Safety and efficacy of pharmaceutical agents can be greatly improved by encapsulation within, or covalent attachment to, a biomaterial carrier. Drug delivery systems must deliver the necessary amount of drug to the targeted site for a necessary period of time, both efficiently and precisely. Various kinds of high-performance biomaterials are being constantly developed for this purpose. Cyclodextrins are potential candidates for such a role, because of their ability to alter physical, chemical, and biological properties of guest molecules through the formation of inclusion complexes. The alpha-, beta-, and gamma-cyclodextrins are widely used natural cyclodextrins, consisting of six, seven, and eight D-glucopyranose residues, respectively, linked by -1,4 glycosidic bonds into a macro cycle. Each cyclodextrin has its own ability to form inclusion complexes with specific guests, an ability, which depends on a proper fit of the guest molecule into the hydrophobic cyclodextrin cavity. The most common pharmaceutical application of cyclodextrins is to enhance the solubility, stability, and bioavailability of drug molecules. Such kinds of ligand-receptor complexes can be used for different applications, e.g., for a transdermal therapeutic system (TTS) or in biofunctional textiles. The aim of this study was the investigation of the influence of the different cyclodextrins on the cell proliferation using HaCaT keratinocytes as an in vitro test system. Moreover, the study was performed to find harmless and nontoxic cyclodextrin concentrations for dermal applications. By means of different independent in vitro tests could be confirmed that alpha-, beta-, and gamma-cyclodextrins in concentrations up to 0.1% (w/v) do not show any antiproliferative influence on HaCaT keratinocytes. Sometimes even proliferative effects could be found. However, all used cyclodextrins (besides gamma-cyclodextrin and its derivatives) in concentrations of 0.5 and 1% (w/v), respectively, exert a cytotoxic influence on the proliferation of HaCaT keratinocytes. On the basis of these findings, the following rank order of cyclodextrins regarding their cytotoxicity was proposed: M-beta-CD > beta-CD > HP-beta-CD > alpha-CD > (gamma-CD). It could be confirmed that beta-CD and M-beta-CD trigger the activity of the effectors caspases -3 and -7. A significant increase of LDH release could be found for beta-CD and methyl-beta-CD in concentrations of 0.5 and 1% (w/v). The calculated cytotoxicity amounted 45 and 79%, respectively. The measurements of the interleukins IL-6 and IL-8 confirmed the findings of the proliferation assays as well as the LDH measurements. These findings may provide further rationale to the use of CDs in topical formulations for dermal and transdermal drug delivery or as raw material to functionalize textiles for medical applications.