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Bladder cancer (BCa) remains a significant source of morbidity and mortality. BCa is one of the most expensive tumors to treat, in part because of a lack of nonsurgical options. The recent advent of immunotherapy, alone or in combination with other compounds, has improved therapeutic options. Resistance to immunotherapy remains common, and many patients do not have durable response. Recent advances indicate immunotherapy efficacy may be tied in part to the endogenous bacteria present in our body, more commonly referred to as the microbiome. Laboratory and clinical data now support the idea that a healthy microbiome is critical to effective response to immunotherapy. At the same time, pathogenic interactions between the microbiome and immune cells can also serve to drive formation of tumors, increasing the complexity of these interactions. Given the rising importance of immunotherapy in BCa, understanding how we might be able to alter the microbiome to improve therapeutic efficacy offers a novel route to improved patient care. The goal of this review is to examine our current understanding of microbial interactions with the immune system and cancer with an emphasis on BCa. We will further attempt to define both current gaps in knowledge and future directions that may yield beneficial results to the field.
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Preclinical modelling is a crucial component of advancing the understanding of cancer biology and therapeutic development. Several models exist for understanding the pathobiology of bladder cancer and evaluating therapeutics. N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced bladder cancer is a commonly used model that recapitulates many of the features of human disease. Particularly in mice, BBN is a preferred laboratory model owing to a high level of reproducibility, high genetic fidelity to the human condition, and its relative ease of use. However, important aspects of the model are often overlooked in laboratory studies. Moreover, the advent of new models has yielded a variety of methodologies that complement the use of BBN. Toxicokinetics, histopathology, molecular genetics and sex can differ between available models and are important factors to consider in bladder cancer modelling.
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Bladder cancer (BCa) incidence is tightly linked to aging. Older patients with BCa present with higher grade tumors and have worse outcomes on Bacillus-Calmette-Guerin (BCG) immunotherapy. Aging is also known to result in changes in the gut microbiome over mammalian lifespan, with recent studies linking alterations in the gut microbiome to changes in tumor immunity. There is limited information on the microbiome in BCa models though, despite known links to aging and immunotherapy. The purpose of this study was to evaluate how aging impacts tumor formation, inflammation, and the microbiome of mice given the model BCa carcinogen N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). We hypothesized old animals would have larger, more inflamed tumors and a shift in their fecal microbiome compared to their younger counterparts. Young (~8-week-old) or old (~78-week-old) C57Bl/6J animals were administered 0.05% BBN in drinking water for 16 weeks and then euthanized or allowed to progress for an additional 4 weeks. After 16 weeks of BBN, old mice had higher bladder to body weight ratio than young mice, and also muscle invasive tumors, which were not seen in their young counterparts. Old animals also had increased innate immune recruitment, but CD4+/CD8+ T cell recruitment did not appear different. BBN dramatically altered the microbiome in both sets of animals as measured by ß-diversity, including changes in multiple genera of bacteria. These data suggest old mice have a differential response to BBN-induced BCa. Given the median age of patients with BCa, understanding how the aged phenotype interacts with BCa is imperative.
