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Posts Tagged ‘inflammatory bowel disease (IBD)’


 

Two New Drugs for Inflammatory Bowel Syndrome Are Giving Patients Hope

Reporter: Stephen J. Williams, Ph.D.

Actavis Receives FDA Approval for VIBERZI (eluxadoline) for the Treatment of Irritable Bowel Syndrome with Diarrhea (IBS-D) in Adults -First in class treatment for IBS-D treats hallmark symptoms of IBS-D; abdominal pain and diarrhea

DUBLIN, May 27, 2015 /PRNewswire/ — Actavis plc (NYSE: ACT) announced today that VIBERZI™ (eluxadoline) was approved by the Food and Drug Administration (FDA) as a twice-daily, oral treatment for adults suffering from irritable bowel syndrome with diarrhea (IBS-D). VIBERZI (eluxadoline) has mixed opioid receptor activity, it is a mu receptor agonist, a delta receptor antagonist, and a kappa receptor agonist.

Logo – http://photos.prnewswire.com/prnh/20130124/NY47381LOGO

“The FDA’s approval of VIBERZI is the first step to providing physicians with a new, evidence-based, treatment option for their adult patients with IBS-D,” said David Nicholson, Executive Vice President, Actavis Global Brands R&D. “At Actavis, we are dedicated to providing new treatment options, and the development of new agents that help address the most bothersome symptoms of IBS-D. We are very pleased to be working with the FDA to advance this IBS-D treatment and we eagerly await DEA scheduling determination later this year.”

IBS-D is a multifactorial disorder marked by recurrent abdominal pain or discomfort and altered bowel function that affects as many as 15 million adult Americans, impacting about twice as many women as men.i,ii,iii There are few treatment options available for IBS-D, particularly options that relieve both the diarrhea and abdominal pain associated with IBS-D.

“The unpredictable symptoms experienced by patients with IBS-D can have a significant impact on everyday life,” said William D. Chey, MD, Nostrant Professor of Gastroenterology at the University of Michigan Health System. “It’s exciting when physicians are able to add an additional treatment option like VIBERZI to their toolbox for patients with IBS-D.”

The FDA has recommended that VIBERZI be classified as a controlled substance. This recommendation has been submitted to the U.S. Drug Enforcement Administration (DEA).  Once VIBERZI receives final scheduling designation, the updated label will be available. Pending final scheduling designation, product launch is anticipated in Q1 2016.

About VIBERZI

VIBERZI is an orally active compound indicated for the treatment of irritable bowel syndrome with diarrhea (IBS-D) in men and women. VIBERZI (eluxadoline) has mixed opioid receptor activity, it is a mu receptor agonist, a delta receptor antagonist, and a kappa receptor agonist.

Efficacy was established in two Phase III clinical studies, demonstrating significant superiority over placebo on the composite endpoint of simultaneous improvement in both abdominal pain and diarrhea at both 75 mg and 100 mg twice daily doses. The primary efficacy responder endpoint was evaluated over the duration of double-blind, placebo-controlled treatment. Response rates were compared based on patients who met the daily composite response criteria (improvement in both abdominal pain and stool consistency on the same day) for at least 50% of the days from weeks 1 to 12 (FDA endpoint) and weeks 1 to 26 (European Medicines Agency endpoint).

The most common adverse events in the two Phase III clinical trials were constipation (7% and 8% for eluxadoline 75 mg and 100 mg; 2% for placebo) and nausea (8% and 7% for eluxadoline 75 mg and 100 mg; 5% for placebo). Rates of severe constipation were less than 1% in patients receiving 75 mg and 100 mg eluxadoline. Rates of discontinuation due to constipation were low for both eluxadoline and placebo (≤2%) and similar rates of constipation occurred between the active and placebo arms beyond 3 months of treatment. A total of 2,426 subjects were enrolled across the two studies.

For more information including full prescribing information about VIBERZI at http://www.actavis.com/Actavis/media/PDFDocuments/VIBERZI_PI.pdf

About IBS-D

Irritable bowel syndrome with diarrhea (IBS-D) is a functional bowel disorder characterized by chronic abdominal pain and frequent diarrhea, which affects approximately 15 million patients in the U.S.  Although the exact cause of IBS-D is not known, symptoms are thought to result from a disturbance in the way the gastrointestinal tract and nervous system interact.

