RESUMEN
The rapidly aging population is consuming more alcohol, leading to increased alcohol-associated acute pancreatitis (AAP) with high mortality. However, the mechanisms remain undefined, and currently there are no effective therapies available. This study aims to elucidate aging- and alcohol-associated spatial transcriptomic signature by establishing an aging AAP mouse model and applying Visium spatial transcriptomics for understanding of the mechanisms in the context of the pancreatic tissue. Upon alcohol diet feeding and caerulein treatment, aging mice (18 months) developed significantly more severe AAP with 5.0-fold increase of injury score and 2.4-fold increase of amylase compared to young mice (3 months). Via Visium spatial transcriptomics, eight distinct tissue clusters were revealed from aggregated transcriptomes of aging and young AAP mice: five acinar, two stromal, and one islet, which were then merged into three clusters: acinar, stromal, and islet for the comparative analysis. Compared to young AAP mice, > 1300 differentially expressed genes (DEGs) and approximately 3000 differentially regulated pathways were identified in aging AAP mice. The top five DEGs upregulated in aging AAP mice include Mmp8, Ppbp, Serpina3m, Cxcl13, and Hamp with heterogeneous distributions among the clusters. Taken together, this study demonstrates spatial heterogeneity of inflammatory processes in aging AAP mice, offering novel insights into the mechanisms and potential drivers for AAP development. KEY MESSAGES: Mechanisms regarding high mortality of AAP in aging remain undefined. An aging AAP mouse model was developed recapturing clinical exhibition in humans. Spatial transcriptomics identified contrasted DEGs in aging vs. young AAP mice. Top five DEGs were Mmp8, Ppbp, Serpina3m, Cxcl13, and Hamp in aging vs. young AAP mice. Our findings shed insights for identification of molecular drivers in aging AAP.
Asunto(s)
Envejecimiento , Pancreatitis , Transcriptoma , Animales , Envejecimiento/genética , Ratones , Pancreatitis/genética , Pancreatitis/inducido químicamente , Pancreatitis/metabolismo , Pancreatitis/patología , Perfilación de la Expresión Génica , Modelos Animales de Enfermedad , Masculino , Inflamación/genética , Ratones Endogámicos C57BL , Etanol/efectos adversos , Pancreatitis Alcohólica/genética , Pancreatitis Alcohólica/metabolismo , Pancreatitis Alcohólica/patología , Enfermedad Aguda , Páncreas/metabolismo , Páncreas/patologíaRESUMEN
Excessive alcohol intake is a major risk factor for pancreatitis, sensitizing the exocrine pancreas to stressors by mechanisms that remain obscure. Impaired autophagy drives nonalcoholic pancreatitis, but the effects of ethanol (EtOH) and alcoholic pancreatitis on autophagy are poorly understood. Here, we find that ethanol reduces autophagosome formation in pancreatic acinar cells, both in a mouse model of alcoholic pancreatitis induced by a combination of EtOH diet and cerulein (a CCK ortholog) and in EtOH+CCK-treated acinar cells (ex vivo model). Ethanol treatments decreased pancreatic level of LC3-II, a key mediator of autophagosome formation. This was caused by ethanol-induced upregulation of ATG4B, a cysteine protease that, cell dependently, regulates the balance between cytosolic LC3-I and membrane-bound LC3-II. We show that ATG4B negatively regulates LC3-II in acinar cells subjected to EtOH treatments. Ethanol raised ATG4B level by inhibiting its degradation, enhanced ATG4B enzymatic activity, and strengthened its interaction with LC3-II. We also found an increase in ATG4B and impaired autophagy in a dissimilar, nonsecretagogue model of alcoholic pancreatitis induced by EtOH plus palmitoleic acid. Adenoviral ATG4B overexpression in acinar cells greatly reduced LC3-II and inhibited autophagy. Furthermore, it aggravated trypsinogen activation and necrosis, mimicking key responses of ex vivo alcoholic pancreatitis. Conversely, shRNA Atg4B knockdown enhanced autophagosome formation and alleviated ethanol-induced acinar cell damage. The results reveal a novel mechanism, whereby ethanol inhibits autophagosome formation and thus sensitizes pancreatitis, and a key role of ATG4B in ethanol's effects on autophagy. Enhancing pancreatic autophagy, particularly by downregulating ATG4B, could be beneficial in mitigating the severity of alcoholic pancreatitis.NEW & NOTEWORTHY Ethanol sensitizes mice and humans to pancreatitis, but the underlying mechanisms remain obscure. Autophagy is important for maintaining pancreatic acinar cell homeostasis, and its impairment drives pancreatitis. This study reveals a novel mechanism, whereby ethanol inhibits autophagosome formation through upregulating ATG4B, a key cysteine protease. ATG4B upregulation inhibits autophagy in acinar cells and aggravates pathological responses of experimental alcoholic pancreatitis. Enhancing pancreatic autophagy, particularly by down-regulating ATG4B, could be beneficial for treatment of alcoholic pancreatitis.
