Gastroenterology

Gastroenterology

Volume 158, Issue 6, May 2020, Pages 1626-1641.e8
Gastroenterology

Original Research
Clinical—Pancreas
Variants That Affect Function of Calcium Channel TRPV6 Are Associated With Early-Onset Chronic Pancreatitis

https://doi.org/10.1053/j.gastro.2020.01.005Get rights and content

Background & Aims

Changes in pancreatic calcium levels affect secretion and might be involved in development of chronic pancreatitis (CP). We investigated the association of CP with the transient receptor potential cation channel subfamily V member 6 gene (TRPV6), which encodes a Ca2+-selective ion channel, in an international cohort of patients and in mice.

Methods

We performed whole-exome DNA sequencing from a patient with idiopathic CP and from his parents, who did not have CP. We validated our findings by sequencing DNA from 300 patients with CP (not associated with alcohol consumption) and 1070 persons from the general population in Japan (control individuals). In replication studies, we sequenced DNA from patients with early-onset CP (20 years or younger) not associated with alcohol consumption from France (n = 470) and Germany (n = 410). We expressed TRPV6 variants in HEK293 cells and measured their activity using Ca2+ imaging assays. CP was induced by repeated injections of cerulein in TRPV6mut/mut mice.

Results

We identified the variants c.629C>T (p.A210V) and c.970G>A (p.D324N) in TRPV6 in the index patient. Variants that affected function of the TRPV6 product were found in 13 of 300 patients (4.3%) and 1 of 1070 control individuals (0.1%) from Japan (odds ratio [OR], 48.4; 95% confidence interval [CI], 6.3–371.7; P = 2.4 × 10–8). Twelve of 124 patients (9.7%) with early-onset CP had such variants. In the replication set from Europe, 18 patients with CP (2.0%) carried variants that affected the function of the TRPV6 product compared with 0 control individuals (P = 6.2 × 10–8). Variants that did not affect the function of the TRPV6 product (p.I223T and p.D324N) were overrepresented in Japanese patients vs control individuals (OR, 10.9; 95% CI, 4.5–25.9; P = 7.4 × 10–9 for p.I223T and P = .01 for p.D324N), whereas the p.L299Q was overrepresented in European patients vs control individuals (OR, 3.0; 95% CI, 1.9–4.8; P = 1.2 × 10–5). TRPV6mut/mut mice given cerulein developed more severe pancreatitis than control mice, as shown by increased levels of pancreatic enzymes, histologic alterations, and pancreatic fibrosis.

Conclusions

We found that patients with early-onset CP not associated with alcohol consumption carry variants in TRPV6 that affect the function of its product, perhaps by altering Ca2+ balance in pancreatic cells. TRPV6 regulates Ca2+ homeostasis and pancreatic inflammation.

Section snippets

Study Participants

A 34-year-old man was referred to our hospital for examination of the cause of his CP. He had developed recurrent pancreatitis attacks since the age of 25 years. He had no history of drinking alcohol or smoking. His parents had no past history of pancreatitis.

For the validation study, we enrolled 300 (152 male and 148 female patients; median age at onset, 25 years; range, 1–70 years) Japanese patients with nonalcoholic CP, including 42 patients with family history of recurrent pancreatitis or

Whole-Exome Sequencing Identified p.A210V and De Novo p.D324N Variants in TRPV6

To identify novel pancreatitis susceptibility genes, we performed WES of a patient with idiopathic CP who developed symptoms at the age of 25 years. We identified 10,278 nonsynonymous or splice site variants in this patient. Among them, 421 variants were novel or rare (minor allele frequency, ≤0.01) in 1000 Genomes28 or in the Human Genetic Variation Database.29 After comparison of the data with the WES data of his clinically unaffected parents and visual inspection of the alignment quality

Discussion

Since the landmark discovery of PRSS1 mutations in patients with hereditary pancreatitis,4 the old concept of Hans Chiari that pancreatic inflammation results from autodigestion50 became the predominant pathophysiological model. The identification of mutations in additional susceptibility genes such as SPINK1, CTRC, CPA1, CEL, and PNLIP7, 8, 9, 10, 11 further underlined the enzyme-centered concept of a disturbed balance of pancreatic digestive enzymes and their specific inhibitors within the

Acknowledgments

The authors are grateful to Makiko Sumii, Yoko Tateda, Miyuki Tsuda, Mami Kikuchi, Makiko Nakagawa, and Kiyotaka Kuroda for the excellent technical assistance. The authors also acknowledge the technical support of the Biomedical Research Core of the Tohoku University Graduate School of Medicine.

CRediT Authorship Contributions

Atsushi Masamune, MD, PhD (Conceptualization: Lead; Data curation: Lead; Investigation: Lead; Methodology: Lead; Project administration: Lead; Resources: Lead; Writing – original draft: Lead; Writing –

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    Conflicts of interest The authors disclose no conflicts.

    Funding This work was supported in part by Grant-in-Aid from the Japan Society for the Promotion of Science (16K15421, 17K15916, 19K17450); Pancreas Research Foundation of Japan; the HIROMI Medical Research Foundation; the Mother and Child Health Foundation; the Smoking Research Foundation, the Ministry of Health, Labour, and Welfare of Japan; Conseil Régional de Bretagne; the Association des Pancréatites Chroniques Héréditaires; the Association de Transfusion Sanguine et de Biogénétique Gaetan Saleun; the Institut National de la Santé et de la Recherche Médicale, France; and the Else Kröner-Fresenius-Stiftung (2017_A108-EKFZ).

    Author names in bold designate shared co-first authorship.

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    Authors share co-senior authorship.

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