Cell-Density-Regulated Chemotactic Responsiveness of Keratinocytes In Vitro

Abstract

Keratinocytes represent the main constituents of the epidermis and have been found to play a regulatory role in a variety of inflammatory skin diseases. The functional activity of keratinocytes is highly heterogeneous, and depends on the cell localization in the epidermal architecture, and the maturation or differentiation state of the cells. Spontaneously proliferating HaCaT cells, showing several similarities to basal epidermal keratinocytes, were found to respond to external chemoattractants, including the chemokines RANTES (regulated on activation normal T cell expressed and secreted) and interleukin-8 and the mu-opioid agonist DAMGO ([d-ala2, N-Me-Phe4, Gly-ol5]enkephalin) in migration assays. The chemotactic responsiveness was highly dependent on the cell density of the monolayer, with greatest chemotactic activity at the highest cell density. Whereas RANTES was found to be the most potent chemoattractant, constitutive RANTES production was also detected in the HaCaT cultures. We found an inverse correlation between constitutive RANTES production and chemotactic responsiveness toward external RANTES, suggesting a possible functional down-modulation of the RANTES receptors, CC chemokine receptor 1 and CC chemokine receptor 5, during culture. Results from confocal laser scanning microscopy showed reduced CC chemokine receptor 1, but not CC chemokine receptor 5, expression by HaCaT cells at low cell densities, which was abolished in the presence of neutralizing antibodies against RANTES. The total CC chemokine receptor 1 pool (surface and intracellular receptors), however, showed no significant change during in vitro culture. Chemotactic responsiveness toward RANTES was directly correlated with the level of CC chemokine receptor 1 surface expression. Taken together these results show that with keratinocyte proliferation and the progressive increase in cell density there are dramatic alterations in keratinocyte function.

Low power millimeter wave irradiation exerts no harmful effect on human keratinocytes in vitro

“…We have previously reported that HaCaT keratinocytes produce constitutive RANTES in a cell-density-regulated manner. We have found that in the logarithmic growth phase of cultures the RANTES production graduately decreased [Szabo et al, 2001b]. In the present study we found that LP-MW irradiation of HaCaT keratinocytes did not significantly modulate constitutive RANTES production.…”

Millimeter wave induced reversible externalization of phosphatidylserine molecules in cells exposed in vitro

“…In order to study this hypothesis, experiments were carried out using the human origin HaCaT keratinocytes, developing multilayers in dense in vitro cultures, thus providing a sufficiently large cell surface area for plasma coagulation within the beam [Szabo et al, 2001]. HaCaT cultures in small Petri dishes (35 Â 10 mm; equivalent dimensions with wells in a sixwell tissue culture plate used for blood exposures) were cooled to 4 8C and covered by cold human plasma before exposure with MMW.…”

Modulation of Peripheral Sensory Neurons by the Immune System: Implications for Pain Therapy

“…In addition, opioids have mitogenic properties in the regeneration of mucosa (Cho et al, 2003), promote reepithelialization and keratinocyte migration (Szabo et al, 2001;Bigliardi et al, 2002;Kü chler et al, 2010), and up-regulate cytokeratin and transforming growth factor-␤ receptors, both important components in wound healing (Tegeder and Geisslinger, 2004; for review, see Bigliardi et al, 2009). In ischemic wounds, locally applied opioids hastened wound closure increased granulation tissue and collagen formation, increased epidermal and dermal organization, and enhanced angiogenesis (Poonawala et al, 2005;Gross et al, 2009).…”