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Butilhidroxibutilnitrosamina , Neoplasias de la Vejiga Urinaria , Humanos , Ratones , Animales , Anciano , Modelos Animales de Enfermedad , Butilhidroxibutilnitrosamina/toxicidad , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Carcinógenos , Envejecimiento , MamíferosRESUMEN
Pyruvate dehydrogenase kinase 4 (PDK4) is a mitochondrial isozyme in the PDK family (PDK1-4) partially responsible for phosphorylation of pyruvate dehydrogenase (PDH). Phosphorylation of PDH is thought to result in a pro-proliferative shift in metabolism that sustains growth of cancer cells. Previous data from our lab indicate the pan-PDK inhibitor dichloroacetate (DCA) or acute genetic knockdown of PDK4 blocks proliferation of bladder cancer (BCa) cells. The goal of this study was to determine the role of PDK4 in an in vivo BCa model, with the hypothesis that genetic depletion of PDK4 would impair formation of BCa. PDK4-/- or WT animals were exposed to N-Butyl-N-(4-hydroxybutyl) nitrosamine (BBN) for 16 weeks, and tumors were allowed to develop for up to 7 additional weeks. PDK4-/- mice had significantly larger tumors at later time points. When animals were treated with cisplatin, PDK4-/- animals still had larger tumors than WT mice. PDK4 expression was assessed in human tissue and in mice. WT mice lost expression of PDK4 as tumors became muscle-invasive. Similar results were observed in human samples, wherein tumors had less expression of PDK4 than benign tissue. In summary, PDK4 has a complex, multifunctional role in BCa and may represent an underrecognized tumor suppressor.
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Macrophage migration inhibitory factor (MIF1) is a pleiotropic cytokine involved in inflammation and cancer. Genetic knockout of Mif1 in the validated N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) model of bladder cancer (BCa) resulted in stage arrest at non-muscle-invasive disease in prior studies. Small-molecule inhibition of MIF1 reduced cancer-associated outcomes, but it did not fully recapitulate genetic models. D-dopachrome tautomerase (gene symbol DDT), commonly referred to as MIF2, is a functional homolog of MIF1, and both MIF1 and MIF2 can bind the cell surface receptor CD74 on multiple cell types to initiate a signaling cascade. It has been proposed that this interaction mediates part of the protumorigenic effects of MIF1 and MIF2 and may explain the discordance in prior studies. We hypothesized that MIF2 functions redundantly with MIF1 in BCa development and progression. The Cancer Genome Atlas (TCGA) analysis indicated MIF and DDT expression were increased in BCa patients compared to control. 4-Iodopyridine (4-IPP), a combined MIF1/MIF2 inhibitor, was more efficacious than ISO-1, a MIF1-only inhibitor, in preventing cellular proliferation in BCa cell lines. To evaluate these findings in vivo, wild-type (WT) and Mif1-/- animals were exposed to 0.05% BBN in drinking water for 16 weeks to initiate tumorigenesis and then evaluated over the subsequent 4 weeks for tumor formation and progression in the presence or absence of 4-IPP. 4-IPP reduced bladder weights in WT animals and bladder weights/tumor stage in Mif1-/- animals. To determine whether MIF1/MIF2 functioned through CD74 in BCa, WT or Cd74-/- animals were used in the same BBN model. Although these animals were partially protected against BBN-induced BCa, 4-IPP did not enhance this effect. In conclusion, our data suggest that MIF2 mechanistically functions in a similar protumorigenic manner to MIF1, and this is at least partially through CD74. Dual inhibition of MIF homologs is more efficacious at reducing tumor burden in this model of BCa. © 2022 The Pathological Society of Great Britain and Ireland.
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Factores Inhibidores de la Migración de Macrófagos , Neoplasias de la Vejiga Urinaria , Animales , Factores Inhibidores de la Migración de Macrófagos/genética , Factores Inhibidores de la Migración de Macrófagos/metabolismo , DDT , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Proliferación Celular , Transducción de SeñalRESUMEN
Ageing is correlated with elevated bladder cancer incidence, morbidity and mortality. Advanced age is also associated with elevated markers of chronic inflammation and perturbations in gut and urinary tract microbiota. One reason for the increased incidence and mortality of bladder cancer in the elderly might be that age-associated changes in multiple microbiomes induce systemic metabolic changes that contribute to immune dysregulation with potentially tumorigenic effects. The gut and urinary microbiomes could be dysregulated in bladder cancer, although the effect of these changes is poorly understood. Each of these domains - the immune system, gut microbiome and urinary microbiome - might also influence the response of patients with bladder cancer to treatment. Improved understanding of age-related alterations to the immune system and gut and urinary microbiomes could provide possible insight into the risk of bladder cancer development and progression in the elderly. In patients with bladder cancer, improved understanding of microbiota might also provide potential targets for therapeutic intervention.
