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Introduction We present a case of a male patient with neurofibromatosis type 1 diagnosed with pancreatic divisum and several gastrointestinal tumors. A 55-year-old man was admitted to the hospital with recurrent chronic pancreatitis, indicating a large mass in the ampulla. In addition, genetic testing revealed two unique germline mutations in the neurofibromin (NF1) gene, and their potential interaction in promoting cancer was further investigated. Conclusion The first similar case was reported in 2020. The current case was distinct from other cases since an additional two NF1 mutations were found in the patient. In conjunction with prior case reports, our findings imply that genetic testing in patients diagnosed with neurofibromatosis type 1 could be helpful in the development of effective treatments. Keywords: NF1, Neuroendocrine tumors, GIST, Pancreas divisum, Periampullary adenoma, ERCP, Author(s): Bin-bin Li[sup.1,2], Hui Zheng[sup.2], Yi-Dan Lou[sup.2,3], Wen-Wei Zhang[sup.1,2] and Song Zheng[sup.1,2,4,5] Introduction Neurofibromatosis type 1 (NF1) is a hereditary condition inherited in an autosomal dominant manner [1, 2]. An [...]

Background The Australian Pancreatic Cancer Screening Program (APCSP) offers endoscopic ultrasound surveillance for individuals at increased risk of pancreatic ductal adenocarcinoma (PDAC) with all participants requiring assessment by a Familial Cancer Service before or after study enrolment. Methods Individuals aged 40-80 years (or 10 years younger than the earliest PDAC diagnosis) were eligible for APCSP study entry if they had 1) [greater than or equai to] two blood relatives with PDAC (at least one of first-degree association); 2) a clinical or genetic diagnosis of Hereditary Pancreatitis or Peutz-Jeghers syndrome irrespective of PDAC family history; or 3) a known PDAC predisposition germline pathogenic variant (BRCA2, PALB2, CDKN2A, or Lynch syndrome) with [greater than or equai to]one PDAC-affected first- or second-degree relative. Retrospective medical record review was conducted for APCSP participants enrolled at the participating Australian hospitals from January 2011 to December 2019. We audited the genetic investigations offered by multiple Familial Cancer Services who assessed APCSP participants according to national guidelines, local clinical protocol and/or the availability of external research-funded testing, and the subsequent findings. Descriptive statistical analysis was performed using Microsoft Excel. Results Of 189 kindreds (285 participants), 50 kindreds (71 participants) had a known germline pathogenic variant at enrolment (BRCA2 n = 35, PALB2 n = 6, CDKN2A n = 3, STK11 n = 3, PRSS1 n = 2, MLH1 n = 1). Forty-eight of 136 (35%) kindreds with no known germline pathogenic variant were offered mutation analysis; 89% was clinic-funded, with increasing self-funded testing since 2016. The relatively low rates of genetic testing performed reflects initial strict criteria for clinic-funded genetic testing. New germline pathogenic variants were detected in five kindreds (10.4%) after study enrolment (BRCA2 n = 3 kindreds, PALB2 n = 1, CDKN2A n = 1). Of note, only eight kindreds were reassessed by a Familial Cancer Service since enrolment, with a further 21 kindreds identified as being suitable for reassessment. Conclusion Germline pathogenic variants associated with PDAC were seen in 29.1% of our high-risk cohort (55/189 kindreds; 82/285 participants). Importantly, 10.4% of kindreds offered genetic testing were newly identified as having germline pathogenic variants, with majority being BRCA2. As genetic testing standards evolve rapidly in PDAC, 5-yearly reassessment of high-risk individuals by Familial Cancer Services is warranted. Keywords: Pancreatic cancer, Cancer screening, Genetics, Genetic testing, Pathogenic variant, Hereditary Cancer syndromes, Author(s): Krithika Murali[sup.1], Tanya M. Dwarte[sup.2,3,4], Mehrdad Nikfarjam[sup.5], Katherine M. Tucker[sup.3,6], Rhys B. Vaughan[sup.7], Marios Efthymiou[sup.7], Allison Collins[sup.8], Allan D. Spigelman[sup.6,9], Lucinda Salmon[sup.1], Amber L. Johns[sup.2], David B. Williams[sup.4], Martin [...]

