Microvillus inclusion disease (MVID) is a congenital diarrheal disorder, caused by inactivating mutations in myosin Vb (MYO5B). MYO5B-deficient mice and cell lines have demonstrated the importance of MYO5B in brush border development; however, the previous models lacked specificity to test intestinal stem cell functions. In the present study, we investigated the effects of progressive MYO5B deficiency originating in intestinal crypt cells utilizing mouse models. Our transcriptomic and multiplex immunostaining datasets demonstrate that MYO5B is critical for intestinal stem cell function. MYO5B-deficient crypts acquire a hyperproliferative phenotype with incomplete cell differentiation in vivo and an elevated organoid formation rate compared to control crypts. An evident disruption in mitochondrial structure and fatty acid metabolism likely underlies these crypt phenotypes. Consistent with mouse models, MVID patient biopsies demonstrate abnormal expansion of the proliferative zone along with villus blunting. These data reveal the direct role of MYO5B in intestinal epithelial progenitor cell functions.

BACKGROUND:Increasingly long and complex informed consents have yielded studies demonstrating comparatively low participant understanding and satisfaction with traditional face-to-face approaches. In parallel, interest in electronic consents for clinical and research genomics has steadily increased, yet limited data are available for trio-based genomic discovery studies. We describe the design, development, implementation, and validation of an electronic iConsent application for trio-based genomic research deployed to support genomic studies of cerebral palsy. METHODS:iConsent development incorporated stakeholder perspectives including researchers, patient advocates, institutional review board members, and genomic data-sharing considerations. The iConsent platform integrated principles derived from prior electronic consenting research and elements of multimedia learning theory. Participant understanding was assessed in an interactive teachback format. The iConsent application achieved nine of ten proposed desiderata for effective patient-focused electronic consenting for genomic research. RESULTS:Overall, participants demonstrated high understanding and retention of key human subjects considerations. Enrollees reported high levels of satisfaction with the iConsent, and we found that participant understanding, iConsent clarity, privacy protections, and study goal explanations were associated with overall satisfaction. CONCLUSIONS:Although opportunities exist to optimize iConsent, we show that such an approach is feasible, can satisfy multiple stakeholder requirements, and can realize high participant satisfaction and understanding while increasing study reach.

BACKGROUND & AIMS: The xenobiotic efflux pump P-glycoprotein is highly expressed on the apical membrane of the gastrointestinal tract, where it regulates the levels of intracellular substrates. P-glycoprote in is altered in disease, but the mechanisms that regulate the levels of P-glycoprotein are still being explored. The molecular motor myosin Vb (Myo5b) traffics diverse cargo to the apical membrane of intestinal epithelial cells. We hypothesized that Myo5b was responsible for the delivery of P-glycoprotein to the apical membrane of enterocytes. METHODS: We used multiple murine models that lack functional Myo5b or the myosin binding partner Rab11a to analyze P-glycoprotein localization. Pig and human tissue were analyzed to determine P-glycoprotein localization in the setting of MYO5B mutations. Intestinal organoids were used to examine P-glycoprotein trafficking and to assay P-glycoprotein function when MYO5 is inhibited. RESULTS: In mice lacking Myo5b or the binding partner Rab11a, P-glycoprotein was improperly trafficked and had decreased presence in the brush border of enterocytes. Immunostaining of a pig model lacking functional Myo5b and human biopsies from a patient with an inactivating mutation in Myo5b also showed altered localization of intestinal P-glycoprotein. Human intestinal organoids expressing the motorless MYO5B tail domain had colocalization with P-glycoprotein, confirming that P-glycoprotein was trafficked by MYO5B in human enterocytes. Inhibition of MYO5 in human intestinal cell lines and organoids resulted in decreased P-glycoprotein capacity. Additionally, inhibition of MYO5 in human colon cancer cells diminished P-glycoprotein activity and increased cell death in response to a chemotherapeutic drug. CONCLUSIONS: Collectively, these data demonstrate that Myo5b is necessary for the apical delivery of P-glycoprotein.

