American Gastroenterological Association (AGA): James W. Freston Conference

CHICAGO – Does eradicating Helicobacter pylori prevent gastric cancer?

The answer is yes, sometimes, but it depends on where you live, and what other bacteria coexist in your gut microbiome.

The overall view is a positive one, Richard M. Peek Jr., MD, said at the at the meeting sponsored by the American Gastroenterological Association. A very large, recent meta-analysis confirms it (Gastroenterology. 2016. doi:10.1053/j.gastro.2016.01.028). Comprising 24 studies and 48,000 subjects, the meta-analysis determined that eradicating the bacteria in infected people cut gastric cancer incidence significantly.

That’s great news – but there’s a big caveat, said Dr. Peek of Vanderbilt University, Nashville. “The benefit was dependent on what your baseline risk was. For those with a high baseline risk, the benefit was tremendous. For those with a low baseline risk, it was not statistically significant.”

There are long-term data suggesting that treating H. pylori sooner rather than later is the way to go. A 2005 study followed more than 700 patients with preneoplastic gastric lesions for 12 years. It found that the treatment effect was cumulative: The longer the patient was free of H. pylori, the more reliably healing occurred (Gut. 2005. doi:10.1136/gut.2005.072009).

At baseline, the patients were randomized to nutritional supplements or to a combination of amoxicillin, metronidazole, and bismuth subsalicylate. At 6 years, the trial was unblinded and all patients were offered treatment. Patients were followed for another 6 years. Those who were H. pylori negative at 12 years had 15% more regression and 14% less progression than subjects who were positive at 12 years. Among those who received anti–H. pylori treatment at the 6-year mark, the effect was smaller and nonsignificant.

Perhaps surprisingly, though, the biggest bang for H. pylori treatment is seen in the antrum of the stomach, not in the corpus. Another meta-analysis, this one of 16 studies, found very consistent reductions in the severity of intestinal metaplasia in the antrum after antibiotic treatment – but no difference at all in corpus metaplasia. The reason for that finding isn’t at all clear, the authors of that paper noted (World J Gastro. 2014. doi:10.3748/wjg.v20.i19.5903).

The bacteria-metaplasia cancer link gets even more complicated when H. pylori is viewed as a contributing member of society, rather than a hermit. The bacterium seems to be a bully in the neighborhood, radically altering the normal gastric microbiome, Dr. Peek said.

In the absence of H. pylori, the gastric microbiome is much more diverse, consisting of about 50% Actinobacteria and 25% Firmicutes species. Bacteroides and Proteobacteria species make up the remainder, with a small population of Cyanobacteria as well. In its presence, Proteobacteria – a gram-negative genus that includes a wide variety of pathogens – almost completely subsume beneficial bacteria.

Researchers saw this change in action in 2011, when a group at the Massachusetts Institute of Technology, Cambridge, inoculated two mouse populations with H. pylori and followed them for gastric neoplasms (Gastroenterology. 2011. doi:10.1053/j.gastro.2010.09.048). All the mice were genetically engineered to overexpress human gastrin, a characteristic that invariably leads them to develop gastric cancers.

One group comprised germ-free mice raised in sterile environments. The control group was free of pathogens, but lived in a conventional environment and so had normal gastric flora. Both groups were inoculated with H. pylori.

By 11 months, the microbiome of the control group was strikingly different. It showed a significant increase in the number of Firmicutes bacteria in the stomach, with an associated decrease in the number and variety of other bacteria including Bacteroides. This was especially interesting when viewed in relation to the rate of gastric neoplasia, Dr. Peek said.

These mice are programmed to develop gastric cancer by 6 months of age – and this is what happened in the control mice, which had H. pylori plus other gastric microbes. But the germ-free mice who were monoinfected with H. pylori showed a much different progression of disease. At 7 months, most showed only a mild hypergastrinemia. Conversely, at 7 months, all of the H. pylori–infected control mice had developed gastric intraepithelial neoplasia, 80% of it high grade. Only 10% of the monoinfected mice developed cancer, and all of it was low grade.

“It looks like there is active collaboration between H. pylori and other bacteria in the stomach,” resulting in this increased cancer risk, Dr. Peek said.

It’s a collaboration that reaches deep into the tumors themselves, he said. “A very interesting study a couple of years ago searched cancer genomes for the presence of bacterial DNA, and found that gastric cancers incorporated the second-highest amount of microbial DNA into their cancer genomes. But it wasn’t just H. pylori. Many other species had integrated their DNA into these tumors.”

That study, published in 2013, was the first to prove that bacterial DNA can impact carcinogenesis. Acute myeloid leukemia showed the highest integration of bacterial DNA, but gastric adenocarcinoma was a close second. Most of the species were of the Proteobacteria lineages (83%), with a third of that represented by Pseudomonas, particularly P. fluorescens and P. aeruginosa. Both of those species have been shown to promote gastric tumorigenesis in rats. All of the DNA integrations occurred in five genes; four of these are already known to be upregulated in gastric cancer (PLOS Comp Biol. 2013;9[6]:e1003107).

