamp or heating pad was used to avoid generalized hypothermia. We collected blood, pancreas, kidney, and lung tissues and analyzed them by histology, immunofluorescence, immunoblot, cytokine and chemokine magnetic bead, and DNA damage assays. The effect of hypothermia on signaling pathways initiated by cerulein and GTL was studied in acinar cells. RESULTS: Rats with pancreatic cooling developed less severe GTL-induced AP compared with rats that received the control balloon. In acinar cells, cooling decreased the lipolysis induced by GTL, increased the micellar form of its fatty acid, lowered the increase in cytosolic calcium, prevented the loss of mitochondrial membrane potential (by 70%-80%), and resulted in a 40%-50% decrease in the uptake of a fatty acid tracer. In rats with AP, cooling decreased pancreatic necrosis by 48%, decreased serum levels of cytokines and markers of cell damage, and decreased markers of lung and renal damage. Pancreatic cooling increased the proportions of rats surviving 6 hours after induction of AP (to 90%, from <10% of rats that received the control balloon). In rats with cerulein-induced AP, pancreatic cooling decreased pancreatic markers of apoptosis and inflammation. CONCLUSIONS: In rats with AP, transgastric local pancreatic hypothermia decreases pancreatic necrosis, apoptosis, inflammation, and markers of pancreatitis severity and increases survival.
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Hipotermia Inducida/métodos , Pancreatitis Aguda Necrotizante/patología , Pancreatitis Aguda Necrotizante/terapia , Animales , Biopsia con Aguja , Ceruletida/efectos adversos , Ceruletida/farmacología , Crioterapia/métodos , Modelos Animales de Enfermedad , Inmunohistoquímica , Masculino , Pancreatitis Aguda Necrotizante/mortalidad , Distribución Aleatoria , Ratas , Ratas Wistar , Sensibilidad y Especificidad , Índice de Severidad de la Enfermedad , Estómago , Tasa de Supervivencia , Factores de TiempoRESUMEN
PURPOSE OF REVIEW: The obesity pandemic poses a unique set of problems for acute pancreatitis - both by increasing acute pancreatitis incidence, and worsening acute pancreatitis severity. This review explores these associations, underlying mechanisms, and potential therapies. RECENT FINDINGS: We review how the obesity associated increase in gallstones, surgical, and endoscopic interventions for obesity management, diabetes, and related medications such as incretin-based therapies and hypertriglyceridemia may increase the incidence of acute pancreatitis. The mechanism of how obesity may increase acute pancreatitis severity are discussed with a focus on cytokines, adipokines, damage-associated molecular patterns and unsaturated fatty acid-mediated lipotoxicity. The role of obesity in exacerbating pancreatic necrosis is discussed; focusing on obesity-associated pancreatic steatosis. We also discuss how peripancreatic fat necrosis worsens organ failure independent of pancreatic necrosis. Last, we discuss emerging therapies including choice of intravenous fluids and the use of lipase inhibitors which have shown promise during severe acute pancreatitis. SUMMARY: We discuss how obesity may contribute to increasing acute pancreatitis incidence, the role of lipolytic unsaturated fatty acid release in worsening acute pancreatitis, and potential approaches, including appropriate fluid management and lipase inhibition in improving acute pancreatitis outcomes.
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Ácidos Grasos Insaturados/metabolismo , Hipertrigliceridemia/complicaciones , Lipasa/antagonistas & inhibidores , Lipólisis/efectos de los fármacos , Obesidad/complicaciones , Pancreatitis/etiología , Inhibidores Enzimáticos/uso terapéutico , Humanos , Hipertrigliceridemia/metabolismo , Hipertrigliceridemia/fisiopatología , Incretinas/efectos adversos , Lipólisis/fisiología , Obesidad/metabolismo , Obesidad/fisiopatología , Pancreatitis/fisiopatología , Índice de Severidad de la EnfermedadRESUMEN
Pseudomonas aeruginosa (PA) is an environmentally ubiquitous, extracellular, opportunistic pathogen, associated with severe infections of immune-compromised host. We demonstrated earlier the presence of both α2,3- and α2,6-linked sialic acids (Sias) on PA (PA(+Sias)) and normal human serum is their source of Sias. PA(+Sias) showed decreased complement deposition and exhibited enhanced association with immune-cells through sialic acid binding immunoglobulin like lectins (Siglecs). Such Sias-siglec-9 interaction between PA(+Sias) and neutrophils helped to subvert host immunity. Additionally, PA(+Sias) showed more resistant to ß-lactam antibiotics as reflected in their minimum inhibitory concentration required to inhibit the growth of 50% than PA(-Sias). Accordingly, we have affinity purified sialoglycoproteins of PA(+Sias). They were electrophoresed and identified by matrix-assisted laser desorption-ionization time-of-flight/time-of-flight mass spectrometry analysis. Sequence study indicated the presence of a few α2,6-linked, α2,3-linked, and both α2,3- and α2,6-linked sialylated proteins in PA. The outer membrane porin protein D (OprD), a specialized channel-forming protein, responsible for uptake of ß-lactam antibiotics, is one such identified sialoglycoprotein. Accordingly, sialylated (OprD(+Sias)) and non-sialylated (OprD(-Sias)) porin proteins were separately purified by using anion exchange chromatography. Sialylation of purified OprD(+Sias) was confirmed by several analytical and biochemical procedures. Profiling of glycan structures revealed three sialylated N-glycans and two sialylated O-glycans in OprD(+Sias). In contrast, OprD(-Sias) exhibit only one sialylated N-glycans. OprD(-Sias) interacts with ß-lactam antibiotics more than OprD(+Sias) as demonstrated by surface plasmon resonance study. Lyposome-swelling assay further exhibited that antibiotics have more capability to penetrate through OprD(-Sias) purified from four clinical isolates of PA. Taken together, it may be envisaged that sialic acids on OprD protein play important role toward the uptake of commonly used antibiotics in PA(+Sias). This might be one of the new mechanisms of PA for ß-lactam antibiotic uptake.
