Helicobacter species and pathogenesis of gallbladder cancer

Raghvendra Raman Mishra, Mallika Tewari and Hari S. Shukla

Varanasi, India

Helicobacter species and pathogenesis of gallbladder cancer

Raghvendra Raman Mishra, Mallika Tewari and Hari S. Shukla

Varanasi, India

(Hepatobiliary Pancreat Dis Int 2010; 9: 129-134)

helicobacter genus;gallbladder cancer;DNA damage;carcinogen;pathogenesis

Introduction

Gallbladder cancer (GBC) is a prominent malignancy of the hepatobiliary tract, being the fi fth most common carcinoma of the gastrointestinal tract in the United States[1]and the third most common malignancy in north India.[2]Primary carcinoma of the gallbladder is more prevalent than other cancers of the extrahepatic biliary tract. It usually remains undetected until it is advanced. GBC is frequently associated with cholelithiasis and cholecystitis in about 75% of cases;but the incidence of documented GBC developing in the presence of cholelithiasis and cholecystitis is only about 0.5%.[3,4]A number of chemical carcinogens structurally similar to naturally-occurring bile acids have been considered to induce GBC. These include methyl cholanthrene, various nitrosamines, and pesticides.[5]The gallbladder stores bile that is released through the common bile duct in the postprandial period. The wall of the gallbladder has three main layers: mucosa(innermost), muscularis (middle) and serosa (outer).GBC starts in the mucosa and spreads through the outer layer as it grows.

Clinical and epidemiological studies have suggested a link between GBC and previous infection with helicobacter and their eradication rates are generally in the 80%-90%range depending on geographical area.[6,7]Kuroki et al[5]demonstrated that the level of epithelial proliferation is higher in helicobacter-positive biliary epithelium than in bacterium-negative epithelium. Several species of helicobacter are believed to play a major role in the causation of GBC.[8]Previously, it was found that H. pylori is not associated with the gallbladder,[9]but in current studies it was found to be associated with the biliary tree[10]and GBC.[11]Multiple studies have demonstrated that infection with helicobacter is associated with 2.7-to 12-fold increases in cancer incidence.[12]Helicobacter species that may colonize the biliary tract have been implicated as a possible cause of hepatobiliary diseases ranging from chronic cholecystitis and primary sclerosing cholangitis to GBC and primary hepatic carcinoma.[13]

The purpose of this review is to bring together the current evidence of the role of various species of helicobacter in the causation of GBC. With GBC rampant in our region in north India, our review also highlights the need for further research on the possible contribution of helicobacter to the etiopathogenesis of GBC.

Historical overview

The genus helicobacter comprises Gram-negative, spiralshaped bacteria that colonize the human gastrointestinal tract. H. pylori was fi rst discovered in the year 1875 by the German scientists in the lining of the human stomach but that could not be grown in culture.[13]In 1982, two Australian scientists, Marshall and Warren, isolated the organism from mucosal specimens of the human stomach. They were the fi rst to successfully culture them as well.[14]Currently, the genus helicobacter has 32 species according to the helicobacter species selector web with validly published names, including helicobacter nemestrinae.[15]This list is rapidly expanding due to novel species being added.[16]

Helicobacter in biliary tract

Risk factors identi fi ed in GBC are cholelithiasis (especially cholesterol gallstone), chronic infections of the gallbladder, obesity, reproductive factors, diet, hepatobiliary anomalies, and environmental exposure to speci fi c chemicals.[3,17,18]Chronic infection is associated with helicobacter[19,20]and salmonella.[3,19]

