Dear Editor

We thank Dr Nandurkar for his interest[1] in our recent papers regarding luminal acidity at the gastro-oesophageal junction following meals.

In our first study, we observed a postprandial region of high acidity in the cardia region and extending across the Z-line.[2] The presence of this postprandial acid pocket at the cardia has been confirmed by two other groups[3,4]

In our more recent study, we observed a high degree of acid exposure when a pH electrode was clipped to the squamous mucosa which is proximal to the Z-line.[4]

Dr Nandurkar wondered why we did not observe more prolonged acid exposure in the second study if the postprandial acid pocket extends into the distal oesophagus. There are several reasons for the apparent differences in the two studies. The first is that the design and methodology of the two studies were different. The first study involved slowly withdrawing a pH probe at 1cm increments every minute from the distal stomach into the oesophagus and was performed after a large fatty meal with the patient in a semirecumbant position. In contrast, the second study employed a static probe fixed to the distal oesophagus and which recorded both fasting and postprandial pH over a 24h ambulatory period. The difference in the duration of the recording of pH in the distal oesophagus between the two studies and the differences in relationship to food intake and difference in posture and mobility make it difficult to directly compare the two studies. In our second study, we did observe that the acid exposure of the most distal oesophagus was more during the postprandial period, consistent with the observations of the first study. However, we acknowledge that the duration of acid exposure of the most distal oesophagus during the postprandial period with the fixed probe was not as great as that observed in our earlier study when the probe was slowly withdrawn from the stomach into the oesophagus.

It is possible that the methodology employed in the first study could exaggerate the degree of proximal extension of the unbuffered acid pocket into the distal oesophagus. This might be due to the pH probe carrying over some acidic juice from the cardia acid pocket into the oesophagus and this taking time to be fully cleared/neutralized. It is always difficult to know the extent to which the technique used to measure physiology may be altering events.

What is clear from both of our studies and the two other groups which have undertaken similar studies is that during the postprandial period, (a) there is a region of relatively unbuffered high acidity in the proximal cardia region of the stomach and (b) this acid frequently encroaches on the distal oesophageal mucosa.[1-4]

References

1. Nandurkar S. Short segment reflux: Acid but no Pocket? [electronic response to Fletcheret al. Studies of acid exposure immediately above the gastro-oesophageal squamocolumnar junction: evidence of short segment reflux] gutjnl.com 2004http://gut.bmjjournals.com/cgi/eletters/53/2/168#343

2. Fletcher J, Wirz A, Young J, Vallance R, McColl K E L. Unbuffered highly acidic gastric juice exists at the gastroesophageal junction after a meal. Gastroenterology, 2001; 121: 775-783.

3. Hila A, Xue S, Knuff D, Katz P O, Castetll D O. Post-prandial gastric acid layer is not only localised below the EG junction. Gastroenterology, 2003; 124: 4: suppl. 1: A410.

4. Iwakiri K, Nind G, Wu Zou D, Sifrim D, Rigda R, Dent J, Holloway R H. Regional variations in postprandial gastric pH and their relationship to acid reflux in healthy volunteers. Gastroenterology, 2003; 124: 4: suppl.1: A412.

4. Fletcher J, Wirz A, Henry E, McColl K E L. Studies of acid exposure immediately above the gastro-oesophageal squamocolumnar junction: evidence of short segment reflux. Gut, 2004; 53: 168-173.

Dear Editor,

I would like to respond to the commentary written by Dr. R. Balfour Sartor entitled, "Does Mycobacterium avium subspecies paratuberculosis cause Crohn's disease?” The answer to this rhetorical question is yes, and I would like to explain why we now have sufficient information to conclude that Mycobacterium avium paratuberculosis (Map) is a major cause of Crohn's disease.

First, I will address the cons Dr. Sartor includes in table 1 of his commentary which lists arguments for and against a Map causation of Crohn's disease.