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Microbioma Gastrointestinal , Microbiota , Neoplasias de la Vejiga Urinaria , Anciano , Envejecimiento , Humanos , Sistema InmunológicoRESUMEN
Acetaminophen (APAP) overdose causes liver injury in animals and humans. Although well-studied in animals, limited longitudinal data exist on cytokine release after APAP overdose in patients. The purpose of this study was to quantify concentrations of cytokines in APAP overdose patients to determine if early cytokine or complement measurements can distinguish between surviving and non-surviving patients. Plasma was obtained from healthy controls, APAP overdose patients with no increase in liver transaminases, and surviving and non-surviving APAP overdose patients with severe liver injury. Interleukin-10 (IL-10), and CC chemokine ligand-2 (CCL2, MCP-1) were substantially elevated in surviving and non-surviving patients, whereas IL-6 and CXC chemokine ligand-8 (CXCL8, IL-8) had early elevations in a subset of patients only with liver injury. Day 1 IL-10 and IL-6 levels, and Day 2 CCL2, levels correlated positively with survival. There was no significant increase in IL-1α, IL-1ß or TNF-α in any patient during the first week after APAP. Monitoring cytokines such as CCL2 may be a good indicator of patient prognosis; furthermore, these data indicate the inflammatory response after APAP overdose in patients is not mediated by a second phase of inflammation driven by the inflammasome.
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Acetaminofén , Enfermedad Hepática Inducida por Sustancias y Drogas , Acetaminofén/toxicidad , Animales , Antiinflamatorios , Humanos , Interleucina-10 , Interleucina-6 , Ligandos , Hígado , Ratones , Ratones Endogámicos C57BLRESUMEN
Ciclopirox (CPX) is an FDA-approved topical antifungal agent that has demonstrated preclinical anticancer activity in a number of solid and hematologic malignancies. Its clinical utility as an oral anticancer agent, however, is limited by poor oral bioavailability and gastrointestinal toxicity. Fosciclopirox, the phosphoryloxymethyl ester of CPX (Ciclopirox Prodrug, CPX-POM), selectively delivers the active metabolite, CPX, to the entire urinary tract following parenteral administration. We characterized the activity of CPX-POM and its major metabolites in in vitro and in vivo preclinical models of high-grade urothelial cancer. CPX inhibited cell proliferation, clonogenicity and spheroid formation, and increased cell cycle arrest at S and G0/G1 phases. Mechanistically, CPX suppressed activation of Notch signaling. Molecular modeling and cellular thermal shift assays demonstrated CPX binding to γ-secretase complex proteins Presenilin 1 and Nicastrin, which are essential for Notch activation. To establish in vivo preclinical proof of principle, we tested fosciclopirox in the validated N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) mouse bladder cancer model. Once-daily intraperitoneal administration of CPX-POM for four weeks at doses of 235 mg/kg and 470 mg/kg significantly decreased bladder weight, a surrogate for tumor volume, and resulted in a migration to lower stage tumors in CPX-POM treated animals. This was coupled with a reduction in the proliferation index. Additionally, there was a reduction in Presenilin 1 and Hes-1 expression in the bladder tissues of CPX-POM treated animals. Following the completion of the first-in-human Phase 1 trial (NCT03348514), the pharmacologic activity of fosciclopirox is currently being characterized in a Phase 1 expansion cohort study of muscle-invasive bladder cancer patients scheduled for cystectomy (NCT04608045) as well as a Phase 2 trial of newly diagnosed and recurrent urothelial cancer patients scheduled for transurethral resection of bladder tumors (NCT04525131).
