If the organs in our bodies could talk, the intestines might be the ones to divulge the most hidden truths about our lifestyle and health. Along the way, their "confessions" could supply crucial information for biomedical and clinical research. Weizmann Institute of Science researchers have now given just this sort of "voice" to the intestine. In a study being published today in Cell , the scientists present a method that can simultaneously identify, through testing a stool sample, all the proteins in the intestine - including those from food, from the person's own body and from the intestinal microbiome. The method thereby makes it possible to decode the interactions among these proteins with unprecedented accuracy and resolution.
The microbiome was the starting point for the research, co-led by Drs. Rafael Valdés-Mas, Avner Leshem and Danping Zheng from Prof. Eran Elinav 's lab, in collaboration with Dr. Alon Savidor of the Nancy and Stephen Grand Israel National Center for Personalized Medicine. "We wanted to go beyond DNA sequencing, the usual method of studying the microbiome," says Elinav, of Weizmann's Systems Immunology Department. "DNA can tell us which bacteria are present in the gut and point to their potential activity. Bacterial proteins, in contrast, can directly reveal whether these bacteria are active, which activity they perform and how their function affects the human body in health and disease."
Identifying proteins is challenging because of their sheer multitude, compounded by similarities among proteins from different species. The 20,000 or so protein-making genes in the human genome, for example, give rise to millions of protein variations; identifying them using existing protein databases can be complicated and highly time-consuming. The new Weizmann Institute method addresses this challenge, in part by combining DNA sequencing and mass spectrometry to generate a smaller, patient-tailored protein map.
""The proteins in the intestines are the 'words' that will one day reveal exactly what our intestines need - and how to provide it"
The method, dubbed IPHOMED - short for Integrated Proteo-genomics of HOst, MicrobiomE and Diet - makes it possible to decode the entirety of microbiome activity by showing which proteins in a stool sample come from which bacterial strains and in what amounts. In addition, it identifies the proteins secreted by the human gut in response to signals coming from the microbiome. Taken together, proteins from these two sources generate an atlas of the body's communication with the microbiome, for example, during exposure to disease-causing bacteria or to antibiotics.
Thus, using the method, Weizmann researchers found that the human gut can secrete dozens of previously unknown antimicrobial peptides that act like natural antibiotics, killing some of the bacteria in the microbiome and ultimately shaping its makeup. This finding might help explain why the composition of each person's microbiome is unique, leading to differences in susceptibility to disease.
No more cheating on the diet
When the researchers thought they'd finished developing the method, it could identify 97 percent of the proteins in each stool sample, which was a high rate, but the failure to consistently characterize the remaining 3 percent seemed puzzling. Further research clarified that these originated neither in the microbiome nor in the body tissues: They came from food.
This revelation suggested that the Weizmann method might be able to meet a dire, long-standing need of nutritional science, that of supplying a noninvasive means of revealing the exact details of a person's diet. To address this challenge, the team created a database of proteins found in hundreds of food products and identified the ones unique to each food item. These developments made it possible to learn with unprecedented precision, from stool samples, what people had eaten. For example, when applied to samples collected from two groups of healthy volunteers, one in Germany and the other in Israel, the method identified a similar level of wheat consumption in both populations, but only the German samples had large amounts of proteins from pork; in contrast, most meat proteins in the Israeli samples came from poultry.
In one of the experiments conducted by the team, volunteers were asked to consume a changing repertoire of food items, including peanuts, on designated days. Not only did the method accurately determine when these food items were eaten, it was so sensitive that it could point to consumption of as few as five peanuts per day.
In another experiment, the researchers were able to accurately track changes in the diet of people with gastrointestinal diseases. In one case, the method correctly identified a child with newly diagnosed celiac disease who had failed to follow his prescribed gluten-free diet.
"Our method could be used to tell whether someone keeps kosher or whether a person is as strictly vegetarian as they proclaim themselves to be," Elinav says with a smile. "But on a more serious note, the traditional diet-tracking method, self-reporting, is notoriously inexact. Knowing in greater precision and detail what people eat, even when their meal is complex and made up of multiple ingredients, can help establish which of the many components of a meal are beneficial to health and which are problematic."
