Gene expression and immunochemical localization of major cytochrome P450 drug-metabolizing enzymes in bovine nasal olfactory and respiratory mucosa

Abstract

Despite tremendous advancement in the characterization of nasal enzyme expression, knowledge of the role of the nasal mucosa in the metabolism of xenobiotics is still inadequate, primarily due to the limited availability of in vitro models for nasal metabolism screening studies. An extensive knowledge of the oxidative and conjugative metabolizing capacity of the cattle (Bos taurus) olfactory and respiratory mucosa can aid in efficient use of these tissues for pre-clinical investigations of the biotransformation and toxicity of therapeutic agents following nasal administration or inhalation. Cows are also exposed to a variety of airborne pollutants and pesticides during their lifetime, the metabolism of which can have profound toxicological and ecological consequences. The aim of the present study was to characterize cytochrome P450 (CYP) enzyme expression in the bovine nasal mucosa. Amplification of the specific genes through real time RT-PCR confirmed expression of several CYP enzymes in bovine hepatic and nasal tissues. The results demonstrate that bovine nasal olfactory and respiratory mucosal and liver tissues express similar populations, families, and distributions of CYP enzymes, as has been previously reported with other species, including humans. Bovine ex vivo tissues can serve as a readily available reference tissue to elucidate preclinical toxicokinetic effects resulting from exposure to substances in the environment or following drug administration.

Modulating nasal mucosal permeation using metabolic saturation and enzyme inhibition techniques

“…CYP450 activity was observed both in the epithelial cell layer and in the submucosal tissues, and the nearly equivalent amounts of 6‐hydroxymelatonin measured following transport across full‐thickness and de‐epithelialized tissues suggests that the CYP450 activity in the submucosal region is not negligible compared to the epithelial layer. Previous work using immunohistochemistry has also demonstrated the significant expression of CYP450 enzymes in both the epithelial and submucosal regions of excised bovine nasal tissues . As a result, the evaluation of the metabolic fate of a drug in the nasal mucosa should include both the epithelial cells along with the submucosal tissues as metabolic loss is likely to be underpredicted if only culture‐based models of the nasal epithelium are used.…”

“…Previous work using immunohistochemistry has also demonstrated the significant expression of CYP450 enzymes in both the epithelial and submucosal regions of excised bovine nasal tissues. [29] As a result, the evaluation of the metabolic fate of a drug in the nasal mucosa should…”

“…This species has also been shown to be well‐suited for investigations of nasal permeation and metabolism . Using real‐time reverse‐transcriptase polymerase chain reaction (RT‐PCR) and enzyme immunohistochemistry, we have previously demonstrated that CYP450 isoforms and corresponding proteins are expressed in the bovine nasal mucosa, and the enzymes are localized both in the epithelial as well as submucosal regions . Examinations of concentration gradient dependent melatonin permeability across the bovine nasal olfactory and respiratory mucosa were carried out to evaluate the metabolic limitations that the nasal route may contribute to limiting the absorption of CYP450 substrates.…”

“…[28] Using real-time reverse-transcriptase polymerase chain reaction (RT-PCR) and enzyme immunohistochemistry, we have previously demonstrated that CYP450 isoforms and corresponding proteins are expressed in the bovine nasal mucosa, and the enzymes are localized both in the epithelial as well as submucosal regions. [29] Examinations of concentration gradient dependent melatonin permeability across the bovine nasal olfactory and respiratory mucosa were carried out to evaluate the metabolic limitations that the nasal route may contribute to limiting the absorption of CYP450 substrates. The effect of the inclusion of CYP450 inhibitor on overall diffusive transfer across the nasal mucosa was studied to evaluate its impact on melatonin's diffusive transfer across the nasal mucosa.…”

Endocytic Uptake of Solid Lipid Nanoparticles by the Nasal Mucosa

“…Lipid-associated delivery systems for nasal administration, including liposomes, solid lipid nanoparticles (SLN), and nanostructured lipid carriers (NLC), have shown promise in the delivery of vaccines, systemically available therapeutics, and as targeting strategies to reach the CNS via the nose-to-brain pathways [1]. These lipid-associated systems can protect their drug or vaccine payloads from enzymatic degradation from the variety of metabolically active enzyme systems in the nasal secretions and mucosa, including cytochrome P450s, monooxygenases, carboxylesterases and a number of conjugative enzyme systems [2][3][4]. Nanoparticles can also enable escape from undesirable, transportermediated efflux; they can mask unsuitable drug properties; and can provide additional pathways for entering the nasal mucosa.…”