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Secretasas de la Proteína Precursora del Amiloide/metabolismo , Antifúngicos/uso terapéutico , Carcinoma de Células Transicionales/tratamiento farmacológico , Ciclopirox/uso terapéutico , Antifúngicos/farmacología , Ciclopirox/farmacología , Humanos , Clasificación del TumorRESUMEN
Cisplatin remains an integral part of the treatment for muscle invasive bladder cancer. A large number of patients do not respond to cisplatin-based chemotherapy and efficacious salvage regimens are limited. Immunotherapy has offered a second line of treatment; however, only approximately 20% of patients respond, and molecular subtyping of tumors indicates there may be significant overlap in those patients that respond to cisplatin and those patients that respond to immunotherapy. As such, restoring sensitivity to cisplatin remains a major hurdle to improving patient care. One potential source of compounds for enhancing cisplatin is naturally derived bioactive products such as phytochemicals, flavonoids and others. These compounds can activate a diverse array of different pathways, many of which can directly promote or inhibit cisplatin sensitivity. The purpose of this review is to understand current drug development in the area of natural products and to assess how these compounds may enhance cisplatin treatment in bladder cancer patients.
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Histological and molecular analyses of urothelial carcinoma often reveal intratumoural and intertumoural heterogeneity at the genomic, transcriptional and cellular levels. Despite the clonal initiation of the tumour, progression and metastasis often arise from subclones that can develop naturally or during therapy, resulting in molecular alterations with a heterogeneous distribution. Variant histologies in tumour tissues that have developed distinct morphological characteristics divergent from urothelial carcinoma are extreme examples of tumour heterogeneity. Ultimately, heterogeneity contributes to drug resistance and relapse after therapy, resulting in poor survival outcomes. Mutation profile differences between patients with muscle-invasive and metastatic urothelial cancer (interpatient heterogeneity) probably contribute to variability in response to chemotherapy and immunotherapy as first-line treatments. Heterogeneity can occur on multiple levels and averaging or normalizing these alterations is crucial for clinical trial and drug design to enable appropriate therapeutic targeting. Identification of the extent of heterogeneity might shape the choice of monotherapy or additional combination treatments to target different drivers and genetic events. Identification of the lethal tumour cell clones is required to improve survival of patients with urothelial carcinoma.
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Carcinoma de Células Transicionales/genética , Carcinoma de Células Transicionales/terapia , Heterogeneidad Genética , Genoma , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/terapia , Humanos , Resultado del TratamientoRESUMEN
Acetaminophen (APAP) overdose-induced acute liver failure is an important clinical problem in the United States and the current antidote N-acetylcysteine, has a short early therapeutic window. Since most patients present late to the clinic, there is need for novel late-acting therapeutic options. Though the neuronal guidance cue netrin-1, has been shown to promote hepatic repair and regeneration during liver ischemia/reperfusion injury, its effect in APAP-induced hepatotoxicity is unknown. In the quest for a late-acting therapeutic intervention in APAP-induced liver injury, we examined the role of netrin-1 in a mouse model of APAP overdose. Male C57BL/6J mice were cotreated with exogenous netrin-1 or vehicle control, along with 300 mg/kg APAP and euthanized at 6, 12, and 24 h. Significant elevations in alanine aminotransferase indicative of liver injury were seen in control mice at 6 h and this was not affected by netrin-1 administration. Also, netrin-1 treatment did not influence mitochondrial translocation of phospho-JNK, or peroxynitrite formation indicating that there was no interference with APAP-induced injury processes. Interestingly however, netrin-1 administration attenuated liver injury at 24 h, as seen by alanine aminotransferase levels and histology, at which time significant elevations in the netrin-1 receptor, adenosine A2B receptor (A2BAR) as well as macrophage infiltration was evident. Removal of resident macrophages with clodronate liposomes or treatment with the A2BAR antagonist PSB1115 blocked the protective effects of netrin-1. Thus, our data indicate a previously unrecognized role for netrin-1 in attenuation of APAP hepatotoxicity by enhancing recovery and regeneration, which is mediated through the A2BAR and involves resident liver macrophages.
