Conflicts of interest The authors have no potential conflicts of interest.
Dr D Nageshwar Reddy, Asian Healthcare Foundation, Institute of Basic Sciences, Asian Institute of Gastroenterology, 6-3-661, Somajiguda, Hyderabad- 500 082, Andhra Pradesh, India. Email: firstname.lastname@example.org
Chronic pancreatitis (CP) is a disease characterized by irreversible destruction and fibrosis of the parenchyma, leading to pancreatic exocrine insufficiency. In developed countries, the etiology for 60% to 70% of CP amongst male patients is alcohol and 25% are classified as idiopathic chronic pancreatitis (ICP). The genetic predisposition to CP could be an inappropriate activation of trypsinogen in the pancreas. Two common haplotypes, c.101A > G (p.N34S) and c.−215G > A, and four intronic alterations of the serine protease inhibitor Kazal type 1 (SPINK1) gene have been found to increase the risk for CP in the Asia Pacific region. Hence, SPINK1 is thought to be a candidate gene for pancreatitis. A loss-of-function alteration in chymotrypsinogen C (CTRC) gene has been shown to be associated with tropical calcific pancreatitis (TCP). Cathepsin B (CTSB) is also found to be associated with TCP. However mutations in cationic and anionic trypsinogen gene do not play an important role in causing CP in Asia Pacific region.
Chronic pancreatitis (CP) is an inflammatory disease which is characterized by irreversible destruction and fibrosis of the parenchyma, leading to pancreatic exocrine insufficiency and progressive endocrine failure leading to diabetes.1 In most developed countries, alcohol abuse causes about 60% to 70% of CP in male patients, and about 25% are classified as idiopathic chronic pancreatitis (ICP). Tropical calcific pancreatitis (TCP, OMIM 608189) is a juvenile form of chronic calcific non-alcoholic pancreatitis, seen almost exclusively in developing countries of the tropical world.2 A recent study in southern India has shown the prevalence of TCP to be 0.02% in the general population.3 TCP has been described as a disease with “pain in childhood, diabetes in puberty and death at the prime of life.”4
TCP was earlier seen only in children, adolescents, or sometimes young adults, who had common characteristics of malnutrition, deficiency signs, cyanotic hue of enlarged lips, bilateral enlargement of parotid glands, pot belly and pedal edema in a few. However, the clinical features and presentation of tropical pancreatitis have changed over a period of time with an older age of onset and a milder form of disease.5–8 The clinical manifestations of TCP are recurrent abdominal pain in childhood, followed by onset of diabetes mellitus a few years later. Prevalence of pancreatic calculi in TCP is nearly 90%, compared with the 30% of alcoholic pancreatitis.9 Histopathological changes include intralobular fibrosis in the early stage and interacinar fibrosis in later stages of the disease.1
Genetic basis of TCP
Genetic predisposition to CP due to heightened oxidative metabolism or depletion of antioxidant/conjugation capacity has been explored without consistent evidence of either.10–12 It was hypothesized that the primary step in the development of pancreatitis could be an inappropriate activation of trypsinogen in the pancreas.13,14 Three different trypsinogens—cationic, anionic and meso, representing 23.1%, 16% and 0.5% of total pancreatic secretory proteins, respectively—have been described in human pancreatic juice. Polymorphism in the respective genes could be the genetic basis of CP.15
It is postulated that 5% of trypsinogens are activated within the normal pancreas. There are safety mechanisms within the pancreas to protect it from premature activation of these enzymes which would otherwise cause autodigestion.16 A protein “pancreatic secretory trypsin inhibitor” (PSTI) or serine protease inhibitor Kazal type1 (SPINK1, OMIM 167790) is present in the secretory granules of acinar cells, which blocks the active site of trypsin in a 1:1 ratio and inhibits tryptic activities (Fig. 1). Two common haplotypes of the serine protease inhibitor Kazal type 1 (SPINK1) gene have been shown to increase the risk for CP. A haplotype comprising the c.101A > G (p.N34S) missense variant and four intronic alterations have been found worldwide. In a recent study, a second haplotype consisting of the c.−215G > A promoter variant and the c.194 + 2T > C intronic alteration has also been observed.17
Other safety mechanisms which inhibit the trypsin that leaks into the interstitial space around the pancreas are trypsin inhibitors, including α1-antitrypsin and β2-microglobulin.16 It has also been hypothesized that the mechanism to prevent trypsin injury inside the acinar cell is to maintain calcium at low levels.18 Trypsinogen activation and trypsin survival are known to be regulated by calcium. Once trypsinogen is secreted into the duct, the calcium-dependent mechanisms utilized by the acinar cell for protection from trypsin are rendered ineffective due to the high level of calcium present in the duct. The duct is, however, protected by maintenance of an alkaline pH and rapid flushing of the zymogens and prematurely activated enzymes out of the pancreas into the duodenum.19
There are extensive genetic studies on SPINK1, which is thought to be a candidate gene for pancreatitis. Forty variants have been reported so far, 14 of which were exclusively found in CP patients but not in controls. This clearly demonstrated the loss-of-function mutations. These findings support the hypothesis that pancreatic secretory trypsin inhibitor (PSTI) is a key negative regulator of prematurely activated trypsin within the pancreatic acinar cells.20 An A > G transition at 101 nucleotide position in the SPINK1 gene leading to substitution of asparagine by serine at codon 34 (N34S) has been reported with its highest frequency (approximately 46%) in an Indian population.21 However, N34S is shown to be in complete linkage disequilibrium with four intronic variants such as, 56 − 37T > C, 87 + 268A > g, 195 − 604G > A, 195–66_-65insTTTT. In spite of being the strongest predictor and an important risk factor in the pathogenesis of TCP, the mechanism of N34S SPINK1mutations contributing to disease phenotype still remains elusive.22