Interestingly, only a few of the sample reads turned up DNA integration with H. pylori.

This reduction in gastric microbial diversity could be an important key to H. pylori’s relation to gastric cancer, Dr. Peek said. He examined this in residents of two towns in Colombia, South America: Tumaco, where the risk of gastric cancer is low, and Tuquerres, where it’s 25 times higher (Sci Rep. 2016. doi:10.1038/srep18594).

What was different was the gastric microbiome of residents. Those living in low-risk Tumaco had much more microbial diversity: 361 varieties, compared with 194 in Tuquerres. And 16 of these groups – representative of what’s usually considered a healthy microbiome – were absent in the high-risk subjects. But Tuquerres residents had two bacteria that weren’t found in Tumaco residents, including Leptorichia wadei, which has been associated with necrotizing enterocolitis.

There was no difference, however, in the prevalence of H. pylori between these high- and low-risk groups.

These new findings illustrate an increasingly complicated interplay of bacteria and gastric cancer, Dr. Peek said. But they also provide a new direction for research.

“We have a framework now where we can move forward and try to understand how some of these other strains impact gastric cancer risk,” he said.

Dr. Peek had no relevant financial disclosures.

[email protected]

On Twitter @Alz_Gal

CHICAGO – Does eradicating Helicobacter pylori prevent gastric cancer?

The answer is yes, sometimes, but it depends on where you live, and what other bacteria coexist in your gut microbiome.

The overall view is a positive one, Richard M. Peek Jr., MD, said at the at the meeting sponsored by the American Gastroenterological Association. A very large, recent meta-analysis confirms it (Gastroenterology. 2016. doi:10.1053/j.gastro.2016.01.028). Comprising 24 studies and 48,000 subjects, the meta-analysis determined that eradicating the bacteria in infected people cut gastric cancer incidence significantly.

That’s great news – but there’s a big caveat, said Dr. Peek of Vanderbilt University, Nashville. “The benefit was dependent on what your baseline risk was. For those with a high baseline risk, the benefit was tremendous. For those with a low baseline risk, it was not statistically significant.”

There are long-term data suggesting that treating H. pylori sooner rather than later is the way to go. A 2005 study followed more than 700 patients with preneoplastic gastric lesions for 12 years. It found that the treatment effect was cumulative: The longer the patient was free of H. pylori, the more reliably healing occurred (Gut. 2005. doi:10.1136/gut.2005.072009).

At baseline, the patients were randomized to nutritional supplements or to a combination of amoxicillin, metronidazole, and bismuth subsalicylate. At 6 years, the trial was unblinded and all patients were offered treatment. Patients were followed for another 6 years. Those who were H. pylori negative at 12 years had 15% more regression and 14% less progression than subjects who were positive at 12 years. Among those who received anti–H. pylori treatment at the 6-year mark, the effect was smaller and nonsignificant.

Perhaps surprisingly, though, the biggest bang for H. pylori treatment is seen in the antrum of the stomach, not in the corpus. Another meta-analysis, this one of 16 studies, found very consistent reductions in the severity of intestinal metaplasia in the antrum after antibiotic treatment – but no difference at all in corpus metaplasia. The reason for that finding isn’t at all clear, the authors of that paper noted (World J Gastro. 2014. doi:10.3748/wjg.v20.i19.5903).

The bacteria-metaplasia cancer link gets even more complicated when H. pylori is viewed as a contributing member of society, rather than a hermit. The bacterium seems to be a bully in the neighborhood, radically altering the normal gastric microbiome, Dr. Peek said.

In the absence of H. pylori, the gastric microbiome is much more diverse, consisting of about 50% Actinobacteria and 25% Firmicutes species. Bacteroides and Proteobacteria species make up the remainder, with a small population of Cyanobacteria as well. In its presence, Proteobacteria – a gram-negative genus that includes a wide variety of pathogens – almost completely subsume beneficial bacteria.

Researchers saw this change in action in 2011, when a group at the Massachusetts Institute of Technology, Cambridge, inoculated two mouse populations with H. pylori and followed them for gastric neoplasms (Gastroenterology. 2011. doi:10.1053/j.gastro.2010.09.048). All the mice were genetically engineered to overexpress human gastrin, a characteristic that invariably leads them to develop gastric cancers.

One group comprised germ-free mice raised in sterile environments. The control group was free of pathogens, but lived in a conventional environment and so had normal gastric flora. Both groups were inoculated with H. pylori.

By 11 months, the microbiome of the control group was strikingly different. It showed a significant increase in the number of Firmicutes bacteria in the stomach, with an associated decrease in the number and variety of other bacteria including Bacteroides. This was especially interesting when viewed in relation to the rate of gastric neoplasia, Dr. Peek said.

