Gut Microbes: Role in Health, IBS, GI and Systemic Disease Mark Pimentel, MD Director, GI Motility Program Cedars-Sinai Medical Center 1
Lacks cellulase cannot breakdown cellulose Depends completely on microbial flora to digest cellulose 50 billion bacteria per 1mm of rumen Anerobic enviroment Byproduct: Methane Kill ALL GI microbes Kill SOME GI microbes 2
Number of Cells 100x more Bacterial cells Bacterial Cells Human Cells Types of Cells Clostridium Lactobacillus Methanobrevibacter E.coli 500 different species Enterococcus 3
Types of Cells Clostridium Lactobacillus Methanobrevibacter E.coli 500 different species Enterococcus Micro-environments Air pocket Intestinal Wall Mucus Layer Oxygen/CO 2 / Nitrogen/H 2 Liquid Phase Anaerobic Solid Phase-Stool 4
SIBO- What is it? Duodenum Small Bowel ~ 0 cfu/ml Colon 10 11 cfu/ml Cecum Jejunum Ileum 10 1 cfu/ml 10 2 cfu/ml 10 3 cfu/ml Breath Testing is abnormal in IBS Forest plot of all age-sex matched studies Type of % % Author Breath Test OR (95% CI) CI) Weight Weight Grover sucrose 2.29 (0.89, 5.87) 5.87) 18.65 18.65 Lupascu Pimentel glucose lactulose 10.89 (3.52, 33.71) 33.71) 16.82 16.82 20.67 (5.29, 80.69) 80.69) 14.68 14.68 Parodi glucose 4.30 (1.24, 14.98) 14.98) 15.71 15.71 Scarpellini Collin lactulose lactulose l 24.27 (7.35, 80.15) 80.15) 16.20 16.20 18.04 (6.55, 49.71) 49.71) 17.94 17.94 Overall (I-squared = 67.9%, p = 0.008) 9.64 (4.26, 21.82) 21.82) 100.00 100.00 NOTE: Weights are from random effects analysis.1.2.5 1 2 5 10 20 Shah, et al Dig Dis Sci, 2010 5
Is Breath Testing Accurate? Orocecal Transit= 5% in cecum 50 40 99Tc + Lactulose ppm 30 20 10 0 0 15 30 45 60 75 90 105 120 135 Issues -95% in small bowel -Time to peak fermentation -Is 0.5g of lactulose enough Yu, et al. Gut 2011 Is Breath Testing Accurate? 37ºC Hydrogen (ppm) Stool 0.5g Lactulose + PBS Winter, et al. ACG, 2012 6
Is Breath Testing Accurate? 99Tc + Lactulose Orocecal Transit= 5% in cecum 60 minutes ppm Winter, et al. ACG 2012 Small Bowel Culture in IBS ercent of Subjects P 50 45 40 35 30 25 20 15 10 5 0 P<0.05 P<0.001 43% 24% 12% 4% >10,000 coliforms >5,000 coliforms Control IBS N=165 IBS, 26 controls Posserud, et al, Gut, 2007;56:802-8. 7
Mechanism: SIBO = IBS (Culture) ercent of Subjects 70 60 50 40 30 20 27.3% 60% P=0.004 004 Non- D-IBS D-IBS P 10 0 N=77 non-d-ibs, N=35 D-IBS Pyleris, et al. DDS, 2012 Mechanism: SIBO = IBS (qpcr) DUODENAL ASPIRATES 6 P<0.05 acteria quantity Log10 B 5 4 3 2 1 P<0.05 Healthy Not-IBS IBS 0 E. coli Klebsiella N=77 non-d-ibs, N=35 D-IBS Pyleris, et al. UEGW, 2012 8