Rosendahl et al.23 identified chymotrypsin C (CTRC, OMIM 601405) as a new pancreatitis-associated gene and discovered that loss-of-function alterations in CTRC gene predispose to pancreatitis by retreating its protective trypsin-degrading activity. The same was shown to be true in TCP patients. A recent study concluded that pancreatitis-associated CTRC mutations can markedly increase the propensity of CTRC to elicit endoplasmic recticulum (ER) stress in pancreatic acinar cells. Thus, carriers of CTRC mutations may be at a higher risk of developing ER stress in the exocrine pancreas, which may contribute to parenchymal damage through acinar cell apoptosis.24
Hereditary pancreatitis (HP) is another entity of chronic pancreatitis. Familial linkage analysis on several affected and unaffected individuals of several generations made it possible to map gene locus on chromosome 7q35 and to demonstrate its association with HP.25,26 Subsequent studies reported a mutation (365G > A) that results in arginine to histidine substitution at 122 position (R122H) in cationic trypsinogen gene [protease, serine, 1 (trypsin 1) (PRSS1), OMIM 276000] to be associated with hereditary pancreatitis.27 Other PRSS1 alterations including A16V, N29T, R116C, R122C and several other genetic alterations have been reported in families with suspected HP or in patients without a family history.28 The current model of PRSS1 mutations suggests that the identified mutations cause enhanced auto-activation of trypsinogen to trypsin, or prevent prematurely activated trypsin from being inactivated by autolysis. Familial aggregations, seen in about 8% of TCP patients, suggest a genetic etiology for TCP.29 However, subsequent studies on PRSS1 have reported its association with HP and CP in western populations but not with TCP.30,31
Triplication (copy number variation) of a 605 kilobase segment containing the PRSS1 and PRSS2 genes has been reported in HP.32 A study by Masson et al.33 revealed the molecular basis of 6% of young ICP patients. However copy number variations with regard PRSS1 and PRSS2 genes were not associated with TCP patients. Although it has been hypothesized that mutations in anionic trypsinogen [protease, serine, 2 (trypsin 2) (PRSS2), OMIM 601564] contribute to the disease by a mechanism similar to that of PRSS1, studies by various groups in ICP and TCP patients did not find associated polymorphisms in PRSS2.34,35 A glycine to arginine change at codon 191 in PRSS2 analyzed in a European population has been demonstrated to play a protective role against CP.36 Further functional studies on purified recombinant G191R protein revealed that generation of a novel tryptic cleavage site within the mutated gene product makes the enzyme hypersensitive to autocatalytic proteolysis, thus playing a protective role in CP. A recent European multicentre study reported the protective role of p.G191R mutation, indicating subjects carrying a heterozygous p.G191R mutation have an approximately 3-fold decreased risk of developing CP compared with carriers of the wild-type allele.37
Cystic fibrosis transmembrane regulator (CFTR, OMIM 602421) gene is associated with alcoholic pancreatitis and ICP, where about 13.4%38 and 25.9%39 of patients in two studies were shown to carry at least one mutation in the gene. An earlier study on western populations revealed an association of CFTR mutations with ICP and the possibility of its interaction with PRSS1 and SPINK1 mutations.40 However, the frequency of CFTR mutations was found to be very low in TCP patients.41
Cathepsin B (CTSB, OMIM 116810) activity is evident as partially purified beef spleen cathepsin B activates trypsinogen to a trypsin-like product. Studies on cationic trypsinogen assigned a central role of cathepsin B in the development of different forms of pancreatitis.42 It was shown that CTSB variants are associated with TCP. Mutations such as L26V and S53G in the propeptide region of the CTSB gene have been found to be associated with TCP. It has been hypothesized that mutations in cathepsin B may cause inept localization of cathepsin B protein in zymogen granules that could lead to premature activation of trypsinogen.43 Type 2 diabetes (T2D)-associated polymorphisms in transcription factor 7 like protein 2 (TCF7L2, OMIM 602228) were screened in TCP and fibro calculus pancreatic diabetes (FCPD) patients. No association was found with FCPD. However, the data suggests that the polymorphisms in TCF7L2 may interact with SPINK1 and CTSB mutations to cause FCPD.44
Chronic pancreatitis shows increased accumulation of extracellular matrix resulting in pancreatic fibrosis. Angiotensin converting enzyme (ACE, OMIM 106180), a zinc metallopeptidase that is a vital enzyme of renin-angiotensin system (RAS), is known to induce proliferation of hepatic stellate cells. It is hypothesized to cause pancreatic fibrosis in TCP patients. A polymorphism in intron-16 of the ACE gene (g.11417-11704del287) is found to be strongly related to the circulating enzyme levels in a dose-dependent manner. However, no association of this polymorphism has been found with TCP.45
Calcium sensing receptor gene (CASR, OMIM 601199) mutations have been suggested to increase the risk of CP, since high intracellular levels of calcium activate trypsinogen within the acinar cells. A combination of CASR and SPINK1 gene mutations predispose to ICP.46 A previous study identified four novel CASR mutations in TCP patients and concluded that the risk of disease may be further increased if there is an associated SPINK1 mutation.47 Tropical calcific pancreatitis is characterized by large ductal calculi. It has been postulated that lithostathine C [encoded by regenerating islet derived protein (Reg) genes] has a role in this. However no polymorphisms in Reg1α gene have been reported in TCP.48–50
There is convincing evidence of a genetic basis for a large majority of patients with chronic pancreatitis in the Asia Pacific region. Unlike in the west, mutations in cationic and anionic trypsinogen gene do not play an important role in this area. Although the genotype is stable, there has been a shift in the phenotype most likely due to environmental factors like alcohol, oxidants and diet.