These mice are programmed to develop gastric cancer by 6 months of age – and this is what happened in the control mice, which had H. pylori plus other gastric microbes. But the germ-free mice who were monoinfected with H. pylori showed a much different progression of disease. At 7 months, most showed only a mild hypergastrinemia. Conversely, at 7 months, all of the H. pylori–infected control mice had developed gastric intraepithelial neoplasia, 80% of it high grade. Only 10% of the monoinfected mice developed cancer, and all of it was low grade.

“It looks like there is active collaboration between H. pylori and other bacteria in the stomach,” resulting in this increased cancer risk, Dr. Peek said.

It’s a collaboration that reaches deep into the tumors themselves, he said. “A very interesting study a couple of years ago searched cancer genomes for the presence of bacterial DNA, and found that gastric cancers incorporated the second-highest amount of microbial DNA into their cancer genomes. But it wasn’t just H. pylori. Many other species had integrated their DNA into these tumors.”

That study, published in 2013, was the first to prove that bacterial DNA can impact carcinogenesis. Acute myeloid leukemia showed the highest integration of bacterial DNA, but gastric adenocarcinoma was a close second. Most of the species were of the Proteobacteria lineages (83%), with a third of that represented by Pseudomonas, particularly P. fluorescens and P. aeruginosa. Both of those species have been shown to promote gastric tumorigenesis in rats. All of the DNA integrations occurred in five genes; four of these are already known to be upregulated in gastric cancer (PLOS Comp Biol. 2013;9[6]:e1003107).

Interestingly, only a few of the sample reads turned up DNA integration with H. pylori.

This reduction in gastric microbial diversity could be an important key to H. pylori’s relation to gastric cancer, Dr. Peek said. He examined this in residents of two towns in Colombia, South America: Tumaco, where the risk of gastric cancer is low, and Tuquerres, where it’s 25 times higher (Sci Rep. 2016. doi:10.1038/srep18594).

What was different was the gastric microbiome of residents. Those living in low-risk Tumaco had much more microbial diversity: 361 varieties, compared with 194 in Tuquerres. And 16 of these groups – representative of what’s usually considered a healthy microbiome – were absent in the high-risk subjects. But Tuquerres residents had two bacteria that weren’t found in Tumaco residents, including Leptorichia wadei, which has been associated with necrotizing enterocolitis.

There was no difference, however, in the prevalence of H. pylori between these high- and low-risk groups.

These new findings illustrate an increasingly complicated interplay of bacteria and gastric cancer, Dr. Peek said. But they also provide a new direction for research.

“We have a framework now where we can move forward and try to understand how some of these other strains impact gastric cancer risk,” he said.

Dr. Peek had no relevant financial disclosures.

[email protected]

On Twitter @Alz_Gal

CHICAGO – Does eradicating Helicobacter pylori prevent gastric cancer?

The answer is yes, sometimes, but it depends on where you live, and what other bacteria coexist in your gut microbiome.

The overall view is a positive one, Richard M. Peek Jr., MD, said at the at the meeting sponsored by the American Gastroenterological Association. A very large, recent meta-analysis confirms it (Gastroenterology. 2016. doi:10.1053/j.gastro.2016.01.028). Comprising 24 studies and 48,000 subjects, the meta-analysis determined that eradicating the bacteria in infected people cut gastric cancer incidence significantly.

That’s great news – but there’s a big caveat, said Dr. Peek of Vanderbilt University, Nashville. “The benefit was dependent on what your baseline risk was. For those with a high baseline risk, the benefit was tremendous. For those with a low baseline risk, it was not statistically significant.”

There are long-term data suggesting that treating H. pylori sooner rather than later is the way to go. A 2005 study followed more than 700 patients with preneoplastic gastric lesions for 12 years. It found that the treatment effect was cumulative: The longer the patient was free of H. pylori, the more reliably healing occurred (Gut. 2005. doi:10.1136/gut.2005.072009).

At baseline, the patients were randomized to nutritional supplements or to a combination of amoxicillin, metronidazole, and bismuth subsalicylate. At 6 years, the trial was unblinded and all patients were offered treatment. Patients were followed for another 6 years. Those who were H. pylori negative at 12 years had 15% more regression and 14% less progression than subjects who were positive at 12 years. Among those who received anti–H. pylori treatment at the 6-year mark, the effect was smaller and nonsignificant.

Perhaps surprisingly, though, the biggest bang for H. pylori treatment is seen in the antrum of the stomach, not in the corpus. Another meta-analysis, this one of 16 studies, found very consistent reductions in the severity of intestinal metaplasia in the antrum after antibiotic treatment – but no difference at all in corpus metaplasia. The reason for that finding isn’t at all clear, the authors of that paper noted (World J Gastro. 2014. doi:10.3748/wjg.v20.i19.5903).