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Proteasas de Cisteína , Pancreatitis Alcohólica , Animales , Humanos , Ratones , Células Acinares/metabolismo , Autofagia , Proteínas Relacionadas con la Autofagia/genética , Proteínas Relacionadas con la Autofagia/metabolismo , Cisteína Endopeptidasas/genética , Cisteína Endopeptidasas/metabolismo , Proteasas de Cisteína/metabolismo , Etanol/farmacología , Pancreatitis Alcohólica/genética , Regulación hacia ArribaRESUMEN
The study of genetic and environmental factors on the risk of acute alcoholic-alimentary pancreatitis (AÐAÐ ) is especially relevant to interpret individual links of pathogenesis, to reduce the incidence by eliminating the impact of harmful factors and improve the quality of life of the population through the introduction of optimal nutrition, and a healthy lifestyle, which is especially important for carriers of risk genotypes. The aim of the research was to study the influence of environmental factors and polymorphic loci rs6580502 of the SPINK1 gene, rs10273639 of the PRSS1 gene, rs213950 of the CFTR gene on the risk of ÐAÐ . Material and methods. Blood DNA samples obtained from 547 patients with AÐAÐ and 573 healthy individuals were used as the material for the study. The groups were comparable by sex and age. All participants were assessed qualitatively and quantitatively for risk factors, smoking and alcohol consumption, the frequency, quantity and regularity of intake of various types of foods, as well as the size and number of portions eaten. Genomic DNA was isolated by the standard phenol-chloroform extraction method, multiplex genotyping of SNPs was performed on a MALDI-TOF MassARRAY-4 genetic analyzer. Results. It was found that the T/T genotype (p=0.0012) of the rs6580502 SPINK1 was associated with an increased risk of AAAP, and the T allele (p=0.0001) and C/T and T/T genotypes (p=0.0001) of the rs10273639 PRSS1, A allele (p=0.01) and A/G and A/A genotypes (p=0.0006) of the rs213950 CFTR were associated with an decreased risk of the disease. The revealed effects of polymorphic loci of candidate genes were enhanced by the effect of alcohol consumption. The risk of AAAP was reduced by fat intake of less than 89 g per day in carriers of the A/G-A/A CFTR genotypes (rs213950), consumption of fresh vegetables and fruits of more than 27 g per day in carriers of the T/C-T/T PRSS1 genotypes (rs10273639), protein intake of more 84 g per day in carriers of T/C-T/T PRSS1 rs10273639 and A/G-A/A CFTR rs213950. The most significant models of gene-environment interactions included risk factors: deficiency in the diet of protein, fresh vegetables and fruits, smoking, and polymorphic variants of the PRSS1 (rs10273639) and SPINK (rs6580502) genes. Conclusion. In order to prevent the development of AAAP, carriers of risk genotypes of candidate genes need not only to exclude or significantly reduce alcohol consumption (in terms of volume, frequency and duration); but also carriers of the A/G-A/A CFTR genotypes (rs213950) need to balance the diet by reducing fat intake to less than 89 g per day and increasing protein intake to more than 84 g per day; carriers of the T/C-T/T PRSS1 (rs10273639) genotypes should increase their intake of fresh vegetables and fruits over 27 g/day and protein over 84 g/day.
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Regulador de Conductancia de Transmembrana de Fibrosis Quística , Interacción Gen-Ambiente , Pancreatitis Alcohólica , Humanos , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Proteínas en la Dieta/uso terapéutico , Frutas , Pancreatitis/etiología , Pancreatitis/genética , Pancreatitis/prevención & control , Pancreatitis Alcohólica/etiología , Pancreatitis Alcohólica/genética , Pancreatitis Alcohólica/prevención & control , Polimorfismo Genético , Polimorfismo de Nucleótido Simple , Calidad de Vida , Tripsina/genética , Inhibidor de Tripsina Pancreática de Kazal/genética , Verduras , Estilo de Vida SaludableRESUMEN
BACKGROUND: Alcohol abuse, a main cause of pancreatitis, has been known to augment NF-κB activation and cell necrosis in pancreatitis. However, the underlying mechanisms are unclear. We recently reported that inhibition of protein kinase D (PKD) alleviated NF-κB activation and severity of experimental pancreatitis. Here we investigated whether PKD signaling mediated the modulatory effects of alcohol abuse on pathological responses in alcoholic pancreatitis. METHODS: Alcoholic pancreatitis was provoked in two rodent models with pair-feeding control and ethanol-containing Lieber-DeCarli diets for up to 8 weeks followed by up to 7 hourly intraperitoneal injections of cerulein at 1 µg/kg (rats) or 3 µg/kg (mice). Effects of PKD inhibition by PKD inhibitors or genetic deletion of pancreatic PKD isoform (PKD3Δpanc mice) on alcoholic pancreatitis parameters were determined. RESULTS: Ethanol administration amplified PKD signaling by promoting expression and activation of pancreatic PKD, resulted in augmented/promoted pancreatitis responses. Pharmacological inhibition of PKD or with PKD3Δpanc mice prevented the augmenting/sensitizing effect of ethanol on NF-κB activation and inflammatory responses, cell necrotic death and the severity of disease in alcoholic pancreatitis. PKD inhibition prevented alcohol-enhanced trypsinogen activation, mRNA expression of multiple inflammatory molecules, the receptor-interacting protein kinase activation, ATP depletion, and downregulation of pro-survival Bcl-2 protein in alcoholic pancreatitis. Furthermore, PKD inhibitor CID755673 or CRT0066101, administrated after the induction of pancreatitis in mouse and rat alcoholic pancreatitis models, significantly mitigated the severity of pancreatitis. CONCLUSION: PKD mediates effect of alcohol abuse on pathological process of pancreatitis and constitutes a novel therapeutic target to treat this disease.
