Photodynamic activity of Temoporfin nanoparticles induces a shift to the M1-like phenotype in M2-polarized macrophages.

The monocyte/macrophage cell lineage reveals an enormous plasticity, which is required for tissue homeostasis, but is also undermined in various disease states, leading to a functional involvement of macrophages in major human diseases such as atherosclerosis and cancer. We recently generated in vivo evidence that crystalline, nonfluorescent nanoparticles of the hydrophobic porphyrin-related photosensitizer Aluminum phthalocyanine are selectively dissolved and thus may be used for specific fluorescent labelling of rejected, but not of accepted xenotransplants. This led us to hypothesize that nanoparticles made of planar photosensitizers such as porphyrins and chlorins were preferentially taken up and dissolved by macrophages, which was verified by in vitro studies. Here, using an in vitro system for macrophage differentiation/polarization of the human monocyte THP-1 cell line, we demonstrate differential uptake/dissolution of Temoporfin-derived nanoparticles in polarized macrophages, which resulted in differential photosensitivity. More importantly, low dose photodynamic sensitization using Temoporfin nanoparticles can be used to trigger M1 re-polarization of THP-1 cells previously polarized to the M2 state. Thus, sublethal photodynamic treatment using Temoporfin nanoparticles might be applied to induce a phenotypic shift of tumor-associated macrophages for the correction of an immunosuppressive microenvironment in the treatment of cancer, which may synergize with immune checkpoint inhibition.

Radiation-induced late effects in two affected individuals of the Lilo radiation accident.

Radiation exposure leads to a risk for long-term deterministic and stochastic late effects. Two individuals exposed to protracted photon radiation in the radiological accident at the Lilo Military site in Georgia in 1997 received follow-up treatment and resection of several chronic radiation ulcers in the Bundeswehr Hospital Ulm, Germany, in 2003. Multi-parameter analysis revealed that spermatogenetic arrest and serum hormone levels in both patients had recovered compared to the status in 1997. However, we observed a persistence of altered T-cell ratios, increased ICAM1 and beta1-integrin expression, and aberrant bone marrow cells and lymphocytes with significantly increased translocations 6 years after the accident. This investigation thus identified altered end points still detectable years after the accident that suggest persistent genomic damage as well as epigenetic effects in these individuals, which may be associated with an elevated risk for the development of further late effects. Our observations further suggest the development of a chronic radiation syndrome and indicate follow-up parameters in radiation victims.

Bath PUVA and saltwater baths followed by UV-B phototherapy as treatments for psoriasis: a randomized controlled trial.

OBJECTIVE: To evaluate the efficacy of psoralens dissolved in a warm-water bath followed by exposure to UV-A irradiation (bath PUVA) or saltwater phototherapy (SW UV-B) compared with tap-water phototherapy (TW UV-B) or UV-B irradiation alone in psoriasis. DESIGN: Multisite, prospective, randomized, controlled trial with 4 parallel groups. SETTING: Total of 102 dermatologic outpatient clinics. PATIENTS: Total of 1241 patients with stable psoriasis vulgaris and a Psoriasis Area and Severity Index score of 7 or greater. INTERVENTIONS: Four-times-weekly UV-B, TW UV-B, SW UV-B, or bath-PUVA with baths preceding UV irradiation over a maximum of 8 weeks. The UV dose was adapted to erythemal response. MAIN OUTCOME MEASURES: Incidence of therapeutic success, defined as a reduction of the Psoriasis Area and Severity Index or affected body surface area of 50% or more. RESULTS: Patients who received TW UV-B had a significantly higher incidence of therapeutic success than did patients treated with UV-B alone (60.7% vs 43.3%; P<.001; number needed to treat, 5.8; 95% confidence interval [CI], 3.9-10.9). Patients who received SW UV-B or bath PUVA had a significantly higher incidence of therapeutic success than did patients treated with TW UV-B (74.9% vs 60.7%; P<.001; number needed to treat, 7.0; 95% CI, 4.6-14.9; and 78.4% vs 60.7%; P<.001; number needed to treat, 5.7; 95% CI, 4.0-9.7, respectively). Bath PUVA was not superior to SW UV-B (78.4% vs 74.9%; P = .34). CONCLUSION: Bath PUVA and SW UV-B are comparably effective treatments in psoriasis and superior to UV-B and TW UV-B.

