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A new height in the study of intestinal bacteria: Harvard University develops human intestinal microbial organ chip technology

via:博客园     time:2019/5/15 12:31:40     readed:427


The microbiome is a collection of large numbers of microorganisms in our body and on the body, which have a profound impact on human health and disease. In particular, the human intestinal flora, which contains the most dense microbes, not only breaks down the release of nutrients, but also is a key factor in the development of many diseases, including infection, inflammatory bowel disease, cancer, metabolism. Sexual diseases, autoimmune diseases and neuropsychiatric diseases.

Most of the human microbiota interactions we know are based on disease states and related studies of bacterial DNA in stool samples, using genomic or metagenome analysis. This is because studying the interaction between microbiota and human intestinal tissue in vitro is a formidable challenge, in large part because symbiotic bacteria kill the human body in a petri dish in a single day due to overgrowth. cell. In addition, many of the commensal microorganisms in the gut are anaerobic, so they require very low oxygen conditions to grow, and this condition can damage human cells.

A research team at Harvard University's Wyss Bio-Inspired Engineering Institute, led by Donald Ingber, founding director of the institute, developed a solution using the “organic-on-a-chip” microfluidic culture technology. The method of this problem. His team is now able to culture stable human microbiota in human intestinal microarrays, which are in direct contact with vascularized human intestinal epithelium for at least 5 days, and they establish an oxygen gradient that maintains intestinal hypoxic conditions. It also provides high levels of oxygen to the endothelium and epithelium that are infected by commensal bacteria. “The anaerobic bowel chip” maintains a stable microbial diversity similar to human feces and a protective physiological barrier formed by human intestinal tissue within a few days. The study was published in Nature Biomedical Engineering.

“The main paradigm shift in the medical world over the past decade has been to recognize the enormous role that microbiota plays in health and disease. This new anaerobic intestinal microarray technology provides a way to study the interaction of clinically relevant human host microbiota at the cellular and molecular levels under in vitro controlled conditions," said Dr. Ingber, who is also a Harvard Medical School. HMS) Professor of Vascular Biology. <RTIgt; By direct contact with microbial populations and differentiated gut tissue, this method can be used to discover specific microorganisms or their metabolites that can cause disease or help prevent these diseases, and because we use patient separation Cells, this method can also be used to personalize drugs. ”

“The early tissue culture system was designed to reproduce the interaction between the human microbiota and the intestinal epithelial cells in vitro, but because they cannot culture these two components in direct contact, they cannot mimic anaerobic bacteria in the intestine. Survival is essential for low oxygen concentrations, so their effectiveness is limited. “The first author, Sasan Jalili-Firoozinezhad, said he is a graduate student at the Ingber team at the Wyss Institute and a Ph.D. student at Professor Joaquim Cabral at the University of Lisbon, Portugal. “More complicated things are: moving along the small intestine toward the colon, and the oxygen content is declining, which also changes the composition of the local microbiota. ”

The anaerobic intestinal chip contains two intestine chips of parallel microchannels separated by a porous membrane. Human intestinal epithelial cells are grown on top of the upper digestive tract membrane, and vascular endothelial cells are grown on the opposite side of the lower digestive tract membrane. The intestinal cells used to line these intestinal chips are either from the cell line or from a human ileal biopsy and amplified by an intermediate organic step. In this step, they form a tiny spherical intestinal structure that is cultured in a chip. These structures were previously broken down into pieces.

To accommodate the complete microbiota, the team put the gut chips into a custom anaerobic chamber, which greatly reduced the oxygen concentration in the upper intestinal epithelial cell channels while maintaining the normal oxygen concentration in the lower endothelial cell channels.

“We created oxygen gradients in both channels that allow the intestinal epithelium to diffuse through the porous membrane to obtain oxygen support,” said co-author Elizabeth Calamari, who designed the device with co-author Dr. Richard Novak. “We installed an optical sensor on the gut chip to report local oxygen concentrations in both channels in real time without disturbing the oxygen gradient. ”

Samples of complex gut microbiota from healthy human feces, stably cultured in gnotobiotic mice, or freshly isolated from infant feces, then injected into the upper epithelial passage where they are in direct contact with the lower intestinal gland The mucus layer naturally secreted by epithelial cells. More importantly, the diversity of the commensal flora growing under hypoxic conditions maintains the abundance observed in the human gut. “We have found through genomic analysis that we can culture more than 200 different bacterial populations for several days, and the ratio of abundance and obligate anaerobic bacteria is similar to that observed in human feces. Importantly, the intact microbiota further enhances the barrier function of the intestinal epithelium, which provides a tight seal and produces a protective mucus layer, which is an important prerequisite for intestinal health. ”

The ability to observe the composition and changes of intact human microbiota in direct contact with human intestinal tissue in vitro for several days provides an opportunity for personalized drug and drug testing. “We can train intestinal tissue and microbiota in specific areas from the same person to discover associations that cause sensitivity or tolerance to specific pathogenic, inflammatory, and systemic diseases,” co-first author Dr. Francesca Gazzaniga said he is a postdoctoral fellow at the Ingber group and co-author Professor Dennis Kasper. “Using anaerobic bowel chips, we can also test the direct effects of drugs on the human microbiota before giving them to people. ”

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