IBS-D can be debilitating and there are limited therapeutic options for managing the chronic symptoms. IBS-D is associated with economic burden in direct medical costs and indirect social costs such as absenteeism and lost productivity, along with decreased quality of life.

About Actavis
Actavis plc (NYSE: ACT), headquartered in Dublin, Ireland, is a unique, global pharmaceutical company and a leader in a new industry model—Growth Pharma. Actavis is focused on developing, manufacturing and commercializing innovative branded pharmaceuticals, high-quality generic and over-the-counter medicines and biologic products for patients around the world.

Actavis markets a portfolio of best-in-class products that provide valuable treatments for the central nervous system, eye care, medical aesthetics, gastroenterology, women’s health, urology, cardiovascular and anti-infective therapeutic categories, and operates the world’s third-largest global generics business, providing patients around the globe with increased access to affordable, high-quality medicines. Actavis is an industry leader in research and development, with one of the broadest development pipelines in the pharmaceutical industry and a leading position in the submission of generic product applications globally.

With commercial operations in approximately 100 countries, Actavis is committed to working with physicians, healthcare providers and patients to deliver innovative and meaningful treatments that help people around the world live longer, healthier lives.

Actavis intends to adopt a new global name – Allergan – pending shareholder approval in 2015.

For more information, visit Actavis’ website at www.actavis.com.

Actavis Cautionary Statement Regarding Forward-Looking Statements

Statements contained in this communication that refer to Actavis’ estimated or anticipated future results, including estimated synergies, or other non-historical facts are forward-looking statements that reflect Actavis’ current perspective of existing trends and information as of the date of this communication. Actual results may differ materially from Actavis’ current expectations depending upon a number of factors affecting Actavis’ business. These factors include, among others, the timing and success of product launches; the difficulty of predicting the timing or outcome of product development efforts and regulatory agency approvals or actions, if any; market acceptance of and continued demand for Actavis’ products; difficulties or delays in manufacturing; and such other risks and uncertainties detailed in Actavis’ periodic public filings with the Securities and Exchange Commission, including but not limited to Actavis plc’s Quarterly Report on Form 10-Q for the quarter ended March 31, 2015 and from time to time in Actavis’ other investor communications. Except as expressly required by law, Actavis disclaims any intent or obligation to update or revise these forward-looking statements.

i Camilleri M. Current and future pharmacological treatments for diarrhea-predominant irritable bowel syndrome. Expert Opinion on Pharmacotherapy. 2013;14:1151.

ii Grundmann O, Yoon SL. Irritable bowel syndrome: epidemiology, diagnosis, and treatment: an update for health-care practitioners. Journal of Gastroenterology and Hepatology. 2010;25:691–699.

iii Eluxadoline Xifaxin Summary Final. November 2014.

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Synergy’s Looming FDA Filing Makes It Pharma of the Month

By James Passeri Follow

| Jan 05, 2016 | 8:39 AM EST  | 0

Keep an eye on Synergy Pharmaceuticals (SGYP) this month: Analysts like it, its shares have waned since a big spike this summer, and the official filing of its star product is expected any day.

When the New York-based pharmaceutical company, which specializes in gastrointestinal therapy, announced that it passed clinical trials on its flagship drug plecanatide this summer, shares rocketed 95%.

But today analysts appear mystified at why the stock has receded 45% from its July high, especially with plecanatide’s new drug application with the Food and Drug Administration expected this month. (It’s currently trading below $6, and the consensus price target is over $13, according to data provided by Bloomberg.)

Synergy should be raking in $600 million from plecanatide, a daily tablet that treats patients with irritable bowel syndrome (IBS), within five years of obtaining FDA approval (expected in 2017, according to equity research firm BTIG. Synergy currently has a market capitalization of just $645 million.

BTIG’s $11 price target is also buoyed by roughly $142 million on the balance sheet, as well as newly appointed management including CFO Gary Sender and COO Troy Hamilton, both former executives at pharma success story Shire (SHPG). Though Shire shares are down just under 4% over the past 12 month, they have rocketed 112% over the past two years.

Synergy also stands to benefit from a growing demand for gastrointestinal treatments, feeding the appetite of Big Pharma for potential acquisitions, according to BTIG.

“With about 45 million Americans suffering from chronic constipation and IBS, and major companies like Allergan(AGN) and Valeant (VRX) focusing their marketing efforts on GI treatments, it seems logical to imagine SGYP as a takeover candidate,” BTIG analyst Timothy Chiang wrote in a November report.