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Alcoholismo , Pancreatitis Alcohólica , Adenosina Trifosfato , Alcoholismo/complicaciones , Alcoholismo/tratamiento farmacológico , Alcoholismo/genética , Animales , Ceruletida , Etanol/toxicidad , Ratones , FN-kappa B/metabolismo , Necrosis , Pancreatitis Alcohólica/tratamiento farmacológico , Pancreatitis Alcohólica/genética , Pancreatitis Alcohólica/metabolismo , Proteína Quinasa C/genética , Proteínas Proto-Oncogénicas c-bcl-2 , ARN Mensajero , Ratas , TripsinógenoRESUMEN
The digestive protease chymotrypsin C (CTRC) protects the pancreas against pancreatitis by degrading potentially harmful trypsinogen. Loss-of-function genetic variants in CTRC increase risk for chronic pancreatitis (CP) with variable effect size, as judged by the reported odds ratio (OR) values. Here, we performed a meta-analysis of published studies on four variants that alter the CTRC amino-acid sequence, are clinically relatively common (global carrier frequency in CP >1%), reproducibly showed association with CP and their loss of function phenotype was verified experimentally. We found strong enrichment of CTRC variants p.A73T, p.V235I, p.K247_R254del, and p.R245W in CP cases versus controls, yielding OR values of 6.5 (95% confidence interval (CI) 2.4-17.8), 4.5 (CI 2.2-9.1), 5.4 (CI 2.6-11.0), and 2.6 (CI 1.6-4.2), respectively. Subgroup analysis demonstrated disease association of variants p.K247_R254del and p.R245W in alcoholic CP with similar effect sizes as seen in the overall CP group. Homozygosity or compound heterozygosity were rare and seemed to be associated with higher risk. We also identified a so far unreported linkage disequilibrium between variant p.K247_R254del and the common c.180C>T (p.G60 =) haplotype. Taken together, the results indicate that heterozygous loss-of-function CTRC variants increase the risk for CP approximately 3-7-fold. This meta-analysis confirms the clinical significance of CTRC variants and provides further justification for the genetic screening of CP patients.
Asunto(s)
Quimotripsina , Pancreatitis Alcohólica , Pancreatitis Crónica , Quimotripsina/genética , Predisposición Genética a la Enfermedad , Humanos , Mutación , Pancreatitis Alcohólica/genética , Pancreatitis Crónica/genéticaRESUMEN
BACKGROUND: Previous genome-wide association studies (GWAS) identified genome-wide significant risk loci in chronic pancreatitis and investigated underlying disease causing mechanisms by simple overlaps with expression quantitative trait loci (eQTLs), a procedure which may often result in false positive conclusions. METHODS: We conducted a GWAS in 584 non-alcoholic chronic pancreatitis (NACP) patients and 6040 healthy controls. Next, we applied Bayesian colocalization analysis of identified genome-wide significant risk loci from both, our recently published alcoholic chronic pancreatitis (ACP) and the novel NACP dataset, with pancreas eQTLs from the GTEx V8 European cohort to prioritize candidate causal genes and extracted credible sets of shared causal variants. RESULTS: Variants at the CTRC (p = 1.22 × 10-21) and SPINK1 (p = 6.59 × 10-47) risk loci reached genome-wide significance in NACP. CTRC risk variants colocalized with CTRC eQTLs in ACP (PP4 = 0.99, PP4/PP3 = 95.51) and NACP (PP4 = 0.99, PP4/PP3 = 95.46). For both diseases, the 95% credible set of shared causal variants consisted of rs497078 and rs545634. CLDN2-MORC4 risk variants colocalized with CLDN2 eQTLs in ACP (PP4 = 0.98, PP4/PP3 = 42.20) and NACP (PP4 = 0.67, PP4/PP3 = 7.18), probably driven by the shared causal variant rs12688220. CONCLUSIONS: A shared causal CTRC risk variant might unfold its pathogenic effect in ACP and NACP by reducing CTRC expression, while the CLDN2-MORC4 shared causal variant rs12688220 may modify ACP and NACP risk by increasing CLDN2 expression.
Asunto(s)
Estudio de Asociación del Genoma Completo , Pancreatitis Alcohólica , Teorema de Bayes , Predisposición Genética a la Enfermedad , Humanos , Proteínas Nucleares , Páncreas , Pancreatitis Alcohólica/genética , Polimorfismo de Nucleótido Simple , Sitios de Carácter Cuantitativo/genética , Inhibidor de Tripsina Pancreática de Kazal/genéticaRESUMEN
BACKGROUND: Genetic mutations in various pancreatic enzymes or their counteracting proteins have been linked to chronic pancreatitis. In particular, variants in the genes encoding pancreatic lipase (PNLIP) and carboxyl ester lipase (CEL) have been associated with pancreatitis. Therefore, we investigated pancreatic phospholipase A2 (PLA2G1B) as a promising candidate gene in patients with chronic pancreatitis. METHODS: We analyzed all coding exons and adjacent intronic regions of PLA2G1B in 416 German patients with non-alcoholic chronic pancreatitis (NACP) and 186 control subjects by direct DNA sequencing. RESULTS: We detected 2 frequent synonymous variants in exon 3: c.222T>C (p.Y74 = ) and c.294G>A (p.S98 = ). The genotype and allele frequencies of these variants were similar between patients and controls (c.222 TC: 9.6% in NACP vs. 9.7% in controls; c.222CC: 0.2% in NACP vs. 0% in controls; c.294 GA: 31.3% in NACP vs. 28.0% in controls; c.294AA: 2.4% in NACP vs. 1.1% in controls). All p-values were non-significant. In addition, we found one synonymous variant, c.138C>T (p.N46 = ) and one non-synonymous variant, c.244A>G (p.S82G), in a single case each. CONCLUSIONS: Our results suggest that genetic alterations in PLA2G1B do not predispose to the development of non-alcoholic chronic pancreatitis.