Microvillus Inclusion Disease (MVID), caused by loss-of-function mutations in the motor protein Myosin Vb (MYO5B), is a severe infantile disease characterized by diarrhea, malabsorption, and acid-base instability, requiring intensive parenteral support for nutritional and fluid management. Human patient-derived enteroids represent a model for investigation of monogenic epithelial disorders but are a rare resource from MVID patients. We developed human enteroids with different loss-of function MYO5B variants and showed that they recapitulated the structural changes found in native MVID enterocytes. Multiplex Immunofluorescence imaging of patient duodenal tissues revealed patient-specific changes in localization of brush border transporters. Functional analysis of electrolyte transport revealed profound loss of Na + /H + exchange (NHE) activity in MVID patient enteroids with near-normal chloride secretion. The chloride channel-blocking anti-diarrheal drug, Crofelemer, dose-dependently inhibited agonist-mediated fluid secretion. MVID enteroids exhibited altered differentiation and maturation versus healthy enteroids. Inhibition of Notch signaling with the γ-secretase inhibitor, DAPT, recovered apical brush border structure and functional Na + /H + exchange activity in MVID enteroids. Transcriptomic analysis revealed potential pathways involved in the rescue of MVID cells including serum- and glucocorticoid-induced protein kinase 2 (SGK2), and NHE regulatory factor 3 (NHERF3). These results demonstrate the utility of patient-derived enteroids for developing therapeutic approaches to MVID. Conflict-of-interest statement:The authors have declared that no conflict of interest exists.

Abstract Background The application of advanced imaging in pediatric research trials introduces the challenge of how to effectively handle and communicate incidental and reportable findings. This challenge is amplified in underserved populations that experience disparities in access to healthcare as recommendations for follow-up care may be difficult to coordinate. Therefore, the purpose of the present report is to describe the process for identifying and communicating findings from a research MRI to low-income Latino children and families. Methods Latino adolescents (n = 86) aged 12–16 years old with obesity and prediabetes underwent a research MRI (3 Tesla Philips Ingenia®) as part of a randomized controlled diabetes prevention trial. The research MRIs were performed at baseline and 6 months to assess changes in whole-abdominal fat distribution and organ fat in response to the intervention. An institutional pathway was developed for identifying and reporting findings to participants and families. The pathway was developed through a collaborative process with hospital administration, research compliance, radiology, and the research team. All research images were reviewed by a board-certified pediatric radiologist who conveyed findings to the study pediatrician for determination of clinical actionability and reportability to children and families. Pediatric sub-specialists were consulted as necessary and a primary care practitioner (PCP) from a free community health clinic agreed to receive referrals for uninsured participants. Results A total of 139 images (86 pre- and 53 post-intervention) were reviewed with 31 findings identified and 23 deemed clinically actionable and reportable. The only reportable finding was severely elevated liver fat (> 10%, n = 14) with the most common and concerning incidental findings being horseshoe kidney (n = 1) and lung lesion (n = 1). The remainder (n = 7) were less serious. Of youth with a reportable or incidental finding, 18 had a PCP but only 7 scheduled a follow-up appointment. Seven participants without a PCP were referred to a safety-net clinic for follow-up. Conclusions With the increased utilization of high-resolution imaging in pediatric research, additional standardization is needed on what, when, and how to return incidental and reportable findings to participants, particularly among historically underrepresented populations that may be underserved in the community. Trial registration Preventing Diabetes in Latino Youth, NCT02615353