Neutrophils and their chemoattractants, the CXC-ELR chemokines keratinocyte cytokine (KC) and macrophage inflammatory protein-2 (MIP-2), play a critical role in pancreatitis. While acute pancreatitis is initiated in acinar cells, it is unclear if these are a source of CXC-ELR chemokines. KC and MIP-2 have NF-[kappa]B, activator protein-1 (AP-1) sites in their promoter regions. However, previous studies have shown increased basal and reduced caerulein-induced AP-1 activation in harvested pancreatic tissue in vitro, which limits interpreting the caerulein-induced response. Moreover, recent studies suggest that NF-[kappa]B silencing in acinar cells alone may not be sufficient to reduce inflammation in acute pancreatitis. Thus the aim of this study was to determine whether acinar cells are a source of KC and MIP-2 and to understand their transcriptional regulation. Primary overnight-cultured murine pancreatic acini were used after confirming their ability to replicate physiological and pathological acinar cell responses. Upstream signaling resulting in KC, MIP-2 upregulation was studied along with activation of the transcription factors NF-[kappa]B and AP-1. Cultured acini replicated critical responses to physiological and pathological caerulein concentrations. KC and MIP-2 mRNA levels increased in response to supramaximal but not to physiological caerulein doses. This upregulation was calcium and protein kinase C (PKC), but not cAMP, dependent. NF-[kappa]B inhibition completely prevented upregulation of KC but not MIP-2. Complete suppression of MIP-2 upregulation required dual inhibition of NF-[kappa]B and AP-1. Acinar cells are a likely source of KC and MIP-2 upregulation during pancreatitis. This upregulation is dependent on calcium and PKC. MIP-2 upregulation requires both NF-[kappa]B and AP-1 in these cells. Thus dual inhibition of NF-[kappa]B and AP-1 may be a more successful strategy to reduce inflammation in pancreatitis than targeting NF-[kappa]B alone. keratinocyte cytokine; macrophage inflammatory protein-2; activator protein-1; nuclear factor-[kappa]B; acinar doi: 10.1152/ajpgi.00177.2010.

Activated pancreatic stellate cells (PSCs) play a pivotal role in pancreatic fibrosis in chronic pancreatitis and pancreatic cancer. Recent studies have suggested a role of IL-33, a newly identified IL-1 family member, in fibrosis. We here examined the expression of IL-33 and the IL-33-mediated regulation of cell functions in PSCs. PSCs were isolated from human and rat pancreas tissues. The expression of IL-33 was examined by Western blotting, PCR, ELISA, and immunostaining. The roles of IL-33 in the regulation of PSC functions were examined by using recombinant IL-33 and small interfering RNA. Activated PSCs expressed IL-33 in the nucleus, and the expression was increased by IL-l[beta], TNF-[alpha] PDGF-BB, and IFN-[gamma], but not TGF-[beta]1. Nuclear IL-33 expression was also observed in the pancreatic acinar and ductal cells. IL-l[beta] induced IL-33 expression mainly through the activation of NF-[kappa]B and ERK pathways and partially through that of p38 MAP kinase, whereas PDGF-BB induced IL-33 expression mainly through the activation of ERK pathway. PSCs expressed soluble ST2, ST2L, and IL-1RAcP, but the expression level of ST2L was relatively low. Recombinant IL-33 did not stimulate key cell functions of PSCs. Decreased IL-33 expression by small interfering RNA resulted in decreased proliferation in response to PDGF-BB. In conclusion, activated PSCs expressed IL-33 in the nucleus. IL-33 might regulate the PDGF-induced proliferation in PSCs. pancreatic cancer; pancreatitis; fibrosis; signal transduction; inflammation doi: 10.1152/ajpgi.00178.2010.