Asunto(s)
Pancreatitis Alcohólica , Pancreatitis Crónica , Frecuencia de los Genes , Pruebas Genéticas , Fosfolipasas A2 Grupo IB/genética , Humanos , Pancreatitis Alcohólica/genética , Pancreatitis Crónica/genética , Análisis de Secuencia de ADNRESUMEN
OBJECTIVE: The carboxyl-ester lipase (CEL) gene contains a variable number of tandem repeats (VNTR) region. It remains unclear whether the number of repeats in the CEL VNTR is related to the risk of pancreatic diseases. The aim of this study was to investigate whether CEL VNTR length is associated with idiopathic chronic pancreatitis (ICP), alcoholic chronic pancreatitis (ACP), or pancreatic cancer in a cohort of Chinese patients. METHODS: CEL VNTRs were genotyped in patients diagnosed with ICP (n = 771), ACP (n = 222), or pancreatic cancer (n = 263), and in healthy controls (n = 927). CEL VNTR lengths were determined using a screening method combining PCR and DNA fragment analysis. RESULTS: Overall, the CEL VNTR lengths ranged from 5 to 22 repeats, with the 16-repeat allele ('normal' size, N) accounting for 73.82% of all observed alleles. The VNTR allele frequencies and genotype distributions were not significantly different between healthy controls and patients with ACP or pancreatic cancer. For the ICP group, allele frequencies did not differ significantly from the controls, while the frequency of the SS genotype (homozygosity for 5-15 repeats) was significantly higher in the patients (4.67%) than in the controls (1.94%) (p = 0.0014; OR = 2.47; 95% CI = 1.39-4.39). CONCLUSIONS: There were no associations between the CEL VNTR length and ACP or pancreatic cancer. However, homozygosity for short VNTR lengths may confer susceptibility to ICP.
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Repeticiones de Minisatélite , Pancreatitis , Carboxilesterasa/genética , Carboxilesterasa/metabolismo , Frecuencia de los Genes , Genotipo , Heterocigoto , Humanos , Lipasa/metabolismo , Repeticiones de Minisatélite/genética , Neoplasias Pancreáticas/genética , Pancreatitis Alcohólica/genética , Neoplasias PancreáticasRESUMEN
The association between a common PRSS1-PRSS2 haplotype and alcoholic chronic pancreatitis (ACP), which was revealed by the first genome-wide association study of chronic pancreatitis (CP), has been consistently replicated. However, the association with non-ACP (NACP) has been controversial. Herein, we sought to clarify this basic issue by means of an allele-based meta-analysis of currently available studies. We then used studies informative for genotype distribution to explore the biological mechanisms underlying the association data and to test for gene-environment interaction between the risk haplotype and alcohol consumption by means of a re-analysis. A literature search was conducted to identify eligible studies. A meta-analysis was performed using the Review Manager software. The association between the risk genotypes and NACP or ACP was tested for the best-fitting genetic model. Gene-environment interaction was estimated by both case-only and multinomial approaches. Five and eight studies were employed for the meta-analysis of ACP and NACP findings, respectively. The risk allele was significantly associated with both ACP (pooled odds ratio (OR) 1.67, 95% confidence interval (CI) 1.56-1.78; p < 0.00001) and NACP (pooled OR 1.28, 95% CI 1.17-1.40; p < 0.00001). Consistent with a dosage effect of the risk allele on PRSS1/PRSS2 mRNA expression in human pancreatic tissue, both ACP and NACP association data were best explained by an additive genetic model. Finally, the risk haplotype was found to interact synergistically with alcohol consumption.
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Pancreatitis Alcohólica/genética , Pancreatitis Crónica/genética , Tripsina/genética , Tripsinógeno/genética , Consumo de Bebidas Alcohólicas/efectos adversos , Consumo de Bebidas Alcohólicas/genética , Expresión Génica , Frecuencia de los Genes , Interacción Gen-Ambiente , Haplotipos , Humanos , Modelos Genéticos , Polimorfismo de Nucleótido SimpleRESUMEN
BACKGROUND: The G-protein-coupled receptor Class C Group 6 Member A (GPRC6A) is activated by multiple ligands and is important for the regulation of calcium homeostasis. Extracellular calcium is capable to increase NLRP3 inflammasome activity of the innate immune system and deletion of this proinflammatory pathway mitigated pancreatitis severity in vivo. As such this pathway and the GPRC6A receptor is a reasonable candidate gene for pancreatitis. Here we investigated the prevalence of sequence variants in the GPRC6A locus in different pancreatitis aetiologies. METHODS: We selected 6 tagging SNPs with the SNPinfo LD TAG SNP Selection tool and the functional relevant SNP rs6907580 for genotyping. Cohorts from Germany, further European countries and China with up to 1,124 patients and 1,999 controls were screened for single SNPs with melting curve analysis. RESULTS: We identified an association of rs1606365(G) with alcoholic chronic pancreatitis in a German (odds ratio (OR) 0.76, 95% confidence interval (CI) 0.65-0.89, p = 8 × 10-5) and a Chinese cohort (OR 0.78, 95% CI 0.64-0.96, p = 0.02). However, this association was not replicated in a combined cohort of European patients (OR 1.18, 95% CI 0.99-1.41, p = 0.07). Finally, no association was found with acute and non-alcoholic chronic pancreatitis. CONCLUSIONS: Our results support a potential role of calcium sensing receptors and inflammasome activation in alcoholic chronic pancreatitis development. As the functional consequence of the associated variant is unclear, further investigations might elucidate the relevant mechanisms.