1) Differences in clinical and pathological responses in Johne's and Crohn's diseases. This argument is not logical. Map causes a different clinical and pathologic state in sheep and cattle.[1] A related mycobacterium, mycobacterium tuberculosis, causes disease with a differing clinical and pathologic presentation in mice, rabbits, monkeys and humans.[2] Yet this variety of interspecies disease presentations is not an argument against a common pathogen for these diseases.

2) Lack of epidemiological support of transmissible infection. Studies have shown that the incidence and prevalence of Crohn's disease has steadily increased in the past 50 years.[3-5] This increase mirrors the corresponding increasing prevalence of Map induced Johne's disease in dairy cattle. Most observers accept that this alarming increase is not likely genetic but rather environmental in origin.

3) No evidence of transmission to humans in contact with animals infected with Map. Map is not efficiently transmitted by proximity to infected animals. It is most efficiently transmitted by consumption of contaminated food or water.

4) Genotypes of Crohn’s disease and bovine Map isolates not similar. The evidence in this area is conflicting. However, at least two studies show that the isolates are similar.[6-7]

5) Variability in detection of Map by PCR and serological testing. There has certainly been variability in the reported rates of detection of Map in Crohn's disease. These differences are largely due to significant methodological differences among studies. For example, some negative studies have used formalin fixed paraffin embedded tissue.[8-9] There is evidence in the literature that formalin fixation causes

fragmentation of DNA with resultant poor amplification of PCR primers.10 Ryan and others showed that when studying the same formalin fixed paraffin embedded samples of tissue, use of PCR systems which amplified small sequences of the IS900 gene detected Map much more reliably than PCR systems which amplified large sequences of this gene.[11]

6) No evidence of mycobacterial cell wall by histochemical staining. I agree with this point, but it is not evidence against a Map causation of the disease. Using in situ hybridization, Sechi and others demonstrated the DNA of the Map organism in human tissue but could not demonstrate a cell wall. The organism most likely exists in a cell wall deficient or spheroplast form in human tissue.[12]

7) No worsening of Crohn's disease with immunosuppressive agents or HIV infection. This argument does not hold for a related mycobacterial infection and therefore cannot be used as evidence against a Map causation in Crohn’s disease. Leprosy is universally understood to be caused by another slow growing mycobacterium and it does not worsen with steroid therapy. In fact, steroids are used therapeutically in some cases of leprosy.[13-14]

8) No documented cell mediated responses to Map in patients with Crohn's disease. The studies which have been performed in this area could not rule out the possibility of anergy.[15]

9) No therapeutic response to traditional antimycobacterial antibiotics. This argument is nonsensical. Most of the studies reported to date have used antibiotics which are not effective against Map in vitro.[16-17] Why then would we expect the antibiotics which were used in these studies to be effective in vivo?

Many of us are exposed to low levels of viable Mycobacterium avium paratuberculosis on a regular basis. Map survives pasteurization [18] and three studies have shown that it may be cultured from retail supplies of pasteurized milk.[19-21] Thus Map joins a long list of pathogens which may be transmitted through milk and the diseases which have been transmitted by milk include tuberculosis, bovine tuberculosis, salmonellosis, brucellosis, Q fever, listeriosis, yersiniosis and toxigenic e.coli.

The most challenging evidence for skeptics that this bacterium is pathogenic is the fact that many patients with Crohn's disease are bacteremic with Map.[22] In medicine we are taught that persistent septicemia is a pathologic state that causes disease. The leaky bowel theory has been proposed as the explanation for the growth of Map in the blood of Crohn's disease. If this explanation is tenable, why are e.coli and candida which are more prevalent in the fecal stream not cultured from the blood of these patients? These patients can clear e.coli and candida but they cannot clear Map, i.e., Map is pathogenic in these patients. Colonization of diseased bowel is also an unlikely explanation considering the fact that there are patients who are bacteremic with Map who have very little gross endoscopic evidence of Crohn’s disease (personal communication from Dr. Saleh Naser). The bacteria were cultured from the Buffy coat in Naser's study and Map thus joins other pathogenic mycobacteria which are intracellular within macrophages. The others include mycobacterium tuberculosis and mycobacterium leprae.[23] A blood- borne mycobacterial infection nicely explains the extraintestinal manifestations of this disease[24] and the presence of granulomas (which are not satisfactorily explained by other theories).[25] The skeptics are no longer refuting the evidence that Map is in the tissue and/or blood of Crohn's patients. However, now they posit some unlikely role for Map rather than understanding that it is a pathogen in a susceptible host.