Reg proteins are normally expressed in pancreatic acinar cells, and the level of several of these proteins was significantly induced upon damage to the endocrine or exocrine pancreas. It has been established that Regl and pancreatic islet neogenesis-associated protein [INGAP, Reg3[delta]] promote the growth or regeneration of the endocrine islet cells. Recent reports suggest that Reg2 is an autoantigen normally expressed in islet [beta]-cells. Reg2 overexpression in vitro offered protection to insulinoma cells. Overexpressed Reg3a increased cyclin D 1 and CDK4 levels and the rate of proliferation in insulinoma cells. Acinar-specific overexpression of INGAP increased [beta]-cell mass and protected the animals from streptozotocin-induced diabetes. Moreover, Reg2 gene expression was induced during pancreatitis. We hypothesized that Reg2 is a secreted protein that promotes the growth, survival, and/or regeneration of pancreatic endocrine and exocrine cells. To test its effectiveness, we used elastase-1 promoter (Ela-Reg2) to develop an acinar cell-specific overexpression of the Reg2 gene. Western blot analysis, real-time PCR, and immunohistochemistry revealed barely detectable levels of endogenous Reg2 in the pancreas of normal wild-type mice and increased Reg2 levels in the pancreas of Ela-Reg2 mice that were similar to or higher than Reg2 levels induced in experimental diabetes or pancreatitis. Compared with wild-type littermates, growth, blood glucose and insulin levels, and glucose tolerance were normal in Ela-Reg2 mice; pancreatic histology revealed no change in endocrine or exocrine tissues. Acinar-specific overexpression of the Reg2 gene offered no protection against streptozotocin-induced [beta]-cell damage and diabetes, in hyperglycemia and weight loss, and no advantage in restoring glucose homeostasis and islet function within 3 mo. Furthermore, serum amylase level and pancreatic histochemistry showed that Reg2 overexpression did not protect acinar cells against caerulein-induced acute pancreatitis. In contrast to INGAP or Reg3[beta], exocrine overexpression of Reg2 offered no protection to the endocrine or exocrine pancreas, indicating clear subtype specificities of the Reg family of proteins. caerulein; immunohistochemistry; pancreatic islets; streptozotocin; transgenic mice doi: 10.1152/ajpgi.00500.2009.

Endogenous trypsin inhibitors are synthesized, stored, and secreted by pancreatic acinar cells. It is believed that they play a protective role in the pancreas by inhibiting trypsin within the cell should trypsinogen become prematurely activated. Rodent trypsin inhibitors are highly homologous to human serine protease inhibitor Kazal-type 1 (SPINK1). The mouse has one pancreatic trypsin inhibitor known as SPINK3, and the rat has two trypsin inhibitors commonly known as pancreatic secretory trypsin inhibitors I and II (PSTI-I and -II). Rat PSTI-I is a 61-amino acid protein that shares 65% sequence identity with mouse SPINK3. It was recently demonstrated that mice with genetic deletion of the Spink3 gene ([Spink3.sup.-/-]) do not survive beyond 15 days and lack normal pancreata because of pancreatic autophagy. We have shown that targeted transgenic expression of the rat Psti1 gene to acinar cells in mice [TgN(Psti1)] protects mice against caerulein-induced pancreatitis. To determine whether the autophagic phenotype and lethality in [Spink3.sup.-/-] mice were due to lack of pancreatic trypsin inhibitor, we conducted breeding studies with [Spink.sup.3+/-] heterozygous mice and TgN(Psti1) mice. We observed that, whereas [Spink3.sup.+/+], [Spink3.sup.+/-], and [Spink3-/-]/TgN(Pstil) mice had similar survival rates, no [Spink3.sup.-/-] mice survived longer than 1 wk. The level of expression of SPINK3 protein in acini was reduced in heterozygote mice compared with wild-type mice. Furthermore, endogenous trypsin inhibitor capacity was reduced in the pancreas of heterozygote mice compared with wild-type or knockout mice rescued with the rat Psti1 gene. Surprisingly, the lesser amount of SPINK3 present in the pancreata of heterozygote mice did not predispose animals to increased susceptibility to caerulein-induced acute pancreatitis. We propose that a threshold level of expression is sufficient to protect against pancreatitis. doi: 10.1152/ajpgi.00431.2009 PSTI-I; pancreatitis; autophagy