Risk of PI-IBS Increases 7-fold After Infectious Gastroenteritis* Protective Effect Study (year/bacteria) Ji (2005/Shigella) Mearin (2005/Salmonella) Wang (2004/Unspecified) Okhuysen (2004/Unspecified) Cumberland (2003/Unspecified) llnyckyj (2003/Unspecified) Parry (2003/Bacterial NOS) Rodriguez (1999/Bacterial NOS) Pooled estimate t Increased Risk 0.1 0.5 1 10 50 OR OR (95% Cl) 2.8 (1.0-7.5) 8.7 (3.3-22.6) 10.7 (2.5-45.6) 10.1 (0.6-181.4) 6.6 (2.0-22.3) 2.7 (0.2-30.2) 9.9 (3.2-30.0) 11.3 (6.3-20.1) 73(4811 7.3 (4.8-11.1) 1) 9.8% IBS in cases vs 1.2% IBS in controls *Systematic review of 8 studies involving 588,061 subjects; follow-up ranged from 3 to 12 months. Halvorsen HA et al. Am J Gastroenterol. 2006;101:1894-1899. Characteristics of Acute Illness Identify Patients at Risk for PI-IBS 10 P=.000 OR for IBS After Acute Gastroenteritis* 8 6 4 2 P=.029 P=.006 P=.013 P=.001 P=.000 0-2 Age Female Diarrhea >7 d Bloody Stools Abdominal Cramps Weight Loss >10 lbs *Identified from multiple logistic regression analysis from 2069 participating in the Walkerton Health Study. Marshall JK et al. Gastroenterology. 2006;131:445-450. 9
Salmonella and PI-IBS Percent with Relative Risk IBS at 1 year (CI) Irritable Bowel 12% 5.2 (2.7-9.8) Non-ulcer dyspepsia 15% 7.8 (3.1-19.7) Nausea was a risk factor for NUD Mearin, et al. Gastro, 2006 NORMAL ACUTE GASTROENTERITIS 90% 10% COMPLETE ~GENETIC SUSCEPTIBILIITY RECOVERY ~ABNORMAL HOST RESPONSE ~TOXIN INTENSITY FUNCTIONAL POST-INFECTIOUS GI DISEASES? IBS 10
Post-infectious IBS = IBS Shah, et al J Neurogastroenterol Motil, 2012 Mechanism: Rat Model (SIBO = IBS) n=33 n=33 C. jejuni Stool = Campy- No Acute Gastroenteritis Stool = Campy+ Acute Gastroenteritis 3 Months After Recovery Stool Consistency Evaluation Stool = Campy- Recovery Bacterial Quantitation by RT-PCR of Duodenum, Jejunum, Ileum Pimentel, et al. Dig Dis Sci, 2008 11
Mechanism: Rat Model (SIBO = IBS) Percent of rats with SI IBO 50 45 40 35 30 25 20 15 10 5 0 27% 21% 17% 6.7% Duodenum Jejunum ileum Total 3 months after Campylobacter jejuni 81-176 infection Pimentel, et al., Dig Dis Sci, 2008 ICC and CDT toxin C. jejuni E. Coli Salmonella Shigella Cytolethal Distending Toxin (CDT) All cause IBS A B Cell G2 Phase Unknown Entry Arrest 12
IBS Mechanism/CdtB causes IBS Campy CDT- P-value Rifaximin P-value Stool % wet weight 60.1±6.8 60.8±3.6 0.47 61.1±3.8 0.33 Consistency 1.51±0.37 1.23±0.27 <0.00001 1.15±0.30 <0.0000001 Standard Deviation 8.4±6.4 4.2±2.4 <0.0001 4.1±2.3 <0.0001 Proportion with normal bowel form all 3 days 17.8% 50.0% <0.01 OR=4.63 59.3% <0.00001 OR=6.7 Note: No significant differences were seen between CDT- and Rifaximin treated arms of the study. Morales, et al. J Neurogastroenterl Motil, 2012 Pimentel, et al. Dig Dis Sci, 2011 IBS Mechanism/Nerve Damage CONTROLS 20x Jee, et al. World J Gastroenterol, 2010 13
IBS Mechanism/Nerve Damage Campy with SIBO 20x Jee, et al. World J Gastroenterol, 2010 IBS Mechanism/Nerve Damage Campy no CdtB 20x Jee, et al. World J Gastroenterol, 2010 14
IBS Mechanism/Autoimmune to Nerves RAT MODEL Pre-immune serum (CONTROL) wab (ANTI- CDTB) Rat Ganglia Rat ICC ckit + wab wab Anti-CdtB is Anti-Glial Anti-CdtB is anti-icc 15
IBS Mechanism/Nerve Damage GANGLIA OF HUMAN ILEAL SECTIONS wab S-100 S-100 + wab Anti-CdtB is also Anti-glial in humans IBS Mechanism/Sequence/Main Food Poisoning Bacterial Toxin Autoimmunity Gut Nerve Damage Bacterial Overgrowth IBS 16
