Fig 1.
Molecular profiling of human sweat gland compartments.
(A) Schematic illustration of a human sweat gland and its main tissue segments. (Left) Diagram of the sweat gland structure in human skin; (center) diagram of a sweat gland cross section in human skin; (upper right) diagram of the cellular arrangement in the duct consisting of luminal and basal cells surrounded by basement membrane; (lower right) diagram of the cellular arrangement in the secretory portion consisting of luminal and myoepithelial cells surrounded by basement membrane. (B) Cross section of sweat gland coiled tubules in the deep dermis of human skin. HE-stained skin cross section shows the sweat glands surrounded by adipocytes and connective tissue. The coiled regions of sweat glands were detected as rounded structures in the hypodermis. Boxed area in the upper panel is magnified in the lower panel. (C) Expression patterns of sweat gland cell markers K8, αSMA, K77, and S100A2 in human sweat glands. K8 and K77 were expressed in parts of the luminal cell layers, and αSMA and S100A2 in parts of the basal cell layers. Insets show magnified views of the sweat glands. (D) Double immunofluorescence detection of sweat gland markers K8, αSMA, K77, and S100A2 in sweat glands. Insets show magnified views. Nuclei (blue) were counterstained with Hoechst 33342. (B–D) Scale bars: 50 μm.
Fig 2.
Histological anatomies of basement membranes, cell surface receptors, and cytoskeleton markers in coiled structure compartments of sweat glands.
Expression patterns of (A) LN332 and LAMA5 (basement membrane markers), (B) ITGB1 and ITGA6 (cell surface receptor markers), and (C) E-cadherin and phalloidin (cytoskeleton markers) in the basal layers of sweat glands. Arrowheads and arrows in (B) indicate secretory portions and ducts, respectively. Black arrowheads and arrows in (C) indicate strong detection of actin filaments in ductal apical and secretory basal sides, respectively. Arrowheads and arrows indicate expression of E-cadherin in ductal basal and secretory luminal cell layers, respectively. Double immunofluorescence detection of LN332 and LAMA5 (D), and ITGB1 and ITGA6 (E) with sweat gland basal cell markers S100A2 and αSMA, and E-cadherin and phalloidin (F) with sweat gland secretory portion markers K8 and αSMA in sweat gland cross sections. (G) Expression patterns of CD31 (blood vessel) and PGP9.5 (nerve fiber) markers in sweat glands. Sweat glands boxed in the upper panels are shown in the lower panels at higher magnification. Nuclei (blue) were counterstained with Hoechst 33342. Scale bars: 50 μm.
Fig 3.
Imaging of the 3D structure of sweat glands.
(A) Neutral red-positive sweat gland coiled organs detected in human skin tissue. The boxed region is shown at higher magnification in the left panel. (B) Visualization by whole-mount staining of an entire sweat gland embedded in human skin. αSMA and S100A2 were detected in sweat glands by double immunofluorescence. Asterisks and double asterisks indicate αSMA-positive secretory portions and S100A2-positive coiled ducts, respectively. Arrowheads and arrows indicate hair follicles and blood vessels, respectively. The dashed line indicates the skin surface. (C) Procedure for whole-mount staining of coiled fragments of sweat glands. The left panel shows neutral red-stained sweat gland organs collected from human skin tissue. (D) Visualization of the basement membrane of the entire sweat gland coiled structure by whole-mount staining for LN332. (E) Double immunofluorescence detection of LN332 and LAMA5 in sweat glands. (F) Imaging of ductal and secretory portions of coiled fragments dissected from sweat glands. Double immunofluorescence detection of K8 and S100A2 in an isolated sweat gland. (Left panel) Projection image of a whole-mount 3D sweat gland stained for K8 and S100A2. (Right panel) Optical section of the whole-mount image. Arrows and arrowheads indicate S100A2-positive ductal and K8-positive secretory portions, respectively. Nuclei (blue) were counterstained with Hoechst 33342 (B, E, and F). Boxed areas in the left panels are shown at higher magnification in the right panels (D, E). Scale bars: 200 μm (A) and 50 μm (B, D–F).
Fig 4.
3D imaging of a sweat gland at single cell resolution.
(A) Whole-mount 3D confocal images of ITGB1 and ITGA6 expression. Arrowhead and arrow indicate the portions comprised of cuboidal basal cells and elongated cells, respectively. (B) Cuboidal cells (left) and highly elongated cells (right) forming sweat gland putative ducts and secretory portions, respectively. The patterns of F-actin-phalloidin complexes were used to distinguish ducts and secretory portions of sweat glands with the former consisting of cuboidal cells (left) and the latter of highly elongated cells (right). Arrowheads and arrows indicate cuboidal and highly elongated cells, respectively. Yellow and red boxed areas in the center panel are magnified in the left and right panels, respectively. (C) Sweat gland stained with phalloidin. Arrows indicate the elongated cells (dashed lines) that were longitudinally and uniformly arranged on the secretory portion. (D) Whole-mount 3D confocal images of S100A2 and αSMA expression. Arrowhead and arrow indicate the portions comprised of cuboidal basal cells and elongated cells, respectively. (E) Double immunofluorescence detection of αSMA and phalloidin in sweat glands. The right panel shows a high magnification view of the left panel. Arrows indicate phalloidin-positive elongated myoepithelial cells expressing αSMA. Arrowheads indicate ducts. Asterisks indicate αSMA-positive blood vessels. (F) Schematic presentation of the 3D coiled structure of sweat glands. Highly elongated myoepithelial cells are longitudinally arranged on the entangled secretory portion of sweat glands. Boxed areas in the left panels are shown at higher magnification in the right panels (A, C–E). Nuclei (blue) were counterstained with Hoechst 33342 (B, D, and E). Scale bars: 50 μm.
Fig 5.
Imaging of sweat gland blood vessels and nerve fibers.
(A) Immunofluorescence detection of CD31 and phalloidin in sweat glands. Arrows and arrowheads indicate secretory portions and ducts of sweat gland tubules, respectively. (B) Double immunofluorescence detection of CD31 and LN332 in sweat glands. Arrowheads indicate blood vessels running parallel to sweat gland tubules. (C) Immunofluorescence detection of PGP9.5 in sweat glands. Arrows indicate nerve fibers wrapping around the tubules of sweat glands. Arrowheads indicate ductal tubules. (D) Immunofluorescence detection of PGP9.5 and αSMA in sweat glands. Lower panel shows optical sections of sweat glands. Arrows indicate nerve fibers that enwrap the αSMA-positive secretory portions. The boxed area in the upper panel is shown at higher magnification in the lower panel (A–C). Nuclei (blue) were counterstained with Hoechst 33342. Scale bars: 50 μm.
Fig 6.
Schematic model for mechanical contraction of sweat glands during sweat secretion.
Tubular secretory portions, but not ducts, of sweat glands have self-entangled coiled structural features. Elongated myoepithelial cells are arranged longitudinally parallel to the entangled secretory tubules. Nerve fibers enwrap myoepithelial cell layers surrounding the secretory portions. In sweat glands, multiple elongated myoepithelial cells synchronously contract their tubular secretory portions for excretion of sweat (upper panel), while stellate-shaped myoepithelial cells individually contract their acinar secretory portions of salivary and mammary glands (lower panel).