Ohta E, Itoh T, Nemoto T, Kumagai J, Ko SB, Ishibashi K, Ohno M, Uchida K, Ohta A, Sohara R, Uchida S, Sasaki S, Rai T. pancreas-specific aquaporin 12 null mice showed increased susceptibility to caerulein-induced acute pancreatitis. Am J Physiol Cell Physiol 297: C1368-C1378, 2009. First published September 2, 2009; doi: 10.1152/ajpcell.00117.2009--Aquaporin 12 (AQPI2) is the most recently identified member of the mammalian AQP family and is specifically expressed in pancreatic acinar cells. In vitro expression studies have revealed that AQP12 is localized at intracellular sites. To determine the physiological roles of AQP12 in the pancreas, we generated knockout mice for this gene (AQP12-KO). No obvious differences were observed under normal conditions between wild-type (WT) and AQP12-KO mice in terms of growth, blood chemistry, pancreatic fluid content, or histology. However, when we induced pancreatitis through the administration of a cholecystokinin-8 (CCK-8) analog, the AQP12-KO mice showed more severe pathological damage to this organ than WT mice. Furthermore, when we analyzed exocytosis in the pancreatic acini using a two-photon excitation imaging method, the results revealed larger exocytotic vesicles (vacuoles) in the acini of AQP12-KO mice at a high CCK-8 dose (100 nM). From these results, we conclude that AQP12 may function in the mechanisms that control the proper secretion of pancreatic fluid following rapid and intense stimulation. pancreatic acinar cell: knockout mouse; pancreatic exocrine function; cholecystokinin; exocytosis doi: 10.1152/ajpcell.00117.2009

The objective of this study was to investigate the role of MAPKAP kinase 2 (MK2) and heat shock protein (HSP) HSP60 in the pathogenesis of a new model of severe acute pancreatitis (AP). MK2 plays a significant role in the regulation of cytokines. It has been shown that induction and expression of several HSPs can protect against experimental pancreatitis. Interplay between both systems seems of high interest. Mice with a homozygous deletion of the MK2 gene were used. Severe AP was induced by combined intraperitoneal injections of cerulein with lipopolysaccharide (LPS). Severity of AP was assessed by biochemical markers and histology. The serum IL-6 and lung myeloperoxidase (MPO) levels were determined for assessing the extent of systemic inflammatory response. Expression of HSP25, HSP60, HSP70, and HSP90 was analyzed by Western blotting. Repeated injections of cerulein alone or cerulein plus LPS (Cer+LPS) resulted in local inflammatory responses in the pancreas and corresponding systemic inflammatory changes with pronounced severity in the Cer+LPS group. Compared with the C57B1 wild-type mice, the MK2-/- mice presented with significant milder pancreatitis and attenuated responses of serum amylase and trypsinogen activity. Furthermore, serum IL-6 was decreased as well as lung MPO activity. Injection of LPS alone displayed neither pancreatic inflammatory responses nor alterations of pancreatic enzyme activities but evidently elevated serum IL-6 levels and increased lung MPO activity. In contrast hereto, in the MK2-/-mice, these changes were much milder. Increased expression of HSP25 and HSP60 occurred after induction of AP. Especially, HSP60 was robustly elevated after Cer+LPS treatment, in both MK2-/- and wild-type mice. Thus the homozygous deletion of the MK2 gene ameliorates the severity of acute pancreatitis and accompanying systemic inflammatory reactions in a new model of severe acute pancreatitis. Our data support the hypothesis that MK2 participates in the multifactorial regulation of early inflammatory responses in AP, independently of the regulation of stress proteins like HSP25 and HSP60 and most likely due to its effect on cytokine regulation. MAP kinases; cytokines; heat shock proteins; inflammation doi: 10.1152/ajpgi.00225.2009.

The transcription factor NF-[kappa]B plays a critical role in inflammatory and cell death responses during acute pancreatitis. Previous studies in our laboratory demonstrated that protein kinase C (PKC) isoforms PKC[delta] and [epsilon] are key regulators of NF-[kappa]B activation induced by cholecystokinin-8 (CCK-8), tumor necrosis factor-[alpha], and ethanol. However, the downstream participants in regulating NF-[kappa]B activation in exocrine pancreas remain poorly understood. Here, we demonstrate that protein kinase D1 (PKD1) is a key downstream target of PKC[delta] and PKC[epsilon] in pancreatic acinar cells stimulated by two major secretagogues, CCK-8 and the cholinergic agonist carbachol (CCh), and that PKD1 is necessary for NF-[kappa]B activation induced by CCK-8 and CCh. Both CCK-8 and CCh dose dependently induced a rapid and striking activation of PKD1 in rat pancreatic acinar cells, as measured by in vitro kinase assay and by phosphorylation at PKD1 activation loop (Ser744/748) or autophosphorylation site (Ser9l6). The phosphorylation and activation of PKD1 correlated with NF-[kappa]B activity stimulated by CCK-8 or CCh, as measured by NF-[kappa]B DNA binding. Either inhibition of PKC[delta] or by isoform-specific inhibitory peptides, genetic deletion of PKC[delta] and [epsilon] in pancreatic acinar cells, or knockdown of PKD1 by using small interfering RNAs in AR42J cells resulted in a marked decrease in PKD1 and NF-[kappa]B activation stimulated by CCK-8 or CCh. Conversely, overexpression of PKD1 resulted in augmentation of CCK-8-and CCh-stimulated NF-[kappa]B activation. Finally, the kinetics of PKD1 and NF-[kappa]B activation during cerulein-induced rat pancreatitis showed that both PKD1 and NF-[kappa]B activation were early events during acute pancreatitis and that their time courses of response were similar. Our results identify PKD1 as a novel early convergent point for PKC[delta] and [epsilon] in the signaling pathways mediating NF-[kappa]B activation in pancreatitis. PKC delta; PKC epsilon; pancreatitis; carbachol; cerulein