Advancing diagnosis and treatment of disease
To explore the use of their new method in the diagnosis and treatment of disease, the scientists applied it to stool samples of patients with inflammatory bowel disease, which is characterized by severe intestinal inflammation that is affected by diet and the microbiome. The analysis of samples from Israeli, German and American study participants enabled decoding, in great molecular detail, the altered interactions between the human gut and intestinal microbiome that drive the origins of this disease. The study led to the discovery of dozens of new proteins that could serve as potential future targets for drugs to treat this currently cureless disease. The researchers also identified human and bacterial proteins that, used together, could be developed into new biomarkers for diagnosing the type of the disease, assessing its severity and tracking its progress. These new probes promise to outperform calprotectin, the only clinically approved biomarker for inflammatory bowel disease.
In addition, using an IPHOMED analysis of the patients' diet, the researchers were able to quantify the patients' compliance with nutritional therapies for intestinal bowel disease and link the level of their adherence to such diets to improved control of inflammation. Moreover, they managed to apply their noninvasive method to detecting disease in the small intestine, the long, thin tube that in healthy people absorbs most of the proteins from food. Because the small intestine is notoriously difficult to visualize and access, this disease could not have been picked up by conventional means.
"Taken together, the proteins in the intestines are the 'words' that will one day allow us to hear exactly what our intestines are telling us and thus learn to give them exactly the help they need," Elinav says. "This ability will help researchers devise personalized nutritional and medical interventions for a wide variety of disorders, particularly those affected by the microbiome, including inflammatory, metabolic, malignant and neurodegenerative diseases."
Science Numbers
Using the most recent high-throughput methods, IPHOMED simultaneously identified up to 15,268 bacterial proteins from the microbiome, up to 528 proteins secreted by the human body and up to 1,041 proteins from food.
Dr. Leshem, who was an MD-PhD student in Elinav's lab during this study, is now a surgical resident at the Tel Aviv Sourasky Medical Center (Ichilov). Study participants also included Yotam Cohen, Dr. Lara Kern, Dr. Niv Zmora, Dr. Yiming He, Shimrit Eliyahu Miller, Tal Yosef Hevroni, Liron Richman, Barbara Raykhel, Shira Allswang, Reut Better, Fernando Slamovitz, Dr. Dragos Ciocan, Uria Mor, Dr. Mally Dori-Bachash, Shahar Molina, Dr. Jotham Suez, Dr. Suhaib K. Abdeen and Dr. Hagit Shapiro of Elinav's lab; Dr. Merav Shmueli and Prof. Yifat Merbl of Weizmann's Systems Immunology Department; Dr. Yishai Levin and Corine Katina of the Nancy and Stephen Grand Israel National Center for Personalized Medicine at the Weizmann Institute; Dr. Ghil Jona of Weizmann's Life Sciences Core Facilities Department; Prof. Alon Harmelin and Dr. Noa Stettner of Weizmann's Veterinary Resources Department; Dr. Aurelia Saftien, Dr. Nyssa Cullin, Kyanna S. Ouyang and Dr. Jens Puschhof of the Microbiome and Cancer Division, German Cancer Research Center (DKFZ), Germany; Prof. Koji Atarashi and Prof. Kenya Honda of the RIKEN Center for Integrative Medical Sciences (IMS) and Keio University, Japan; Prof. Nitsan Maharshak, Prof. Oren Shibolet and Prof. Zamir Halpern of the Tel Aviv Sourasky Medical Center (Ichilov), Israel; Prof. Dror S. Shouval and Prof. Raanan Shamir of Schneider Children's Medical Center, Israel; Dr. Michal Kori of the Kaplan Medical Center, Israel; Prof. Minhu Chen of the First Affiliated Hospital, Sun Yat-Sen University, China; and Dr. Wolfgang Lieb, Dr. Corinna Bang and Prof. Andre Franke of the University Hospital of Schleswig-Holstein, Christian-Albrechts-University, Germany.
Prof. Eran Elinav's research is supported by the Helen and Martin Kimmel Award for Innovative Investigation; the Leona M. and Harry B. Helmsley Charitable Trust; Miel de Botton; Dan Andreae; the Rising Tide Foundation; the Sagol Center for Research on the Aging Brain; and the Sagol Weizmann-MIT Bridge Program. Prof. Elinav is head of the Belle S. and Irving E. Meller Center for the Biology of Aging and the incumbent of the Sir Marc and Lady Tania Feldmann Professorial Chair of Immunology. The Vera Rosenberg Schwartz Research Fellow Chair supports a staff scientist in his lab.
