Melanogenesis Is Directly Affected by Metabolites of Melatonin in Human Melanoma Cells.

Melatonin (N-acetyl-5-methoxytryptamine, MEL), its kynurenic (N1-acetyl-N2-formyl-5-methoxykynurenine, AFMK) and indolic derivatives (6-hydroxymelatonin, 6(OH)MEL and 5-methoxytryptamine, 5-MT) are endogenously produced in human epidermis. Melatonin, produced by the pineal gland, brain and peripheral organs, displays a diversity of physiological functions including anti-inflammatory, immunomodulatory, and anti-tumor capacities. Herein, we assessed their regulatory effect on melanogenesis using amelanotic (A375, Sk-Mel-28) and highly pigmented (MNT-1, melanotic) human melanoma cell lines. We discovered that subjected compounds decrease the downstream pathway of melanin synthesis by causing a significant drop of cyclic adenosine monophosphate (cAMP) level, the microphthalmia-associated transcription factor (MITF) and resultant collapse of tyrosinase (TYR) activity, and melanin content comparatively to N-phenylthiourea (PTU, a positive control). We observed a reduction in pigment in melanosomes visualized by the transmission electron microscopy. Finally, we assessed the role of G-protein-coupled seven-transmembrane-domain receptors. Obtained results revealed that nonselective MT1 and MT2 receptor antagonist (luzindole) or selective MT2 receptor antagonist (4-P-PDOT) did not affect dysregulation of the melanin pathway indicating a receptor-independent mechanism. Our findings, together with the current state of the art, provide a convenient experimental model to study the complex relationship between metabolites of melatonin and the control of pigmentation serving as a future and rationale strategy for targeted therapies of melanoma-affected patients.

Deciphering the Functional Role of RIPK4 in Melanoma.

The receptor-interacting protein kinase 4 (RIPK4) plays an important role in the development and maintenance of various tissues including skin, but its role in melanoma has not been reported. Using patient-derived cell lines and clinical samples, we show that RIPK4 is expressed in melanomas at different levels. This heterogenous expression, together with very low level of RIPK4 in melanocytes, indicates that the role of this kinase in melanoma is context-dependent. While the analysis of microarray data has revealed no straightforward correlation between the stage of melanoma progression and RIPK4 expression in vivo, relatively high levels of RIPK4 are in metastatic melanoma cell lines. RIPK4 down-regulation by siRNA resulted in the attenuation of invasive potential as assessed by time-lapse video microscopy, wound-healing and transmigration assays. These effects were accompanied by reduced level of pro-invasive proteins such as MMP9, MMP2, and N-cadherin. Incubation of melanoma cells with phorbol ester (PMA) increased PKC-1ß level and hyperphosphorylation of RIPK4 resulting in degradation of RIPK4. Interestingly, incubation of cells with PMA for short and long durations revealed that cell migration is controlled by the NF-κB signaling in a RIPK4-dependent (RIPK4high) or independent (RIPK4low) manner depending on cell origin (distant or lymph node metastasis) or phenotype (mesenchymal or epithelial).

The role of RIPK4 in epidermis physiology / Rola bialka RIPK4 w fizjologii naskórka.

Proteins which regulate morphogenesis of the epidermis ensure its proper construction and function and mutations or abnormal expression of those proteins impact epidermal function. One recently described protein is Receptor Interacting Serine/Threonine Kinase 4 (RIPK4). Mutations in RIPK4 cause the autosomal-recessive form of Bartsocas-Papas syndrome and Popliteal Pterygium Syndrome the Aslan type. In mice, deletion of Ripk4 (Ripk4-/-) leads to premature death of neonates caused by numerous skin adhesions. RIPK4 regulates development and maintenance of differentiation and proliferation homeostasis of keratinocytes, as well as inflammation. RIPK4 also appears to act as a tumor suppressor in skin, since expression is decreased in squamous skin carcinoma and inducible deletion in mice facilitates induction and growth of papillomas and squamous skin carcinomas in chemical carcinogenesis. The review describes the RIP family, the engagement of RIPK4 in differentiation of the epidermis and consequences resulting from its improper expression.

A melanin-independent interaction between Mc1r and Met signaling pathways is required for HGF-dependent melanoma.