I agree with Dr. Sartor that clearance of Map infection with resolution of the patient's Crohn's disease would be excellent evidence that Map is causative of the disease. However, to obtain this evidence under current conditions will be a nearly impossible task. Unless patients carefully avoid dairy products, drink only Map free water and consume Map free beef, they are at continual risk of contracting the infection. Until Map is removed from the food chain and water supplies, attempts to clear the bacterium with antibiotics will be difficult and in many cases futile. Future antibiotic trials should include the recently developed diarylquinoline compound which has been found to have activity against most mycobacteria.[26]

I agree with Dr. Sartor that additional Map related studies are necessary to further elucidate the role of this bacterium in Crohn’s disease. Such studies should include a Map feeding and inoculation study in primates to see whether we can induce Crohn's disease in primates who are infected with the organism. Additional studies could include in vitro studies of the differences in various subjects’ macrophages or neutrophils to kill Map.

I disagree with Dr. Sartor that additional studies will resolve this classic scientific revolution. The only study which could resolve this controversy would be a Map feeding study in a human population. Of course, this study could never be performed.

Because this controversy can not be resolved in short order, we must use the available information to carry out proper public health measures. We need government mandated programs for the testing and culling of Map infected animals from the beef and dairy cattle herds, improved methods of pasteurization, and we should eliminate Map from drinking water (Map is resistant to chlorination). This task is daunting but a similar task has been carried out effectively in the control and elimination of tuberculosis from dairy herds. The cost will be high but will be offset by the economic benefits for the agricultural industry which annually faces large losses due to Johne's disease. In the United States alone, losses to the cattle industry have been estimated at $1.5 billion per year.[27] More importantly, we will save potential future Map infected Crohn's patients from contracting what we now know is a preventable disease.

References

1. Chiodini, R.J., Van Kruiningen, H.J., and Merkal, R.S. Ruminant paratuberculosis (Johne’s disease): the current status and future prospects. Cornell Vet. 1984; 74: 218-62.

2. Capuano, S.V., Croix, D.A., Pawar, S., et al. Experimental Mycobacterium tuberculosis infection of cynomolgus macaques closely resembles the various manifestations of human m. tuberculosis infection. Infect Immun 2003; 71: 5831-44.

3. Loftus, E.V., Silverstein, M.D., Sandborn, W.J., et al. Crohn’s disease in Olmsted County, Minnesota, 1940-1993: incidence, prevalence and survival. Gastroenterology 1998; 114: 1161—8.

4. Tsironi, E., Feakins, R.M., Roberts, C.S.J., et al. Incidence of inflammatory bowel disease is rising and abdominal tuberculosis is falling in Bangladeshis in East London, United Kingdom. American Journal of Gastroenterology 2004; 99: 1749-55.

5. Desai, H.G. and Gupte, P.A. Increasing incidence of Crohn’s disease in India: is it related to improved sanitation? Indian Journal of Gastroenterology 2005;24: 23-4.

6. Overduin, P., Schouls, L., Roholl, P. et al. Use of multilocus variable-number tandem-repeat analysis for typing mycobacterium avium subsp. Paratuberculosis. J Clin Microbiol 2004; 42: 5022-28.

7. Ghadiali, A.H., Strother, M., Naser S.A., et al. Mycobacterium avium subsp. Paratuberculosis strains isolated from Crohn’s disease patients and animal species exhibit similar polymorphic locus patterns. J Clinical Microbiol 2004; 42: 5345-5348.

8. Fujita, H., Yoshinobu, E., Ishige, I., et al. Quantitative analysis of bacterial DNA from mycobacteria spp., bacteroides vulgatus, and escheria coli in tissue samples from patients with inflammatory bowel disease. J Gastroenterol 2002; 37: 509-16.