A pragmatic randomized controlled trial on the effectiveness of highly concentrated saline spa water baths followed by UVB compared to UVB only in moderate to severe psoriasis.

BACKGROUND: There is a lack of sufficiently large randomized trials evaluating the effectiveness of saline spa balneophototherapy compared to ultraviolet B (UVB) only. OBJECTIVE: The study aimed to evaluate whether highly concentrated saline spa water baths followed by UVB (HC-SSW-UVB) are superior to UVB only in moderate to severe psoriasis. METHODS: One hundred and sixty (160) adults with a Psoriasis Area and Severity Index (PASI) of >10 from 4 German spa centers were randomly allocated to HC-SSW-UVB (local sodium chloride concentration between 25% and 27%) or UVB only 3 a week until remission (PASI < 5) or for a maximum of 6 weeks. Reduction of PASI > or = 50% (PASI-50) at the end of the intervention period was defined as primary outcome. Only persons receiving at least 1 intervention were included into the primary analysis. RESULTS: Participants allocated to HC-SSW-UVB attained to a statistically significantly higher rate of PASI-50 than patients allocated to UVB only (68/79 [86%] versus 38/71 [54%]; p < 0.001; number needed to treat, 3.1; 95% confidence interval, 2.1-6.0). Postintervention analysis did not yield a clear hint of a persisting effect. CONCLUSIONS: The study indicates that HC-SSW-UVB are superior to routine UVB at the end of a 6-week treatment course.

Numerical abnormalities of the Cyclin D1 gene locus on chromosome 11q13 in non-melanoma skin cancer.

Deregulation of the cell-cycle G1-restriction point control via abnormalities of Rb-pathway components is a frequent event in the formation of cancer. The aim of this study was to evaluate numerical aberrations of the Cyclin D1 (CCND1, PRAD1, bcl-1) gene locus at chromosome 11q13 in basal cell carcinomas (BCCs) and squamous cell carcinomas (SCCs) of the skin and to compare it with the Cyclin D1 protein expression. Fluorescence in situ hybridization with DNA-probes specific for the Cyclin D1 gene locus and the centromere of chromosome 11 as well as immunostaining for Cyclin D1 protein was applied on 5 microm serial paraffin sections. Six of the 30 (20%) SCCs showed additional Cyclin D1 gene copies and 2/30 (6.6%) cases had a loss of the Cyclin D1 gene locus in relation to the centromere 11 number. In contrast, only one of the 14 BCCs (7%) showed one additional Cyclin D1 gene copy in relation to the centromere 11 number. None of the BCCs demonstrated aneusomy for chromosome 11 in contrast to SCCs, where it was found in 21/30 (70%) cases. Twenty-six of the 30 (86.6%) cutaneous SCCs and 13/14 (93%) BCCs expressed Cyclin D1 protein. All SCCs and the BCC with additional Cyclin D1 gene copies showed positivity for Cyclin D1 protein. Both SCCs with less Cyclin D1 gene copies than centromere 11 signals showed a weak protein expression. Our findings suggest that numerical abnormalities of the Cyclin D1 gene locus could result in an altered gene-dose effect, possibly leading to an aberrant expression in affected tumor cells. This might result in deregulation of cell cycle control, eventually leading to uncontrolled cell cycle progression.

Additional Cyclin D(1) gene copies associated with chromosome 11 aberrations in cutaneous malignant melanoma.

Cyclin D1 (CCND1, PRAD1, bcl-1) is associated with progression through G1 and entry into S phase of the cell cycle, as a partner of cyclin-dependent kinases (CDKs). The aim of this study was to evaluate cytogenetic aberrations of the gene locus at chromosome 11q13 in cutaneous malignant melanoma. Fluorescence in situ hybridisation (FISH) was applied on metaphase spreads of short-term primary cell cultures as well as on 5 microm paraffin tissue sections of primary melanomas and melanoma metastases, and on melanoma cell lines (C32, WM 266-4, HT 144, RPMI 7951, SK MEL 28). For FISH analysis DNA probes specific for Cyclin D1, centromere 11 and painting probes for whole chromosome 11 were used. FISH revealed additional Cyclin D1 gene copies in relation to centromere 11 copy numbers in 9/19 (47%) primary melanomas and in 7/20 (35%) melanoma metastases. The melanoma cell line SK MEL 28 also showed extra Cyclin D1 gene copies. A 24-colour FISH (mFISH) investigation revealed translocated chromosome 11 material to chromosomes 12, 14, 15, 20, 21, 22 and Y in these cases, respectively. Positivity for Cyclin D1 protein was detected by immunostaining in 17/19 (89%) primary melanomas (10 weak, 7 strong expression) and in 9/12 (75%) melanoma metastases (5 weak and 4 strong). All primary melanomas and 3/4 examined metastases with additional Cyclin D1 gene copies concomitantly revealed positive protein staining. Based on these results Cyclin D1 could well be involved in the pathogenesis of a subset of malignant melanoma. Additional studies will have to elucidate the role of further genes located at chromosome 11q13.

