The Underappreciated Majority

MUSC researchers aim to link the microbiome to health and disease

By Matthew Greseth

Our darkest fiction is full of Orwellian dystopias, shadowy cabals, and mind-controlling supervillains. But it turns out that the brainless, microscopic, single-celled organisms that live inside us have been pulling on our strings all along.

- Ed Yong, I contain Multitudes: The Microbes Within Us and a Grander View of Life

 Microbes were once seen exclusively as foreign invaders and antibiotics as our principal weapon against them. In truth, we mostly live in harmony with our microbiome — the totality of bacteria, fungi and viruses that colonize virtually every niche in the human body. Indeed, bacterial cells outnumber our own cells by a factor of ten.1 These commensal bacteria do not cause disease and often serve vital biological functions. In fact, when the balance of microbes is off, we become more susceptible to disease, suggesting that rebalancing the microbiome could have therapeutic benefit.

The past 15 years have seen an explosion of microbiome research. This was in part due to clinical successes of microbiome-related interventions, such as fecal transplants for controlling Clostridium difficile, and in part to advances in techniques for culturing and sequencing microbes and in interpreting the large datasets that result.¹

To ensure its clinicians and researchers can help create new microbiome-based therapies, MUSC is ramping up its microbiome infrastructure, providing support in sequencing the microbiota and in analyzing the resulting datasets.

Building a microbiome infrastructure at MUSC

Studying the microbiome presents a major obstacle — how do you identify and classify the plethora of individual bacterial species that constitute the microbiome? The first step is to purify the DNA. The South Carolina Clinical and Translational Research (SCTR) Institute Nexus Research Laboratory, led by manager Amy Gandy, MS, provides processing of patient samples to extract the DNA, which contains human and microorganismal DNA. Following isolation, the Center for Genomic Medicine specifically amplifies the microorganismal DNA and then identifies the organisms present through sequencing. These data enable researchers, often with the help of bioinformaticists who specialize in analyzing large datasets, to compare microbiomes and identify differences for further investigation.

“At some point, we should be able to manipulate the microbiome community - then we start to change how we practice medicine.”

-Zhihai Li, M.D., Ph.D.

One current study is headed by data scientist and microbiome researcher Alexander V. Alekseyenko, Ph.D., associate professor in the Biomedical Informatics Center and founding director of the MUSC Program for Human Microbiome Research, who received a $100,000 grant from SCTR to enhance infrastructure for microbiome research. Although SCTR has performed microbiome testing for some time, this grant allows the purchase of state-of-the-art equipment. Now, SCTR will be able to extract and quantify DNA in a more standardized, automated fashion, providing greater availability and turnaround times.

In addition, MUSC supports basic microbiome research through a gnotobiotic animal core, directed by Caroline Westwater, Ph.D., professor in the College of Dental Medicine. Animals in this facility, one of only a handful in the country, are germ-free (no microbiota) or gnotobiotic (defined microbiota). This facility enables researchers to define what microorganisms are present (or absent) in order to identify the causative agent of disease.

Understanding the role of the microbiome in health and disease

MUSC researchers are already tapping into these resources to explore the role of the microbiome in human health and disease in areas as diverse as infections, lupus, skeletal health and cancer.

Clostridium difficile

C. difficile infections are a major complication for cancer and transplant patients, who are often given antibiotic regimens that destroy the normal microbiota and allow C. difficile to overpopulate. Antibiotics have had limited success in treating C. difficile, and antibiotic resistance is a major problem for a number of hospital-acquired infections. Fecal transplants provide a promising alternative. In C. difficile patients, transplantation of fecal matter from a healthy gut allows for the re-establishment of a healthy microbiome and has a 90 percent cure rate, compared with 20 percent using antibiotics.2 MUSC offers fecal transplants to infected patients under the supervision of Scott R. Curry, M.D., assistant professor in the College of Medicine, who is also tasked with C. difficile surveillance. By tagging each case with a zip code, Curry tracks the movement of each case so that appropriate cleaning measures can be taken.

Candida albicans

Candida albicans is a commensal fungus found on the skin and mucous membranes of 40 to 60 percent of healthy adults. C. albicans becomes infectious when there is a shift in the make-up of the microbiome, known as dysbiosis. The research of Caroline Westwater, Ph.D., aims to better understand the mechanism of this shift.

“I look at the interface between the host and the microbe. There is a battle between those two things all the time. For most of us, that battle reaches a stalemate. But in some individuals it shifts and the outcome is disease,” explains Westwater.

By focusing on host-microbial interactions, the Westwater lab has identified one possible mechanism that protects the host from disease — the host synthesizes a novel peptide that exhibits antifungal properties. Westwater is trying to better understand how the peptide controls C. albicans and whether other molecules mimicking this peptide could be even more effective.