9. Baksh, F.K., Finkelstein, S.D., Ariyanayagam-Baksh, S.M., et al. Mod Pathol 2004; 17, 1289-94.

10. Goelz,S.E., Hamilton S.R. and Vogelstein, B. Purification of DNA from a formalin fixed and paraffin embedded human tissue. Biochem Biophys Res Commun 1985; 130: 118-26.

11. Ryan, P., Bennett, M.W., Aarons, S., et al. PCR detection of mycobacterium paratuberculosis in Crohn’s disease granulomas isolated by laser capture microdissection. Gut 2002; 51: 665-70.

12. Sechi, L.A., Manuela, M., Francesco, T. et al. Identification of mycobacterium avium subsp. Paratuberculosis in biopsy specimens from patients with Crohn’s disease identified by in situ hybridization. J Clin Microbiol 2001; 39: 4514-17.

13. Marlowe, S.N., Hawksworth, R.A., Butlin, C.R., et al. Clinical outcomes in a randomized controlled study comparing azathioprine and prednisolone versus prednisolone alone in the treatment of severe leprosy type 1 reactions in Nepal. Trans R Soc Trop Med Hyg 2004; 98: 602-9.

14. Richardus, J.H., Withigton, S.G., Anderson, A.M. et al. Adverse events of standardized regimens of corticosteroids for prophylaxis and treatment of nerve function impairment in leprosy: results from the ‘TRIPOD’ trials. Leprosy Review 2003; 74: 319-27.

15. Ibbotson, J.P., Lowes, J.R., Chahal, H. et al. Mucosal cell- mediated immunity to mycobacterial, enterobacterial and other microbial antigens in inflammatory bowel disease. Clin Exp Immunol 1992; 87: 224- 30.

16. Hermon-Taylor, J. The causation of Crohn’s disease and treatment with antimicrobial drugs. It J. Gastroenterol Hepatol 1998; 30: 607-10.

17. Rastogi, Nalin, Goh, K.S. and Labrousse, V. Activity of clarithromycin compared with those of other drugs against mycobacterium paratuberculosis and further enhancement of its extracellular and intracellular activities by ethambutol. Antimicrobl Agents Chemother 1992; 36: 2843-46.

18. Grant, I.R., Ball, H.J. and Rowe, M.T. Effect of higher pasteurization temperatures, and longer holding times at 72 C, on the inactivation of mycobacterium paratuberculosis in milk. Lett Applied Microbiol 1999; 28: 461-65.

19. Grant, I.R., Ball, H.J. and Rowe, M.T. Incidence of mycobacterium paratuberculosis in bulk raw and commercially pasteurized cow’s milk from approved dairy processing establishments in the United Kingdom. Appl Environ Microbiol 2002; 68: 2428-35.

20. Ellingson, J.L.E., Anderson, J.L., Koziczkowski, J.J., et al. Detection of viable mycobacterium avium subsp. Paratuberculosis in retail pasteurized whole milk by two culture methods and PCR. J Food Prot 2005; 68: 966-72.

21. Ayele, W.Y., Svastova, P., Roubal, P. et al. Mycobacterium avium subspecies paratuberculosis cultured from locally and commercially pasteurized cow’s milk in the Czech Republic. Appl Environ Microbiol 2005; 71: 1210-14.

22. Naser S.A., Ghobrial, G., Romero, C., et al. Culture of mycobacterium avium subspecies paratuberculosis from the blood of patients with Crohn’s disease. Lancet 2004; 364: 1039-44.

23. Harrison’s Principles of Internal Medicine, 15th Edition, McGraw- Hill 2001; 1024 and1035.

24. Greenstein, A.J. et al. The extraintestinal manifestations of Crohn’s disease and ulcerative colitis: a study of 700 patients. Medicine 1976; 55:401.

25. Robbins Pathologic Basis of Disease, 6th Edition, Saunders 1999; 816.

26. Andries, K., Verhasselt, P., Guillemont, J., et al. A diarylquinoline drug active on the ATP synthase of mycobacterium tuberculosis. Science 2005; 307: 223-7.