BACKGROUND & AIMS: Microvillus inclusion disease (MVID) is caused by inactivating mutations in the myosin VB gene (MY05B). MVID is a complex disorder characterized by chronic, watery, life-threatening diarrhea that usually begins in the first hours to days of life. We developed a large animal model of MVID to better understand its pathophysiology. METHODS: Pigs were cloned by transfer of chromatin from swine primary fetal fibroblasts, which were edited with TALENs and single-strand oligonucleotide to introduce a P663-L663 substitution in the endogenous swine MY05B (corresponding to the P660L mutation in human MY05B, associated with MVID) to fertilized oocytes. We analyzed duodenal tissues from patients with MVID (with the MY05B P660L mutation) and without (controls), and from pigs using immunohistochemistry. Enteroids were generated from pigs with MY05B(P663L) and without the substitution (control pigs). RESULTS: Duodenal tissues from patients with MVID lacked MY05B at the base of the apical membrane of intestinal cells; instead MY05B was intracellular. Intestinal tissues and derived enteroids from MY05B(P663L) piglets had reduced apical levels and diffuse subapical levels of sodium hydrogen exchanger 3 and SGLT1, which regulate transport of sodium, glucose, and water, compared with tissues from control piglets. However, intestinal tissues and derived enteroids from MY05B(P663L) piglets maintained CFTR on apical membranes, like tissues from control pigs. Liver tissues from MY05B(P663L) piglets had alterations in bile salt export pump, a transporter that facilitates bile flow, which is normally expressed in the bile canaliculi in the liver. CONCLUSIONS: We developed a large animal model of MVID that has many features of the human disease. Studies of this model could provide information about the functions of MY05B and MVID pathogenesis, and might lead to new treatments.

Dyslipidemia is central to the pathogenesis of many of the obesity related morbidities, which is at least in part mediated by increased lipid absorption, which in turn depends on intestinal bile acids.In the Zucker rat model of obesity, ileal bile acid absorption is known to be increased secondary to the increased cellular expression of the apical Na-dependent bile acid co-transporter (ASBT; slc10a2).ASBT is exclusively expressed in mature villus cells in the the terminal ileum.However, during obesity whether ASBT distribution may be increased caudo-orally in the small intestine and farther down the villus-crypt axis to enhance bile acid absorption and thus, dyslipidemia, is unknown.Hyothesis: In obesity, ASBT expression is increased both caudo-orally as well as along the villus-crypt axis.Aim: Determine if ASBT expression is increased along the caudo-oral length of the small intestine as well as the villus-crypt axis during obesity Methods: Obese (lepr fa ) Zucker rats (OZR) were used as a model of obesity with leans as control (LZR).Western blot and immunofluorescence studies were performed with rat specific antibodies.Results: ASBT expression was found to be significantly increased along the caudo-oral length of the small intestine in OZR compared to LZR (n=4, p<0.05).Approximately 50% more of the proximal ileum demonstrated ASBT expression in OZR compared to LZR.Transcription factor GATA4, which may be responsible for the intestine-specific induction of ASBT expression in proximal intestine, was also increased in obesity (n=3, p<0.05).Further, immunofluorescence studies demonstrated that ASBT expression along the entire villus-crypts axis was profoundly increased in OZR compared to LZR (n=4, p<0.05).Densitometric analysis showed that the highest expression of ASBT was seen at the villus tip and it gradually decreased towards the villus base, in both LZR and OZR.However, the relative expression of ASBT was found to be significantly increased throughout the entire villus-crypt axis in OZR compared to LZR.These studies demonstrated that in obesity, along with the previously known stimulation of ASBT in villus cells, ASBT expression is also increased along the caudo-oral length of the small intestine as well as along the villus-crypt axis.Conclusions: Increased bile acid absorption that occurs via ASBT during obesity occurs at three levels, enhanced expression: in villus cells, along the villus-crypt axis as well as caudo-orally.This enhanced ASBT expression at multiple levels is likely mediated by transcription factor GATA4.Thus, the dyslipidemia of obesity resulting from increased intestinal lipid assimilation that requires increased bile acids is result of increased ASBT expression at the villus cell, villus-crypt axis and caudo-oral levels of the small intestine.