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Acetaminofén/toxicidad , Antídotos/farmacología , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológico , Fallo Hepático Agudo/tratamiento farmacológico , Hígado/efectos de los fármacos , Netrina-1/farmacología , Animales , Modelos Animales de Enfermedad , Masculino , Ratones , Ratones Endogámicos C57BL , Estados UnidosRESUMEN
Bladder cancer (BCa) is a common solid tumor marked by high rates of recurrence, especially in non-muscle invasive disease. Prostaglandin E2 (PGE2) is a ubiquitously present lipid mediator responsible for numerous physiological actions. Inhibition of cyclooxygenase (COX) enzymes by the non-steroidal anti-inflammatory (NSAID) class of drugs results in reduced PGE2 levels. NSAID usage has been associated with reductions in cancers such as BCa. Clinical trials using NSAIDs to prevent recurrence have had mixed results, but largely converge on issues with cardiotoxicity. The purpose of this review is to understand the basic science behind how and why inhibitors of PGE2 may be effective against BCa, and to explore alternate therapeutic modalities for addressing the role of PGE2 without the associated cardiotoxicity. We will address the role of PGE2 in a diverse array of cancer-related functions including stemness, immunosuppression, proliferation, cellular signaling and more.
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Antiinflamatorios no Esteroideos/uso terapéutico , Ciclooxigenasa 2/química , Dinoprostona/metabolismo , Prostaglandina-E Sintasas/antagonistas & inhibidores , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/metabolismo , Animales , Ensayos Clínicos como Asunto , Humanos , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
Cholestasis is a result of obstruction of the biliary tracts. It is a common cause of liver pathology after exposure to toxic xenobiotics and during numerous other liver diseases. Accumulation of bile acids in the liver is thought to be a major driver of liver injury during cholestasis and can lead to eventual liver fibrosis and cirrhosis. As such, current therapy in the field of chronic liver diseases with prominent cholestasis relies heavily on increasing choleresis to limit accumulation of bile acids. Many of these same diseases also present with autoimmunity before the onset of cholestasis though, indicating the inflammation may be an initiating component of the pathology. Moreover, cytotoxic inflammatory mediators accumulate during cholestasis and can propagate liver injury. Anti-inflammatory biologics and small molecules have largely failed clinical trials in these diseases though and as such, targeting inflammation as a means to address cholestatic liver injury remains debatable. The purpose of this review is to understand the different roles that inflammation can play during cholestatic liver injury and attempt to define how new therapeutic targets that limit or control inflammation may be beneficial for patients with chronic cholestatic liver disease.
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Colestasis/inmunología , Animales , Ácidos y Sales Biliares/efectos adversos , Ácidos y Sales Biliares/metabolismo , Conductos Biliares/metabolismo , Colestasis/etiología , Colestasis/genética , Colestasis/metabolismo , Humanos , Hígado/inmunologíaRESUMEN
Though mitochondrial oxidant stress plays a critical role in the progression of acetaminophen (APAP) overdose-induced liver damage, the influence of mitochondrial bioenergetics on this is not well characterized. This is important, since lifestyle and diet alter hepatic mitochondrial bioenergetics and an understanding of its effects on APAP-induced liver injury is clinically relevant. Pyruvate dehydrogenase (PDH) is critical to mitochondrial bioenergetics, since it controls the rate of generation of reducing equivalents driving respiration, and pyruvate dehydrogenase kinase 4 (PDK4) regulates (inhibits) PDH by phosphorylation. We examined APAP-induced liver injury in PDK4-deficient (PDK4-/-) mice, which would have constitutively active PDH and hence elevated flux through the mitochondrial electron transport chain. PDK4-/- mice showed significant protection against APAP-induced liver injury when compared to wild type (WT) mice as measured by ALT levels and histology. Deficiency of PDK4 did not alter APAP metabolism, with similar APAP-adduct levels in PDK4-/- and WT mice, and no difference in JNK activation and translocation to mitochondria. However, subsequent amplification of mitochondrial dysfunction with release of mitochondrial AIF, peroxynitrite formation and DNA fragmentation were prevented. Interestingly, APAP induced a rapid decline in UCP2 protein levels in PDK4-deficient mice. These data suggest that adaptive changes in mitochondrial bioenergetics induced by enhanced respiratory chain flux in PDK4-/- mice render them highly efficient in handling APAP-induced oxidant stress, probably through modulation of UCP2 levels. Further investigation of these specific adaptive mechanisms would provide better insight into the control exerted by mitochondrial bioenergetics on cellular responses to an APAP overdose.