27. Stabel, J.R., Johne’s disease: a hidden threat. J Dairy Sci 1998; 81: 283-88.

Dear Editor,

The recent paper by Thomas and colleagues[1] in the May 2005 issue of Gut provides further support for an involvement of increased colonic deoxycholic acid formation and absorption in cholesterol gallstone pathogenesis. This paper and others in the series[2-4] from Professor R. Hermon Dowling’s laboratory at Guy’s Hospital, London, was ably placed in perspective with the accompanying comprehensive commentary[5] by Dr. Alan F. Hofmann. He summarized critically the decade-long body of work from the Dowling group and others on the connection between slow intestinal propulsion and the biliary tract in octreotide-treated acromegaly. The salient findings of Dowling’s work may have relevance perhaps to other subsets of gallstone-prone individuals.

Acromegalic patients treated with octreotide display dysfunctional gallbladder motility and sluggish intestinal transit rates. Surprisingly, neither Thomas and colleagues[1] nor Hofmann[5] considered that slower small in addition to slower large intestinal transit times may have important pro-lithogenic consequences per se, in that small intestinal hypomotility may augment biliary cholesterol levels by promoting increased cholesterol absorption from the small intestine. Octreotide does not appear to have any appreciable effect on plasma lipoprotein catabolism and “reverse” cholesterol transport or hepatic cholesterol biosynthesis,[6] hence it is unlikely to promote biliary cholesterol hypersecretion from lipoprotein cholesterol or de novo biosynthesis.

We propose therefore, that heightened cholesterol absorption from the small intestine could provide an important source of the sterol for biliary cholesterol hypersecretion. Because octreotide abolishes cholecystokinin (CCK) release from the small intestine,[7,8] attacks of clinical, presumtively cholelithogenic cholecystitis and cholangitis, well documented in patients with use of this long-acting somatostatin analogue, have customarily been attributed solely to gallbladder hypomotility.[9] However, the loss of post-prandinal CCK release via octreotide,[9] is likely to have an additional pathophysiologic effect in humans. Not only is the contractile response of the gallbladder abolished, but we showed recently that small intestinal propulsion is also slowed significantly in CCK-1 (previously known as CCK- A) receptor knockout mice.[10] Hence by inference, a physiological function of diurnal CCK levels in the healthy mouse and presumably human is to accelerate small intestinal motility, in addition to contracting the gallbladder and relaxing Oddi’s sphincter.

Any degree of disruption of the physiologic motors of the small intestine extends the residence time of cholesterol-rich mixed bile salt micelles in the intestinal lumen. This allows more time for monomeric diffusion and cholesterol absorption by small intestinal enterocytes from the cholesterol-rich mixed micellar reservoir within the chyme. As a result, any deficit in CCK release will induce significant increases in fractional cholesterol absorption from the upper small intestine as we have shown in the CCK-1 receptor knockout mouse compared to wild-type mice.[10] Not surprisingly, there are reports in the literature that a dysfunctional CCK-1 receptor may cause cholesterol gallstones in humans.[11,12] Because the CCK-1 receptor plays such a crucial role in the physiological regulation of gallbladder and small intestinal motility, it is likely that in addition to gallbladder hypomotility, gallstone patients with a dysfunctional CCK-1 receptor also display increased absorption of dietary cholesterol from the dysmotile small intestine.

In his commentary[5] Dr. Hofmann made the point that in several large clinical studies,[13-15] investigators failed to find a strong correlation between biliary deoxycholate levels and cholesterol saturation of bile. This fact alone should prompt an alternative or additional explanation for cholesterol gallstones by the occurence of slow transit constipation in humans. Perhaps the putative increase in cholesterol absorption from the gut may be the “missing link” in the augmented gallstone risk from diminished intestinal motility from any cause. As evidenced by human and animal studies, deoxycholate does not promote cholesterol absorption from the small intestine as efficiently as does its conjugated cholate precursor.[16,17] Moreover, from a physical chemical perspective, higher levels of deoxycholate conjugates in bile may actually counteract augmented cholesterol absorption for reasons of small intestinal hypomotility.