Melanocortin 1 receptor (MC1R) signaling stimulates black eumelanin production through a cAMP-dependent pathway. MC1R polymorphisms can impair this process, resulting in a predominance of red phaeomelanin. The red hair, fair skin and UV sensitive phenotype is a well-described melanoma risk factor. MC1R polymorphisms also confer melanoma risk independent of pigment. We investigated the effect of Mc1r deficiency in a mouse model of UV-induced melanoma. C57BL/6-Mc1r+/+-HGF transgenic mice have a characteristic hyperpigmented black phenotype with extra-follicular dermal melanocytes located at the dermal/epidermal junction. UVB induces melanoma, independent of melanin pigmentation, but UVA-induced and spontaneous melanomas are dependent on black eumelanin. We crossed these mice with yellow C57BL/6-Mc1re/e animals which have a non-functional Mc1r and produce predominantly yellow phaeomelanin. Yellow C57BL/6-Mc1re/e-HGF mice produced no melanoma in response to UVR or spontaneously even though the HGF transgene and its receptor Met were expressed. Total melanin was less than in C57BL/6-Mc1r+/+-HGF mice, hyperpigmentation was not observed and there were few extra-follicular melanocytes. Thus, functional Mc1r was required for expression of the transgenic HGF phenotype. Heterozygous C57BL/6-Mc1re/+-HGF mice were black and hyperpigmented and, although extra-follicular melanocytes and skin melanin content were similar to C57BL/6-Mc1r+/+-HGF animals, they developed UV-induced and spontaneous melanomas with significantly less efficiency by all criteria. Thus, heterozygosity for Mc1r was sufficient to restore the transgenic HGF phenotype but insufficient to fully restore melanoma. We conclude that a previously unsuspected melanin-independent interaction between Mc1r and Met signaling pathways is required for HGF-dependent melanoma and postulate that this pathway is involved in human melanoma.

Tetraacetyl riboflavin derivative mediates caspase 3/7 activation via MAPK in A431 cells upon blue light influence.

An acetylated riboflavin derivative, 3-methyl-tetraacetyl riboflavin (3MeTARF), is a compound with high photostability and photophysical properties similar to riboflavin, including the ability to photogenerate singlet oxygen. In the present study, we compared the effects of irradiation on A431 cancer cells with blue LED light (438 nm) in the presence of 3MeTARF and riboflavin on MAPK phosphorylation, apoptosis, caspase 3/7 activation and PARP cleavage. We observed that photogenerated oxidative stress in this reaction activates MAPK by increasing phosphorylation of p38 and JNK proteins. Preincubation of cells with inhibitors specific for phosphorylation of p38 and JNK proteins (SB203580, SP600125), respectively, results in decreased caspase 3/7 activation and PARP cleavage. We showed that the tetraacetyl derivative more effectively activates MAPK and skin cancer cell death compared to riboflavin. These data, together with results of our previous study, support the hypothesis that 3MeTARF, of riboflavin, might be more useful and desirable as a compound for use in photodynamic oxidation processes, including its therapeutic potential.

RIPK4 downregulation impairs Wnt3A-stimulated invasiveness via Wnt/ß-catenin signaling in melanoma cells and tumor growth in vivo.

PURPOSE: The role of Wnt signaling in oncogenesis and drug resistance is well known. Receptor-interacting protein kinase (RIPK4) contributing to the increased activity of many signaling pathways, including Wnt/ß-catenin, may be an important target for designing new drugs for metastatic melanoma, but its role in melanoma is not fully understood. METHODS: We tested the effect of genetic manipulation of RIPK4 (CRISPR/Cas9) on xenograft growth. In addition, immunohistochemistry was used to detect active ß-catenin, Ki67 and necrosis in xenografts. Wnt signaling pathway activity was examined using Western blot and Top-Flash. The effect of RIPK4 knockout on melanoma cells in vitro stimulated Wnt3A on wound overgrowth, migration and invasion ability was then evaluated. RESULTS: Our study showed that CRISPR/Cas9-mediated RIPK4 knockout (KO) significantly reduced tumor growth in a mouse model of melanoma, particularly of WM266.4 cells. RIPK4 KO tumors exhibited lower percentages of Ki67+ cells as well as reduced necrotic area and decreased levels of active ß-catenin. In addition, we observed that RIPK4 knockout impaired Wnt3A-induced activation of LRP6 and ß-catenin, as manifested by a decrease in the transcriptional activity of ß-catenin in Top-Flash in both tested melanoma cell lines, A375 and WM266.4. Prolonged incubation (48 h) with Wnt3A showed reduced level of MMP9, C-myc, and increased SOX10, proteins whose transcription is also dependent on ß-catenin activity. Moreover, RIPK4 knockout led to the inhibition of scratch overgrowth, migration and invasion of these cells compared to their controls. CONCLUSION: RIPK4 knockdown inhibits melanoma tumor growth and Wnt3A stimulated migration and invasion indicating that RIPK4 might be a potential target for melanoma therapy.

