Claudin-18 Deficiency Results in Alveolar Barrier Dysfunction and Impaired Alveologenesis in Mice

Abstract

Claudins are a family of transmembrane proteins that are required for tight junction formation. Claudin (CLDN)-18.1, the only known lung-specific tight junction protein, is the most abundant claudin in alveolar epithelial type (AT) 1 cells, and is regulated by lung maturational agonists and inflammatory mediators. To determine the function of CLDN18 in the alveolar epithelium, CLDN18 knockout (KO) mice were generated and studied by histological, biochemical, and physiological approaches, in addition to whole-genome microarray. Alveolar epithelial barrier function was assessed after knockdown of CLDN18 in isolated lung cells. CLDN18 levels were measured by quantitative PCR in lung samples from fetal and postnatal human infants. We found that CLDN18 deficiency impaired alveolar epithelial barrier function in vivo and in vitro, with evidence of increased paracellular permeability and architectural distortion at AT1-AT1 cell junctions. Although CLDN18 KO mice were born without evidence of a lung abnormality, histological and gene expression analysis at Postnatal Day 3 and Week 4 identified impaired alveolarization. CLDN18 KO mice also had evidence of postnatal lung injury, including acquired AT1 cell damage. Human fetal lungs at 23-24 weeks gestational age, the highest-risk period for developing bronchopulmonary dysplasia, a disease of impaired alveolarization, had significantly lower CLDN18 expression relative to postnatal lungs. Thus, CLDN18 deficiency results in epithelial barrier dysfunction, injury, and impaired alveolarization in mice. Low expression of CLDN18 in human fetal lungs supports further investigation into a role for this tight junction protein in bronchopulmonary dysplasia.

Claudin-18–mediated YAP activity regulates lung stem and progenitor cell homeostasis and tumorigenesis

“…In addition to alterations in epithelial permeability and ion transport, we observed increased lung cellularity from embryonic day 18 (E18) onward ( Figure 1A and Supplemental Figure 1A; supplemental material available online with this article; https://doi. org/10.1172/JCI90429DS1) with areas of more marked alveolar wall hypercellularity ( Figure 1A), as well as airspace enlargement ( Figure 1A and Supplemental Figure 1, A and B) as previously reported (25 -/-cells was maintained following passage, indicating vs. 1.2% ± 0.3%, respectively) ( Figure 2, E and F). TUNEL assay shows low levels of apoptosis in both Cldn18 -/-and WT mice (0.2% vs. 0.1%, respectively) (Supplemental Figure 8), indicating that increased cell number is the result of increased proliferation rather than decreased apoptosis.…”

“…Deletion of the stomach-specific Cldn18.2 isoform in mice leads to loss of TJ strands and increased paracellular H + leakage in the stomach, resulting in atrophic gastritis and metaplasia but without evidence of tumor formation (23). Recently generated Cldn18 -/-mice with deletion of both isoforms showed increased lung permeability to ions and solutes, consistent with known roles of claudins in regulation of barrier function (24,25).…”

The relative balance of GM-CSF and TGF-β1 regulates lung epithelial barrier function

“…Consistent with this observation, claudin-4-deficient mice have a relatively mild lung phenotype (23). Moreover, claudin-4 upregulation does not rescue the decrease in AEC barrier function found in claudin-18 knockout mice (31,32). These data further underscore the need to understand how multiple components of the tight junction proteome interact to form a fully functional lung epithelial barrier.…”

Evolution of the vertebrate claudin gene family: insights from a basal vertebrate, the sea lamprey

“…This was surprising, because Cldn18 has not been reported to be expressed during neural crest development in any other vertebrate. Claudin 18 knockout mice are viable and exhibit changes in cell architecture and differentiation of alveolar epithelium, but no neural crest phenotype, suggesting that it is not involved in neural crest development in mice (LaFemina et al, 2014, Li et al, 2014. In chick, claudin 18 expression has been reported in Hensen's node, epiblast, neural and non-neural ectoderm, pharynx, eye and limb ectoderm, but not in the migrating neural crest (Collins et al, 2013).…”