Connexin26 Modulates Radiation-Induced Skin Damage by Regulating Chemokine CCL27 through MAPK Signaling.

Connexin26 (Cx26) plays an important role in ionizing radiation-induced damage, and CC chemokine ligand 27 (CCL27) regulates the skin immune response. However, the relationship between Cx26 and CCL27 in radiation-induced skin damage is unclear. After X-ray irradiation, clonogenic survival and micronucleus formation were assessed in immortalized human keratinocytes (HaCaT). Proteins in the mitogen activated protein kinase (MAPK) signaling pathway and CCL27-related proteins were detected by immunoblotting. HaCaTCx26-/- cells were constructed to verify the effects of Cx26 on CCL27 secretion. A mouse model was established to examine the expression of CCL27 and skin inflammation in vivo. The degree of skin injury induced by 6 MV of X rays was closely related to CCL27. The phosphorylation of ERK, p38 and NF-κB was significantly increased in irradiated cells. The secretion of CCL27 was significantly decreased in HaCaT wild-type cells relative to HaCaTCx26-/- cells. Whereas cell survival fractions decreased, and the micronuclei formation rate increased as a function of increasing X-ray dose in HaCaT cells, the opposite trend occurred in HaCaTCx26-/- cells. Our findings show that Cx26 likely plays a role in the activation of the MAPK and NF-κB/COX-2 signaling pathways and regulates the secretion of CCL27 in keratinocytes after X-ray radiation-induced skin damage.

Human chemokines as antimicrobial peptides with direct parasiticidal effect on Leishmania mexicana in vitro.

Chemokines and chemokine receptor-mediated effects are important mediators of the immunological response and cure in human leishmaniasis. However, in addition to their signalling properties for leukocytes, many chemokines have also been shown to act directly as antimicrobial peptides on bacteria and fungi. We screened ten human chemokines (CXCL2, CXCL6, CXCL8, CXCL9, CXCL10, CCL2, CCL3, CCL20, CCL27, CCL28) for antimicrobial effects on the promastigote form of the protozoan parasite Leishmania mexicana, and observed direct parasiticidal effects of several, CCL28 being the most potent. Damage to the plasma membrane integrity could be visualised by entrance of propidium iodide, as measured with flow cytometry, and by scanning electron microscopy, which showed morphological changes and aggregation of cells. The findings were in concordance with parasiticidal activity, measured by decreased mitochondrial activity in an MTT-assay. This is the first report of direct antimicrobial activity by chemokines on parasites. This component of immunity against Leishmania parasites identified here warrants further investigation that might lead to new insight in the mechanisms of human infection and/or new therapeutic approaches.

In vivo imaging of cutaneous T-cell lymphoma migration to the skin.

Cutaneous T-cell lymphoma (CTCL) is characterized by the accumulation of malignant CD4(+) T cells in the skin. Although the expression of adhesion molecules and chemokine receptors on CTCL cells has been studied extensively on ex vivo isolated cells, very little is known about the dynamics and mechanisms of CTCL trafficking in vivo. However, detailed knowledge of the molecular cues mediating CTCL migration may be used to interfere with their homing to the skin. We made use of real-time intravital epifluorescence video and two-photon microscopy to visualize malignant T cells from Sezary syndrome (SS), a leukemic variant of CTCL, in dermal microvessels in mouse ear skin. We found that SS cells rolled along dermal venules in a P-selectin- and E-selectin-dependent manner at ratios similar to CD4(+) memory T cells from normal donors. We furthermore show that the chemokine CCL17/TARC, but not CCL27/CTACK, was sufficient to induce the arrest of SS cells in the microvasculature. However, a combination of both chemokines was required to induce extravasation of SS cells. Together, our experiments delineate the molecular adhesion cascade operant in SS cell homing to the skin in vivo.

NK cells are migrated and indispensable in the anti-tumor activity induced by CCL27 gene therapy.

Natural killer (NK) cells have been demonstrated could play an important role in the treatment of a number of tumors in mice. In the present study, chemokine CCL27, which be considered only selectively chemoattracts cutaneous lymphocyte antigen positive memory T cells and Langerhans cells, firstly demonstrated that it could induce the accumulation of NK cells into tumor by the intratumoral injection of CCL27-encoding fiber-mutant vector, AdRGD-CCL27. Experiments using spleen cell fractionation and RT-PCR showed CCL27 receptor, mCCR10, was strongly expressed in NK cells, suggesting the accumulation of NK cells in tumor was attributed to chemoattractant activity of CCL27 itself. Moreover, the combination of AdRGD-CCL27 and AdRGD-IL-12 induced the synergistic anti-tumor activity via NK-dependent manner and induced more NK cells infiltration into tumor nodule than that induced by AdRGD-CCL27 alone or AdRGD-IL-12 alone.