Screening And Validation Of Susceptible Genes For Chronic Pancreatitis

Section 1: Comprehensive Screening for PRSS1, SPINK1, CFTR, CTRC and CLDN2 Gene Mutations in Chinese Pediatric Patients with Idiopathic Chronic PancreatitisObjective: This study was designed to investigate the mutation frequecncies of PRSS1, SPINK1, CFTR, CTRC, and CLDN2 genes in Chinese children with idiopathic chronic pancreatitis and further evaluate the correlations with clinical characteristics.Methods: Related clinical data and peripheral blood samples were collected. The PRSS1, SPINK1, CFTR, CTRC and CLDN2 gene mutations were detected using DNA sequencing and analyzed by correlating with certain clinical characteristics.Results: From January 1997 to December 2008, a total of 75 idiopathic chronic pancreatitis children(40 boys and 35 girls) were recruited. The median age at first onset was 11.9 years. Frequency of PRSS1 and SPINK1 mutations was 9.3% and 57.3%, respectively, with an overall frequency of 66.6%(50/75). Seven patients had heterozygous mutations in PRSS1, including N29I(n=1), R122 H or R122C(n=6). Forty-three patients had IVS3+2T>C(10 homozygous and 33 heterozygous) in SPINK1. In addition, one patient had a novel deletion of CFTR. Another patient had a novel missense in CLDN2 exon 2(c.592A>C mutation). No patients carried CTRC missense mutation. Clinically, patients with SPINK1 IVS3+2T>C mutation had a higher rate of pancreatic duct stones, pancreatic pseudocyst, and pancreatic calcification than those without(P<0.05).Conclusion: SPINK1 mutations were more commonly associated with Chinese idiopathic chronic pancreatitis children. SPINK1 IVS3+2T>C mutation may play an important role in the pathogenesis of Chinese pediatric idiopathic chronic pancreatitis. However, further study is needed to confirm and to investigate the potential molecular mechanism.Section 2: Clinical Features and Endoscopic Treatment of Chinese Patients with Hereditary PancreatitisObjective: Hereditary pancreatitis has been rarely investigated in China. We aimed to describe the clinical features and mutation frequency of Chinese patients with hereditary pancreatitis from single center and evaluate short-term and long-term outcomes of endoscopic treatments for such patients.Methods: Demographic and clinical data were collected. All the patients were followed up and evaluated 3 months after each discharge and in July 2013. Mutations in SPINK1, PRSS1 and CFTR genes were analyzed.Results: A total of 24 inpatients with hereditary pancreatitis(male 13; female 11) from 21 different families participated in this study. Mean age at first onset and at diagnosis were 25.5(6-46) years and 30.0(12-54) years, respectively. The predominant radiological feature was pancreatic calcifications(95.8%). Forty-one ERCP procedures were successfully performed in 21 cases and the other 3 patients underwent surgery due to no alleviation of the symptoms. In the final long-term follow-up, 21 patients got complete or incomplete remission after ERCP and/or surgery. Genetic analyses were available in 20 patients and mutation rates of R122 H, N29 I and A16 V in PRSS1 gene were 60.0%(n=12)?25.0%(n=5) and 5.0%(n=1).Conclusion: As compared with previous studies from western countries, our patient cohort had a relatively higher frequency of R122 H mutation, but showed a much lower surgery rate, indicating that endoscopic interventions may be recommended to be the first-line treatment and is recommended to be repeated and combined with ESWL if necessary.Section 3: Molecular Profiling of CPA1 gene in Chronic PancreatitisObjective: Chronic pancreatitis was proved to be associated with certain CPA1 genetic mutations, which was rarely reported in China. This present study aimed to describe the genetic profile of CPA1 gene and its clinical significance, which is a recently identified candidate for chronic pancreatitis, in a Chinese cohort.Methods: A total of 316 patients with sporadic non-alcoholic chronic pancreatitis and 256 healthy volunteers as controls were enrolled. Peripheral blood samples were collected and genomic DNA was extracted. All the exons of CPA1 gene were sequenced and the correlation between the mutations and clinical characteristics were analyzed.Results: The sequencing results revealed that a total of 14 variants in 10 exons including 10 missense mutationa and 4 synonymous mutations, were detected, but only the frequency of synonymous c.237 G>C variant was statistically significant between patients and controls(88.3% vs. 76.2%, P=0.000). The frequency of missense variants was 3.1% (10/316) in patient cohort and 3.5%(9/256) in controls, respectively. The correlation analysis of CPA1 variants with clinicoapathological characteristics including the onset age, symptoms, radiographical features and the incidence of post-ERCP pancreatitis were conducted and no statistically significant difference was found.Conclusion: The present study showed that the profiling of CPA1 gene in Chinese chronic pancreatitis population was different from that in other countries, which may be due to the geographical difference and the proportion of adult patients in the study cohort.Section 4: Pilot Investigation on the De novo Mutations in Adolescents with Idiopathic Chronic PancreatitisObjective: To enrich the understanding on the spectrum of adolescent idiopathic chronic pancreatitis related genetic mutations and explain why the detection rates of previously known pathogenic variants was low in our cohort, further providing new insight into clarifying the specific pathogenesis of idiopathic chronic pancreatitis.Methods: Adolescent patients diagnosed with idiopathic chronic pancreatitis were included. The blood samples from the patients and their parents were collected, and exon trapping combined with deep sequencing was applied to detect the de novo mutations by comparing the sequencing between the offsprings and the parents.Results: A total of 50 adolescent patients with idiopathic chronic pancreatitis were initially enrolled and 32 patients were excluded due to incomplete information and failure to obtain the consent from their parents. First-generation sequencing showed that 4 patients carried mutations in PRSS1 or SPINK1 gene and they were devoid of the second-generation sequencing. The deep sequencing analysis on 14 patients did not reveal any pathogenic candidates for young idiopathic chronic pancreatitis with healthy parents.Conclusion: Considering that the pathogenesis of chronic pancreatitis is complicated, future research should be devoted to including more participants, screening and validating the idiopathic chronic pancreatitis related de novo mutations, thus enriching the genetic study on chronic pancreatitis.