The points we make here are not offered to diminish the putative role of deoxycholic acid in cholesterol gallstone pathogenesis, for which there is abundant evidence from the work of the Dowling laboratory.[1-4] Nonetheless, we argue here that increased fractional absorption of cholesterol from a hypomotile small intestine may result in delivery of more cholesterol to the liver in chylomicron remnants, and hence may play a crucial role in hepatic cholesterol hypersecretion and biliary lithogenicity.[18] Consequently, in the setting of a hypomotile gallbladder, lithogenic bile will have a greatly prolonged residence time, facilitating phase separation of cholesterol crystals and their agglomerate into gallstones.

Two decades ago, Ponz de Leon and colleagues[19] showed that metaclopramide acceleration of small intestinal transit in humans significantly decreased intestinal cholesterol absorption, however they failed to show, in the opposite experiment, that slowing of small intestinal transit with atropine augmented cholesterol absorption. Clearly, the small intestinal dysmotility and cholesterol absorption issues raised here need to be proven in humans but this unique group of patients affords an easily testable cohort.[10] A salutary result would afford an intellectually satisfying explanation for the profound lithogenicity of bile not otherwise accounted for in this group of patients.

Sincerely yours,

Martin C. Carey, M.D., D.Sc.
Department of Medicine
Brigham & Women’s Hospital
Harvard Medical School
Boston, MA 02115, USA

David Q.-H. Wang, M.D., Ph.D.
Department of Medicine
Beth Israel Deaconess Medical Center
Harvard Medical School
Boston, MA 02215, USA

References

1. Thomas LA, Veysey MJ, Murphy GM, et al. Octreotide induced prolongation of colonic transit increases faecal anaerobic bacteria, bile acid metabolising enzymes, and serum deoxycholic acid in patients with acromegaly. Gut 2005;54:630-5.

2. Hussaini SH, Murphy GM, Kennedy C, et al. The role of bile composition and physical chemistry in the pathogenesis of octreotide-associated gallbladder stones. Gastroenterology 1994;107:1503-13.

3. Veysey MJ, Thomas LA, Mallet AI, et al. Prolonged large bowel transit increases serum deoxycholic acid: a risk factor for octreotide induced gallstones. Gut 1999;44:675-81.

4. Thomas LA, Veysey MJ, Bathgate T, et al. Mechanism for the transit- induced increase in colonic deoxycholic acid formation in cholesterol cholelithiasis. Gastroenterology 2000;119:806-15.

5. Hofmann AF. Increased deoxycholic acid absorption and gall stones in acromegalic patients treated with octreotide: more evidence for a connection between slow transit constipation and gall stones. Gut 2005;54:575-8.

6. Olivecrona H, Ericsson S, Angelin B. Growth hormone treatment does not alter biliary lipid metabolism in healthy adult men. J Clin Endocrinol Metab 1995;80:1113-7.

7. Lembcke B, Creutzfeldt W, Schleser S, et al. Effect of the somatostatin analogue sandostatin (SMS 201-995) on gastrointestinal, pancreatic and biliary function and hormone release in normal men. Digestion 1987;36:108- 24.

8. Milenov K, Rakovska A, Kalfin R. Effects of cholecystokinine on the gallbladder motility: interaction with somatostatin and vasoactive intestinal peptide. Acta Physiol Pharmacol Bulg 1995;21:67-76.

9. Fisher RS, Rock E, Levin G, Malmud L. Effects of somatostatin on gallbladder emptying. Gastroenterology 1987;92:885-90.

10. Wang DQ-H, Schmitz F, Kopin AS, Carey MC. Targeted disruption of the murine cholecystokinin-1 receptor promotes intestinal cholesterol absorption and susceptibility to cholesterol cholelithiasis. J Clin Invest 2004;114:521-8.

11. Miller LJ, Holicky EL, Ulrich CD, Wieben ED. Abnormal processing of the human cholecystokinin receptor gene in association with gallstones and obesity. Gastroenterology 1995;109:1375-80.