The bacteria-metaplasia cancer link gets even more complicated when H. pylori is viewed as a contributing member of society, rather than a hermit. The bacterium seems to be a bully in the neighborhood, radically altering the normal gastric microbiome, Dr. Peek said.

In the absence of H. pylori, the gastric microbiome is much more diverse, consisting of about 50% Actinobacteria and 25% Firmicutes species. Bacteroides and Proteobacteria species make up the remainder, with a small population of Cyanobacteria as well. In its presence, Proteobacteria – a gram-negative genus that includes a wide variety of pathogens – almost completely subsume beneficial bacteria.

Researchers saw this change in action in 2011, when a group at the Massachusetts Institute of Technology, Cambridge, inoculated two mouse populations with H. pylori and followed them for gastric neoplasms (Gastroenterology. 2011. doi:10.1053/j.gastro.2010.09.048). All the mice were genetically engineered to overexpress human gastrin, a characteristic that invariably leads them to develop gastric cancers.

One group comprised germ-free mice raised in sterile environments. The control group was free of pathogens, but lived in a conventional environment and so had normal gastric flora. Both groups were inoculated with H. pylori.

By 11 months, the microbiome of the control group was strikingly different. It showed a significant increase in the number of Firmicutes bacteria in the stomach, with an associated decrease in the number and variety of other bacteria including Bacteroides. This was especially interesting when viewed in relation to the rate of gastric neoplasia, Dr. Peek said.

These mice are programmed to develop gastric cancer by 6 months of age – and this is what happened in the control mice, which had H. pylori plus other gastric microbes. But the germ-free mice who were monoinfected with H. pylori showed a much different progression of disease. At 7 months, most showed only a mild hypergastrinemia. Conversely, at 7 months, all of the H. pylori–infected control mice had developed gastric intraepithelial neoplasia, 80% of it high grade. Only 10% of the monoinfected mice developed cancer, and all of it was low grade.

“It looks like there is active collaboration between H. pylori and other bacteria in the stomach,” resulting in this increased cancer risk, Dr. Peek said.

It’s a collaboration that reaches deep into the tumors themselves, he said. “A very interesting study a couple of years ago searched cancer genomes for the presence of bacterial DNA, and found that gastric cancers incorporated the second-highest amount of microbial DNA into their cancer genomes. But it wasn’t just H. pylori. Many other species had integrated their DNA into these tumors.”

That study, published in 2013, was the first to prove that bacterial DNA can impact carcinogenesis. Acute myeloid leukemia showed the highest integration of bacterial DNA, but gastric adenocarcinoma was a close second. Most of the species were of the Proteobacteria lineages (83%), with a third of that represented by Pseudomonas, particularly P. fluorescens and P. aeruginosa. Both of those species have been shown to promote gastric tumorigenesis in rats. All of the DNA integrations occurred in five genes; four of these are already known to be upregulated in gastric cancer (PLOS Comp Biol. 2013;9[6]:e1003107).

Interestingly, only a few of the sample reads turned up DNA integration with H. pylori.

This reduction in gastric microbial diversity could be an important key to H. pylori’s relation to gastric cancer, Dr. Peek said. He examined this in residents of two towns in Colombia, South America: Tumaco, where the risk of gastric cancer is low, and Tuquerres, where it’s 25 times higher (Sci Rep. 2016. doi:10.1038/srep18594).

What was different was the gastric microbiome of residents. Those living in low-risk Tumaco had much more microbial diversity: 361 varieties, compared with 194 in Tuquerres. And 16 of these groups – representative of what’s usually considered a healthy microbiome – were absent in the high-risk subjects. But Tuquerres residents had two bacteria that weren’t found in Tumaco residents, including Leptorichia wadei, which has been associated with necrotizing enterocolitis.

There was no difference, however, in the prevalence of H. pylori between these high- and low-risk groups.

These new findings illustrate an increasingly complicated interplay of bacteria and gastric cancer, Dr. Peek said. But they also provide a new direction for research.

“We have a framework now where we can move forward and try to understand how some of these other strains impact gastric cancer risk,” he said.

Dr. Peek had no relevant financial disclosures.

[email protected]

On Twitter @Alz_Gal

CHICAGO – A new model of gastroesophageal reflux disease (GERD) paints it as a disease caused by inflammatory molecules, rather than a reaction to an acid-inflicted wound.

And rather than esophagitis due to GERD being a top-down process, from surface epithelium to submucosa, multiple lines of evidence now suggest it is a bottom-up phenomenon sparked by activation of a hypoxia-inducible factor that occurs when esophageal epithelium is exposed to acidic bile salts, Rhonda Souza, MD, said at the meeting sponsored by the American Gastroenterological Association.

Michele G. Sullivan/Frontline Medical News

“We’re proposing that reflux is a cytokine-mediated injury,” said Dr. Souza of the University of Texas Southwestern Medical Center, Dallas, and the Dallas VA Medical Center. “The reflux of acid and bile doesn’t destroy the epithelial cells directly, but induces them to produce proinflammatory cytokines. These cytokines attract lymphocytes first, which induce the basal cell proliferation characteristic of GERD. Ultimately, it’s these inflammatory cells that mediate the epithelial injury – not the direct caustic effects of gastric acid.”