Asunto(s)
Porinas/metabolismo , Infecciones por Pseudomonas/tratamiento farmacológico , Pseudomonas aeruginosa/metabolismo , Ácidos Siálicos/metabolismo , Antibacterianos/administración & dosificación , Humanos , Polisacáridos/metabolismo , Porinas/química , Porinas/genética , Infecciones por Pseudomonas/metabolismo , Infecciones por Pseudomonas/microbiología , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/patogenicidad , beta-Lactamas/administración & dosificación , beta-Lactamas/químicaRESUMEN
L-Asparaginase-II from Escherichia coli (EcA) is a central component in the treatment of acute lymphoblastic leukemia (ALL). However, the therapeutic efficacy of EcA is limited due to immunogenicity and a short half-life in the patient. Here, we performed rational mutagenesis to obtain EcA variants with a potential to improve ALL treatment. Several variants, especially W66Y and Y176F, killed the ALL cells more efficiently than did wild-type EcA (WT-EcA), although nonleukemic peripheral blood monocytes were not affected. Several assays, including Western blotting, annexin-V/propidium iodide binding, comet, and micronuclei assays, showed that the reduction in viability of leukemic cells is due to the increase in caspase-3, cytochrome c release, poly(ADP-ribose) polymerase activation, down-regulation of anti-apoptotic protein Bcl-XL, an arrest of the cell cycle at the G0/G1 phase, and eventually apoptosis. Both W66Y and Y176F induced significantly more apoptosis in lymphocytes derived from ALL patients. In addition, Y176F and Y176S exhibited greatly decreased glutaminase activity, whereas K288S/Y176F, a variant mutated in one of the immunodominant epitopes, showed reduced antigenicity. Further in vivo immunogenicity studies in mice showed that K288S/Y176F was 10-fold less immunogenic as compared with WT-EcA. Moreover, sera obtained from WT-EcA immunized mice and ALL patients who were given asparaginase therapy for several weeks recognized the K288S/Y176F mutant significantly less than the WT-EcA. Further mechanistic studies revealed that W66Y, Y176F, and K288S/Y176F rapidly depleted asparagine and also down-regulated the transcription of asparagine synthetase as compared with WT-EcA. These highly desirable attributes of these variants could significantly advance asparaginase therapy of leukemia in the future.
Asunto(s)
Antineoplásicos , Asparaginasa , Epítopos de Linfocito B , Proteínas de Escherichia coli , Mutación Missense , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Sustitución de Aminoácidos , Animales , Antineoplásicos/inmunología , Antineoplásicos/farmacología , Asparaginasa/genética , Asparaginasa/inmunología , Asparaginasa/farmacología , Caspasa 3/genética , Caspasa 3/inmunología , Caspasa 3/metabolismo , Línea Celular Tumoral , Citocromos c/genética , Citocromos c/inmunología , Citocromos c/metabolismo , Epítopos de Linfocito B/genética , Epítopos de Linfocito B/inmunología , Epítopos de Linfocito B/farmacología , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/inmunología , Proteínas de Escherichia coli/farmacología , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Mutagénesis , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Proteína bcl-X/genética , Proteína bcl-X/inmunología , Proteína bcl-X/metabolismoRESUMEN
Sialic acids (Sias) are nine-carbon keto sugars primarily present on the terminal residue of cell surface glycans. Sialic acid binding immunoglobulins (Ig)-like lectins (siglecs) are generally expressed on various immune cells. They selectively recognize different linkage-specific sialic acids and undertake a variety of cellular functions. Many pathogens either synthesize or acquire sialic acids from the host. Sialylated pathogens generally use siglecs to manipulate the host immune response. The present review mainly deals with the newly developed information regarding mechanism of acquisition of sialic acids by pathogens and their biological relevance especially in the establishment of successful infection by impairing host innate immunity. The pathogens which are unable to synthesize sialic acids might adsorb these from the host as a way to engage the inhibitory siglecs. They promote association with the immune cells through sialic acids-siglec dependent manner. Such an association plays an important role to subvert host's immunity. Detailed investigation of these pathways has been discussed in this review. Particular attention has been focused on Pseudomonas aeruginosa (PA) and Leishmania donovani.