BACKGROUND & AIMSInactivating mutations in MYO5B cause microvillus inclusion disease (MVID), but the physiological cause of the diarrhea associated with this disease is unclear. We investigated whether loss of MYO5B results in aberrant expression of apical enterocyte transporters.METHODSWe studied alterations in apical membrane transporters in MYO5B-knockout mice, as well as mice with tamoxifen-inducible, intestine-specific disruption of Myo5b (VilCreERT2;Myo5bflox/flox mice) or those not given tamoxifen (controls). Intestinal tissues were collected from mice and analyzed by immunostaining, immunoelectron microscopy, or cultured enteroids were derived. Functions of brush border transporters in intestinal mucosa were measured in Ussing chambers. We obtained duodenal biopsy specimens from individuals with MVID and individuals without MVID (controls) and compared transporter distribution by immunocytochemistry.RESULTSCompared to intestinal tissues from littermate controls, intestinal tissues from MYO5B-knockout mice had decreased apical localization of SLC9A3 (also called NHE3), SLC5A1 (also called SGLT1), aquaporin (AQP) 7, and sucrase isomaltase, and subapical localization of intestinal alkaline phosphatase and CDC42. However, CFTR was present on apical membranes of enterocytes from MYO5B knockout and control mice. Intestinal biopsies from patients with MVID had subapical localization of NHE3, SGLT1, and AQP7, but maintained apical CFTR. After tamoxifen administration, VilCreERT2;Myo5bflox/flox mice lost apical NHE3, SGLT1, DRA, and AQP7, similar to germline MYO5B knockout mice. Intestinal tissues from VilCreERT2;Myo5bflox/flox mice had increased CFTR in crypts and CFTR localized to the apical membranes of enterocytes. Intestinal mucosa from VilCreERT2;Myo5bflox/flox mice given tamoxifen did not have an intestinal barrier defect, based on Ussing chamber analysis, but did have decreased SGLT1 activity and increased CFTR activity.CONCLUSIONSAlthough trafficking of many apical transporters is regulated by MYO5B, trafficking of CFTR is largely independent of MYO5B. Decreased apical localization of NHE3, SGLT1, DRA, and AQP7 might be responsible for dysfunctional water absorption in enterocytes of patients with MVID. Maintenance of apical CFTR might exacerbate water loss by active secretion of chloride into the intestinal lumen.

15 [25%], 2 [8.7%], 5 [8.5%], 4 [11.1%]) and infections (22 [36.7%], 6 [26.1%], 22 [37.3%], and 10 [27.8%]).Two malignancies (Hodgkin's disease and malignant neoplasm of thymus) occurred in the 12 mg BID arm.No deaths occurred.Conclusions: Dosedependent improvements in clinical and endoscopic outcomes and markers of inflammation with UPA were observed with 36-week treatment in patients who responded to a 16-week induction regimen.The overall safety profile of UPA is consistent with other studies in rheumatoid arthritis.

Objectives Microvillus inclusion disease (MVID) is a severe form of neonatal diarrhea, caused mainly by mutations in MYO5B . Inactivating mutations in MYO5B causes depolarization of enterocytes in the small intestine, which gives rise to chronic, unremitting secretory diarrhea. While the pathology of the small intestine in MVID patients is well described, little is known about extraintestinal effects of MYO5B mutation. Methods We examined stomach, liver, pancreas, colon, and kidney in Navajo MVID patients, who share a single homozygous MYO5B - P660L (1979C>T p.Pro660Leu, exon 16). Sections were stained for markers of the apical membrane to assess polarized trafficking. Results Navajo MVID patients showed notable changes in H/K-ATPase-containing tubulovesicle structure in the stomach parietal cells. Colonic mucosa was morphologically normal, but did show losses in apical ezrin and Syntaxin 3. Hepatocytes in the MVID patients displayed aberrant canalicular expression of the essential transporters MRP2 and BSEP. The pancreas showed small fragmented islets and a decrease in apical ezrin in pancreatic ducts. Kidney showed normal primary cilia. Conclusions These findings indicate that the effects of the P660L mutation in MYO5B in Navajo MVID patients are not limited to the small intestine, but that certain tissues may be able to compensate functionally for alterations in apical trafficking.