Fibroblast activation protein: differential expression and serine protease activity in reactive stromal fibroblasts of melanocytic skin tumors.

Growth and metastasis of solid neoplasms require the recruitment of a supporting tumor stroma. A highly consistent trait of tumor stromal fibroblasts in most epithelial cancers is the induction of fibroblast activation protein (FAP), a member of the serine protease family. Recently it was demonstrated that FAP has both dipeptidyl peptidase and collagenolytic activity capable of degrading gelatin and type I collagen. In this study, we describe the expression and enzyme activity of FAP in benign and malignant melanocytic skin tumors. FAP-positive fibroblasts were detected immunohistochemically in the reactive stroma of all melanocytic nevi tested. In primary and metastatic melanomas an upregulation of FAP expression in the reactive mesenchyme could be observed. Whereas 30% of the nevi revealed additional FAP expression on subsets of melanocytic cells, melanoma cells from primary and metastatic melanomas were FAP negative. This may indicate a possible role for FAP in the control of tumor cell growth and proliferation during melanoma carcinogenesis. Consistent with this in vivo expression pattern FAP enzyme activity could be detected by a specific immunocapture assay in extracts of melanocytic nevi and melanoma metastases, whereas no significant activity was detectable in normal adult skin. Strong protein expression of FAP was observed in patterned structures restricted to a subset of the melanoma metastases. Our findings that these FAP-positive structures showed no overlap with endothelial cell surface markers, nor with various melanoma antigens, suggest that FAP is a marker for specific stromal-cell-derived patterns in cutaneous melanoma metastases.

Management of cutaneous radiation injuries: diagnostic and therapeutic principles of the cutaneous radiation syndrome.

The cutaneous symptoms that appear after radiation exposure are caused by a combination of inflammatory processes and alteration of cellular proliferation as a result of a specific pattern of transcriptionally activated proinflammatory cytokines and growth factors. The symptoms follow a time course consisting of prodromal erythema, manifestation, chronic stage, and late stage; these symptoms are referred to as cutaneous radiation syndrome (CRS). The time course depends on several factors such as the applied radiation dose, radiation quality, individual radiation sensitivity, extent of contamination and absorption, and volume of skin exposed. For diagnosis of CRS, the following procedures are used: 7.5 to 20 MHz B-scan sonography, thermography, capillary microscopy, profilometry, nuclear magnetic resonance imaging, bone scintigraphy, and histology. Based on the results of previous experimental and clinical research, pharmacotherapy of CRS can include topical or systemic application of corticosteroids, gamma interferon, pentoxifylline and vitamin E, and superoxide dismutase. The treatment varies according to the stage of CRS. Due to the complexity of the clinical manifestations of radiation disease in most patients, interdisciplinary treatment at specialized centers is necessary. In most cases, dermatologists are asked to provide lifelong therapy and follow-up of the patients.

Thyroid examination in highly radiation-exposed workers after the Chernobyl accident.