Digestive trypsins undergo proteolytic breakdown during their transit in the human alimentary tract, which has been assumed to occur through trypsin-mediated cleavages, termed autolysis. Autolysis was also postulated to play a protective role against pancreatitis by eliminating prematurely activated intrapancreatic trypsin. However, autolysis of human cationic trypsin is very slow in vitro, which is inconsistent with the documented intestinal trypsin degradation or a putative protective role. Here we report that degradation of human cationic trypsin is triggered by chymotrypsin C, which selectively cleaves the [Leu.sup.81]-[Glu.sup.82] peptide bond within the [Ca.sup.2+] binding loop. Further degradation and inactivation of cationic trypsin is then achieved through tryptic cleavage of the [Arg.sup.122]-[Val.sup.123] peptide bond. Consequently, mutation of either [Leu.sup.81] or [Arg.sup.122] blocks chymotrypsin C-mediated trypsin degradation. Calcium affords protection against chymotrypsin C-mediated cleavage, with complete stabilization observed at 1 mM concentration. Chymotrypsin C is highly specific in promoting trypsin degradation, because chymotrypsin B1, chymotrypsin B2, elastase 2A, elastase 3A, or elastase 3B are ineffective. Chymotrypsin C also rapidly degrades all three human trypsinogen isoforms and appears identical to enzyme Y, the enigmatic trypsinogen-degrading activity described by Heinrich Rinderknecht in 1988. Taken together with previous observations, the results identify chymotrypsin C as a key regulator of activation and degradation of cationic trypsin. Thus, in the high [Ca.sup.2+] environment of the duodenum, chymotrypsin C facilitates trypsinogen activation, whereas in the lower intestines, chymotrypsin C promotes trypsin degradation as a function of decreasing luminal [Ca.sup.2+] concentrations. chronic pancreatitis | digestive enzymes | trypsinogen degradation

Activation of the inhibitor of NF-[kappa]B kinase/NF-[kappa]B (IKK/NF-[kappa]B) system and expression of proinflammatory mediators are major events in acute pancreatitis. However, the in vivo consequences of IKK activation on the [...]

The article defines the microRNA expression pattern that can differentiate ductal adenocarcinoma of pancreas from normal pancreas and chronic pancreatitis. The findings suggest that though miRNA might be able to distinguish between long- and short-term survivors, the findings need to be validated in other experiments.