The involvement of RIPK4 in TNF-α-stimulated IL-6 and IL-8 production by melanoma cells.

PURPOSE: The receptor-interacting protein kinase (RIPK4) has an oncogenic function in melanoma, regulates NF-κB and Wnt/ß-catenin pathways, and is sensitive to the BRAF inhibitors: vemurafenib and dabrafenib which lead to its decreased level. As its role in melanoma remains not fully understood, we examined the effects of its downregulation on the transcriptomic profile of melanoma. METHODS: Applying RNA-seq, we revealed global alterations in the transcriptome of WM266.4 cells with RIPK4 silencing. Functional partners of RIPK4 were evaluated using STRING and GeneMANIA databases. Cells with transient knockdown (via siRNA) and stable knockout (via CRISPR/Cas9) of RIPK4 were stimulated with TNF-α. The expression levels of selected proteins were assessed using Western blot, ELISA, and qPCR. RESULTS: Global analysis of gene expression changes indicates a complex role for RIPK4 in regulating adhesion, migration, proliferation, and inflammatory processes in melanoma cells. Our study highlights potential functional partners of RIPK4 such as BIRC3, TNF-α receptors, and MAP2K6. Data from RIPK4 knockout cells suggest a putative role for RIPK4 in modulating TNF-α-induced production of IL-8 and IL-6 through two distinct signaling pathways-BIRC3/NF-κB and p38/MAPK. Furthermore, increased serum TNF-α levels and the correlation of RIPK4 with NF-κB were revealed in melanoma patients. CONCLUSION: These data reveal a complex role for RIPK4 in regulating the immune signaling network in melanoma cells and suggest that this kinase may represent an alternative target for melanoma-targeted adjuvant therapy.

Low androgen/progesterone or high oestrogen/androgen receptors ratio in serous ovarian cancer predicts longer survival.

The treatment of ovarian cancer (OC) remains one of the greatest challenges in gynaecological oncology. The presence of classic steroid receptors in OC makes hormone therapy an attractive option; however, the response of OC to hormone therapy is modest. Here, we compared the expression patterns of progesterone (PGR), androgen (AR) and oestrogen alpha (ERα) receptors between serous OC cell lines and non-cancer ovarian cells. These data were analysed in relation to steroid receptor expression profiles from patient tumour samples and survival outcomes using a bioinformatics approach. The results showed that ERα, PGR and AR were co-expressed in OC cell lines, and patient samples from high-grade and low-grade OC co-expressed at least two steroid receptors. High AR expression was negatively correlated, whereas ERα and PGR expression was positively correlated with patient survival. AR showed the opposite expression pattern to that of ERα and PGR in type 1 (SKOV-3) and 2 (OVCAR-3) OC cell lines compared with non-cancer (HOSEpiC) ovarian cells, with AR downregulated in type 1 and upregulated in type 2 OC. A low AR/PGR ratio and a high ESR1/AR ratio were associated with favourable survival outcomes in OC compared with other receptor ratios. Although the results must be interpreted with caution because of the small number of primary tumour samples analysed, they nevertheless suggest that the evaluation of ERα, AR and PGR by immunohistochemistry should be performed in patient biological material to plan future clinical trials.

Functional melanocortin 1 receptor Mc1r is not necessary for an inflammatory response to UV radiation in adult mouse skin.

The G-protein-coupled receptor, Mc1r, plays a major role in pigment production and has been reported to be important in the inflammatory response. We have investigated the effect of deficiency in Mc1r on UV-induced inflammation. Mice on the same genetic background were used - C57BL/6-c (albino), C57BL/6 (black), C57BL/6-Mc1r(e/e) deficient (yellow). FACS analysis of disaggregated skin showed a similar dose-dependent increase in Ly6G(+) and CD11b(+) cells in response to UV radiation in all groups. No differences in UV-induced edema or in DNA damage were detected between groups. The contact hypersensitivity response, neonatal immune tolerance and UV immunosuppression were all similar in C57BL/6 and C57BL/6-Mc1r(e/e) mice. We conclude that the absence of Mc1r does not impair the inflammatory response to UV radiation or the generation of immunosuppression.

