Chronic pancreatitis is a persistent inflammatory disease of the pancreas. The digestive protease trypsin plays a fundamental role in the pathogenesis. Here we analyzed the gene encoding the trypsindegrading enzyme chymotrypsin C (CTRC) in German subjects with idiopathic or hereditary chronic pancreatitis. Two alterations, p.R254W and p.K247_R254del, were significantly overrepresented in the pancreatitis group and were present in 30/901 (3.3%) affected individuals but only in 21/2,804 (0.7%) controls (OR=4.6; CI=2.6−8.0; P=1.3×10 −7 ). A replication study identified these two variants in 10/348 (2.9%) individuals with alcoholic chronic pancreatitis but only in 3/432 (0.7%) subjects with alcoholic liver disease (OR=4.2; CI=1.2−15.5; P=0.02). CTRC variants were also found in 10/71 (14.1%) Indian subjects with tropical pancreatitis but only in 1/84 (1.2%) control (OR=13.6; CI=1.7 −109.2; P=0.0028). Functional analysis of the CTRC variants revealed impaired activity and/or reduced secretion. The results indicate that loss-of-function alterations in CTRC predispose to pancreatitis by diminishing its protective trypsin-degrading activity.Chronic pancreatitis is a continuing inflammatory disorder characterized by permanent destruction of the pancreatic parenchyma leading to maldigestion and diabetes mellitus due to exocrine and endocrine insufficiency. Penetrating insight into the pathomechanism came from relatively recent studies investigating the genes encoding cationic trypsinogen (PRSS1; OMIM 276000), anionic trypsinogen (PRSS2; OMIM 601564), and the pancreatic secretory trypsin inhibitor (SPINK1; OMIM 167790). Gain-of-function variants in PRSS1 have been linked to autosomal dominant hereditary pancreatitis and subsequently also to idiopathic chronic pancreatitis 1-4 . Recently, triplication of the PRSS1 locus has been observed in a subset of families with hereditary pancreatitis 5 . In vitro biochemical studies revealed that the majority of disease predisposing PRSS1 variants increase autocatalytic conversion of trypsinogen to active trypsin and probably promote premature intrapancreatic trypsin activation in vivo 6,7 . Consistent with the central pathophysiological role of trypsin, p.N34S and other loss-offunction alterations in the trypsin inhibitor SPINK1 predispose to idiopathic, tropical, and alcoholic chronic pancreatitis 8-15 . In contrast to pathogenic PRSS1 and SPINK1 variations, the p.G191R PRSS2 variant affords protection against chronic pancreatitis due to rapid autodegradation 16 . Taken together, genetic and biochemical evidence defines a pathological pathway in which a sustained imbalance between intrapancreatic trypsinogen activation and trypsin inactivation results in the development of chronic pancreatitis ( Supplementary Fig. 1).Because trypsin degradation serves as a protective mechanism against pancreatitis, we hypothesized that loss of function in trypsin degrading enzymes increases the risk for pancreatitis. We recently demonstrated that chymotrypsin C (CTRC) degrades all human tryps...

The gene SCN9A is responsible for three human pain disorders. Nonsense mutations cause a complete absence of pain, whereas activating mutations cause severe episodic pain in paroxysmal extreme pain disorder and primary erythermalgia. This led us to investigate whether single nucleotide polymorphisms (SNPs) in SCN9A were associated with differing pain perception in the general population. We first genotyped 27 SCN9A SNPs in 578 individuals with a radiographic diagnosis of osteoarthritis and a pain score assessment. A significant association was found between pain score and SNP rs6746030; the rarer A allele was associated with increased pain scores compared to the commoner G allele (P = 0.016). This SNP was then further genotyped in 195 pain-assessed people with sciatica, 100 amputees with phantom pain, 179 individuals after lumbar discectomy, and 205 individuals with pancreatitis. The combined P value for increased A allele pain was 0.0001 in the five cohorts tested (1277 people in total). The two alleles of the SNP rs6746030 alter the coding sequence of the sodium channel Nav1.7. Each was separately transfected into HEK293 cells and electrophysiologically assessed by patch-clamping. The two alleles showed a difference in the voltagedependent slow inactivation (P = 0.042) where the A allele would be predicted to increase Nav1.7 activity. Finally, we genotyped 186 healthy females characterized by their responses to a diverse set of noxious stimuli. The A allele of rs6746030 was associated with an altered pain threshold and the effect mediated through C-fiber activation. We conclude that individuals experience differing amounts of pain, per nociceptive stimulus, on the basis of their SCN9A rs6746030 genotype.Na v 1.7 | nociception | pain | SCN9A | single nucleotide polymorphisms

Screening for variants in TPMT did not reduce the proportions of patients with hematologic ADRs during thiopurine treatment for IBD. However, there was a 10-fold reduction in hematologic ADRs among variant carriers who were identified and received a dose reduction, compared with variant carriers who did not, without differences in treatment efficacy. ClinicalTrials.gov number: NCT00521950.

