| Abstract |
Thermoregulation of the human body mainly relies on sweating. Thereby,
aqueous sweat fluid secreted by eccrine sweat glands onto the skin surface
evaporates and cools the skin. Although this is a natural, highly
efficient process providing humans with an evolutionary advantage,
appearance of wet patches on clothes is mostly undesired today. Especially
in case of sweat-related disorders such as hyperhidrosis, where excessive
sweating occurs, this poses a high burden for the individual. Underlying
dysregulation of sweating is, up to now, only partly understood and just
few alleviating agents are available. For some of them the mechanism of
action is well investigated, whereas the one of the most common
antiperspirant ingredient, aluminum chlorohydrate (ACH), is still only
partly disclosed. It is assumed to encompass physical blockage of the
eccrine sweat gland.
To intensify the knowledge of the sweating mechanism on a cellular level
and to elucidate possible physiological effects of ACH, a cell-based in
vitro test procedure was developed. As sweating is mainly a process of ion
fluxes between eccrine sweat gland cells, gland lumen, and surrounding
tissue, herein established methodical test system relies on monitoring of
intracellular changes of calcium, potassium, sodium, and chloride ions
using cultured primary human eccrine sweat gland cells. Employing this
novel procedure ACH was demonstrated to also evoke physiological reactions
in human eccrine sweat gland cells. Strikingly, a distinct class of
substances, Cl(-)-containing ammonium solutions, elicited the same
characteristic ion changes as ACH. With further testing, polyols were
identified as another class of substances dysregulating the ion
equilibrium in vitro. For both substance classes the antiperspirant effect
was verified in humans. Strengthening herein developed reliable in vitro
test system, even proposals for underlying cellular mode of action of
these agents are possible. This highlights the capabilities of these
methods and contributes significantly to understanding sweating on a
cellular basis.
Adding to the latter aspect, an organotypic three-dimensional model of the
human eccrine sweat gland was developed in this work to facilitate
detailed scrutiny of cell-cell-interactions between the cell types of
secretory coil and reabsorbing duct. In a further step, those cells were
successfully integrated into newly designed in vitro dermal equivalents
comparable to the natural environment in human skin. Both these in vitro
models emphasized cellular interdependency of coil and duct cells in
developing certain proteins and revealed some alterations in protein
expression of cultured cells compared to native eccrine sweat gland cells.
Those deviances were also apparent in herein generated eccrine sweat gland
duct cell line. After transduction with simian virus 40 large T antigen-
containing lentiviral vector and overcoming of a short proliferation
crisis, transduced eccrine duct cells exhibited an extended lifespan with
stable growth suggesting their immortalized state. As duct cells represent
the primary target of topically applied products and, so far, no
immortalized duct cell line is available for research, this newly
generated and described transduced duct cell line represents an important
tool for standardization of cellular material in future in vitro sweat
gland research. It should facilitate more detailed elucidation of
physiological sweating processes and pose a defined source of cellular
material for generation of organotypic sweat gland models.
Results of this PhD thesis add significantly to understanding the
mechanism of sweating including required cellular interactions. With the
help of newly generated eccrine sweat gland duct cell line standardization
of these in vitro approaches is feasibly in future, which allows for
further detailed investigation of perspiration and cellular
interdependency with relevance for treating dermal disorders.
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