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Acetaminofén/envenenamiento , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Sobredosis de Droga/complicaciones , Hígado/patología , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/metabolismo , Animales , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Modelos Animales de Enfermedad , Sobredosis de Droga/etiología , Metabolismo Energético/efectos de los fármacos , Femenino , Humanos , Hígado/citología , Hígado/efectos de los fármacos , Masculino , Ratones , Ratones Noqueados , Mitocondrias/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/genética , Proteína Desacopladora 2/metabolismoRESUMEN
Cancer cells use alterations of normal metabolic processes to sustain proliferation indefinitely. Transcriptional and posttranscriptional control of the pyruvate dehydrogenase kinase (PDK) family is one way in which cancer cells alter normal pyruvate metabolism to fuel proliferation. PDKs can phosphorylate and inactivate the pyruvate dehydrogenase complex (PDHC), which blocks oxidative metabolism of pyruvate by the mitochondria. This process is thought to enhance cancer cell growth by promoting anabolic pathways. Inhibition of PDKs induces cell death through increased PDH activity and subsequent increases in ROS production. The use of PDK inhibitors has seen widespread success as a potential therapeutic in laboratory models of multiple cancers; however, gaps still exist in our understanding of the biology of PDK regulation and function, especially in the context of individual PDKs. Efforts are currently underway to generate PDK-specific inhibitors and delineate the roles of individual PDK isozymes in specific cancers. The goal of this review is to understand the regulation of the PDK isozyme family, their role in cancer proliferation, and how to target this pathway therapeutically to specifically and effectively reduce cancer growth.
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Neoplasias/enzimología , Neoplasias/metabolismo , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/metabolismo , Complejo Piruvato Deshidrogenasa/metabolismo , Transducción de Señal , Animales , Humanos , Terapia Molecular Dirigida , Neoplasias/tratamiento farmacológicoRESUMEN
OBJECTIVE: Regulation of TF (tissue factor):FVIIa (coagulation factor VIIa) complex procoagulant activity is especially critical in tissues where plasma can contact TF-expressing cells. One example is the liver, where hepatocytes are routinely exposed to plasma because of the fenestrated sinusoidal endothelium. Although liver-associated TF contributes to coagulation, the mechanisms controlling the TF:FVIIa complex activity in this tissue are not known. Approach and Results: Common bile duct ligation in mice triggered rapid hepatocyte TF-dependent intrahepatic coagulation coincident with increased plasma bile acids, which occurred at a time before observable liver damage. Similarly, plasma TAT (thrombin-antithrombin) levels increased in cholestatic patients without concurrent hepatocellular injury. Pathologically relevant concentrations of the bile acid glycochenodeoxycholic acid rapidly increased hepatocyte TF-dependent procoagulant activity in vitro, independent of de novo TF synthesis and necrotic or apoptotic cell death. Glycochenodeoxycholic acid increased hepatocyte TF activity even in the presence of the phosphatidylserine-blocking protein lactadherin. Interestingly, glycochenodeoxycholic acid and taurochenodeoxycholic acid increased the procoagulant activity of the TF:FVIIa complex relipidated in unilamellar phosphatidylcholine vesicles, which was linked to an apparent decrease in the Km for FX (coagulation factor X). Notably, the zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, a bile acid structural analog, did not increase relipidated TF:FVIIa activity. Bile acids directly enhanced factor X activation by recombinant soluble TF:FVIIa complex but had no effect on FVIIa alone. CONCLUSIONS: The results indicate that bile acids directly accelerate TF:FVIIa-driven coagulation reactions, suggesting a novel mechanism whereby elevation in a physiological mediator can directly increase TF:FVIIa procoagulant activity.