CONTEXT: Radioactive contamination from the Chernobyl nuclear accident that happened on the morning of 26th April 1986 had a major impact on thyroid health in the Belarus region. OBJECTIVE: Observational study of a cohort of 99 adults, most strongly exposed to ionizing radioactivity. DESIGN, SETTING AND PATIENTS: Observational study performed between 1998 and 2000. The cohort comprised 99 workers (92 male) of the Chernobyl nuclear power plant. Examination including physical examination, ultrasonography of the thyroid gland and measurement of serum free thyroxin (fT(4)), free triiodothyronine (fT(3)) and TSH. Anti-thyroperoxidase (anti-TPO), antithyroglobulin (anti-Tg) antibodies and thyroid stimulating immunoglobulin were also determined. MAIN OUTCOME MEASURES: The impact of exposure to high-dose radiation, including radioactive iodine, on the thyroid gland was examined. RESULTS: Levels of fT(4) in all probands were within the normal World Health Organization-defined range. Elevated levels of fT(3) were found in two workers (2%), high titres of anti-TPO and anti-Tg antibodies were present in four subjects (4%). Mild hypothyroidism was present in one patient. Enlargement of the thyroid gland was observed in 17 workers (17%). There was no evidence of clinically overt thyroid cancer. CONCLUSIONS: The Chernobyl accident showed surprisingly little impact on the thyroid in a cohort of workers strongly exposed to radiation. Our data suggest an age-dependent heterogeneity in response to the short-lived radioiodine isotopes and favours long-term follow-up analysis.

The effect of amelogenins (Xelma) on hard-to-heal venous leg ulcers.

With an aging population venous ulceration is likely to become an increasing problem. Despite improvements in care and the widespread introduction of compression bandaging, the mainstay of current management, a significant proportion of venous leg ulcers remain hard to heal. Therefore, a single-blinded, randomized multicenter study was performed to compare wound size reduction using amelogenin proteins (Xelma) formulated into a solution which forms a temporary extracellular matrix on contact with the wound bed. Propylene glycol alginate 7% served as a control. Patients were randomized to receive either amelogenin protein or control treatment. The investigational products were applied weekly under soft silicone secondary dressings for up to a maximum of 12 weeks. Compression therapy was maintained throughout the investigation. Wound size reduction was measured by tracing and all wounds were photographed. In total 123 patients were recruited, 62 patients in the amelogenin group, and 61 in the control group, respectively. Subgroup analyses were performed for ulcers with a size>10 cm2 at baseline and for ulcers of duration of >12 months. The wound size reduction was greatest in the group treated with amelogenin (33.8 vs. 25.6%, n=117), this difference being greatest for larger ulcers (25 vs. 7.9% for ulcers>10 cm(2), n=61) and those of long duration (29.3 vs. 10.9% for ulcers>12-month duration, n=61). We conclude that this product may be clinically useful in the treatment of these venous leg ulcers.

Expression of stromal cell markers in distinct compartments of human skin cancers.

BACKGROUND: The importance of changes in the supporting tumor stroma for cancer initiation and progression is well established. The characteristics of an activated tumor stroma, however, are not completely understood. In an effort to better characterize the desmoplastic response to human skin tumors, we evaluated the expression pattern of three stromal cell markers, fibroblast-activation protein (FAP), endoglyx-1, and endosialin, in a series of melanocytic and epithelial skin tumors. METHODS: Immunohistochemistry using antibodies against FAP, endoglyx-1, and endosialin was carried out in skin samples obtained from 43 patients. Furthermore, microarray data from an independent set of human skin cancers were analyzed. RESULTS: FAP-positive fibroblasts were detected in all tumor tissues tested, including cases of melanocytic nevi, melanoma metastases, basal cell carcinomas, and squamous cell carcinomas. In cutaneous melanoma metastases, we identified different compartments within the stromal response on the basis of the regions of FAP expression. Endoglyx-1 expression was confined to normal and tumor blood vessel endothelium including 'hot spots' of neoangiogenesis within the cutaneous melanoma metastases. Endosialin was selectively induced in subsets of small- and medium-sized tumor blood vessels in melanoma metastases and squamous cell carcinomas. CONCLUSIONS: These data describe novel aspects of stromal marker expression in distinct compartments of human skin tumors and may point to potential targets for novel therapeutic strategies aimed at the tumor stroma.

PUVA-gel vs. PUVA-bath therapy for severe recalcitrant palmoplantar dermatoses. A randomized, single-blinded prospective study.