Primary Outcome (4 weeks after Tx) Efficacy Outcome Study Primary SGA-IBS Weekly Odds Ratio 1.53 1.45 1.49 (95% CI) p-value (1.10,2.12) 0.0125 (1.05,2.01) 0.0263 (1.18,1.88) 0.0008 Key Secondary IBS Bloating Weekly 1.62 1.49 1.56 (1.16,2.27) (1.08,2.06) (1.23,1.96) 0.0045 0.0167 0.0002 Other Secondary SGA-IBS Daily IBS Bloating Daily IBS Ab Pain Daily 1.76 1.59 1.61 1.41 1.76 1.52 1.45 1.46 1.42 (1.26,2.47) 0.0009 (1.13,2.24) 0.0072 (1.28,2.04) <0.0001 (1.01,1.97) 0.0486 (1.26,2.44) 0.0008 (1.21,1.92) 0.0004 (1.05,2.02) 0.0255 (1.05,2.03) 0.0232 (1.13,1.78) 0.0028 FDA Proposed Ab Pain & Stool Daily (FDA) Ab Pain Daily (FDA) Stool Consist. Daily (FDA) 1.40 1.55 1.47 1.48 1.46 1.46 1.80 1.57 1.67 (1.02,1.92) 0.0401 (1.12,2.13) 0.0077 (1.17,1.84) 0.0009 (1.08,2.03) 0.0157 (1.06,2.00) 0.0194 (1.17,1.83) 0.0009 (1.25,2.59) 0.0015 (1.12,2.21) 0.0096 (1.31,2.14) <0.0001 Pimentel, et al NEJM, 2011 0.0 0.5 1.0 1.5 2.0 2.5 Odds Ratio and 95% CI Favors Placebo Favors Rifaximin Durability of Response (3 months) Efficacy Odds Study (95% CI) p-value Outcome Ratio 1.35 (1.00, 1.82) 0.0477 Primary SGA-IBS 1.52 (1.13, 2.03) 0.0053 Weekly 1.44 (1.17, 1.77) 0.0007 Key Secondary IBS Bloating Weekly 1.28 156 1.56 1.42 (0.95, 1.73) (1.16, 16 2.09) (1.15, 1.75) 0.1042 0.0031 0031 0.0011 Other Secondary SGA-IBS Daily IBS Bloating Daily IBS Ab Pain Daily 1.60 1.47 1.48 1.50 1.67 1.53 1.35 1.35 1.31 (1.18, 2.18) (1.09, 1.99) (1.20, 1.83) (1.10, 2.04) (1.24, 2.25) (1.24, 1.89) (1.00, 1.83) (1.01, 1.81) (1.06, 1.61) 0.0025 0.0127 0.0003 0.0103 0.0008 <0.0001 0.0495 0.0435 0.0118 FDA Proposed Ab Pain & Stool Daily (FDA) Ab Pain Daily (FDA) Stool Consist. Daily (FDA) 1.36 1.44 1.40 1.31 1.37 1.33 1.70 1.48 1.58 (1.01, 1.83) (1.08, 1.92) (1.14, 1.72) (0.98, 1.75) (1.03, 1.83) (1.09, 1.64) (1.24, 2.33) (1.09, 2.00) (1.27, 1.97) 0.0396 0.0141 0.0014 0.0725 0.0298 0.0058 0.0009 0.0114 <0.0001 0.0 0.5 1.0 1.5 2.0 2.5 Odds Ratio and 95% CI Pimentel, et al NEJM, 2011 Favors Placebo Favors Rifaximin 17
Antibiotic Resistance Conventional Antibiotics Recurrence SIBO 40% Eradicated 25% Eradicated Rifaximin Recurrence 70% Eradicated 100% Eradicated Yang, et al. Dig Dis Sci, 2007. Harm with Drugs in IBS DRUG NNT NNH Benefit to Harm Desipramine 8 18.3 2.3 Alosetron 7.5 19.4 2.6 Rifaximini i 10 8971 897.1 NNT=Number needed to treat. NNH=Number needed to harm. NNH=number of subjects treated for one subject to withdraw due to an adverse event that was greater than the equivalent in placebo. Shah, et al. Am J Med, 2012. 18
Harm with Drugs in IBS Drug Desipramine Alosetron Rifaximin Excess of Placebo Only Dry mouth, flushing, constipation, insomnia, decreased appetite, palpitations Constipation, Abdominal pain/discomfort Bad Taste in mouth Shah, et al. Am J Med, in press. Compared to other Drugs +12.5% +13% +10% -16% -17% Net value -3.5% -4% +10% Shah, et al. DDW, 2012 19