Dr. Souza and her colleagues, including Dr. Stuart Spechler and Dr. Kerry Dunbar, also of UT Southwestern and the Dallas VA Medical Center, have been building this case for several years, beginning with a surgical rat model of GERD. Their histologic findings in this model have been recapitulated in human cell lines and, most recently, in a clinical trial of 12 patients (JAMA. 2016. doi:10.1001/jama.2016.5657).

The rat model, published in 2009 (Gastroenterology. 2009. doi:10.1053/j.gastro.2009.07.055) provided one of the first very early looks at the pathogenesis of acute GERD.

Rats underwent esophagoduodenostomy, a procedure that left the stomach in place so that both gastric and duodenal contents could reflux into the esophagus, thus ensuring immediate esophageal exposure to acid and bile acids. But the investigators were puzzled as to why it took weeks to see changes in the esophageal surface. “The epithelial mucosa stayed intact for far longer than it should have – up to 3 weeks – if acid simply caused a caustic injury as the mechanism of cell death and replacement,” Dr. Souza said.

What she did see, however, was a rapid migration of T cells into the submucosa. “By postoperative week 3, we observed profound basal cell and papillary hyperplasia, but the surface cells were still intact, so this hyperplasia was not due to the death of surface cells.”

The team proceeded to an in vitro model using esophageal squamous cell lines established from endoscopic biopsies obtained from GERD patients. When the squamous cells were exposed in culture to acidic bile salts, the cells ramped up their production of several proinflammatory cytokines, including interleukin-8 and interleukin-1b. The production of proinflammatory cytokines released into the surrounding media were potent recruitment signals for lymphocytes.

The researchers saw this same signaling response in their rat model. “We saw a dramatic increase in IL-8 by postoperative week 2. It was in the intracellular spaces between cells at the epithelial surface and in the cell cytoplasm, and we also saw it in the submucosa and in the lamina propria.”

Acute reflux esophagitis has been almost impossible to observe in humans, Dr. Souza said, because most patients don’t seek medical attention until they’ve had months or years of acid reflux symptoms. By then, the injury response to gastroesophageal reflux has become chronic and well established.

The human study, published in May, confirmed the findings in the rat model. It comprised 12 patients with severe GERD who had been on twice-daily proton pump inhibitor (PPI) therapy for at least 1 month. Successful PPI treatment heals reflux esophagitis rapidly, and healing was endoscopically confirmed at baseline in all these patients. Then, however, they gave up their medication so that the damage would begin again. Dr. Souza and her colleagues could travel back in time, clinically speaking, and track the histopatholgic changes as they occurred. Within 2 weeks, esophagitis had reappeared in every patient: Three had the least-severe LA (Los Angeles) grade A, four had LA grade B, and five had LA grade C esophagitis, with extensive mucosal breaks.

“We know from older studies that within 6 months of going off of PPIs, most patients with reflux esophagitis develop it again, but we weren’t sure we would get this response within 2 weeks. It was surprising that not only did everyone get it, but that a few were so severe,” Dr. Souza said.

Biopsies at weeks 1 and 2 showed the same kind of inflammatory signaling seen in the rats. Again, the responding cells were almost exclusively lymphocytes; neutrophils and eosinophils were very rare or absent in all specimens. The team also observed basal cell and papillary hyperplasia and areas of spongiosis, even though the surface cells were still intact.

The lymphocyte-predominant response is the key to this new pathogenic theory, Dr. Souza wrote in her JAMA paper.

“If the traditional notion were true, that acute GERD is caused by refluxed acid directly inflicting lethal, chemical injury to surface epithelial cells, then basal cell and papillary hyperplasia would have been expected only in areas with surface erosions, and the infiltrating inflammatory cells would have been granulocytes primarily.”

She also suggested that PPIs may be healing esophagitis not simply by preventing acid reflux, but by exerting anti-inflammatory properties.

“Cytokines like IL-8 may also have proliferative effects which might have contributed to esophageal basal cell and papillary hyperplasia observed in the absence of surface erosions. In esophageal epithelial cells in culture, PPIs inhibit secretion of IL-8 through acid-independent mechanisms. This observation raises the interesting possibility that anti-inflammatory PPI effects, independent of their effects on acid inhibition, might contribute to GERD healing by PPIs.”

Dr. Souza said she continues to investigate, focusing now on how the initial insult of acidic bile salts on esophageal epithelium stimulates this inflammatory response. The key may be in a small protein called hypoxia-inducible factor-2 alpha (HIF-2a), one of a family of transcription factors that enable cells to respond to hypoxic stress.