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Conductos Biliares/cirugía , Colestasis Intrahepática/metabolismo , Colestasis Intrahepática/fisiopatología , Factor VIIa/metabolismo , Factor X/metabolismo , Animales , Ácidos y Sales Biliares/metabolismo , Coagulación Sanguínea/fisiología , Trastornos de la Coagulación Sanguínea/fisiopatología , Pruebas de Coagulación Sanguínea , Células Cultivadas , Modelos Animales de Enfermedad , Hepatocitos/metabolismo , Humanos , Cinética , Ligadura/métodos , Ratones , Ratones Endogámicos C57BL , Fosfatidilserinas/metabolismo , Distribución AleatoriaRESUMEN
Cholestasis results in blockage of bile flow whether the point of obstruction occurs extrahepatically or intrahepatically. Bile acids are a primary constituent of bile, and thus one of the primary outcomes is acute retention of bile acids in hepatocytes. Bile acids are normally secreted into the biliary tracts and then released into the small bowel before recirculating back to the liver. Retention of bile acids has long been hypothesized to be a primary cause of the associated liver injury that occurs during acute or chronic cholestasis. Despite this, a surge of papers in the last decade have reported a primary role for inflammation in the pathophysiology of cholestatic liver injury. Furthermore, it has increasingly been recognized that both the constituency of individual bile acids that make up the greater pool, as well as their conjugation status, is intimately involved in their toxicity, and this varies between species. Finally, the role of bile acids in drug-induced cholestatic liver injury remains an area of increasing interest. The purpose of this review is to critically evaluate current proposed mechanisms of cholestatic liver injury, with a focus on the evolving role of bile acids in cell death and inflammation.
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Ácidos y Sales Biliares/metabolismo , Colestasis/metabolismo , Citocinas/metabolismo , Animales , Sistema Biliar/metabolismo , Sistema Biliar/patología , Colestasis/patología , HumanosRESUMEN
PURPOSE: Bladder cancer is a "Warburg-like" tumor characterized by a reliance on aerobic glycolysis and expression of pyruvate kinase M2 (PKM2). PKM2 oscillates between an active tetramer and an inactive dimer. We aim to further characterize PKM2, in particular PKM2 dimer, as a urinary biomarker of bladder cancer and a potential target for treatment. METHODS: HTB-9, HTB-5, and UM-UC3 bladder cancer cells were assessed for proliferation under differential glucose levels using the hexosaminidase assay. Western blot and Blue-native analysis was performed for protein expression of PKM2. Shikonin, an herb that is known to bind and inhibit PKM2, was utilized to determine if PKM2 has a role in glucose usage and cellular proliferation in bladder cancer cells by caspase activity assay. Institutional review board approval was obtained to collect healthy control and bladder cancer patient urine samples. The ScheBo M2-PK EDTA Plasma Test was performed on urine samples to assess urine Tumor M2-PK values. RESULTS: The three bladder cancer cell lines tested all demonstrate statistically significant increases in proliferation when exposed to higher level of glucose (200mg/dL). Similarly, low doses of glucose (25mg/dL) result in reduced proliferation. Increased cell growth in higher glucose concentration correlated with up-regulation of PKM2 protein expression. Shikonin, a PKM2 inhibitor, reduced cell proliferation and switched PKM2 isoforms from the dimer to tetramer. Lastly, dimer PKM2 (Tumor-M2PK) levels were assessed in the urine samples from bladder cancer (Bca) patients and healthy controls. Tumor M2-PK significantly correlated with the presence of BCa in our subjects. CONCLUSIONS: Our studies demonstrate the potential of PKM2, specifically the dimer (Tumor-M2PK) as a target of drug therapy and as a urinary marker for bladder cancer.