BackgroundCyclooxygenase-2 (COX-2, PTGS2) is an enzyme involved in the synthesis of prostaglandins and thromboxanes, which are regulators of biologic processes such as inflammation, cell proliferation and angiogenesis. COX-2 over-expression was reported in many (pre) malignant tissues, but data strongly vary and seem to depend on the methodology used.MethodsNormal colorectal mucosa and paired cancerous tissue from 60 patients with colorectal cancer was investigated for the levels of COX-2 mRNA by real-time quantitative Polymerase Chain Reaction (qPCR). COX-2 levels were expressed relative to either: tissue weight or levels of the housekeeping genes beta-2 microglobulin (B2M) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH).ResultsCOX-2 mRNA levels, normalized with respect to tissue weight or mRNA levels of the housekeeping genes B2M or GAPDH, were over-expressed in 80%, 70% and 40% of the colorectal tumor tissues, as compared to the paired adjacent normal colorectal mucosa samples, respectively. Highest mRNA COX-2 ratios tumor/normal were measured when expressed per mg tissue (mean ratio 21.6). When normalized with respect to the housekeeping genes B2M or GAPDH, mean tumor/normal ratios were 16.1 and 7.5, respectively.ConclusionExpression of COX-2 mRNA levels per mg tissue is most simple in comparison to normalization with respect to the housekeeping genes B2M or GAPDH. Levels of COX-2 mRNA are found over-expressed in almost 80% of the colorectal tumors, compared to paired adjacent normal colorectal mucosa, suggesting a role of COX-2 as a potential biomarker for cancer risk, whereas inhibitors of COX-2 could be of value in chemoprevention of colon cancer.

Polycystic liver disease (PCLD, OMIM 174050) is a dominantly inherited condition characterized by the presence of multiple liver cysts of biliary epithelial origin. Fine mapping established linkage to marker D19S581 (Z(max) = 9.65; theta = 0.01) in four large Dutch families with PCLD. We identified a splice-acceptor site mutation (1138-2A-->G) in PRKCSH in three families, and a splice-donor site mutation (292+1G-->C) in PRKCSH segregated completely with PCLD in another family. The protein encoded by PRKCSH, here named hepatocystin, is predicted to localize to the endoplasmic reticulum. These findings establish germline mutations in PRKCSH as the probable cause of PCLD.

Gain-of-function mutations of Na V 1.7 have been shown to produce two distinct disorders: Na V 1.7 mutations that enhance activation produce inherited erythromelalgia (IEM), characterized by burning pain in the extremities; Na V 1.7 mutations that impair inactivation produce a different, nonoverlapping syndrome, paroxysmal extreme pain disorder (PEPD), characterized by rectal, periocular, and perimandibular pain. Here we report a novel Na V 1.7 mutation associated with a mixed clinical phenotype with characteristics of IEM and PEPD, with an alanine 1632 substitution by glutamate (A1632E) in domain IV S4 -S5 linker. Patch-clamp analysis shows that A1632E produces changes in channel function seen in both IEM and PEPD mutations: A1632E hyperpolarizes (Ϫ7 mV) the voltage dependence of activation, slows deactivation, and enhances ramp responses, as observed in Na V 1.7 mutations that produce IEM. A1632E depolarizes (ϩ17mV) the voltage dependence of fast inactivation, slows fast inactivation, and prevents full inactivation, resulting in persistent inward currents similar to PEPD mutations. Using current clamp, we show that A1632E renders dorsal root ganglion (DRG) and trigeminal ganglion neurons hyperexcitable. These results demonstrate a Na V 1.7 mutant with biophysical characteristics common to PEPD (impaired fast inactivation) and IEM (hyperpolarized activation, slow deactivation, and enhanced ramp currents) associated with a clinical phenotype with characteristics of both IEM and PEPD and show that this mutation renders DRG and trigeminal ganglion neurons hyperexcitable. These observations indicate that IEM and PEPD mutants are part of a physiological continuum that can produce a continuum of clinical phenotypes.

Primary erythermalgia is a rare disorder characterized by recurrent attacks of red, warm and painful hands, and/or feet. We previously localized the gene for primary erythermalgia to a 7.94 cM region on chromosome 2q. Recently, Yang et al identified two missense mutations of the sodium channel alpha subunit SCN9A in patients with erythermalgia. The presence of voltage-gated sodium channels in sensory neurons is thought to play a crucial role in several chronic painful neuropathies. We examined four different families and two sporadic cases and detected missense sequence variants in SCN9A to be present in primary erythermalgia patients. A total of five of six mutations were located in highly conserved regions. One family with autosomal dominantly inherited erythermalgia was double heterozygous for two separate SCN9A mutations. These data establish primary erythermalgia as a neuropathic disorder and offers hope for treatment of this incapacitating painful disorder.

The single-nucleotide polymorphisms rs10273639 at the PRSS1-PRSS2 locus and rs7057398 and rs12688220 at the CLDN2-MORC4 locus are associated with CP and strongly associate with ACP, but only rs7057398 with NACP in female patients.