Dual Action of Curcumin as an Anti- and Pro-Oxidant from a Biophysical Perspective.

Curcumin, a natural polyphenol widely used as a spice, colorant and food additive, has been shown to have therapeutic effects against different disorders, mostly due to its anti-oxidant properties. Curcumin also reduces the efficiency of melanin synthesis and affects cell membranes. However, curcumin can act as a pro-oxidant when blue light is applied, since upon illumination it can generate singlet oxygen. Our review aims to describe this dual role of curcumin from a biophysical perspective, bearing in mind its concentration, bioavailability-enhancing modifications and membrane interactions, as well as environmental conditions such as light. In low concentrations and without irradiation, curcumin shows positive effects and can be recommended as a beneficial food supplement. On the other hand, when used in excess or irradiated, curcumin can be toxic. Therefore, numerous attempts have been undertaken to test curcumin as a potential photosensitizer in photodynamic therapy (PDT). At that point, we underline that curcumin-based PDT is limited to the treatment of superficial tumors or skin and oral infections due to the weak penetration of blue light. Additionally, we conclude that an increase in curcumin bioavailability through the using nanocarriers, and therefore its concentration, as well as its topical use if skin is exposed to light, may be dangerous.

Riboflavin and Its Derivates as Potential Photosensitizers in the Photodynamic Treatment of Skin Cancers.

Riboflavin, a water-soluble vitamin B2, possesses unique biological and physicochemical properties. Its photosensitizing properties make it suitable for various biological applications, such as pathogen inactivation and photodynamic therapy. However, the effectiveness of riboflavin as a photosensitizer is hindered by its degradation upon exposure to light. The review aims to highlight the significance of riboflavin and its derivatives as potential photosensitizers for use in photodynamic therapy. Additionally, a concise overview of photodynamic therapy and utilization of blue light in dermatology is provided, as well as the photochemistry and photobiophysics of riboflavin and its derivatives. Particular emphasis is given to the latest findings on the use of acetylated 3-methyltetraacetyl-riboflavin derivative (3MeTARF) in photodynamic therapy.

The Role of Singlet Oxygen in Photoreactivity and Phototoxicity of Curcumin.

Curcumin is a plant-derived yellow-orange compound widely used as a spice, dye and food additive. It is also believed to have therapeutic effects against different disorders. On the other hand, there are data showing its phototoxicity against bacteria, fungi and various mammalian cells. Since the mechanism of its phototoxic action is not fully understood, we investigated here the phototoxic potential of curcumin in liposomal model membranes and in HaCaT cells. First, detection of singlet oxygen (1 O2 ) luminescence proved that curcumin generates 1 O2 upon blue light irradiation in organic solvent and in liposomes. Then, HPLC-EC(Hg) measurements revealed that liposomal and cellular cholesterol is oxidized by 1 O2 photogenerated by curcumin. Enrichment of liposome membranes with curcumin significantly increased the oxygen photo-consumption rate compared to the control liposomes as determined by EPR oximetry. Cytotoxicity measurements, mitochondrial membrane potential analyses and protein hydroperoxides detection confirmed strong phototoxic effects of curcumin in irradiated HaCaT cells. These data show that since curcumin is advertised as a valuable dietary supplement, or a component of cosmetics for topical use, caution should be recommended especially when skin is exposed to light.

Vemurafenib and Dabrafenib Downregulates RIPK4 Level.

Vemurafenib and dabrafenib are BRAF kinase inhibitors (BRAFi) used for the treatment of patients with melanoma carrying the V600E BRAF mutation. However, melanoma cells develop resistance to both drugs when used as monotherapy. Therefore, mechanisms of drug resistance are investigated, and new molecular targets are sought that could completely inhibit melanoma progression. Since receptor-interacting protein kinase (RIPK4) probably functions as an oncogene in melanoma and its structure is similar to the BRAF protein, we analyzed the impact of vemurafenib and dabrafenib on RIPK4 in melanomas. The in silico study confirmed the high similarity of BRAF kinase domains to the RIPK4 protein at both the sequence and structural levels and suggests that BRAFi could directly bind to RIPK4 even more strongly than to ATP. Furthermore, BRAFi inhibited ERK1/2 activity and lowered RIPK4 protein levels in BRAF-mutated melanoma cells (A375 and WM266.4), while in wild-type BRAF cells (BLM and LoVo), both inhibitors decreased the level of RIPK4 and enhanced ERK1/2 activity. The phosphorylation of phosphatidylethanolamine binding protein 1 (PEBP1)-a suppressor of the BRAF/MEK/ERK pathway-via RIPK4 observed in pancreatic cancer did not occur in melanoma. Neither downregulation nor upregulation of RIPK4 in BRAF- mutated cells affected PEBP1 levels or the BRAF/MEK/ERK pathway. The downregulation of RIPK4 inhibited cell proliferation and the FAK/AKT pathway, and increased BRAFi efficiency in WM266.4 cells. However, the silencing of RIPK4 did not induce apoptosis or necroptosis. Our study suggests that RIPK4 may be an off-target for BRAF inhibitors.