Whether or not this leads to a buyout or another stock surge, Synergy certainly can be counted on for a healthy dose of small-cap volatility as its chief product takes the final steps toward reaching its customers.

 

 

Synergy Pharmaceuticals Announces Successful End-of-Phase 2 Meeting with FDA for Plecanatide in Irritable Bowel Syndrome with Constipation

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Pivotal Phase 3 IBS-C Program to be Initiated in the Fourth Quarter of 2014

NEW YORK– Synergy Pharmaceuticals Inc. (NASDAQ:SGYP) today announced that it has successfully completed an End-of-Phase 2 meeting with the U.S. Food and Drug Administration (FDA) on its lead drug plecanatide for the treatment of irritable bowel syndrome with constipation (IBS-C). Agreement was reached with the FDA for the plecanatide pivotal phase 3 IBS-C clinical development program that is scheduled to begin in the fourth quarter of this year.

“We are very pleased with the outcome of our meeting with the FDA and have a clear path forward to start the IBS-C registration program with plecanatide this year,” said Dr. Gary S. Jacob, Chairman and CEO of Synergy. “The pivotal phase 3 IBS-C trials will include both 3.0 mg and 6.0 mg plecanatide, which are consistent with the doses currently being evaluated in our phase 3 chronic idiopathic constipation (CIC) program. Plecanatide has demonstrated a clinical dose-response for efficacy with an excellent tolerability profile that is observed across trials. This is an important advantage as we look to bring two doses to market in both indications and provide physicians with options for addressing individual patient needs.”

Synergy’s pivotal phase 3 IBS-C clinical development program will consist of two registration trials, each including 1,050 patients who will receive either placebo, 3.0 mg or 6.0 mg plecanatide. IBS-C patients successfully completing either of the 12-week placebo-controlled registration trials will be offered enrollment into a long-term safety trial in order to complement and support the ongoing long-term safety database for the CIC indication.

About Plecanatide

Plecanatide is Synergy’s lead uroguanylin analog in late-stage clinical development to treat patients with CIC and IBS-C. Uroguanylin is a natural gastrointestinal (GI) hormone produced by humans in the small intestine and plays a key role in regulating the normal functioning of the digestive tract through its activity on the guanylate cyclase-C (GC-C) receptor. The GC-C receptor is known to be a primary source for stimulating a variety of beneficial physiological responses. Orally administered plecanatide mimics uroguanylin’s functions by binding to and activating the GC-C receptor to stimulate fluid and ion transit required for normal bowel function. Synergy has successfully completed a phase 2b trial of plecanatide in 951 patients with CIC and is currently enrolling patients in two pivotal phase 3 CIC trials. The company also recently announced positive top-line data results from a phase 2b dose-ranging study with plecanatide in patients with IBS-C.

About Synergy Pharmaceuticals

Synergy Pharmaceuticals (NASDAQ:SGYP) is a biopharmaceutical company focused on the development of novel therapies based on the natural human hormone, uroguanylin, to treat GI diseases and disorders. Synergy has created two unique analogs of uroguanylin – plecanatide and SP-333 – designed to mimic the natural hormone’s activity on the GC-C receptor and target a variety of GI conditions. SP-333 is currently in phase 2 development for opioid-induced constipation and is also being explored for ulcerative colitis. For more information, please visit www.synergypharma.com.

 

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Contribution to Inflammatory Bowel Disease (IBD) of bacterial overgrowth in gut on a chip

Larry H. Bernstein, MD, FCAP, Curator

LPBI

 

Contributions of microbiome and mechanical deformation to intestinal bacterial overgrowth and inflammation in a 

human gut-on-a-chip 
Gut-On-a-Chip Holds Clues for Treating Inflammatory Bowel Diseases
Greg Watry
Human intestinal epithelial cells cultured in the Wyss Institute's human gut-on-a-chip form differentiated intestinal villi when cultured in the presence of lifelike fluid flow and rhythmic, peristalsis-like motions. Here the villi are visible using a traditional microscope (left) or a confocal microscope (right); when the same villi are stained with fluorescent antibodies, it clearly reveals the nuclei in the intestinal cells (blue) and their specialized apical membranes when they contact the intestinal lumen (green). Credit: Wyss Institute at Harvard University
Human intestinal epithelial cells cultured in the Wyss Institute’s human gut-on-a-chip form differentiated intestinal villi when cultured in the presence of lifelike fluid flow and rhythmic, peristalsis-like motions. Here the villi are visible using a traditional microscope (left) or a confocal microscope (right); when the same villi are stained with fluorescent antibodies, it clearly reveals the nuclei in the intestinal cells (blue) and their specialized apical membranes when they contact the intestinal lumen (green). Credit: Wyss Institute at Harvard University

Roughly the size of a computer memory stick and made of clear flexible polymer, the human gut-on-a-chip was created by Harvard Univ.’s Wyss Institute in 2012. Three years later, researchers are utilizing the technology in hopes of creating new therapies for inflammatory bowel diseases (IBD).