Under normal oxygen conditions, HIF proteins are low, their levels regulated by an enzyme called prolyl hydroxylase. This enzyme is inactivated under hypoxic conditions, or in the presence of reactive oxygen species. HIF factors then rise and, among other functions, stimulate a strong inflammatory response. Inflamed tissues like those seen in esophagitis are frequently hypoxic, Dr. Souza said, and this state could be activating HIFs.

She examined HIF levels in her 12-patient cohort. These results were presented earlier this year at the Digestive Disease Weekmeeting in San Diego.

“At weeks 1 and 2, we found large associations between HIF-2a and increases in a number of proinflammatory cytokines including IL-8 and intercellular adhesion molecule–1,” a protein that facilitates leukocyte migration. Preliminary studies of HIF-2a inhibition in esophageal squamous cells in culture exposed to acidic bile salts show promising results as a potential therapeutic strategy to reduce proinflammatory cytokine expression. It is conceivable that anti-inflammatory therapies directed at HIF-2a may be on the horizon for the prevention and treatment of reflux esophagitis, she added.

Neither Dr. Souza nor her colleagues had any relevant financial disclosures.

[email protected]

On Twitter @Alz_Gal

CHICAGO – A new model of gastroesophageal reflux disease (GERD) paints it as a disease caused by inflammatory molecules, rather than a reaction to an acid-inflicted wound.

And rather than esophagitis due to GERD being a top-down process, from surface epithelium to submucosa, multiple lines of evidence now suggest it is a bottom-up phenomenon sparked by activation of a hypoxia-inducible factor that occurs when esophageal epithelium is exposed to acidic bile salts, Rhonda Souza, MD, said at the meeting sponsored by the American Gastroenterological Association.

Michele G. Sullivan/Frontline Medical News

“We’re proposing that reflux is a cytokine-mediated injury,” said Dr. Souza of the University of Texas Southwestern Medical Center, Dallas, and the Dallas VA Medical Center. “The reflux of acid and bile doesn’t destroy the epithelial cells directly, but induces them to produce proinflammatory cytokines. These cytokines attract lymphocytes first, which induce the basal cell proliferation characteristic of GERD. Ultimately, it’s these inflammatory cells that mediate the epithelial injury – not the direct caustic effects of gastric acid.”

Dr. Souza and her colleagues, including Dr. Stuart Spechler and Dr. Kerry Dunbar, also of UT Southwestern and the Dallas VA Medical Center, have been building this case for several years, beginning with a surgical rat model of GERD. Their histologic findings in this model have been recapitulated in human cell lines and, most recently, in a clinical trial of 12 patients (JAMA. 2016. doi:10.1001/jama.2016.5657).

The rat model, published in 2009 (Gastroenterology. 2009. doi:10.1053/j.gastro.2009.07.055) provided one of the first very early looks at the pathogenesis of acute GERD.

Rats underwent esophagoduodenostomy, a procedure that left the stomach in place so that both gastric and duodenal contents could reflux into the esophagus, thus ensuring immediate esophageal exposure to acid and bile acids. But the investigators were puzzled as to why it took weeks to see changes in the esophageal surface. “The epithelial mucosa stayed intact for far longer than it should have – up to 3 weeks – if acid simply caused a caustic injury as the mechanism of cell death and replacement,” Dr. Souza said.

What she did see, however, was a rapid migration of T cells into the submucosa. “By postoperative week 3, we observed profound basal cell and papillary hyperplasia, but the surface cells were still intact, so this hyperplasia was not due to the death of surface cells.”

The team proceeded to an in vitro model using esophageal squamous cell lines established from endoscopic biopsies obtained from GERD patients. When the squamous cells were exposed in culture to acidic bile salts, the cells ramped up their production of several proinflammatory cytokines, including interleukin-8 and interleukin-1b. The production of proinflammatory cytokines released into the surrounding media were potent recruitment signals for lymphocytes.

The researchers saw this same signaling response in their rat model. “We saw a dramatic increase in IL-8 by postoperative week 2. It was in the intracellular spaces between cells at the epithelial surface and in the cell cytoplasm, and we also saw it in the submucosa and in the lamina propria.”

Acute reflux esophagitis has been almost impossible to observe in humans, Dr. Souza said, because most patients don’t seek medical attention until they’ve had months or years of acid reflux symptoms. By then, the injury response to gastroesophageal reflux has become chronic and well established.

The human study, published in May, confirmed the findings in the rat model. It comprised 12 patients with severe GERD who had been on twice-daily proton pump inhibitor (PPI) therapy for at least 1 month. Successful PPI treatment heals reflux esophagitis rapidly, and healing was endoscopically confirmed at baseline in all these patients. Then, however, they gave up their medication so that the damage would begin again. Dr. Souza and her colleagues could travel back in time, clinically speaking, and track the histopatholgic changes as they occurred. Within 2 weeks, esophagitis had reappeared in every patient: Three had the least-severe LA (Los Angeles) grade A, four had LA grade B, and five had LA grade C esophagitis, with extensive mucosal breaks.