Ultraviolet Radiation-Induced Mitochondrial Disturbances Are Attenuated by Metabolites of Melatonin in Human Epidermal Keratinocytes.

Melatonin (N-acetyl-5-methoxytryptamine) is recognized as an effective antioxidant produced by the pineal gland, brain and peripheral organs, which also has anti-inflammatory, immunomodulatory, and anti-tumour capacities. Melatonin has been reported as a substance that counteracts ultraviolet radiation B (UVB)-induced intracellular disturbances. Nevertheless, the mechanistic actions of related molecules including its kynurenic derivatives (N1-acetyl-N2-formyl-5-methoxykynurenine (AFMK)), its indolic derivatives (6-hydroxymelatonin (6(OH)MEL) and 5-methoxytryptamine (5-MT)) and its precursor N-acetylserotonin (NAS) are only poorly understood. Herein, we treated human epidermal keratinocytes with UVB and assessed the protective effect of the studied substances in terms of the maintenance of mitochondrial function or their radical scavenging capacity. Our results show that UVB caused the significant elevation of catalase (CAT) and superoxide dismutase (Mn-SOD), the dissipation of mitochondrial transmembrane potential (mtΔΨ), a reduction in ATP synthesis, and the enhanced release of cytochrome c into cytosol, leading subsequently to UVB-mediated activation of the caspases and apoptosis (appearance of sub-G1 population). Our findings, combined with data reported so far, indicate the counteracting and beneficial actions of melatonin and its molecular derivatives against these deleterious changes within mitochondria. Therefore, they define a path to the development of novel strategies delaying mitochondrial aging and promoting the well-being of human skin.

Pheomelanin in the skin of Hymenochirus boettgeri (Amphibia: Anura: Pipidae).

Pheomelanin is supposed to be the first type of melanin found in vertebrates, in contrast to the main type - eumelanin. Our study aimed at detecting pheomelanin in the skin of Hymenochirus boettgerii. We employed electron paramagnetic resonance (EPR) spectroscopy, and transmission electron microscopy (TEM), supplemented with standard histology and immunochemistry. We identified pheomelanin in the dorsal skin of adult frogs (not only in the dermis, but also in the epidermis) and in the dorsal tadpole. Our work identifies Hymenochirus boettgerii as a model in the basic study on the mechanism, evolution and role of melanogenesis in animals, including human.

Vitamin D endocrine system in breast cancer.

Vitamin D is a steroid hormone of great importance in the human body. It is produced in the skin from 7-dehydrocholesterol, upon UV radiation. In order to exert its functions, vitamin D has to be hydroxylated (via CYP27A1 and CYP27B1 hydroxylases), which is followed by its interaction with the vitamin D receptor (VDR) or retinoic acid-related orphan receptors α or γ (RORα and RORγ). By binding with the vitamin D response elements (VDRE) located in the promoter regions, the vitamin D ligand-receptor complex may regulate vitamin D-related genes. Recently, vitamin D has acquired a great interest for its plausible association with cancer development. This review discusses the potential role of vitamin D, its analogues, and enzymes involved in its metabolism with breast cancer incidence and outcome. According to the literature, alterations in the vitamin D endocrine system, both at the mRNA and protein level, have an impact on breast cancer incidence and prognosis. Moreover, specific enzymes participating in vitamin D metabolism may serve as therapeutic targets. Notably, treatment with vitamin D analogues also gives promising results in experimental research. However, given the fact that breast cancer is heterogenous disease, further studies are needed to thoroughly elucidate the potential of vitamin D and enzymes involved in its metabolism in breast cancer development, progression and therapy. Therefore, plausible effects of vitamin D in cancer therapy or prevention have been the principal aim of numerous studies.