The Centers for Disease Control and Prevention estimates that between 1 and 1.3 million people suffer from IBD, including such diseases as ulcerative colitis and Crohn’s disease. With origins still mysterious, IBD is currently incurable.

“It has not been possible to study…human intestinal inflammatory diseases, because it is not possible to independently control these parameters in animal studies or in vitro models,” wrote the researchers in Proceedings of the National Academy of the Sciences. “In particular, given the recent recognition of the central role of the intestinal microbiome in human health and disease, including intestinal disorders, it is critical to incorporate commensal microbes into experimental models, however, this has not been possible using conventional culture systems.”

Additionally, static in vitro methods fail to replicate the pathophysiology of human IBD.

But the hollow-channeled microfluidic gut-on-a-chip successfully simulates the human intestine’s physical structure, microenvironment, peristalsis-like motion, and fluid flow.

“With our human gut-on-a-chip, we can not only culture the normal gut microbiome for extended times, but we can also analyze contributions of pathogens, immune cells, and vascular and lymphatic endothelium, as well as model specific diseases to understand the complex pathophysiological responses of the intestinal tract,” said Donald Ingber, founding director of the Wyss Institute.

The device was “used to co-culture multiple commensal microbes in contact with living human intestinal epithelial cells for more than a week in vitro and to analyze how gut microbiome, inflammatory cells, and peristalsis-associated mechanical deformations independently contribute to intestinal bacterial overgrowth and inflammation,” the researchers wrote.

Thus far, use of the device has yielded two interesting observations.

Four proteins—called cytokines—work together to trigger an inflammatory responses that exacerbate the bowel, the researchers found. Potentially, this new discovery could lead to the development of treatments that block the cytokine interaction.

Another observation, the researchers noted, is that “by ceasing peristalsis-like motions while maintaining luminal flow, lack of epithelial deformation was shown to trigger bacterial overgrowth similar to that observed in patients with ileus and inflammatory bowel disease,” according to the researchers.

The researchers believe the micro-device may one day be applicable to precision medicine. Eventually, a custom treatment may arise from scientists using a patient’s gut microbiota and cells on a human gut-on-a-chip.

 

 

Contributions of microbiome and mechanical deformation to intestinal bacterial overgrowth and inflammation in a human gut-on-a-chip
Hyun Jung Kima,1, Hu Lia,2, James J. Collinsa,b,c,d,e,f,3, and Donald E. Ingbera,g,h,
http://www.pnas.org/content/early/2015/12/09/1522193112.full.pdf

A human gut-on-a-chip microdevice was used to coculture multiple commensal microbes in contact with living human intestinal epithelial cells for more than a week in vitro and to analyze how gut microbiome, inflammatory cells, and peristalsis-associated mechanical deformations independently contribute to intestinal bacterial overgrowth and inflammation. This in vitro model replicated results from past animal and human studies, including demonstration that probiotic and antibiotic therapies can suppress villus injury induced by pathogenic bacteria. By ceasing peristalsis-like motions while maintaining luminal flow, lack of epithelial deformation was shown to trigger bacterial overgrowth similar to that observed in patients with ileus and inflammatory bowel disease. Analysis of intestinal inflammation on-chip revealed that immune cells and lipopolysaccharide endotoxin together stimulate epithelial cells to produce four proinflammatory cytokines (IL-8, IL-6, IL-1β, and TNF-α) that are necessary and sufficient to induce villus injury and compromise intestinal barrier function. Thus, this human gut-on-a-chip can be used to analyze contributions of microbiome to intestinal pathophysiology and dissect disease mechanisms in a controlled manner that is not possible using existing in vitro systems or animal models.