Lupus

Lupus is a systemic autoimmune disease caused by the production of autoantibodies that attack the self, resulting in inflammation of vital organs. Researchers have identified over 50 genetic risk factors for lupus; however, these changes explain the cause of only 20 to 30 percent of lupus cases. This led Diane L. Kamen, M.D., M.S.C.R., associate professor of Medicine in the Division of Rheumatology, to examine the potential contributions of environmental factors to lupus.

Over the past eight years, Kamen’s group has worked with lupus patients to analyze how diet may influence the development of lupus among people at risk, particularly African American women. This large observational study explores whether a Western diet influences the risk of lupus and the severity of lupus. Kamen’s research team has taken this one step further to determine if there are changes in the microbiome that cause lupus risk and severity.

To address this question, Kamen’s group is collecting blood and stool samples from patients with lupus and controls without lupus every two years. When combined with the already collected dietary data, this information provides a large, complex dataset to determine if there is a correlation between the microbiome and the production of autoantibodies, which appear before disease symptoms. Associations between the microbiome and autoantibody production could be a predictor, and potentially a treatment, for lupus.

“That’s the thing we’re really hoping: if we can find a connection between the microbiome and the development of autoimmune disease, then we can develop safe methods to prevent autoimmune diseases like lupus,” says Kamen.

Skeletal health

The young adult skeleton begins a state of slow, continuous deterioration around age 30, which is currently unexplained. The field of osteoimmunology, focused on the interface of the skeletal and immune systems, was recently expanded to include the microbiome.

“The gut microbiota is a critical regulator of the systemic immune response that has profound effects on bone remodeling in the young adult skeleton,” says Chad M. Novince, D.D.S., Ph.D., assistant professor in the Colleges of Dental Medicine and Medicine who studies the impact of the microbiome on skeletal metabolism.

In a study published in Scientific Reports in 2017 (doi:10.1038/ s41598-017-06126-x), Novince and colleagues examined the contribution of the microbiome to skeletal remodeling and showed, for the first time, a signaling pathway that links the gut, liver and skeleton.3 With the gnotobiotic animal core, Novince’s team monitored young adult specific-pathogen-free and germ-free mice to discern the immunomodulatory effects of the commensal gut microbiota on physiologic bone remodeling. The healthy gut microbiota was found to produce metabolites that travel to the liver, where they have profound effects on bone maintenance. Consequently, the healthy gut microbiota plays a heretofore underappreciated role in bone remodeling, inducing bone loss. These data advance our understand- ing of skeletal physiology and have significant implications for the prevention of skeletal deterioration in health and disease.

Cancer

As we age, we may enter a chronic proinflammatory state that dampens immune activation and may allow tumor cells to proliferate. One theory suggests that this is caused by an age-related shift in the microbiome. Several researchers at MUSC, including Zihai Li, M.D., Ph.D., professor and chair of the Department of Microbiology and Immunology and co-leader of the Cancer Immunology Program at the Hollings Cancer Center, and Bei Liu, M.D., M.P.H., associate professor in the Department of Microbiology and Immunology, are studying the interplay between the immune system and the microbiome in colitis and colon cancer.

The protein CNPY2 activates the endoplasmic reticulum (ER) stress pathway and is highly expressed in the gut. Li’s group showed that mice lacking CNPY2 exhibited a reduction in ER stress activa- tion and protection from inflammation-induced colitis. Further examination of this pathway, and its impact on the gut microbiome, may facilitate the development of CNPY2 inhibitors that provide a novel treatment for colitis and possibly colon cancer.

Liu’s work is part of the MIST (Mucosal Immunology Studies Team) initiative, a very prestigious nationwide program focused on identifying the most basic aspects of immunity. Liu’s group showed that mice lacking the chaperone protein grp94, specifically in dendritic cells, spontaneously develop colitis and colon cancer due to dysbiosis of the gut flora. They found the prevalence of the com- mensal bacterium, Akkermansia muciniphila, expanded to constitute up to 50 percent of the microbiome. Working with the gnotobiotic animal core, they will define A. muciniphila’s direct contribution to colitis and colon cancer. In the future, Liu will expand these studies to humans and determine if specific commensal bacteria could serve as predictive markers for colon cancer.

Looking forward

Currently, research is focused on establishing associations between the microbiome and various diseases. As the field progresses, research will need to move beyond correlative studies to developing microbiome-based therapeutic innovations.

“Right now we are still doing this survey kind of work, trying to describe this kind of correlation,” says Li. “At some point, we should be able to manipulate the microbiome community – then we start to change how we practice medicine.”

Moving forward, MUSC is positioned to lead that change. Clinicians and researchers have already begun to define the role of the microbiome in both health and disease and are ready to move towards treating disease through modulation of the microbiome.

References

1 Cani, PD. Gut. 2018;0:1-10.
2 Murphy, B., et al. medpace.com
3 Novince, CM., et al. Sci Rep. 2017. 18;7(1):5747.