12. Miyasaka K, Takata Y, Funakoshi A. Association of cholecystokinin A receptor gene polymorphism with cholelithiasis and the molecular mechanisms of this polymorphism. J Gastroenterol 2002;37(Suppl 14):102-6.

13. Gustafsson U, Sahlin S, Einarsson C. Biliary lipid composition in patients with cholesterol and pigment gallstones and gallstone-free subjects: deoxycholic acid does not contribute to formation of cholesterol gallstones. Eur J Clin Invest 2000;30:1099-106.

14. LaRusso NF, Szczepanik PA, Hofmann AF. Effect of deoxycholic acid ingestion on bile acid metabolism and biliary lipid secretion in normal subjects. Gastroenterology 1977;72:132-40.

15. Jüngst D, Muller I, Kullak-Ublick GA, et al. Deoxycholic acid is not related to lithogenic factors in gallbladder bile. J Lab Clin Med 1999;133:370-7.

16. Sama C, LaRusso NF. Effect of deoxycholic, chenodeoxycholic, and cholic acids on intestinal absorption of cholesterol in humans. Mayo Clin Proc 1982;57:44-50.

17. Wang DQ-H, Tazuma S, Cohen DE, Carey MC. Feeding natural hydrophilic bile acids inhibits intestinal cholesterol absorption: studies in the gallstone-susceptible mouse. Am J Physiol 2003;285:G494-502.

18. Wang DQ-H, Zhang L, Wang HH. High cholesterol absorption efficiency and rapid biliary secretion of chylomicron remnant cholesterol enhance cholelithogenesis in gallstone-susceptible mice. Biochim Biophys Acta 2005;1733:90-9.

19. Ponz de Leon M, Iori R, Barbolini G, et al. Influence of small-bowel transit time on dietary cholesterol absorption in human beings. N Engl J Med 1982;307:102-3.

Correspondence to: Dr. Martin C. Carey
Brigham & Women’s Hospital
Department of Medicine
Gastroenterology Division, Thorn 1430
Boston, MA 02115, USA
Email: mccarey@rics.bwh.harvard.edu

Conflict of Interest: None declared

Dear Sir,

Surgery (resection or transplantation) is the first treatment choice for early HCC, but unfortunately it is feasible only in a minority of patients.[1] Local percutaneous ablation has therefore dramatically increase in importance and radio-frequency thermal ablation (RFTA) has showed in recent reports global or disease-free survivals better than those reported for percutaneous ethanol injection (PEI).[2] The recent reports in GUT apparently also supports a superiority, albeit small, of RFTA on PEI or acetic acid injection in a randomised prospective controlled study.[3] Nevertheless, not all agree on RFTA efficacy [4], major complications are described [5], with an incidence rate of major complications significantly higher than in the ethanol or acetic acid injection procedures.[3]

We are carrying out a multicentre prospective study on efficiency and complications of RFTA in HCC in North-East Italy with 399 HCC patients having being recruited and a centralized radiological assessment.

Overall, 133 patients (33%) experienced some complication, 5 of which were severe (1.3%), with 2 casualties (0.5%) one being procedure-related, in relation to an intestinal perforation and the second being a cerebral infarction.

Last, but definitely not least, some author has reported rapid intra-hepatic progression after RFTA [6,7], a fact that in our minds, deserve special attention. We therefore would like to document 9 cases of rapid, unexpected spread of HCC after RFTA observed in our multicentric study, The nine cases (2.3% of the series) are hereby reported: 8 males and 1 female, etiology was HCV in 6 and HBV in 3, 6 had complete response at the CT scanning performed after 1 month, 2 90% and 1 70%, 1 nodule was treated in 7 cases and 2 in 2, the size ranging from 1,8 to 4,6 cm (all patients had CT scanning prior to the procedure). Grading was G1 in all instances and AFP values were normal in 4 cases. New lesions were:

1. satellites + controlateral HCC at 1 month;
2. three x 3 cm nodes at 3 months;
3. multifocality at 4 months;
4. 3 nodes at 1 month;
5. multiple, up to 6 cm at 3 months;
6. multiple at 1 month;
7. thirteen nodes at 5 months;
8. 1 8 cm node at 6 months
9. multifocality at 5 months.