“We know from older studies that within 6 months of going off of PPIs, most patients with reflux esophagitis develop it again, but we weren’t sure we would get this response within 2 weeks. It was surprising that not only did everyone get it, but that a few were so severe,” Dr. Souza said.

Biopsies at weeks 1 and 2 showed the same kind of inflammatory signaling seen in the rats. Again, the responding cells were almost exclusively lymphocytes; neutrophils and eosinophils were very rare or absent in all specimens. The team also observed basal cell and papillary hyperplasia and areas of spongiosis, even though the surface cells were still intact.

The lymphocyte-predominant response is the key to this new pathogenic theory, Dr. Souza wrote in her JAMA paper.

“If the traditional notion were true, that acute GERD is caused by refluxed acid directly inflicting lethal, chemical injury to surface epithelial cells, then basal cell and papillary hyperplasia would have been expected only in areas with surface erosions, and the infiltrating inflammatory cells would have been granulocytes primarily.”

She also suggested that PPIs may be healing esophagitis not simply by preventing acid reflux, but by exerting anti-inflammatory properties.

“Cytokines like IL-8 may also have proliferative effects which might have contributed to esophageal basal cell and papillary hyperplasia observed in the absence of surface erosions. In esophageal epithelial cells in culture, PPIs inhibit secretion of IL-8 through acid-independent mechanisms. This observation raises the interesting possibility that anti-inflammatory PPI effects, independent of their effects on acid inhibition, might contribute to GERD healing by PPIs.”

Dr. Souza said she continues to investigate, focusing now on how the initial insult of acidic bile salts on esophageal epithelium stimulates this inflammatory response. The key may be in a small protein called hypoxia-inducible factor-2 alpha (HIF-2a), one of a family of transcription factors that enable cells to respond to hypoxic stress.

Under normal oxygen conditions, HIF proteins are low, their levels regulated by an enzyme called prolyl hydroxylase. This enzyme is inactivated under hypoxic conditions, or in the presence of reactive oxygen species. HIF factors then rise and, among other functions, stimulate a strong inflammatory response. Inflamed tissues like those seen in esophagitis are frequently hypoxic, Dr. Souza said, and this state could be activating HIFs.

She examined HIF levels in her 12-patient cohort. These results were presented earlier this year at the Digestive Disease Weekmeeting in San Diego.

“At weeks 1 and 2, we found large associations between HIF-2a and increases in a number of proinflammatory cytokines including IL-8 and intercellular adhesion molecule–1,” a protein that facilitates leukocyte migration. Preliminary studies of HIF-2a inhibition in esophageal squamous cells in culture exposed to acidic bile salts show promising results as a potential therapeutic strategy to reduce proinflammatory cytokine expression. It is conceivable that anti-inflammatory therapies directed at HIF-2a may be on the horizon for the prevention and treatment of reflux esophagitis, she added.

Neither Dr. Souza nor her colleagues had any relevant financial disclosures.

[email protected]

On Twitter @Alz_Gal

CHICAGO – A new model of gastroesophageal reflux disease (GERD) paints it as a disease caused by inflammatory molecules, rather than a reaction to an acid-inflicted wound.

And rather than esophagitis due to GERD being a top-down process, from surface epithelium to submucosa, multiple lines of evidence now suggest it is a bottom-up phenomenon sparked by activation of a hypoxia-inducible factor that occurs when esophageal epithelium is exposed to acidic bile salts, Rhonda Souza, MD, said at the meeting sponsored by the American Gastroenterological Association.

Michele G. Sullivan/Frontline Medical News

“We’re proposing that reflux is a cytokine-mediated injury,” said Dr. Souza of the University of Texas Southwestern Medical Center, Dallas, and the Dallas VA Medical Center. “The reflux of acid and bile doesn’t destroy the epithelial cells directly, but induces them to produce proinflammatory cytokines. These cytokines attract lymphocytes first, which induce the basal cell proliferation characteristic of GERD. Ultimately, it’s these inflammatory cells that mediate the epithelial injury – not the direct caustic effects of gastric acid.”

Dr. Souza and her colleagues, including Dr. Stuart Spechler and Dr. Kerry Dunbar, also of UT Southwestern and the Dallas VA Medical Center, have been building this case for several years, beginning with a surgical rat model of GERD. Their histologic findings in this model have been recapitulated in human cell lines and, most recently, in a clinical trial of 12 patients (JAMA. 2016. doi:10.1001/jama.2016.5657).

The rat model, published in 2009 (Gastroenterology. 2009. doi:10.1053/j.gastro.2009.07.055) provided one of the first very early looks at the pathogenesis of acute GERD.