Helicobacter DNA has been identi fi ed in bile.[21]There is a low incidence of helicobacter in the gallbladder epithelium of Mexican patients with gallstone disease.Some studies have even questioned its association with gallstone pathogenesis.[22]Silva et al[23]investigated the presence of helicobacter species by culture and nested polymerase chain reaction (PCR) of 16S rRNA genes in gallbladder tissue and bile from 46 Brazilian subjects with and 18 without cholelithiasis. The control group was mainly composed of liver donors and patients who had been subjected to cholecystectomy as part of the surgical treatment for morbid obesity. No helicobacter species were grown from the bile or gallbladder tissues of control subjects but helicobacter DNA was detected in the gallbladder tissue and bile in 31.3% and 42.9% of patients with cholelithiasis, respectively. In a logistic regression model, cholelithiasis was positively and independently associated with the female gender,increasing age, and the presence of helicobacter DNA in the gallbladder tissue. The presence of helicobacter DNA in the bile was not associated with cholelithiasis.A signi fi cant association between the presence of helicobacter DNA in the gallbladder epithelium and histological cholecystitis, even after adjusting for gender and age, was also observed. The sequence of the 16S rRNA genes was ＞99% identical to that of H. pylori.These results strongly support the hypothesis that helicobacter is associated with the pathogenesis of human cholelithiasis and cholecystitis.[23]On the other hand,the presence of 16S rRNA genes speci fi c to H. pylori, H.hepaticus, and H. bilis, the H. pylori urease A gene, and the H. pylori 26K protein gene in bile were determined by PCR and sequencing analysis.[24]DNA fragments of helicobacter species other than H. pylori, H. hepaticus,and H. bilis are commonly detectable in the bile of patients with extrahepatic biliary diseases, whether benign or malignant, implying that these helicobacter species play a major role in the pathophysiology of cholesterol gallstone formation in humans[25]and some are frequently associated with gallbladder disease.[26]

Role of helicobacter in the etiopathogenesis of GBC

Helicobacter DNAs have been detected in human bile,hepatobiliary tissue, and primary biliary cirrhosis by PCR assays.[27-29]This was also reported in bile and gallbladder tissue from patients with chronic cholecystitis in Chile,and H. bilis infection in bile is associated with biliary tract and gallbladder cancers in two high-risk populations, the Japanese and the Thais.[30]The bacterium discovered was identi fi ed as a known helicobacter organism, H. bilis, by phylogenetic analysis.[31]However, the pathogenicity of helicobacter species in hepatobiliary carcinogenesis has not been fully elucidated.

There are several pathways by which helicobacter induces cancerous change. One mechanism involves the enhanced production of free radicals close to helicobacter and an increased rate of host cell mutation.The proposed mechanism called the perigenetic pathway[32]involves enrichment of the transformed host cell phenotype by means of alterations in cell proteins such as adhesion proteins. It has been proposed that helicobacter induces in fl ammation and locally high levels of tumor necrosis factor alpha (TNF-α) and interlukin-6.According to the anticipated perigenetic mechanism,signaling molecules associated with in fl ammation,such as TNF-α, alter gastric epithelial cell adhesion and lead to the dispersion and movement of mutated epithelial cells without the need of additional mutation in tumor suppressor genes. The antibody responses toH. hepaticus and H. bilis proteins remain high following absorption in patients with hepatobiliary disease, and these fi ndings should stimulate further investigations to establish whether helicobacter species play a role in the pathogenesis of biliary carcinoma-like disease in humans and other mammals.[33]

Table. Chronological worldwide studies of the last decade on association of helicobacter with gallbladder diseases

H. canis was found to be associated with hepatitis,[34]H. hepaticus with liver carcinoma,[35]and H. bilis with chronic cholecystitis.[36]These fi ndings have highlighted the possibility of new types of infectious agents in the causation of hepatobiliary disease. Helicobacter genes were found in all patients with primary liver carcinoma,whereas only a small fraction of those without primary liver carcinoma had helicobacter genes according to the study by Stanley et al.[37]Although only a part of a small study, these results raise the possibility that H. pylori may be a factor in primary liver cancer. Several studies have shown that H. pylori plays a role in the induction of uremia and the subsequent development of hepatic encephalopathy in patients with cirrhosis.[38,39]Although not con fi rmed by other studies, a putative mechanism is that elevated ammonia levels occur secondary to the effects of bacterial urease. Current research focuses on the role of H. pylori in the formation of intrahepatic stones and induction of biliary epithelial in fl ammation.As evidenced, sequencing of PCR-ampli fi ed 16S rRNA gene fragments has also been used to show that H. bilis can infect the gallbladder of humans with chronic cholecystitis.[40]

Helicobacter infection and its diagnosis

It is estimated that two-thirds of the world population are infected by this bacterium. Actual infection rates vary from nation to nation, but it is common, probably due to poor sanitary conditions. Bacterial colonization of the human gastrointestinal tract has been a subject of study for decades. It has long been known that the colon and, to a lesser extent, the small bowel are populated with a complex microbial ecosystem made up largely of anaerobic bacteria which thrive in the highly reduced environment of the bowel. What has become clear over the last 20 years is that the stomach of humans and a wide range of other animals also have a microbial ecology that consists primarily of one or more ureaseproducing helicobacter species. Furthermore, helicobacter species are commonly present as part of the enteric and hepatobiliary biota in humans[41]and a variety of other animals. In short, members of the helicobacter genus are ubiquitous colonizers of the enteric mucosal surface,which forms a critical interface between an organism and its environment.