Retreatment with Rifaximin Percent of subjects who respo onded 100 90 80 70 60 50 40 30 20 10 0 38/40 17/18 54/65 6/7 3/4 1st RTX 2nd RTX 3rd RTX 4th RTX 5th RTX Pimentel, et al. Dig Dis Sci, 2011. Retreatment with Rifaximin before an number of months relapse Media 12 10 8 6 4 2 0 N=63 Range=0.5-45 mo N=46 Range=1-45 months N=21 Range=2-37 mo N=6 Range=1-18 mo N=4 Range=2-24 mo 1st RTX 2nd RTX 3rd RTX 4th RTX 5th RTX Note: RTX is the retreatment number. In this figure, the time (in months) refers to the number of months of wellness before the treatment number in each column. For example, for 1 RTX, it would be the number of months the patient was well after the first rifaximin administration and prior to the 1 RTX. Note: Rare patients were missing accurate dates and thus were not included here. Note: The median is a more accurate depiction of the duration of benefit since the ranges were wide. Pimentel, et al. Dig Dis Sci, 2011. 20
Complexities of Gas Production Methane Hydrogen 70 Hydrogen Producers H 2 M illio n P a r t s P e r M 60 50 40 20 Methane 10 Producers 0 4H 2 1CH 4 30 0 15 30 45 60 75 90 105 120 135 150 165 180 Time (Minutes) 70 on Parts Per Milli 60 50 40 30 20 10 0 0 15 30 45 60 75 90 105 120 135 150 165 180 Time (Minutes) H 2 H 2 S Producers 5H 2 1H 2 S P a rts P e r M illio n 70 60 50 40 30 20 10 0 0 15 30 45 60 75 90 105 120 135 150 165 180 Time (Minutes) Methane- Important in C-IBS Meta-analysis of studies Author Year OR (95% (95% CI) CI) % % Weight Weight Peled 1987 Fiedorek 1990 Pimentel 2003 Pimentel 2003 Majewski 2007 Bratten 2008 Parodi 2009 Hwang 2009 Attaluri 2009 Overall (I-squared = 64.6%, p = 0.004) 0.83 (0.20, (0.20, 3.56) 3.56) 4.32 (1.60, (1.60, 11.68) 11.68) 5.58 (2.22, (2.22, 14.03) 14.03) 44.23 (2.48, (2.48, 788.51) 788.51) 1.81 (0.70, (0.70, 4.67) 4.67) 2.22 (1.14, (1.14, 4.33) 4.33) 1.89 (0.79, (0.79, 4.51) 4.51) 47.67 (8.73, (8.73, 260.41) 260.41) 3.70 (2.06, (2.06, 6.66) 6.66) 3.51 (2.00, (2.00, 6.16) 6.16) 8.43 12.03 12.68 3.18 12.46 15.14 13.17 6.99 15.92 100.00 8.43 12.03 12.68 3.18 12.46 15.14 13.17 6.99 15.92 100.00 NOTE: Weights are from random effects analysis.1.2.5 1 2 5 10 20 Kunkel, et al. Dig Dis Sci, 2011. 21
Methane as a diagnostic test C-IBS Not C-IBS Methane 22 6 28 No Methane 2 26 28 24 32 Sensitivity=0.92, Specificity=0.81, Positive Predictive Value=0.79, Negative Predictive Value=0.93 Pimentel, et al Dig Dis Sci. 2010. Methanobrevibacter smithii- the cause of constipation Hydrogen Producing IBS Stool Methane Producing IBS Stool Kim, et al. DDW, 2010. 22
Methanobrevibacter smithii or methane as a test- dictates treatment Percen nt eradicating CH4 CH4 Eradication Clinical Response 100 P=0.001 001 90 P=0.01 01 90 80 80 70 60 50 40 30 20 10 0 Neomycin Rifaximin Neo+Rifax Percent with Clinical Response 70 60 50 40 30 20 10 0 Neomycin Rifaximin Neo+Rifax Low, et al. Gastroenterol and Hepatol 2010 IBS/SIBO WORKSHEET 3.0 Molecular Mimicry Acute Gastroenteritis Immune response Anti-CDT Ab Other Ab React to neural elements/myenteritis Reduced DMP-ICC Immunity to Infection STRESS PROKINETIC SMALL BOWEL COLON CRF DYSMOTILITY METHANE SIBO/ IBS HYDROGEN SLOW TRANSIT C-IBS ANTIBIOTIC D- AND M-IBS 23