BACKGROUND:Successful treatment of patients with inflammatory bowel disease (IBD) requires regular intake of medication. Nonadherence to treatment is associated with increased frequency of relapses, morbidity, and cost.METHODS:Pediatric patients with IBD taking oral medication and with access to text messaging (TM) services were included. Children were randomized by age, sex, medication administration responsibility (self vs parent), and disease activity (Pediatric Crohn Disease Activity Index or Pediatric Ulcerative Colitis Activity Index) into TM intervention and standard of care. Prospectively, the interventional group received 2-way TM reminders about medication administration. Failure to confirm intake by the patient resulted in a TM alert to the caregiver and weekly compliance reports were sent to patients, caregivers, and healthcare providers. Patients' medical records were reviewed and an adherence Morisky questionnaire completed at recruitment, 6 and 12 months.RESULTS:A total of 51 children were randomized (21 TM and 30 control). The age, sex, diagnosis (ulcerative colitis/Crohn), activity index, ethnicity, insurance, and Morisky score at baseline were similar in both groups. Morisky score improved by 1 and 0.8 points, respectively in the TM group at 6 and 12 months, whereas it did not change in the control group (P = 0.0131 and P = 0.1687, prospectively).CONCLUSIONS:TM may be effective in promoting adherence in children with IBD. Larger and longer multicenter studies are required to confirm this finding.

Rab11a is a key component of the apical recycling endosome that aids in the trafficking of proteins to the luminal surface in polarized epithelial cells. Utilizing conditional Rab11a-knockout specific to intestinal epithelial cells, and human colonic epithelial CaCo2-BBE cells with stable Rab11a knockdown, we examined the molecular and pathological impact of Rab11a deficiency on the establishment of apical cell polarity and microvillus morphogenesis. We demonstrate that loss of Rab11a induced alterations in enterocyte polarity, shortened microvillar length and affected the formation of microvilli along the lateral membranes. Rab11a deficiency in enterocytes altered the apical localization of syntaxin 3. These data affirm the role of Rab11a in apical membrane trafficking and the maintenance of apical microvilli in enterocytes.

Purpose of review This review will focus on therapeutic considerations and recent advances in treatment of caustic ingestion injuries. Recent findings A retrospective study suggests that it may be safe to advance the endoscope beyond the first circumferential burn to allow for a more complete assessment of extent of injury. A randomized controlled prospective study suggested that a 3-day course of high-dose methylprednisolone might reduce the occurrence of esophageal stricture formation. Balloon dilatation has been shown to be as effective as other bougienage techniques with lower risk of perforations. Recent studies indicate that esophageal dilatation can be safely performed as early as 5–15 days after initial ingestion and may decrease risk for long-term stricture formation. The use of adjunctive treatment, such as topical mitomycin C and esophageal stents, shows promise in reducing the reoccurrence of stricture formation after dilatation. Summary Caustic ingestion remains a significant problem in children, despite continued efforts to educate the public about ways to avoid this preventable accident. Because there are few good quality therapeutic trials in children, many of the current recommendations regarding treatment are based on expert opinion. Large, prospective, multicenter, controlled treatment trials are needed to identify the best protocols to prevent serious complications.

Current data suggest significant variations in the practice of pediatric endoscopy are prevalent. We aimed to determine if specific endoscopist characteristics (e.g. sex, colonoscopy volume, year graduated from medical school) were associated with quality indicators in children undergoing colonoscopy.

BACKGROUND AND AIMS:Currently, there are no quality measures specific to children undergoing GI endoscopy. We aimed to determine the baseline quality of pediatric colonoscopy by using the Pediatric Endoscopy Database System-Clinical Outcomes Research Initiative (PEDS-CORI), a central registry. METHODS:We conducted prospective data collection by using a standard computerized report generator and central registry (PEDS-CORI) to examine key quality indicators from 14 pediatric centers between January 2000 and December 2011. Specific quality indicators, including bowel preparation, ileal intubation rate, documentation of American Society of Anesthesiologists Physical Status Classification System (ASA) class, and procedure time, were compared during the study period. RESULTS:We analyzed 21,807 colonoscopy procedures performed in patients with a mean age of 11.5 ± 4.8 years. Of the 21,807 reports received during the study period, 56% did not include bowel preparation quality, and 12.7% did not include ASA classification. When bowel preparation was reported, the quality was described as excellent, good, or fair in 90.3%. The overall ileal intubation rate was 69.4%, and 15.6% reported cecal intubation only, calculated to be 85% cecum or ileum intubation. Thus, 15% of colonoscopy procedures did not report reaching the cecum or ileum. When excluding the proportion of procedures not intended to reach the ileum (31.5%), the overall ileal intubation rate increased to 84.0%. The rate of ileum examination varied from 85% to 95%, depending on procedure indication. CONCLUSIONS:Colonoscopy reports from our central registry revealed significant variations and inconsistent documentation in pediatric colonoscopy. Our study identifies areas for quality improvement and highlights the need for developing accepted quality measures specific to pediatric endoscopy.