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Pancreatitis/genética , Receptores Acoplados a Proteínas G/genética , Adulto , Anciano , Pueblo Asiatico , ADN/genética , Europa (Continente) , Femenino , Predisposición Genética a la Enfermedad , Variación Genética , Estudio de Asociación del Genoma Completo , Humanos , Masculino , Persona de Mediana Edad , Pancreatitis Alcohólica/genética , Polimorfismo de Nucleótido Simple , Receptores Sensibles al Calcio/genética , Factores de Riesgo , Transducción de Señal/genética , Población BlancaRESUMEN
Alcoholic pancreatitis is a multifactorial, progressive, inflammatory disorder of the pancreas. Alcohol initiates pancreatitis and promotes its progression in the context of genetic susceptibility and/or other environmental risk factors such as smoking. Genetic mutations can cause digestive enzyme misfolding, which induces endoplasmic reticulum (ER) stress and elicits pancreatitis. Here, we tested the hypothesis that alcohol synergizes with misfolding in promoting ER stress and thereby accelerates chronic pancreatitis progression. To this end, we fed an ethanol-containing diet to CPA1 N256K mice, which carry the human p.N256K CPA1 mutation and develop spontaneous chronic pancreatitis. Inexplicably, CPA1 N256K mice suffered generalized seizures after 2-3 wk of ethanol feeding, which resulted in high mortality and the early termination of the study. Analysis of CPA1 N256K mice euthanized after 3-3.5 wk of ethanol feeding revealed more severe chronic pancreatitis associated with significantly increased Hspa5 [ER chaperone immunoglobulin heavy chain-binding protein (BiP)] mRNA levels when compared with CPA1 N256K mice on a control liquid diet. In contrast, ethanol feeding of C57BL/6N mice for 4 wk increased Hspa5 levels to a lesser degree and caused no pancreatitis. We conclude that ethanol feeding synergizes with the misfolding CPA1 mutant in promoting ER stress and thereby accelerates progression of chronic pancreatitis in CPA1 N256K mice.NEW & NOTEWORTHY Alcoholic pancreatitis is a multifactorial, progressive, inflammatory disorder of the pancreas. This study demonstrates that alcohol synergizes with digestive enzyme misfolding in promoting endoplasmic reticulum stress and thereby accelerates progression of chronic pancreatitis.
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Carboxipeptidasas A/metabolismo , Etanol/toxicidad , Páncreas/efectos de los fármacos , Pancreatitis Alcohólica/genética , Animales , Peso Corporal , Carboxipeptidasas A/genética , Ingestión de Alimentos , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Etanol/administración & dosificación , Predisposición Genética a la Enfermedad , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Pancreatitis Alcohólica/patologíaRESUMEN
BACKGROUND: Genetic predisposition plays an important role in the development of alcoholic pancreatitis (AP), with previous studies suggesting that genetics variants in certain genes, such asCYP2E1 and CTRC, partially explain individual susceptibility to this disease. Therefore, the aim of this work was to conduct a systematic review and meta-analysis of existing studies that analyzed how polymorphisms within CYP2E1 and CTRC genes influence the risk of AP. MATERIAL AND METHODS: We performed a systematic review of studies that analyzed the genotype distribution of CYP2E1 and CTRC allelic variants among patients with AP and a group of controls. A meta-analysis was conducted using a random effects model. Odds ratios (ORs) and their confidence intervals (CIs) were calculated. RESULTS: The T allele of theCTRC 180 C > T variant was significantly more prevalent among patients with AP compared to all controls (OR = 1.79, 95% CI = 1.43-2.24; P < 0.00001) and healthy subjects (OR = 1.84, 95% CI = 1.46-2.31; P < 0.00001). The Trp variant of CTRC Arg254Trp polymorphism was also more prevalent in patients with AP; however, these results were not significant after excluding one study. We found no clear evidence that CYP2E1-DraI or of CYP2E1-RsaI/PstI polymorphisms modulate the risk of developing AP. CONCLUSIONS: Our meta-analysis supports that the T allele ofCTRC 180C > T polymorphisms modulates the risk of alcoholic pancreatitis. No clear evidence was found for the remaining SNPs being associated with this disease.
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Quimotripsina/genética , Citocromo P-450 CYP2E1/genética , Estudios de Asociación Genética/métodos , Predisposición Genética a la Enfermedad/genética , Variación Genética/genética , Pancreatitis Alcohólica/genética , Estudios de Casos y Controles , Predisposición Genética a la Enfermedad/epidemiología , Humanos , Pancreatitis Alcohólica/diagnóstico , Pancreatitis Alcohólica/epidemiología , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
BACKGROUND/OBJECTIVES: Carboxyl ester lipase is a pancreatic enzyme encoded by CEL, an extremely polymorphic human gene. Pathogenic variants of CEL either increases the risk for chronic pancreatitis (CP) or cause MODY8, a syndrome of pancreatic exocrine and endocrine dysfunction. Here, we aimed to characterize a novel duplication allele of CEL (CEL-DUP2) and to investigate whether it associates with CP or pancreatic cancer. METHODS: The structure of CEL-DUP2 was determined by a combination of Sanger sequencing, DNA fragment analysis, multiplex ligation-dependent probe amplification and whole-genome sequencing. We developed assays for screening of CEL-DUP2 and analyzed cohorts of idiopathic CP, alcoholic CP and pancreatic cancer. CEL protein expression was analyzed by immunohistochemistry. RESULTS: CEL-DUP2 consists of an extra copy of the complete CEL gene. The allele has probably arisen from non-allelic, homologous recombination involving the adjacent pseudogene of CEL. We found no association between CEL-DUP2 carrier frequency and CP in cohorts from France (cases/controls: 2.5%/2.4%; P = 1.0), China (10.3%/8.1%; P = 0.08) or Germany (1.6%/2.3%; P = 0.62). Similarly, no association with disease was observed in alcohol-induced pancreatitis (Germany: 3.2%/2.3%; P = 0.51) or pancreatic cancer (Norway; 2.5%/3.2%; P = 0.77). Notably, the carrier frequency of CEL-DUP2 was more than three-fold higher in Chinese compared with Europeans. CEL protein expression was similar in tissues from CEL-DUP2 carriers and controls. CONCLUSIONS: Our results support the contention that the number of CEL alleles does not influence the risk of pancreatic exocrine disease. Rather, the pathogenic CEL variants identified so far involve exon 11 sequence changes that substantially alter the protein's tail region.