Analysis of photoreactivity and phototoxicity of riboflavin's analogue 3MeTARF.

Recent studies focus on usage of blue light of λ = 450 nm in combination with photosensitizers to treat surface skin disorders, including cancers. In search of convenient therapeutic factor we studied riboflavin analogue 3-methyl-tetraacetylriboflavin (3MeTARF) as potential sensitizer. Riboflavin (Rfl) itself, non -toxic in the darkness, upon absorption of UVA and blue light, may act as photosensitizer. However, Rfl efficiency is limited due to its susceptibility to photodecomposition. Riboflavin's acetylated analogue, 3MeTARF, bears substituents in ribose chain, which inhibit intramolecular processes leading to degradation. Upon excitation, this compound, reveals higher photochemical resistance, remaining a good singlet oxygen generator. Thus, being more stable as the sensitizer, might be much more efficient in photodynamic processes. The objective of undertaken study was to elucidate mechanisms of 3MeTARF photoreactivity under the irradiation with blue light in comparison to its mater compound, riboflavin. We approached this goal by using spectroscopic methods, like direct singlet oxygen phosphorescence detection at 1270 nm, EPR spin trapping and oximetry. Additionally, we tested both riboflavin and 3MeTARF phototoxicity against melanoma cells (WM115) and we studied mechanism of photodynamic cell death, as well. Moreover, 3MeTARF induces apoptosis in melanoma cells at ten times lower concentration than riboflavin itself. Our studies confirmed that 3MeTARF remains stable upon blue light activation and is more efficient photosensitizer than Rfl.

Resveratrol enhances apoptosis induced by the heterocyclic aromatic amines in p53-wt LoVo cells, but not in p53-deficient HaCaT cells.

In the present study, we investigated the influence of resveratrol on PhIP treated human colon cancer cells and compared the effect to HaCaT cells considered as normal, human keratinocytes. Our results show that resveratrol decreases DNA damage in both cell types, it increases the sensitivity of LoVo cells to apoptosis and has no effect on PhIP-treated HaCaT cells. We confirm that PhIP-induced apoptosis is p53 and caspase 3/7 dependent. Interestingly, normal cells such as HaCaT, which lack functional p53 are more resistant to PhIP treatment.

Photodynamic activity of platinum(IV) chloride surface-modified TiO2 irradiated with visible light.

The visible light-induced phototoxicity of titanium dioxide modified with platinum(IV) chloride complexes, [TiO2/PtCl4], was tested. In vitro experiments with the mouse melanoma cells (S-91) have demonstrated phototoxicity of the [TiO2/PtCl4] material. Detection of efficiently generated various reactive oxygen species (.OH, O2. -, H2O2, 1O2) and also reactive chlorine species has proven the photodynamic activity of the tested material, induced by visible light (lambda>455 nm). The cellular death (recognized as a necrosis) is a result of the cell membrane peroxidation.

Deficient inflammatory response to UV radiation in neonatal mice.

Mechanisms of juvenile susceptibility to cancer are not well understood. The immune response in neonates favors nonresponsiveness or T(H)2-dominant responses, raising the question of a role for neonatal immunity in this susceptibility. We have investigated the postulate that the inflammatory response differs in neonatal and adult skin. We found no inflammatory infiltrate into neonatal mouse skin in response to UV irradiation as a function of time, dose, or wavelength, although UV-induced DNA damage was readily detected. In contrast, UV irradiation of adult mice initiated a dose- and time-dependent influx of inflammatory cells, chiefly CD11b(+)Ly6G(+) neutrophils, into the skin, detected by immunohistochemistry and quantitated by FACS analysis. This inflammatory response was initiated by UVB (290-320 nm) but not by UVA (320-400 nm). Further, in neonates, in contrast to adults, neither topical trinitrochlorobenzene (TNCB) nor i.p. thioglycollate initiated an inflammatory infiltrate. Conversely, topical TNCB applied to neonates was tolerogenic, resulting in a subsequent antigen-specific decrease of the contact-hypersensitivity response in adults. Neonatal blood contained abundant neutrophils, which exhibited impaired chemotaxis to the chemokine growth-related oncogene-alpha but efficient chemotaxis to the bacterial product fMLP, concomitant with decreased expression of CXCR2 but normal levels of CD11b. We propose this neonatal deficiency in the inflammatory response is a significant, previously unrecognized factor in neonatal immune tolerance and may contribute to neonatal susceptibility to cancer, including melanoma and other UV-induced cancers.