Lessons from the Germ Free Mouse Reduced organ weight Reduced cardiac output Reduced O2 consumption Increased food intake Reduced mediastinal and sympathetic lymph nodes Reduced mucosal associated lymphoid tissue Reduced serum immunoglobulins Tannaock, Am J Clin Nutrition 2001 Defense of Host by detecting and sampling Nutrients for Host Information for Host immune development IMMUNE REGULATION GUT FLORA DEFENSE AGAINST PATHOGENS NUTRITION/ METABOLISM 24
Specific Host Microbe Interaction Modulate Immune Response Modulate other Bacteria Impact Appetite Regulation Digest the Indigestible Vitamin Production and Metabolism Produce Peptide Mediators Gut Microbes Generate Heat and change BMR Gas Production and Effects Inflammatory Response Germ Free Animals Germ Free -Low body weight -Increased oral intake + = Add Fecal Flora Gain 60% more body weight with reduced oral intake Backhed F Proc Natl Acad Sci U S A. 2004 Nov 2 101(44) 15718 23 25
Fecal Transplant Germ Free -Low body weight -Increased oral intake + = Add Fecal Flora (Bacteroidetes>Firmicutes) Gain weight with reduced oral intake Germ Free -Low body weight -Increased oral intake + = Add Fecal Flora from Obese mice (Firmicutes>Bacteroidetes) Far greater weight gain Turnbaugh Nature 444, 1027-1031 (21 December 2006) Energy lost in stool Bomb calorimetry of the fecal gross energy content (kcal g -1 )of lean (+/+) and obese (ob/ob) conventionally raised ( *P<0.05, ***P<0.001) In addition ob/ob cecal microbiota = increased Methanogenic archaea Turnbaugh Nature 444, 1027-1031 (21 December 2006) 26
Methanogen Model No Methanobrevibacter smithii Indigestible Substrate H 2 Intoxicates self Methanobrevibacter smithii Indigestible Substrate H 2 H 2 H 2 H 2 CH 4 Energy Digestible Substrates ENERGY DIGESTIBLE SUBSTRATE = Syntroph = Methanogen Body Mass Index and PCOS Mathur R et al Digestive Disease Sciences. 55(4):1085, 201 27
Percent Body Fat and PCOS Mathur R, et al Digestive Disease Sciences. 55(4):1085, 2010 Methane in Obese Patients Mathur et al, Gastro and Hep, 2012 28
Methane in Non-obese Patients BMI 40 38 36 34 32 30 28 26 24 22 20 Multivariate analysis controlling for age, sex, diabetes, anti depressant and other confounding (P<0.001) Normal Breath test N=792 Positive Hydrogen Breath Test Only Methane Methane and Hydrogen Positive Mathur R, et al. JCEM in press 2013 Methane and Glucose Metabolism Glucose AUC (mg/dl) 800 n=5 750 700 650 600 550 500 No methane P=0.03 Methane 29
M smithii and Body Weight in New Animal Model Mathur R, et al. Obesity 2013 Conclusions The basis for IBS may be acute gastroenteritis and SIBO Antimicrobial therapies benefit IBS subjects Autoimmunity may play a role in IBS and gut microbe changes Bacteria are important in obesity Methanogens have a significant future role in constipation and metabolic syndrome 30