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Lipasa/genética , Pancreatitis Crónica/epidemiología , Pancreatitis Crónica/genética , Adulto , Anciano , Alelos , ADN/genética , Femenino , Duplicación de Gen , Frecuencia de los Genes , Pruebas Genéticas , Heterocigoto , Humanos , Masculino , Persona de Mediana Edad , Páncreas/patología , Neoplasias Pancreáticas/epidemiología , Neoplasias Pancreáticas/genética , Pancreatitis Alcohólica/epidemiología , Pancreatitis Alcohólica/genética , Pancreatitis Crónica/patología , RiesgoRESUMEN
INTRODUCTION: Chronic pancreatitis (CP) may be caused by oxidative stress. An important source of reactive oxygen species (ROS) is the methylglyoxal-derived formation of advanced glycation endproducts (AGE). Methylglyoxal is detoxified by Glyoxalase I (GLO1). A reduction in GLO1 activity results in increased ROS. Single nucleotide polymorphisms (SNPs) of GLO1 have been linked to various inflammatory diseases. Here, we analyzed whether common GLO1 variants are associated with alcoholic (ACP) and non-alcoholic CP (NACP). METHODS: Using melting curve analysis, we genotyped a screening cohort of 223 ACP, 218 NACP patients, and 328 controls for 11 tagging SNPs defined by the SNPinfo LD TAG SNP Selection tool and the functionally relevant variant rs4746. For selected variants the cohorts were extended to up to 1,441 patient samples. RESULTS: In the ACP cohort, comparison of genotypes for rs1937780 between patients and controls displayed an ambiguous result in the screening cohort (p = 0.08). However, in the extended cohort of 1,441 patients no statistically significant association was found for the comparison of genotypes (p = 0.11), nor in logistic regression analysis (p = 0.214, OR 1.072, 95% CI 0.961-1.196). In the NACP screening cohort SNPs rs937662, rs1699012, and rs4746 displayed an ambiguous result when patients were compared to controls in the recessive or dominant model (p = 0.08, 0.08, and 0.07, respectively). Again, these associations were not confirmed in the extended cohorts (rs937662, dominant model: p = 0.07, logistic regression: p = 0.07, OR 1.207, 95% CI 0.985-1.480) or in the replication cohorts for rs4746 (Germany, p = 0.42, OR 1.080, 95% CI 0.673-1.124; France, p = 0.19, OR 0.90, 95% CI 0.76-1.06; China, p = 0.24, OR 1.18, 95% CI 0.90-1.54) and rs1699012 (Germany, Munich; p = 0.279, OR 0.903, 95% CI 0.750-1.087). CONCLUSIONS: Common GLO1 variants do not increase chronic pancreatitis risk.
Asunto(s)
Predisposición Genética a la Enfermedad , Lactoilglutatión Liasa/genética , Pancreatitis Alcohólica/genética , Pancreatitis Crónica/genética , Femenino , Estudios de Asociación Genética , Genotipo , Productos Finales de Glicación Avanzada/genética , Humanos , Masculino , Persona de Mediana Edad , Estrés Oxidativo/genética , Pancreatitis Alcohólica/metabolismo , Pancreatitis Alcohólica/patología , Pancreatitis Crónica/metabolismo , Pancreatitis Crónica/patología , Polimorfismo de Nucleótido Simple/genética , Piruvaldehído/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factores de RiesgoRESUMEN
BACKGROUND: Genetic mutations are one of the etiological factors that predispose people to develop chronic pancreatitis. OBJECTIVES: The aim of our study was to examine the effect of p.Trp55*, p.Arg254Trp and c.738_761del mutations in the chemotrypsin gene (CTRC) on the development of alcoholic chronic pancreatitis (ACP) in order to answer the questions whether these mutations vary between gender groups, whether they were related to the age when ACP was first diagnosed, and whether they affected the morphological changes in the pancreas and the course of ACP. MATERIAL AND METHODS: The study included 124 patients with ACP, 52 with nonalcoholic pancreatitis and 52 controls. The p.Trp55*, c.738_761del and p.Arg254Trp mutations in the CTRC gene were tested by the polymerase chain reaction (PCR). RESULTS: The c.738_761del and p.Arg254Trp mutations occurred in 3.07% and 1.31% of cases, respectively. None of the examined patients were found to have the p.Trp55* mutation. The frequency of detected mutations did not significantly differ between the study groups. The c.738_761del mutation was detected more frequently in women than in men. No significant differences were found in the age at ACP onset, morphological changes affecting the pancreas, or in the course of ACP between the patients with and without the 2 examined mutations. The c.738_761del mutation was significantly more frequent in the diabetic patients than in the non-diabetics. The patients with this mutation more frequently required surgery than those without the c.738_761del mutation. CONCLUSIONS: No relationship between the c.738_761del and p.Arg254Trp mutations and the development of APC was found. The c.738_761del mutation was more frequent in females than in males. Neither mutation affected the patient's age at ACP onset or its course. In contrast to p.Arg254Trp, the c.738_761del mutation correlated with diabetes development and the need for surgery in the course of ACP.
