Immunolocalization and translocation of aquaporin-5 water channel in sweat glands

doi:10.1016/j.jdermsci.2013.01.013

Journal of Dermatological Science

Volume 70, Issue 1, April 2013, Pages 26-33

https://doi.org/10.1016/j.jdermsci.2013.01.013 Get rights and content

Abstract

Background

Aquaporin-5 (AQP5) is a member of the water channel protein family. Although AQP5 was shown to be present in sweat glands, the presence or absence of regulated intracellular translocation of AQP5 in sweat glands remained to be determined.

Objective

We investigated whether AQP5 in sweat glands translocated during sweating, and also sought to determine the intracellular signal that triggers this translocation.

Methods

Immunofluorescent analyses of AQP5 in mouse and human sweat glands were performed. Madin-Darby Canine kidney (MDCK) cell lines stably expressing human AQP5 were generated, and the regulated translocation of AQP5 in the polarized cells was assessed by immunofluorescent analysis and biotinylation assays.

Results

AQP5 showed rapid translocation to the apical membranes during sweating. In human eccrine sweat glands, immunoreactive AQP5 was detected in the apical membranes and the intercellular canaliculi of secretory coils, and in the basolateral membranes of the clear cells. Treatment of human AQP5-expressing MDCK cells with calcium ionophore A23187 resulted in a twofold increase of AQP5 in the apical membranes within 5 min.

Conclusion

The regulated AQP5 translocation may contribute to sweat secretion by increasing the water permeability of apical plasma membranes of sweat glands.

Introduction

Secretion of fluid is the principal function of eccrine sweat glands. An eccrine sweat gland is a single tubular structure consisting of a secretory portion and a ductal portion. In the secretory portion, primary fluid, i.e., primary sweat, is secreted onto the lumen by active salt transport followed by movement of water, and the fluid transits through the duct where salt reabsorption occurs [1], [2]. Dyshidrosis, including hyperhidrosis, hypohidrosis and anhidrosis, is a serious problem that can have a severe impact on daily life. For example, primary focal hyperhidrosis is a disorder of excessive sweating that occurs in the palms, soles, axillae and craniofacial region. This condition results in occupational, psychological and physical impairment and potential social stigmatization [3], [4]. Patients have several treatment options [5], but these treatments have complications and limitations [6]. Patients with obstinate dyshidrosis or who experience serious side effects are desperate for new treatments. In order to develop new treatments, further understanding of the regulation of sweating, especially the regulation of fluid movement, is required.

Aquaporins (AQPs) are a family of integral membrane channel proteins that allow the rapid movement of water across the plasma membrane. Thirteen members of the AQP family (AQP0–AQP12) have been identified in mammals to date, and these proteins are expressed in various fluid-transporting epithelia with a distinct tissue-specific pattern [7]. Although there have been a few reports regarding the presence of AQP5 in sweat glands [8], [9], [10], [11], [12], [13], its involvement in sweating is not well understood.

In the present study, we focused on translocation of AQP5 in sweat glands. By immunofluorescent studies of AQP5 in the sweat glands, we found that AQP5 showed apical translocation during sweating. We then generated Madin-Darby canine kidney (MDCK) cell lines stably expressing human AQP5 (hAQP5), and found that the intracellular calcium might mediate this translocation.

Section snippets

Antibodies and chemical reagents

Primary antibodies used were: rabbit monoclonal anti-AQP5 (ab93230, Abcam, Cambridge, MA, USA), goat polyclonal anti-AQP5 (sc-9890, Santa Cruz Biotechnology, CA, USA), mouse monoclonal anti-Na⁺/K⁺ATPase (sc-21712, Sigma–Aldrich, St. Louis, MO, USA), and rabbit anti-anoctamin-1 (ANO1) antibody generated in our laboratory using the antigen peptide (NH₂ single bond C + GDGSPVPSYEYHGDALCOOH, corresponding to amino acid residues 941–956 of mouse ANO1). Secondary antibodies used for immunofluorescence were Alexa

AQP5 translocated from the non-apical region to the apical membranes in cells of mouse sweat glands under sweating condition

AQP5 was reported to translocate to the apical membranes of cells of rat parotid glands with cevimeline, a muscarinic receptor agonist [15]. We therefore assessed changes in the subcellular localization of AQP5 in sweat glands during sweating in mice. To determine whether the mice were sweating or not through their paws, we used a modified Minor method. In the control anesthetized mice, spots which correspond to sweat secretion, were scarcely observed on the paws (Fig. 1a). To make the mice

Discussion

AQP5 water channel is expressed in several secretory epithelia, including salivary glands [24], airway submucosal glands [25], lacrimal glands [26], and sweat glands, as well as in alveolar type I epithelial cells [25], [27]. Aquaporin-2 (AQP2) is a water channel in the collecting ducts of the kidney that translocates from intracellular membranes to plasma membranes in response to vasopressin [28], [29], [30]. AQP5 has 63% identity to AQP2 [31], and was shown to translocate from the cytoplasm

Funding sources

This study was supported by Health and Labor Sciences Research Grants from Ministry of Health, Labor and Welfare of Japan: Research on intractable diseases (Nos. 0309001, 1211202), Grant-in-Aid for Scientific Research (A) from the Japan Society for the Promotion of Science (Nos. 20249047, 22249032), Takeda Science Foundation, and Salt Science Research Foundation (1026).

Acknowledgments

We thank Chiyako Miyagishi and Motoko Chiga of Tokyo Medical and Dental University for excellent technical assistance.

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Immunolocalization and translocation of aquaporin-5 water channel in sweat glands

Abstract

Background

Objective

Methods

Results

Conclusion

Introduction

Section snippets

Antibodies and chemical reagents

AQP5 translocated from the non-apical region to the apical membranes in cells of mouse sweat glands under sweating condition

Discussion

Funding sources

Acknowledgments

J Am Acad Dermatol

Ann Thorac Surg

Life Sci

J Am Acad Dermatol

Cell

Biochim Biophys Acta

Biochem Biophys Res Commun

Biochem Biophys Res Commun

J Biol Chem

Exp Eye Res

J Biol Chem

Biochem Biophys Res Commun

The physiology, pharmacology, and biochemistry of the eccrine sweat gland

Rev Physiol Biochem Pharmacol

The secretion of salt and water by the eccrine sweat gland

Arch Dermatol

Botulinum A toxin improves life quality in severe primary focal hyperhidrosis

Eur J Neurol

The illness intrusiveness rating scale: a measure of severity in individuals with hyperhidrosis

Qual Life Res

Aquaporin water channels in mammals

Clin Exp Nephrol