 

Significance The main advance of this study is the development of a microengineered model of human intestinal inflammation and bacterial overgrowth that permits analysis of individual contributors to the pathophysiology of intestinal diseases, such as ileus and inflammatory bowel disease, over a period of weeks in vitro. By studying living human intestinal epithelium, with or without vascular and lymphatic endothelium, immune cells, and mechanical deformation, as well as living microbiome and pathogenic microbes, we identified previously unknown contributions of specific cytokines, mechanical motions, and microbiome to intestinal inflammation, bacterial overgrowth, and control of barrier function. We provide proof-of-principle to show that the microfluidic gut-on-a-chip device can be used to create human intestinal disease models and gain new insights into gut pathophysiology.

 

Various types of inflammatory bowel disease (IBD), such as Crohn’s disease and ulcerative colitis, involve chronic inflammation of human intestine with mucosal injury and villus destruction (1), which is believed to be caused by complex interactions between gut microbiome (including commensal and pathogenic microbes) (2), intestinal mucosa, and immune components (3). Suppression of peristalsis also has been strongly associated with intestinal pathology, inflammation (4, 5), and small intestinal bacterial overgrowth (5, 6) in patients with Crohn’s disease (7) and ileus (8). However, it has not been possible to study the relative contributions of these different potential contributing factors to human intestinal inflammatory diseases, because it is not possible to independently control these parameters in animal studies or in vitro models. In particular, given the recent recognition of the central role of the intestinal microbiome in human health and disease, including intestinal disorders (2), it is critical to incorporate commensal microbes into experimental models; however, this has not been possible using conventional culture systems. Most models of human intestinal inflammatory diseases rely either on culturing an intestinal epithelial cell monolayer in static Transwell culture (9) or maintaining intact explanted human intestinal mucosa ex vivo (10) and then adding live microbes and immune cells to the apical (luminal) or basolateral (mucosal) sides of the cultures, respectively. These static in vitro methods, however, do not effectively recapitulate the pathophysiology of human IBD. For example, intestinal epithelial cells cultured in Transwell plates completely fail to undergo villus differentiation, produce mucus, or form the various specialized cell types of normal intestine. Although higher levels of intestinal differentiation can be obtained using recently developed 3D organoid cultures (11), it is not possible to expose these cells to physiological peristalsis-like motions or living microbiome in long-term culture, because bacterial overgrowth occurs rapidly (within ∼1 d) compromising the epithelium (12). This is a major limitation because establishment of stable symbiosis between the epithelium and resident gut microbiome as observed in the normal intestine is crucial for studying inflammatory disease initiation and progression (13), and rhythmical mechanical deformations driven by peristalsis are required to both maintain normal epithelial differentiation (14) and restrain microbial overgrowth in the intestine in vivo (15).

Thus, we set out to develop an experimental model that would overcome these limitations. To do this, we adapted a recently described human gut-on-a-chip microfluidic device that enables human intestinal epithelial cells (Caco-2) to be cultured in the presence of physiologically relevant luminal flow and peristalsislike mechanical deformations, which promotes formation of intestinal villi lined by all four epithelial cell lineages of the small intestine (absorptive, goblet, enteroendocrine, and Paneth) (12, 16). These villi also have enhanced barrier function, drug-metabolizing cytochrome P450 activity, and apical mucus secretion compared with the same cells grown in conventional Transwell cultures, which made it possible to coculture a probiotic gut microbe (Lactobacillus rhamnosus GG) in direct contact with the intestinal epithelium for more than 2 wk (12), in contrast to static Transwell cultures (17) or organoid cultures (11) that lose viability within hours under similar conditions. In the present study, we leveraged this human gut-on-a-chip to develop a disease model of small intestinal bacterial overgrowth (SIBO) and inflammation. We analyzed how probiotic and pathogenic bacteria, lipopolysaccharide (LPS), immune cells, inflammatory cytokines, vascular endothelial cells and mechanical forces contribute individually, and in combination, to intestinal inflammation, villus injury, and compromise of epithelial barrier function. We also explored whether we could replicate the protective effects of clinical probiotic and antibiotic therapies on-chip to demonstrate its potential use as an in vitro tool for drug development, as well as for dissecting fundamental disease mechanisms.