Microvillus inclusion disease (MVID) is an autosomal recessive disorder that typically presents in the neonatal period with severe intractable watery diarrhea that persists despite total bowel rest. Pathological findings include pathognomonic microvillus inclusion bodies seen by electron microscopy, varying degrees of villous atrophy, and secretory granules accumulated in the apical cytoplasm of enterocytes (1). Treatment involves total parental nutrition (TPN) and small bowel transplantation. Recently, multiple mutations in myosin Vb (MYO5B) have been implicated in the etiology of MVID (2,3). Several multicenter reports have examined the clinicopathological findings in MVID (4,5). Although MVID primarily affects the intestinal tract, other organ systems are affected either secondary to complications from the disease or by the disease itself. Several studies have described a specific MVID-associated liver disease, similar to progressive familial and benign recurrent intrahepatic cholestasis. In a polarized hepatocyte epithelial cell line, one study proposed that bile canalicular biogenesis requires functional myosin Vb (6). This may be involved in the pathogenesis of MVID-associated liver disease, but further research is needed to confirm this. TPN-related cholestasis may also occur. There are no reports of pancreatic dysfunction or hyperglycemia with MVID. We report the first case of diabetes in a child with MVID (MYO5B mutation). An 11-year-old Navajo boy was diagnosed as having MVID within the first month of life. He was completely TPN-dependent and had not undergone small bowel transplantation. He had a history of minimally elevated transaminases but no cholestasis. Besides recurrent central-line infections, there were no other complications associated with MVID. He was hospitalized because of multiple daily episodes of emesis along with polyuria, polydipsia, and drinking >10 L of fluids per day. The routine glucose infusion rate (GIR) of his TPN was between 18 and 20 mg · kg−1 · min−1 and he had no hyperglycemia for >1 year while on this GIR. He had no fevers or changes in stool pattern. Endoscopic biopsies revealed abnormalities consistent with MVID. Serial measurements of lipase and amylase were normal during the last 3 years including at the time of presentation. An abdominal computed tomography scan revealed a normal-appearing pancreas. Fecal elastase was not obtained because the results would have been difficult to interpret in the face of a secretory diarrhea. A serum trypsinogen level was not obtained. Liver tests were abnormal with an elevated aspartate aminotransferase of 60 U/L (normal: 15–37 U/L) and elevated γ-glutamyl transferase of 604 U/L (normal: 15–85 U/L). An alanine aminotransferase of 52 U/L (normal: 30–65 U/L) and total bilirubin of 0.8 mg/dL (normal: 0–1 mg/dL) were normal. Blood cultures were negative. A urine analysis revealed 4+ glucose without ketonuria. Blood glucose was elevated at 429 mg/dL. Monitoring of blood glucose levels demonstrated a peak blood sugar of 1028 mg/dL. He did not develop ketonuria or metabolic acidosis. Hemoglobin A1c level was elevated at 7.7% (normal: <5.7%). Glucose and insulin levels were drawn before starting a 12-hour TPN cycle and 2 hours after TPN administration (GIR 18.3 mg · kg−1 · min−1) was started. Results demonstrated a fasting glucose of 67 mg/dL (normal: 65–99 mg/dL) and insulin of <2 μIU/mL (normal: <17 μIU/mL). Two-hour levels revealed glucose 222 mg/dL (normal: 65–139 mg/dL) and insulin 16 μIU/mL (normal: 31–88 μIU/mL). Glutamic acid decarboxylase-65 and insulinoma antigen-2 antibodies were both negative in evaluating for type 1 diabetes mellitus. Insulin was titrated to achieve euglycemia. He was ultimately given regular insulin infused with his TPN. During hospitalization at a GIR of 9 mg · kg−1 · min−1, he still had hyperglycemia while receiving 25 U/day insulin. At discharge, the blood glucose levels ranged from 80 to 200 mg/dL while receiving 25 U/day insulin (0.8 U · kg−1 · day−1). DISCUSSION We have described the first reported patient with MVID who developed diabetes. Pancreatic dysfunction, insulin resistance, and hyperglycemia in children with MVID have not been previously reported. In general, male patients with neonatal-onset secretory diarrhea and diabetes should be evaluated for immunodysregulation, polyendocrinopathy, enteropathy, X-linked syndrome and its variants; however this patient had a known MYO5B mutation confirming a diagnosis of MVID, and an evaluation for IPEX was not performed. A functional myosin Vb may be necessary for optimal glucose transport into myocytes (7). Glucose transporter type 4 (GLUT4), primarily found in adiposities and myocytes and stimulated by insulin, requires a functional actin cytoskeleton for translocation to the cell surface where glucose can be transported into the cell. In vitro, myosin Vb is important in transporting vesicular cargo along actin tracks. Myosin Vb has been shown to interact with Rab8a, a guanosine diphosphate–/guanosine-5′-triphosphate–, for optimal GLUT4 trafficking in myocytes (7). Myocytes transfected with a green fluorescent protein-chimera of myosin Vb tail showed a 40% reduced GLUT4 cell surface concentration compared with untransfected cells after being stimulated by insulin (7). A dysfunctional myosin Vb motor protein could lead to decreased GLUT4 expression on the cell surface, leading to decreased glucose transport into the cell and resulting in hyperglycemia. If this change were to occur, one would expect a compensatory hyperinsulinemia to overcome the decreased GLUT4 expression and achieve euglycemia. In this patient, a modified 2-hour intravenous glucose tolerance test revealed a markedly elevated 2-hour glucose level without a significant insulin response, indicating possible pancreatic β cell dysfunction leading to insulinopenia. Myosin Vb is known to be present in the pancreas, but its specific role is yet to be determined (8). GLUT2, primarily found in pancreatic β cells, is essential to the insulin secretory pathway, but it is unknown whether myosin Vb interacts with GLUT2. If myosin Vb was found to be important for GLUT2 trafficking in pancreatic β cells, then this could account for the inadequate insulin response to hyperglycemia seen in this patient. In conclusion, hyperglycemia is a potential complication of MVID. We hypothesize that patients with MVID may have trafficking problems with GLUT4 and GLUT2, which lead to the development of diabetes, especially under stressful conditions. Further research into the role of myosin Vb in pancreatic β cells and GLUT4 trafficking in myocytes may uncover a mechanism for hyperglycemia in MVID.