Helicobacter species isolated from the bile and gallbladder tissue, such as H. pullorum,[42]H. canis,[43]H.cholecystus,[44]H. rappini,[45]H. hepaticus, and H. bilis,[46-48]have been associated with biliary diseases. In the past few years, the presence of DNA from species of helicobacter,including the well-known human pathogen H. pylori, has been identi fi ed in the bile and biliary epithelium obtained from patients with hepatobiliary diseases.[49-52]

For the diagnosis of helicobacter-associated diseases,it is important to consider the cost of the biomedical test.Investigation of helicobacter species in a sample can be done by molecular analysis (PCR with speci fi c primers,denaturizing grade gel electrophoresis, and DNA sequencing),[53]microbiological analysis, histopathological analysis, tissue culture analysis, and biochemical analysis. All these tests are successfully used in the case of H. pylori.

The studies published in the last decade on the association of helicobacter with gallbladder diseases are summarized in the Table.

Conclusions

Helicobacter DNA has been found in bile and gallbladder tissue of humans with biliary diseases. Other than H.pylori, different helicobacter species are also present in the human biliary tree but the number of microorganisms can differ according to species, with several intestinal species being present in higher numbers than H. pylori.This difference may be explained by the fact that intestinal helicobacter species (such as H. rappini, H. bilis,H. canis, H. cholecystus and H. pullorum) are resistant to bile in vitro, a property that may emphasize protection against the deleterious effect of bile in vivo and adapt them better to the biliary environment. In fact, all of these species have been seen in the biliary system of several animal species.

To the present it is still unclear whether helicobacter species present in the human biliary tree play a signi fi cant role in the pathogenesis of diseases of the biliary tract, especially GBC although the available data do not rule out its possibility. This paper certainly paves a path for further research on helicobacter species involved in the etiopathogenesis of GBC.

Funding: None.

Ethical approval: Not needed.

Contributors: MRR wrote the main body of the article under the supervision of TM. SHS and TM provided advice on medical aspects. SHS is the guarantor.Competing interest: No bene fi ts in any form have been received or will be received from a commercial party related directly or indirectly to the subject of this article.

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BACKGROUND: Gallbladder cancer (GBC) is a rare disease but a leading cause of cancer-related death worldwide. A number of etiological factors have been implicated in the causation of GBC and pathogenic infection by bacteria is one of these.

DATA SOURCES: A PubMed search on "helicobacter", "gallbladder cancer", and "biliary tract malignancies" was done on the topic, and the relevant data were collected, reviewed,and analyzed.

RESULTS: Helicobacter is an epsilon proteobacterium that infects the mucosal lining of the human gastrobiliary system.Infection with helicobacter is an important risk factor for the development of cancer and the bacterium has been categorized as a group-Ⅰcarcinogen by the International Agency for Research on Cancer (IARC). These microbes enter the human body by means of contaminated food and water. Thereby they invade the tissues and produce chemical carcinogens that lead to DNA damage and subsequently a series of gene mutations transform normal cells into cancer cells. In this review, we focus our attention on the role of helicobacter in the causation of biliary tract malignancies.

CONCLUSIONS: The review attempts to summarize the current available data on the role of helicobacter in the causation of GBC. There are accountable data available to suggest the role of helicobacter species in the causation of GBC although larger studies are urgently required for con fi rmation.

Author Af fi liations: Department of Surgical Oncology, Institute of Medical Sciences, Banaras Hindu University, Varanasi 221005, India (Mishra RR,Tewari M and Shukla HS)

Prof. Hari S. Shukla, MS, FRCSEd, PhD, Department of Surgical Oncology, Institute of Medical Sciences, Banaras Hindu University,7 SKG Colony, Lanka, Varanasi 221005, India (Tel: 0091-542-2367718; Fax:0091-542-2368856; Email: harishukla@usa.net)

? 2010, Hepatobiliary Pancreat Dis Int. All rights reserved.

September 21, 2009

Accepted after revision March 4, 2010