Asunto(s)
Quimotripsina/genética , Predisposición Genética a la Enfermedad , Mutación/genética , Pancreatitis Alcohólica/genética , Pancreatitis/genética , Población Blanca/genética , Femenino , Humanos , Masculino , Pancreatitis Alcohólica/etnología , Pancreatitis Crónica/etnología , Pancreatitis Crónica/genética , Polonia , Reacción en Cadena de la Polimerasa , Inhibidor de Tripsina Pancreática de Kazal , Población Blanca/etnologíaRESUMEN
OBJECTIVES: Rare pathogenic variants in the SPINK1, PRSS1, CTRC, and CFTR genes have been strongly associated with a risk of developing chronic pancreatitis (CP). However, their potential impact on the age of disease onset and clinical outcomes, as well as their potential interactions with environmental risk factors, remain unclear. These issues are addressed here in a large Chinese CP cohort. METHODS: We performed targeted next-generation sequencing of the four CP-associated genes in 1061 Han Chinese CP patients and 1196 controls. To evaluate gene-environment interactions, the patients were divided into three subgroups, idiopathic CP (ICP; n = 715), alcoholic CP (ACP; n = 206), and smoking-associated CP (SCP; n = 140). The potential impact of rare pathogenic variants on the age of onset of CP and clinical outcomes was evaluated using the Kaplan-Meier model. RESULTS: We identified rare pathogenic genotypes involving the SPINK1, PRSS1, CTRC, and/or CFTR genes in 535 (50.42%) CP patients but in only 71 (5.94%) controls (odds ratio = 16.12; P < 0.001). Mutation-positive patients had significantly earlier median ages at disease onset and at diagnosis of pancreatic stones, diabetes mellitus and steatorrhea than mutation-negative ICP patients. Pathogenic genotypes were present in 57.1, 39.8, and 32.1% of the ICP, ACP, and SCP patients, respectively, and influenced age at disease onset and clinical outcomes in all subgroups. CONCLUSIONS: We provide evidence that rare pathogenic variants in the SPINK1, PRSS1, CTRC, and CFTR genes significantly influence the age of onset and clinical outcomes of CP. Extensive gene-environment interactions were also identified.
Asunto(s)
Edad de Inicio , Interacción Gen-Ambiente , Genotipo , Pancreatitis Crónica/genética , Adolescente , Adulto , Pueblo Asiatico/genética , Cálculos/diagnóstico , Quimotripsina/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Diabetes Mellitus/diagnóstico , Predisposición Genética a la Enfermedad , Humanos , Estimación de Kaplan-Meier , Persona de Mediana Edad , Mutación , Enfermedades Pancreáticas/diagnóstico , Pancreatitis Alcohólica/genética , Pancreatitis Crónica/diagnóstico , Fumar/efectos adversos , Esteatorrea/diagnóstico , Tripsina/genética , Inhibidor de Tripsina Pancreática de Kazal/genética , Adulto JovenRESUMEN
Excessive alcohol consumption leads to severe alcoholic hepatitis (sAH) or chronic alcoholic pancreatitis (CAP) only in a subset of patients. We aimed to characterize the intestinal microbiota profiles of alcoholic patients according to the presence and nature of the complications observed: sAH or CAP. Eighty two alcoholic patients were included according to their complications: CAP (N = 24), sAH (N = 13) or no complications (alcoholic controls, AC, N = 45). We analyzed the intestinal microbiota by high-throughput sequencing. Bacterial diversity was lower in patients with CAP, who had a global intestinal microbiota composition different from that of AC. The intestinal microbiota composition of these two groups differed for 17 genera, eight of which were more frequent in patients with CAP (e.g. Klebsiella, Enterococcus and Sphingomonas). There was no significant difference in bacterial diversity between the sAH and CAP groups. However, 16 taxa were more frequent in sAH patients, and 10 were more frequent in CAP patients. After adjustment for confounding factors sAH patients were found to have higher levels of Haemophilus. For alcoholic patients, specific intestinal microbiota signatures are associated with different complications. Patients with CAP and sAH also display specific dysbiosis relative to AC.
Asunto(s)
Disbiosis/epidemiología , Heces/microbiología , Microbioma Gastrointestinal/genética , Hepatitis Alcohólica/microbiología , Pancreatitis Alcohólica/microbiología , Adolescente , Adulto , Anciano , Estudios de Casos y Controles , Femenino , Francia/epidemiología , Hepatitis Alcohólica/genética , Hepatitis Alcohólica/patología , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Pancreatitis Alcohólica/genética , Pancreatitis Alcohólica/patología , Estudios Prospectivos , ARN Ribosómico 16S/genética , Adulto JovenRESUMEN
Pancreatic stellate cells (PSCs) play a critical role in fibrogenesis during alcoholic chronic pancreatitis (ACP). Transforming growth factor-beta1 (TGF-ß1) is a key regulator of extracellular matrix production and PSC activation. Endotoxin lipopolysaccharide (LPS) has been recognized as a trigger factor in the pathogenesis of ACP. This study aimed to investigate the mechanisms by which LPS modulates TGF-ß1 signalling and pancreatic fibrosis. Sprague-Dawley rats fed with a Lieber-DeCarli alcohol (ALC) liquid diet for 10 weeks with or without LPS challenge during the last 3 weeks. In vitro studies were performed using rat macrophages (Mφs) and PSCs (RP-2 cell line). The results showed that repeated LPS challenge resulted in significantly more collagen production and PSC activation compared to rats fed with ALC alone. LPS administration caused overexpression of pancreatic TLR4 or TGF-ß1 which was paralleled by an increased number of TLR4-positive or TGF-ß1-positive Mφs or PSCs in ALC-fed rats. In vitro, TLR4 or TGF-ß1 production in Mφs or RP-2 cells was up-regulated by LPS. LPS alone or in combination with TGF-ß1 significantly increased type I collagen and α-SMA production and Smad2 and 3 phosphorylation in serum-starved RP-2 cells. TGF-ß pseudoreceptor BAMBI production was repressed by LPS, which was antagonized by Si-TLR4 RNA or by inhibitors of MyD88/NF-kB. Additionally, knockdown of Bambi with Si-Bambi RNA significantly increased TGF-ß1 signalling in RP-2 cells. These findings indicate that LPS increases TGF-ß1 production through paracrine and autocrine mechanisms and that LPS enhances TGF-ß1 signalling in PSCs by repressing BAMBI via TLR4/MyD88/NF-kB activation.
