A G542X cystic fibrosis mouse model for examining nonsense mutation directed therapies

McHugh, Daniel R.; Steele, M.; Valerio, Dana M.; Miron, Alexander; Mann, Rachel J.; LePage, David F.; Conlon, Ronald A.; Cotton, Calvin U.; Drumm, Mitchell L.; Hodges, Craig A.

doi:10.1371/journal.pone.0199573

Cited by 47 publications

(47 citation statements)

References 67 publications

(82 reference statements)

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“…Affinity-purified polyclonal antibodies were raised against recombinant peptides of mouse SGLT1 (NP_062784.3; amino acid residues 576-610) and of mouse CFTR (NP_066388.1; 634-675 and 1438-1476), respectively, using the same method as described previously. 21 Specificity of each antibody was confirmed by immunostaining in Sglt1-knockout (a gift from Dr. Koepsell) 22 or Cftr null (G542X mutation generated in Case Western CF Mouse Model Core) 23 mouse tissues ( Supplementary Figures 1 and 2).…”

Section: Antibody Productionmentioning

confidence: 99%

Lysophosphatidic Acid Increases Maturation of Brush Borders and SGLT1 Activity in MYO5B-deficient Mice, a Model of Microvillus Inclusion Disease

et al. 2020

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BACKGROUND & AIM: Myosin VB (MYO5B) is an essential trafficking protein for membrane recycling in gastrointestinal epithelial cells. The inactivating mutations of MYO5B cause the congenital diarrheal disease, microvillus inclusion disease (MVID). MYO5B deficiency in mice causes mislocalization of SGLT1 and NHE3, but retained apical function of CFTR, resulting in malabsorption and secretory diarrhea. Activation of lysophosphatidic acid (LPA) receptors can improve diarrhea, but the effect of LPA on MVID symptoms is unclear. We investigated whether LPA administration can reduce the epithelial deficits in MYO5B-knockout mice. METHODS: Studies were conducted with tamoxifen-induced, intestine-specific knockout of MYO5B (VilCre ERT2 ;Myo5b flox/flox) and littermate controls. Mice were given LPA, an LPAR2 agonist (GRI977143), or vehicle for 4 days after a single injection of tamoxifen. Apical SGLT1 and CFTR activities were measured in Üssing chambers. Intestinal tissues were collected, and localization of membrane transporters was evaluated by immunofluorescence analysis in tissue sections and enteroids. RNA sequencing and enrichment analysis were performed with isolated jejunal epithelial cells. RESULTS: Daily administration of LPA reduced villus blunting, frequency of multivesicular bodies, and levels of cathepsins in intestinal tissues of MYO5B-knockout mice compared with vehicle administration. LPA partially restored the brush border height and the localization of SGLT1 and NHE3 in small intestine of MYO5B-knockout mice and enteroids. The SGLT1-dependent short-circuit current was increased and abnormal CFTR activities were decreased in jejunum from MYO5B-knockout mice given LPA compared with vehicle. CONCLUSIONS: LPA may regulate a MYO5B-independent trafficking mechanism and brush border maturation, and therefore be developed for treatment of MVID.

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Section: Antibody Productionmentioning

confidence: 99%

Lysophosphatidic Acid Increases Maturation of Brush Borders and SGLT1 Activity in MYO5B-deficient Mice, a Model of Microvillus Inclusion Disease

et al. 2020

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“…The CFTR gene is expressed in abundance throughout the intestine (17). Mouse models of CF have demonstrated that CFTR dysfunction leads to mucus accumulation within the intestinal lumen, disturbed motility, small bowel bacterial overgrowth, and inflammation with an abnormal innate immune response (18).…”

Section: Discussionmentioning

confidence: 99%

Mucosal Abnormalities in Children With Congenital Chloride Diarrhea—An Underestimated Phenotypic Feature?

et al. 2020

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Objectives and Study: Congenital chloride diarrhea (CCD) is a rare, autosomal recessive disorder caused by mutations in the SLC26A3 gene encoding a transmembrane chloride/bicarbonate ion exchanger mainly expressed in the apical brush border of the ileal and colonic epithelium. Lifelong, secretory, chloride-rich diarrhea and hypochloremic, hypokalemic metabolic alkalosis are characteristic. Histological evidence of bowel inflammation is not typically described in CCD and has only been reported in a few patients. Methods: We report four cases of CCD who received adequate resuscitation with appropriate replacement of their fecal salt and water losses. Three had associated inflammatory bowel changes at endoscopy. The index case of CCD who developed frankly bloodstained diarrhea aged 7 months was found to have histologically confirmed colitis at endoscopy. An electronic search of the hospital database to identify all patients with confirmed CCD was performed. A further three children underwent de novo diagnostic evaluation and treatment. A retrospective case note review was undertaken to determine the incidence and subtype of inflammatory bowel disease (IBD) by clinical, endoscopic, and histological means. Results: Four children with genetically confirmed CCD were identified, two being female. The first girl had a granulomatous colitis with ulceration. She went into remission with a combination of steroids and azathioprine. Immunosuppression was subsequently discontinued without a further flare of colitis. A second girl was found to have patchy inflammatory changes in the small bowel and focal active colitis. A third patient, a boy, demonstrated mild inflammatory changes in the small bowel with apoptotic debris and mild inflammation in the colon. A fourth patient did not develop intestinal inflammation. Conclusion: Our case series highlights the potential association of CCD with panenteric inflammation. While our cohort was small, CCD is rare and three out of four children referred to our tertiary referral center were affected. While early diagnosis and adequate salt replacement therapy are crucial in CCD management, the clinician should also be aware of bowel inflammation as a potential cause of failure of CCD therapy to control bowel symptomatology. Further insight is needed to understand the underlying patho-mechanism giving rise to bowel inflammation in this group.

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“…A mouse carrying the G542X nonsense mutation in the CFTR locus was generated by CRISPR nucleases showing common manifestations of CF determined by the absence of CFTR ion channel, such as intestinal obstruction and reduced growth [98].…”

Section: Genome Editing For the Development Of Cf Modelsmentioning

confidence: 99%

Gene Therapy for Cystic Fibrosis: Progress and Challenges of Genome Editing

Maule

Arosio

Cereseto

2020

IJMS

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Since the early days of its conceptualization and application, human gene transfer held the promise of a permanent solution to genetic diseases including cystic fibrosis (CF). This field went through alternated periods of enthusiasm and distrust. The development of refined technologies allowing site specific modification with programmable nucleases highly revived the gene therapy field. CRISPR nucleases and derived technologies tremendously facilitate genome manipulation offering diversified strategies to reverse mutations. Here we discuss the advancement of gene therapy, from therapeutic nucleic acids to genome editing techniques, designed to reverse genetic defects in CF. We provide a roadmap through technologies and strategies tailored to correct different types of mutations in the cystic fibrosis transmembrane regulator (CFTR) gene, and their applications for the development of experimental models valuable for the advancement of CF therapies.

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A G542X cystic fibrosis mouse model for examining nonsense mutation directed therapies

Cited by 47 publications

References 67 publications

Lysophosphatidic Acid Increases Maturation of Brush Borders and SGLT1 Activity in MYO5B-deficient Mice, a Model of Microvillus Inclusion Disease

Lysophosphatidic Acid Increases Maturation of Brush Borders and SGLT1 Activity in MYO5B-deficient Mice, a Model of Microvillus Inclusion Disease

Mucosal Abnormalities in Children With Congenital Chloride Diarrhea—An Underestimated Phenotypic Feature?

Gene Therapy for Cystic Fibrosis: Progress and Challenges of Genome Editing

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