BACKGROUND/PURPOSE: In order to avoid unwanted effects of systemic psoralen and ultraviolet A (PUVA) therapy, various topical PUVA treatment modalities have been developed and are being increasingly used. However, up to now very few controlled studies comparing the therapeutic efficacy of different topical photochemotherapy modalities are available. Thus, the aim of our study was to compare the clinical efficacy of conventional PUVA-bath therapy to topical PUVA-gel therapy in patients with recalcitrant dermatoses of the palms and soles. METHODS: Twenty patients with severe palmoplantar dermatoses or localized psoriatic plaques were enrolled in our observer-blinded, randomized half-sided study. The treatment modalities compared were: (i) aqueous 8-methoxypsoralen (8-MOP)-containing gel plus broadband UVA irradiation (PUVA-gel therapy) and (ii) 8-MOP bath of the hands and/or feet plus broadband UVA (PUVA-bath therapy). RESULTS: On the body half, which was randomized to PUVA-gel therapy, the median Area and Severity Index for palmoplantar dermatoses (ASIppd) decreased from 28 (range 6-56) to 1.5 (range 1-37, P = 0.00) after a median 33 (13-49) irradiations compared with a reduction from 26.5 (range 6-52.5) to 1.5 (range 0-38, P = 0.00) for PUVA-bath therapy. Both improvements of ASIppd scores were found to be statistically significant, with no significant difference between PUVA-gel and PUVA-bath therapy. Severe phototoxic reactions such as strong erythema, blistering and/or pain were not observed in any patient. CONCLUSION: PUVA-gel therapy seems to be an effective therapeutic alternative to conventional PUVA-bath therapy in treating localized dermatoses of the palms and soles. The advantage of PUVA-gel therapy is reduced organizational efforts and expenses.

The IKK-2/Ikappa Balpha /NF-kappa B pathway plays a key role in the regulation of CCR3 and eotaxin-1 in fibroblasts. A critical link to dermatitis in Ikappa Balpha -deficient mice.

Tumor necrosis factor (TNF)-alpha-induced phosphorylation of the IkappaB proteins by the IkappaB kinase (IKK) complex containing IKK-2 and subsequent degradation of the IkappaB proteins are prerequisites for NF-kappaB activation, resulting in the stimulation of a variety of pro-inflammatory target genes. The C-C chemokine eotaxin-1 is a potent chemoattractant for eosinophils and Th2 lymphocytes, may play an important role in the pathogenesis of atopic dermatitis, and acts via binding to its receptor CCR3. To investigate the role of NF-kappaB signaling in the regulation of these genes, we stably expressed a transdominant mutant of IkappaBalpha and a constitutively active mutant of IKK-2 in mouse NIH3T3 fibroblasts. The transdominant IkappaBalpha mutant completely inhibited TNF-alpha-mediated induction of both eotaxin-1 and CCR3, whereas expression of constitutively active IKK-2 was sufficient to drive almost full expression of these two genes in the absence of TNF-alpha. Moreover, we observed elevated expression levels of CCR3 and eotaxin-1 protein levels in the skin of IkappaBalpha-deficient mice characterized by a widespread dermatitis. Finally, using dermal fibroblasts derived from IkappaBalpha-deficient mice, we observed elevated basal expression, enhanced inducibility by TNF-alpha, and attenuated down-regulation upon TNF-alpha withdrawal of both CCR3 and eotaxin-1 mRNA levels. These results demonstrate that the IKK-2/IkappaBalpha/NF-kappaB pathway plays a critical role for CCR3 and eotaxin-1 expression in fibroblasts and suggests a critical link to the pathogenesis of atopic dermatitis.

Expression of c-myc and bcl-2 in primary and advanced cutaneous melanoma.

Apoptosis is an important co-factor in the pathogenesis of a plethora of malignancies. Enhanced c-myc activation can result either in proliferation or apoptosis. Coexpression with antiapoptotic bcl-2, which abrogates the apoptotic function of c-myc might lead to an enormous growth advantage of cells. In order to elucidate the role of c-myc and bcl-2 as well as the coexpression of both genes in human melanoma, their expression was studied in four samples of normal skin (SK), 15 surgical margins (SM), 20 benign melanocytic nevi (MN), 20 primary melanomas (MM), and 30 melanoma metastases (MMET) by RT-PCR. These results were compared with immunohistochemistry (IH) in 7 SK, 7 SM, 26 MN, 50 MM, and 34 MMET. Similar results were found with both methods. However, MMET expressed c-myc (PCR 28/30, IH 23/34) as well as bcl-2 (PCR 27/30, IH 24/34) more frequently. Primary melanomas showed a similar expression pattern as SM and nevi. Moreover, in contrast to SK, SM, MN, and MM coexpression of bcl-2 and c-myc was found more frequently in MMET (PCR 25/30, p < 0.01, IH 19/34, p < 0.01). These results indicate that coexpression of c-myc and bcl-2 appears to be associated with advanced melanoma and contributes to the malignant phenotype.