This rapid unexplained and biologically unclear progression of HCC following RFTA treatment is most unusual considering the natural history of the disease. It could be prompted by increased intra-tumoural pressure with intravascular spread, seeding due to artero-venus fistula or to the expandable hooks needles. Suggested risk factors have been high AFP levels, location near major portal branches and poor tumour differentiation.[5,6] Also, increased liver concentrations of growth factors (TGF-ALFA and BETA-FGF) have been documented in rat liver after thermal coagulation, with increased HCC growth.[8]

Our data show that side effects are relatively frequent following RFTA, with a mortality rate lower than 1% but still present. Our series additionally confirm that “explosive HCC spread” may occur after RFTA, in our experience with no correlation with AFP, differentiation and location of the tumour, and frequently even after total ablation.

Why do these patients develop this severe, (dramatic if observed in patients in the waiting list for OLTx!)[9] and unexpected complication, what is its prevalence, whether we can identify patients at risk is open to a discussion that we think should be exhaustive before abandoning PEI as obsolete and considering RFTA as the treatment of early disease not eligible for surgery. Let’s wait for the end of the honey-moon effect of this new procedure!

References

1. Christians K K, Pitt H A, Rilling WS , Franco J, Quiroz FA, Adams MB, Wallace JR, Quebbeman EJ. Hepatocellular carcinoma: Multi-modality management. Surgery 2001; 130 (4): 554-560

2. Lencioni R, Cioni D, Crocetti L, Bartolozzi C. Percutaneous ablation of hepatocellular carcinoma: state-of-the-art. Liver Transpl. 2004 Feb;10(2 Suppl 1):S91-7.

3. Lin SM, Lin CJ, Lin CC, HSU CW, Chen YC. Randomised controlled trial comparing percutaneous radiofrequency thermal ablation, percutaneous ethanol injection, and percutaneous acetic acid injection to treat hepatocellular carcinoma of 3 cm or less. Gut 2005; 54: 1151-6.

4. Mazzaferro V, Battiston C, Perrone S, Pulvirenti A, Regalia E, Romito R, Sarli D, Schiavo M, Garbagnati F, Marchiano A, Spreafico C, Camerini T, Mariani L, Miceli R, Andreola S.. Radiofrequency ablation of small hepatocellular carcinoma in cirrhotic patients awaiting liver transplantation: a prospective study. Ann Surg. 2004;240:900-9.

5. De Baère T, Risse O, Kuoch V, Domain C, Sengel C, Smayra T, Gamal El Din M, Letoublon C, Elias D. Adverse Event During Radiofrequency Treatment of 582 Hepatic Tumours. AJR 2003; 181: 695-700

6. Ruzzante A, De Manzoni G, Molfetta M, Pachera S, Genco B, Donataccio M, Guglielmi A. Rapid progression of hepatocellular carcinoma after Radiofrequency Ablation. World J Gastroenterol 2004; 10(8) 1137-1140

7. Nicoli N, Casaril A, Hilal MA, Mangiante G, Marchiori L, Ciola M, Invernizzi L, Campagnaro T, Mansueto G. A case of rapid intrahepatic dissemination of hepatocellular carcinoma after radiofrequency thermal ablation. Am J Surg. 2004 Aug;188(2):165-7.

8. Ohno T, Kawano K, Yokoyama H, Tahara K, Sasaki A, Aramaky M, Kitano S. Microwave coagulation therapy accelerates growth of cancer in rat liver. Journal of Hepatology 2002; 36: 774-779

9. Cillo U, Vitale A, Bassanello M, Boccagni P, Brolese A, Zanus G, Burra P, Fagiuoli S, Farinati F, Rugge M, D'Amico DF. Liver transplantation for the treatment of moderately or well-differentiated hepatocellular carcinoma. Ann Surg. 2004 Feb;239(2):150-9.