Rats underwent esophagoduodenostomy, a procedure that left the stomach in place so that both gastric and duodenal contents could reflux into the esophagus, thus ensuring immediate esophageal exposure to acid and bile acids. But the investigators were puzzled as to why it took weeks to see changes in the esophageal surface. “The epithelial mucosa stayed intact for far longer than it should have – up to 3 weeks – if acid simply caused a caustic injury as the mechanism of cell death and replacement,” Dr. Souza said.

What she did see, however, was a rapid migration of T cells into the submucosa. “By postoperative week 3, we observed profound basal cell and papillary hyperplasia, but the surface cells were still intact, so this hyperplasia was not due to the death of surface cells.”

The team proceeded to an in vitro model using esophageal squamous cell lines established from endoscopic biopsies obtained from GERD patients. When the squamous cells were exposed in culture to acidic bile salts, the cells ramped up their production of several proinflammatory cytokines, including interleukin-8 and interleukin-1b. The production of proinflammatory cytokines released into the surrounding media were potent recruitment signals for lymphocytes.

The researchers saw this same signaling response in their rat model. “We saw a dramatic increase in IL-8 by postoperative week 2. It was in the intracellular spaces between cells at the epithelial surface and in the cell cytoplasm, and we also saw it in the submucosa and in the lamina propria.”

Acute reflux esophagitis has been almost impossible to observe in humans, Dr. Souza said, because most patients don’t seek medical attention until they’ve had months or years of acid reflux symptoms. By then, the injury response to gastroesophageal reflux has become chronic and well established.

The human study, published in May, confirmed the findings in the rat model. It comprised 12 patients with severe GERD who had been on twice-daily proton pump inhibitor (PPI) therapy for at least 1 month. Successful PPI treatment heals reflux esophagitis rapidly, and healing was endoscopically confirmed at baseline in all these patients. Then, however, they gave up their medication so that the damage would begin again. Dr. Souza and her colleagues could travel back in time, clinically speaking, and track the histopatholgic changes as they occurred. Within 2 weeks, esophagitis had reappeared in every patient: Three had the least-severe LA (Los Angeles) grade A, four had LA grade B, and five had LA grade C esophagitis, with extensive mucosal breaks.

“We know from older studies that within 6 months of going off of PPIs, most patients with reflux esophagitis develop it again, but we weren’t sure we would get this response within 2 weeks. It was surprising that not only did everyone get it, but that a few were so severe,” Dr. Souza said.

Biopsies at weeks 1 and 2 showed the same kind of inflammatory signaling seen in the rats. Again, the responding cells were almost exclusively lymphocytes; neutrophils and eosinophils were very rare or absent in all specimens. The team also observed basal cell and papillary hyperplasia and areas of spongiosis, even though the surface cells were still intact.

The lymphocyte-predominant response is the key to this new pathogenic theory, Dr. Souza wrote in her JAMA paper.

“If the traditional notion were true, that acute GERD is caused by refluxed acid directly inflicting lethal, chemical injury to surface epithelial cells, then basal cell and papillary hyperplasia would have been expected only in areas with surface erosions, and the infiltrating inflammatory cells would have been granulocytes primarily.”

She also suggested that PPIs may be healing esophagitis not simply by preventing acid reflux, but by exerting anti-inflammatory properties.

“Cytokines like IL-8 may also have proliferative effects which might have contributed to esophageal basal cell and papillary hyperplasia observed in the absence of surface erosions. In esophageal epithelial cells in culture, PPIs inhibit secretion of IL-8 through acid-independent mechanisms. This observation raises the interesting possibility that anti-inflammatory PPI effects, independent of their effects on acid inhibition, might contribute to GERD healing by PPIs.”

Dr. Souza said she continues to investigate, focusing now on how the initial insult of acidic bile salts on esophageal epithelium stimulates this inflammatory response. The key may be in a small protein called hypoxia-inducible factor-2 alpha (HIF-2a), one of a family of transcription factors that enable cells to respond to hypoxic stress.

Under normal oxygen conditions, HIF proteins are low, their levels regulated by an enzyme called prolyl hydroxylase. This enzyme is inactivated under hypoxic conditions, or in the presence of reactive oxygen species. HIF factors then rise and, among other functions, stimulate a strong inflammatory response. Inflamed tissues like those seen in esophagitis are frequently hypoxic, Dr. Souza said, and this state could be activating HIFs.

She examined HIF levels in her 12-patient cohort. These results were presented earlier this year at the Digestive Disease Weekmeeting in San Diego.

“At weeks 1 and 2, we found large associations between HIF-2a and increases in a number of proinflammatory cytokines including IL-8 and intercellular adhesion molecule–1,” a protein that facilitates leukocyte migration. Preliminary studies of HIF-2a inhibition in esophageal squamous cells in culture exposed to acidic bile salts show promising results as a potential therapeutic strategy to reduce proinflammatory cytokine expression. It is conceivable that anti-inflammatory therapies directed at HIF-2a may be on the horizon for the prevention and treatment of reflux esophagitis, she added.

Neither Dr. Souza nor her colleagues had any relevant financial disclosures.

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