Byline: Ioannis Virlos (1), Emanuela Mazzon (2), Ivana Serraino (3), Tiziana Genovese (3), Rosanna Paola (3), Christoph Thiemerman (4), Ajith Siriwardena (1), Salvatore Cuzzocrea (3) Keywords: Nuclear factor IoB; Calpain I inhibitor; Intercellular adhesion molecule 1; Inducible nitric oxide synthase; Poly(ADP-ribose) synthetase; Cerulein Abstract: Objective Nuclear factor-IoB (NF-IoB) is a transcription factor which plays a pivotal role in the induction of genes involved in the response to injury and inflammation. Calpain I inhibitor is a potent antioxidant which is an effective inhibitor of NF-IoB. This study examined whether the postulate that calpain I inhibitor attenuates experimental acute pancreatitis. Design and setting In a murine model we measured NF-IoB activation, expression of intercellular adhesion molecule (ICAM) 1, nitrotyrosine, inducible nitric oxide synthase (iNOS), nuclear enzyme poly(ADP-ribose) synthetase (PARS), myeloperoxidase, malondialdehyde, amylase and lipase and determined histological evidence of lung and pancreas injury in four groups: control (saline only), cerulein, calpain I inhibitor plus cerulein and calpain I inhibitor plus saline. Measurements and results Intraperitoneal injection of cerulein in mice resulted in severe, acute pancreatitis characterised by oedema, neutrophil infiltration, tissue haemorrhage and necrosis and elevated serum levels of amylase and lipase. Infiltration of pancreatic and lung tissue with neutrophils (measured as increase in myeloperoxidase activity) was associated with enhanced lipid peroxidation (increased tissue levels of malondialdehyde). Immunohistochemical examination demonstrated a marked increase in immunoreactivity for nitrotyrosine, iNOS and PARS in the pancreas and lung of cerulein-treated mice. In contrast, pre-treatment with calpain I inhibitor markedly reduced: the degree of pancreas and lung injury upregulation/expression of ICAM-1 staining for iNOS, nitrotyrosine and PARS and lipid peroxidation. Additionally, calpain I inhibitor treatment significantly prevented the activation of NF-IoB as suggested by the inhibition of IIoB-[alpha] degradation in the pancreas tissues after cerulein administration. Conclusions Taken together, our results clearly demonstrate that prevention of the activation of NF-IoB by calpain I inhibitor ameliorates experimental murine acute pancreatitis. Author Affiliation: (1) Hepatobiliary Unit, Department of Surgery, Manchester Royal infirmary, Manchester, UK (2) Department of Biomorphology, School of Medicine, University of Messina, Italy (3) Institute of Pharmacology, School of Medicine, University of Messina, Torre Biologica-Policlinico Universitario, Via C. Valeria-Gazzi, 98100, Messina, Italy (4) Department of Experimental Medicine and Nephrology, William Harvey Research Institute, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Charterhouse Square, London, UK Article History: Registration Date: 05/05/2004 Received Date: 24/01/2004 Accepted Date: 03/05/2004 Online Date: 27/05/2004

Impaired lung function in severe acute pancreatitis is the primary cause of morbidity and mortality in this condition. Preprotachykinin-A (PPT-A) gene products substance P and neurokinin (NK)-A have been shown to play important roles in neurogenic inflammation. Substance P acts primarily (but not exclusively) via the NK1 receptor. NKA acts primarily via the NK2 receptor. Earlier work has shown that knockout mice deficient in NK1 receptors are protected against acute pancreatitis and associated lung injury. NK1 receptors, however, bind other peptides in addition to substance P, not all of which are derived from the PPT-A gene. To examine the role of PPT-A gene products in acute pancreatitis, the effect of PPT-A gene deletion on the severity of acute pancreatitis and the associated lung injury was investigated. Deletion of PPT-A almost completely protected against acute pancreatitis-associated lung injury, with a partial protection against local pancreatic damage. These results show that PPT-A gene products are critical proinflammatory mediators in acute pancreatitis and the associated lung injury. multiple organ dysfunction syndrome; caerulein; substance P; neurogenic inflammation; systemic inflammatory response syndrome

Background: Mutations in the PRSS1 gene explain most occurrences of hereditary pancreatitis (HP) but many HP families have no PRSS1 mutation. Recently, an association between the mutation N34S in the [...]

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene may increase the risk of pancreatitis even in people who do not have cystic fibrosis. Pancreatitis is an inflammation of the pancreas. Researchers tested 134 patients with chronic pancreatitis for 22 different mutations of the CFTR gene. Thirteen percent of the patients had a CFTR mutation. The most common mutation was the deltaF508 mutation, which is also common in cystic fibrosis patients. However, none of the patients had any symptoms of cystic fibrosis.

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene may increase the risk of pancreatitis. Pancreatitis is an inflammation of the pancreas. Researchers tested 27 people with pancreatitis of unknown cause for 18 different CFTR mutations. Ten had at least one mutation, and in three, mutations affected both copies of the gene. These three patients had abnormal chloride transport, which is one feature of cystic fibrosis. However, they had no other symptoms of cystic fibrosis.