Researchers have identified a novel mutation in a patient with microvillus inclusion disease (MVID) and rapidly developed a specific mouse model to find insights into the disease process.
MVID is characterized by severe diarrhea, generally beginning within a few hours of birth. The condition is caused by inactivating mutations in the gene myosin VB (MYO5B). Affected individuals usually require lifetime use of total parenteral nutrition or small-bowel transplantation.
More than 100 MYO5B mutations have been identified in MVID patients, most of whom inherit two unique mutant alleles. This can lead to variability in phenotypes, which single-mutation animal models have been unable to mimic. Generally, patients have atrophy of microvilli on enterocytes as well as inclusion bodies within enterocytes that contain microvilli.
In the study, published in Cellular and Molecular Gastroenterology and Hepatology, researchers describe genetic sequencing of MYO5B mutations in a patient and both parents. One mutation is predicted to lead to protein truncation (c.1821delG), and the other (c.1555G>A) appeared to be inherited from the patient’s mother. The patient suffered from severe diarrhea after birth and was intermittently feeding intolerant.
The researchers conducted a range of diagnostic tests and biopsies. One particularly interesting finding was expression of the A-kinase anchoring protein 350 in the vicinity of the inclusion, wrote Andreanna Burman and her coauthors at Vanderbilt University, Nashville, Tenn. This protein takes part in a protein-mediating scaffolding pathway, suggesting that it could play a role in the development of the inclusions.
The researchers used multiplexed immunofluorescence (MxIF) staining of a biopsy of the duodenum to look for expression of a range of proteins. They found a striking decrease in enterocytes that express glucose transporter 2, implying that malabsorption might be due at least in part to enterocytes that fail to mature.
The patient also had internalization of apical nutrient transporters, which has been reported in other MVID patients. The researchers also developed a mouse model that incorporated the patient’s novel compound heterozygous genotype, which they cross-bred to produce a tamoxifen-inducible mouse model.
After the tamoxifen injection, the patient-mimicking animals exhibited a severe watery diarrhea, losing 19% of their body weight by day 4. The animals’ intestines showed a similar phenotype to that of the patient.
The apical sodium transporters sodium-glucose cotransporter 1 (SGLT1), apical sodium-dependent bile transporter, and NHE3 were internalized away from the apical membrane of the enterocytes in the patient-mimicking model animals. This structural difference may explain the limited absorption of sodium and water and the resultant watery diarrhea. The researchers also noted disruption of the actinin-4+ terminal web structure, as well as increased SGLT1 localization with lysosome-associated membrane protein 1+ lysosomes within the mouse model enterocytes. This association may indicate degradation of mislocalized proteins, the authors noted, which has also been seen in expanded RAB7+ vesicles within MVID patient tissues.
Other signs pointed to the expansion of immature cells in the upper crypt and lower villus, as well as faster shedding of enterocytes in the villus than in control animals. Scanning electron microscope images also showed immature and disorganized microvilli in the enterocytes of the patient-mimicking mice.
Taken together, “these findings are consistent with a deficit in enterocyte maturation in Myo5b(G519R) mice and in the patient with the MYO5B(G519R) mutation,” the authors wrote.
The authors suggest that their approach could be used more generally to quickly create mouse models of patient-specific monogenic congenital disorders.
The authors disclosed no conflicts of interest. The research was funded by the National Foundation of Science, National Institutes of Health, Vanderbilt Digestive Diseases Research Center Pilot and Feasibility grant, an American Physiological Society John F. Perkins, Jr. Research Career Enhancement Award, and a gift from the Christine Volpe Fund.