 

Fig. 1. The human gut-on-a-chip microfluidic device and changes in phenotype resulting from different culture conditions on-chip, as measured using genome-wide gene profiling. (A) A photograph of the device. Blue and red dyes fill the upper and lower microchannels, respectively. (B) A schematic of a 3D cross-section of the device showing how repeated suction to side channels (gray arrows) exerts peristalsis-like cyclic mechanical strain and fluid flow (white arrows) generates a shear stress in the perpendicular direction. (C) A DIC micrograph showing intestinal basal crypt (red arrow) and villi (white arrow) formed by human Caco-2 intestinal epithelial cells grown for ∼100 h in the gut-on-achip under medium flow (30 μL/h) and cyclic mechanical stretching (10%, 0.15 Hz). (Scale bar, 50 μm.) (D) A confocal immunofluorescence image showing a horizontal cross-section of intestinal villi similar to those shown in Fig. 1C, stained for F-actin (green) that labels the apical brush border of these polarized intestinal epithelial cells (nuclei in blue). (Scale bar, 50 μm.) (E) Hierarchical clustering analysis of genome-wide transcriptome profiles (Top) of Caco-2 cells cultured in the static Transwell, the gut-on-a-chip (with fluid flow at 30 μL/h and mechanical deformations at 10%, 0.15 Hz) (Gut Chip), or the mechanically active gut-on-a-chip cocultured with the VSL#3 formulation containing eight probiotic gut microbes (Gut Chip + VSL#3) for 72 h compared with normal human small intestinal tissues (Duodenum, Jejunum, and Ileum; microarray data from the published GEO database). The dendrogram was generated based on the averages calculated across all replicates, and all branches in the cluster have the approximately unbiased (AU) P value equal to 100. The y axis next to the dendrogram represents the metric for Euclidean distance between samples. Corresponding pseudocolored GEDI maps analyzing profiles of 650 metagenes between samples described above (Bottom).

 

Fig. 2. Reconstitution of pathological intestinal injury induced by interplay between nonpathogenic or pathogenic enteroinvasive E. coli bacteria or LPS endotoxin with immune cells. (A) DIC images showing that the normal villus morphology of the intestinal epithelium cultured on-chip (Control) is lost within 24 h after EIEC (serotype O124:NM) are added to the apical channel of the chip (+EIEC; red arrows indicate bacterial colonies). (B) Effects of GFP-EC, LPS (15 μg/mL), EIEC, or no addition (Control) on intestinal barrier function (Left). Right shows the TEER profiles in the presence of human PBMCs (+PBMC). GFP-EC, LPS, and EIEC were added to the apical channel (intestinal lumen) at 4, 12, and 35 h, respectively, and PBMCs were subsequently introduced through the lower capillary channel at 44 h after the onset of experiment (0 h) (n = 4). (C) Morphological analysis of intestinal villus damage in response to addition of GFP-EC, LPS, and EIEC in the absence (−PBMC) or the presence of immune components (+PBMC). Schematics (experimental setup), phase contrast images (horizontal view, taken at 57 h after onset), and fluorescence confocal micrographs (vertical cross-sectional views at 83 h after onset) were sequentially displayed. F-actin and nuclei were coded with magenta and blue, respectively. (D) Quantification of intestinal injury evaluated by measuring changes in lesion area (Top; n = 30) and the height of the villi (Bottom; n = 50) in the absence (white) or the presence (gray) of PBMCs. Intestinal villi were grown in the gut-on-a-chip under trickling flow (30 μL/h) with cyclic deformations (10%, 0.15 Hz) during the preculture period for ∼100 h before stimulation (0 h, onset). Asterisks indicate statistical significance compared with the control at the same time point (*P < 0.001, **P < 0.05). (Scale bars, 50 μm.)

 

Recapitulating Organ-Level Intestinal Inflammatory Responses. During inflammation in the intestine, pathophysiological recruitment of circulating immune cells is regulated via activation of the underlying vascular endothelium. To analyze this organ-level inflammatory response in our in vitro model, a monolayer of human microvascular endothelial cells (Fig. 3 C and D and Fig. S6 A and C) or lymphatic endothelial cells (Fig. S6 B and C) was cultured on the opposite (abluminal) side of the porous ECM-coated membrane in the lower microchannel of the device to effectively create a vascular channel (Fig. 3C). To induce intestinal inflammatory responses, LPS (Fig. 3 C and D) or TNF-α (Fig. S6) was flowed through the upper epithelial channel for 24 h, and then PBMCs were added to the vascular channel for 1 h without flow (Fig. 3 C and D). Treatment with both LPS (or TNF-α) and PBMCs resulted in the activation of intercellular adhesion molecule-1 (ICAM-1) expression on the surface of the endothelium (Fig. 3 C and D, Left, and Fig. S6) and a significant increase (P < 0.001) in the number of PBMCs that adhered to the surface of the capillary endothelium compared with controls (Fig. 3D). These results are consistent with our qPCR results, which also showed up-regulation of genes involved in immune cell trafficking (Fig. S5). Neither addition of LPS nor PBMCs alone was sufficient to induce ICAM-1 expression in these cells (Fig. 3D), which parallels the effects of LPS and PBMCs on epithelial production of inflammatory cytokines (Fig. 3A) as well as on villus injury (Fig. 2 B and D).