Microvillus inclusion disease (MVID) is a severe form of congenital diarrhea that arises from inactivating mutations in the gene encoding myosin Vb (MYO5B). We have examined the association of mutations in MYO5B and disruption of microvillar assembly and polarity in enterocytes. Stable MYO5B knockdown (MYO5B-KD) in CaCo2-BBE cells elicited loss of microvilli, alterations in junctional claudins, and disruption of apical and basolateral trafficking; however, no microvillus inclusions were observed in MYO5B-KD cells. Expression of WT MYO5B in MYO5B-KD cells restored microvilli; however, expression of MYO5B-P660L, a MVID-associated mutation found within Navajo populations, did not rescue the MYO5B-KD phenotype but induced formation of microvillus inclusions. Microvilli establishment required interaction between RAB8A and MYO5B, while loss of the interaction between RAB11A and MYO5B induced microvillus inclusions. Using surface biotinylation and dual immunofluorescence staining in MYO5B-KD cells expressing mutant forms of MYO5B, we observed that early microvillus inclusions were positive for the sorting marker SNX18 and derived from apical membrane internalization. In patients with MVID, MYO5B-P660L results in global changes in polarity at the villus tips that could account for deficits in apical absorption, loss of microvilli, aberrant junctions, and losses in transcellular ion transport pathways, likely leading to the MVID clinical phenotype of neonatal secretory diarrhea.