Can the immersion time of PUVA bath therapy be shortened?

Up to now, there are only a few data available concerning the influence of bathing time on skin phototoxicity. We compared the erythemal responses of normal skin to bath PUVA with 8-methoxypsoralen (8-MOP) after 5, 10 and 20 min immersion time. Currently, 20 min is the routinely performed immersion time in many European countries, including Germany, while in other countries bathing times are shorter. The minimal phototoxic dose (MPD) following immersion times of 5 min and 10 min in a warm water bath (37 degrees C) containing 1 mg/l 8-MOP was compared to the MPD following 20 min immersion time in a half-sided manner in a total of 24 patients. Our results revealed that an immersion time of 5 min did not yield a detectable erythema after 72 h. In contrast, both 10 and 20 min PUVA baths induced visible erythemas with a significantly higher median MPD following 10 min immersion (2.25 J/cm2) compared to 20 min baths (1.5 J/cm2). As an erythemal response of 8-MOP PUVA bath seems reduced after shorter immersion times, comparative studies on the clinical efficacy using shorter time regimens have to be conducted before conclusive recommendations for clinical PUVA-bathing time can be given.

Psoralen plus ultraviolet-A-bath photochemotherapy as an adjunct treatment modality in cutaneous chronic graft versus host disease.

BACKGROUND: Cutaneous chronic graft versus host disease (GVHD) is a severe complication following allogeneic stem cell (PBSCT) and bone marrow transplantation (BMT). Immunosuppressive therapy consists of prednisone, cyclosporine-A, azathioprine or mycophenolate mofetil (MMF). Treatment of patients refractory to immunosuppression represents a major problem. METHODS: We report six patients suffering from severe chronic GVHD of the skin who did not respond to immunosuppressive therapy or relapsed after reduction of glucocorticosteroids. Patients were treated with psoralen plus ultraviolet (PUVA)-bath photochemotherapy three times weekly following a standardized treatment protocol under continued treatment with prednisone and/or MMF. One patient was additionally pretreated with ultraviolet-A1 (UV-Al). RESULTS: After a median of 14, 5 treatment sessions, skin lesions improved. Out of six patients, three showed a complete remission. In all patients, systemic immunosuppressive therapy could be reduced. In sclerodermic lesions, skin thickness returned to the levels of normal skin after 25 treatments confirmed by 20 MHz ultrasound evaluation. In a follow-up ranging from 2 to 21 months (median 10, 3 months), skin conditions remained stable. CONCLUSION: Psoralen plus ultraviolet-A-bath represents an effective adjunct treatment option for extensive chronic and sclerodermic cutaneous GVHD offered by dermatologists. This is of high interest in patients suffering from cutaneous GVHD resistant to conventional immunosuppressive therapy and should be included to the menu of topical treatment options for chronic cutaneous GVHD.

Delayed effects of accidental cutaneous radiation exposure: fifteen years of follow-up after the Chernobyl accident.

BACKGROUND: During the Chernobyl accident in 1986, 237 individuals were identified as having the most severe exposure to ionizing radiation. In the period between 1998 and 2000, 99 long term survivors out of this group were reassessed for radiation-induced cutaneous lesions. OBJECTIVE: To identify sequelae of accidental cutaneous irradiation. METHODS: Detailed dermatologic examinations, including biopsies of suspicious cutaneous lesions for histopathologic examination and 20 MHz sonography, were performed in all patients. RESULTS: Twenty-two of the 99 patients displayed radiation-induced cutaneous lesions. Epidermal atrophy, telangiectases, and pigment alterations were present in all these individuals. Keratotic lesions were found in 14 patients. Cutaneous fibrosis was documented in 8 individuals by the use of 20 MHz sonography, while a radiation ulcer was found in 5. In one patient, two basal cell carcinomas were found. CONCLUSION: The life-long follow-up of irradiated persons is of great importance in order to identify cutaneous neoplasms at an early treatable stage.