Evaluating Antiinflammatory Probiotic and Antibiotic Therapeutics On-Chip. To investigate how the gut microbiome modulates these inflammatory reactions, we cocultured the human intestinal villi with the eight strains of probiotic bacteria in the VSL#3 formulation that significantly enhanced intestinal differentiation (Fig. 1E and Fig. S1B). To mimic the in vivo situation, we colonized our microengineered gut on a chip with the commensal microbes (VSL#3) first and then subsequently added immune cells (PBMCs), pathogenic bacteria (EIEC), or both in combination. The VSL#3 microbial cells inoculated into the germ-free lumen of the epithelial channel primarily grew as discrete microcolonies in the spaces between adjacent villi (Fig. 4A and Movie S3) for more than a week in culture (Fig. S7A), and no planktonic growth was detected. These microbes did not overgrow like the EIEC (Fig. 2A and Movie S2), although occasional microcolonies also appeared at different spatial locations in association with the tips of the villi (Fig. S7 B and C). The presence of these living components of the normal gut microbiome significantly enhanced (P < 0.001) intestinal barrier function, producing more than a 50% increase in TEER relative to control cultures (Fig. 4B) without altering villus morphology (Fig. 4C). This result is consistent with clinical studies suggesting that probiotics, including VSL#3, can significantly enhance intestinal barrier function in vivo (18).

To mimic the effects of antibiotic therapies that are sometimes used clinically in patients with intestinal inflammatory disease (29), we identified a dose and combination of antibiotics (100 units per mL penicillin and 100 μg/mL streptomycin) that produced effective killing of both EIEC and VSL#3 microbes in liquid cultures (Fig. S9) and then injected this drug mixture into the epithelial channel of guton-a-chip devices infected with EIEC. When we added PBMCs to these devices 1 h later, intestinal barrier function (Fig. 4B) and villus morphology (Fig. 4C) were largely protected from injury, and there was a significant reduction in lesion area (Fig. 4D). Thus, the gut-on-a-chip was able to mimic suppression of injury responses previously observed clinically using other antibiotics that produce similar bactericidal effects.

Analyzing Mechanical Contributions to Bacterial Overgrowth. Finally, we used the gut-on-a-chip to analyze whether physical changes in peristalsis or villus motility contribute to intestinal pathologies, such as the small intestinal bacterial overgrowth (SIBO) (5, 6) observed in patients with ileus (8) and IBD (7). When the GFPEC bacteria were cultured on the villus epithelium under normal flow (30 μL/h), but in the absence of the physiological cyclic mechanical deformations, the number of colonized bacteria was significantly higher (P < 0.001) compared with gut chips that experienced mechanical deformations (Fig. 5A). Bacterial cell densities more than doubled within 21 h when cultured under conditions without cyclic stretching compared with gut chips that experienced physiological peristalsis-like mechanical motions, even though luminal flow was maintained constant (Fig. 5B). Thus, cessation of epithelial distortion appears to be sufficient to trigger bacterial overgrowth, and motility-induced luminal fluid flow is not the causative factor as assumed previously (7).

 

Discussion One of the critical prerequisites for mimicking the living human intestine in vitro is to establish a stable ecosystem containing physiologically differentiated intestinal epithelium, gut bacteria, and immune cells that can be cultured for many days to weeks. Here we leveraged a mechanically active gut-on-a-chip microfluidic device to develop an in vitro model of human intestinal inflammation that permits stable long-term coculture of commensal microbes of the gut microbiome with intestinal epithelial cells. The synthetic model of the human living intestine we built recapitulated the minimal set of structures and functions necessary to mimic key features of human intestinal pathophysiology during chronic inflammation and bacterial overgrowth including epithelial and vascular inflammatory processes and destruction of intestinal villi.

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