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Biomarcadores de Tumor/orina , Proteínas Portadoras/orina , Proteínas de la Membrana/orina , Piruvato Quinasa/orina , Hormonas Tiroideas/orina , Neoplasias de la Vejiga Urinaria/orina , Adulto , Anciano , Biomarcadores de Tumor/química , Proteínas Portadoras/química , Estudios de Casos y Controles , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Medicamentos Herbarios Chinos/farmacología , Femenino , Glucosa/metabolismo , Glucólisis , Humanos , Masculino , Proteínas de la Membrana/química , Persona de Mediana Edad , Naftoquinonas/farmacología , Estructura Cuaternaria de Proteína , Piruvato Quinasa/química , Hormonas Tiroideas/química , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/patología , Proteínas de Unión a Hormona TiroideRESUMEN
BACKGROUND: One of the major issues in the field of acute liver failure (ALF) is the lack of reliable biomarkers that predict outcome. Many cases present with very limited treatment options and prognostic indicators are invaluable. We tested whether leukocyte cell derived chemotaxin 2 can be used as a prognostic biomarker to predict patient survival either alone or in combination with other routine clinical parameters. METHODS: Serum samples and associated clinical data from came from two independent sources, the Acute Liver Failure Study Group (ALFSG) registry and the University of Kansas Medical Center. We analyzed a total of 61 cases, each with individual time points collected over a period of 0 to 7 days after hospital admission. Analysis was developed to compare responses in survivors vs. non-survivors. RESULTS: The data indicate that survivors had significantly lower serum levels of leukocyte cell derived chemotaxin 2 compared to non-survivors (P=0.03). Further, it was able to predict patient survival when taken together with either international normalized ratio (INR) alone (71% concordance) or INR and bilirubin (76% concordance) or INR and serum albumin (77% concordance). Furthermore, when we analyzed data for each day, serum Lect2 and INR taken together were able to predict survival at day three after hospital admission with 86.3% concordance. CONCLUSIONS: These studies have revealed test batteries consisting of easily available serum tests that are concordant with survival status of ALF patients early during the clinical course.
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Cholestasis can be induced by obstruction of bile ducts or intrahepatic toxicity of drugs and chemicals. However, the mode of cell death during cholestasis, i.e., apoptosis or necrosis, has been controversial. There are fundamental reasons for the controversies, both of which are discussed here, namely the design of experiments and the use of parameters with limited specificity for a certain mode of cell death. Based on the assumption that cholestatic liver injury is caused by accumulation of bile acids, rodent (mainly rat) hepatocytes have been exposed to hydrophobic, glycine-conjugated bile acids, which resulted in apoptotic cell death. The problems with this experimental design are that in rodents bile acids are predominantly taurine conjugated and rodent hepatocytes are never exposed to these levels of glycine-conjugated bile acids. In contrast, taurine-conjugated bile acids trigger inflammatory gene activation in rodent hepatocytes and a necro-inflammatory injury in vivo. On the other hand, human hepatocytes are more resistant to glycine-conjugated bile acids and die by necrosis when exposed to high biliary levels of these bile acids. In this chapter, we describe multiple assays including the caspase activity assay, which is specific for apoptosis, and the general cell death assays alanine aminotransferase or lactate dehydrogenase activities in cell culture medium or plasma. An increase in these enzyme activities without caspase activity indicates necrotic cell death. Thus, both the experimental design and the selection of cell death parameters are critical for the relevance of the experiments for the human pathophysiology.