Asunto(s)
Fibrosis/tratamiento farmacológico , Fibrosis/genética , Proteínas de la Membrana/genética , Pancreatitis Alcohólica/genética , Factor de Crecimiento Transformador beta1/genética , Alcoholes/toxicidad , Animales , Colágeno/biosíntesis , Fibrosis/inducido químicamente , Fibrosis/patología , Regulación de la Expresión Génica/genética , Humanos , Lipopolisacáridos/administración & dosificación , Macrófagos/efectos de los fármacos , Factor 88 de Diferenciación Mieloide/genética , Páncreas/efectos de los fármacos , Páncreas/metabolismo , Páncreas/patología , Células Estrelladas Pancreáticas/efectos de los fármacos , Células Estrelladas Pancreáticas/metabolismo , Pancreatitis Alcohólica/inducido químicamente , Pancreatitis Alcohólica/patología , Ratas , Transducción de Señal , Proteína Smad2/genética , Receptor Toll-Like 4/genéticaRESUMEN
OBJECTIVE: Alcohol-related pancreatitis is associated with a disproportionately large number of hospitalisations among GI disorders. Despite its clinical importance, genetic susceptibility to alcoholic chronic pancreatitis (CP) is poorly characterised. To identify risk genes for alcoholic CP and to evaluate their relevance in non-alcoholic CP, we performed a genome-wide association study and functional characterisation of a new pancreatitis locus. DESIGN: 1959 European alcoholic CP patients and population-based controls from the KORA, LIFE and INCIPE studies (n=4708) as well as chronic alcoholics from the GESGA consortium (n=1332) were screened with Illumina technology. For replication, three European cohorts comprising 1650 patients with non-alcoholic CP and 6695 controls originating from the same countries were used. RESULTS: We replicated previously reported risk loci CLDN2-MORC4, CTRC, PRSS1-PRSS2 and SPINK1 in alcoholic CP patients. We identified CTRB1-CTRB2 (chymotrypsin B1 and B2) as a new risk locus with lead single-nucleotide polymorphism (SNP) rs8055167 (OR 1.35, 95% CI 1.23 to 1.6). We found that a 16.6 kb inversion in the CTRB1-CTRB2 locus was in linkage disequilibrium with the CP-associated SNPs and was best tagged by rs8048956. The association was replicated in three independent European non-alcoholic CP cohorts of 1650 patients and 6695 controls (OR 1.62, 95% CI 1.42 to 1.86). The inversion changes the expression ratio of the CTRB1 and CTRB2 isoforms and thereby affects protective trypsinogen degradation and ultimately pancreatitis risk. CONCLUSION: An inversion in the CTRB1-CTRB2 locus modifies risk for alcoholic and non-alcoholic CP indicating that common pathomechanisms are involved in these inflammatory disorders.
Asunto(s)
Quimotripsina/genética , Pancreatitis Alcohólica , Adulto , Anciano , Europa (Continente)/epidemiología , Femenino , Predisposición Genética a la Enfermedad , Humanos , Masculino , Persona de Mediana Edad , Pancreatitis Alcohólica/epidemiología , Pancreatitis Alcohólica/genética , Polimorfismo de Nucleótido SimpleRESUMEN
AIMS: Oxidative stress may contribute to the development of chronic pancreatitis (CP). The enzymes manganese superoxide dismutase 2 (MnSOD, SOD2) and catalase (CAT) counteract free radical activity within the mitochondria and the cytosol. Moreover, CAT activity contributes to the transformation of ethanol to acetaldehyde, a toxic intermediate product of ethanol metabolism, which has been associated with pancreatic damage. Common functional polymorphisms have been described in the MnSOD gene [rs4880, NM_000636.3:c.47 T > C, alanine (ALA) to valine (Val)] and in the CAT promoter region [rs1001179, NG_013339.1:g.4760 C > T]. We investigated whether these polymorphisms are associated with alcoholic CP. METHODS: We genotyped 470 patients with alcoholic CP for these MnSOD and CAT polymorphisms. We also analysed these variants in 357 healthy control subjects, and in an additional control group of 113 individuals with non-alcoholic CP. We used the age at onset of CP as marker of disease severity and investigated whether different genotypes are associated with different ages at onset. In patients with alcoholic CP, we investigated whether an interaction exists between smoking behaviour and genotypes by comparing genotype distributions in smokers and non-smokers. RESULTS: We did not observe significant differences of genotype frequencies between patient groups and controls. In patient groups, we did not find significant differences in the ages at onset between different genotypes. We did not observe an interaction between these polymorphisms. We did not find an association of these variants with smoking behaviour. CONCLUSIONS: The investigated MnSOD and CAT polymorphisms do not predispose to the development of alcoholic CP. SHORT SUMMARY: Patients with alcoholic pancreatitis and controls were genotyped for polymorphisms in oxidative stress genes. There were no significant differences of genotype frequencies between patients and controls, and no association with the age at onset of disease was observed. The polymorphisms are not associated with the development of alcoholic pancreatitis.