Gene Targeting of the Cysteine Peptidase Cathepsin H Impairs Lung Surfactant in Mice

Abstract

BackgroundThe 11 human cysteine cathepsins are proteases mainly located in the endolysosomal compartment of all cells and within the exocytosis pathways of some secretory cell types. Cathepsin H (Ctsh) has amino- and endopeptidase activities. In vitro studies have demonstrated Ctsh involvement in the processing and secretion of the pulmonary surfactant protein B (SP-B). Furthermore, Ctsh is highly expressed in the secretory organelles of alveolar type II pneumocytes where the surfactant proteins are processed.Methodology/Principal FindingsHence, we generated Ctsh null mice by gene targeting in embryonic stem cells to investigate the role of this protease in surfactant processing in vivo. The targeting construct contains a ß-galactosidase (lacZ) reporter enabling the visualisation of Ctsh expression sites. Ctsh-deficiency was verified by northern blot, western blot, and measurement of the Ctsh aminopeptidase activity. Ctsh −/− mice show no gross phenotype and their development is normal without growth retardation. Broncho-alveolar lavage (BAL) from Ctsh −/− mice contained lower levels of SP-B indicating reduced SP-B secretion. The BAL phospholipid concentration was not different in Ctsh+/+ and Ctsh −/− mice, but measurement of surface tension by pulsating bubble surfactometry revealed an impairment of the tension reducing function of lung surfactant of Ctsh −/− mice.Conclusions/SignificanceWe conclude that cathepsin H is involved in the SP-B production and reduced SP-B levels impair the physical properties of the lung surfactant. However, Ctsh defiency does not reproduce the severe phenotype of SP-B deficient mice. Hence, other proteases of the secretory pathway of type II pneumocytes, i.e. cathepsins C or E, are still able to produce surfactant of sufficient quality in absence of Ctsh.

A mechanistic target of rapamycin complex 1/2 (mTORC1)/V-Akt murine thymoma viral oncogene homolog 1 (AKT1)/cathepsin H axis controls filaggrin expression and processing in skin, a novel mechanism for skin barrier disruption in patients with atopic dermatitis

“…To investigate the effect of Ctsh reduction in vivo , we examined newborn mouse skin from Ctsh −/− and Ctsh +/− mice (Fig 5, A and B ) 19 using histology. Although there was no change in epidermal thickness, the cornified layer was significantly thinner in both the Ctsh +/− and Ctsh −/− mice.…”

“…CTSH is expressed ubiquitously, and in addition to being involved in bulk protein degradation, it does display cell-specific functions, such as its role in the processing and secretion of surfactant protein C in type II pneumocytes 19, 55. Ctsh-deficient mice have reduced lung surfactant levels, which might interfere with breathing mechanisms, causing respiratory complications 19 .…”

“…Ctsh knockout and heterozygote mice were generated, as previously described, 19 and backcrossed onto the C57BL/6J background for eight generations. Ctsh −/− and Ctsh +/− mice and wild-type (WT) littermate control animals were bred under specific pathogen-free conditions in accordance with the German law for Animal Protection (Tierschutzgesetz), as published on May 25, 1998.…”

Spatially-Resolved Proteomics: Rapid Quantitative Analysis of Laser Capture Microdissected Alveolar Tissue Samples

“…The receptor GPR116 (also known as Adgrf5) is known to be present in surfactant producing Type II cells58 and participates in the regulation of surfactant homeostasis585960. GPR116 as well as all the quantifiable surfactant proteins in our dataset (SP-A, SP-B, SP-D) and associated surfactant protein maturation enzymes (including Cathepsin H, Napsin-A, convertase ES-2)616263 were found to increase through development consistent with prior surfactant protein biochemical and transcriptome observations6465 (Fig. 6B).…”

Papain-like peptidases: structure, function, and evolution

“…As an aminopeptidase, it has been shown to process several neuropeptides (147,148) as well as granzyme B (149) . It is also involved in surfactant protein B maturation in the lung, but not indispensable for this process (150) . Interestingly, cathepsin H lacking the mini chain is capable of acting as an endopeptidase (151) .…”