[Intralesional Cryosurgery in the Treatment of Keloids - A Differentiation of Keloid Types for the Improvement of Patient Selection]. / Keloidbehandlung mit intraläsionaler Kryochirurgie.

The overproduction of altered collagen fibers and the overexpression of Tumor Growth Factor-ß must be blocked in order to interrupt the growth process within a keloid scar. This can barely be achieved with the classical therapeutic methods. The results of keloid treatment are difficult to predict and the recurrence rate is usually over 50 %. In addition, some of the procedures used (e. g. irradiation) may induce additional health risks. Intralesional cryosurgery offers a therapeutic alternative that has been evaluated since more than a decade. Our own experience in more than one thousand keloid treatments allows a critical evaluation of the classification in those keloid types, which are recommended to be treated with the technique and those, which may not respond.

A prospective case-control study of non-healing wounds of the lower limbs - the value of biopsies for ulcerating carcinoma.

BACKGROUND: In some leg ulcer patients there is cancer that is responsible for lack of healing of such a wound. AIM: This study was aimed at prospective analysis of histopathology of non-healing wounds (NHWs) in the patient presenting with high and low suspicion for ulcerating carcinoma. MATERIAL AND METHODS: Forty patients with NHWs were enrolled and had been prospectively divided into two groups: 25 patients with high suspicion for ulcerating carcinoma according to their medical history and physical examination, and the second group of 15 patients without suspicion for malignancy (control group). All NHWs were photographed and underwent biopsies. RESULTS: In the control group biopsies did not reveal cancers. On the contrary, in 10 patients (40%) from high suspicion group biopsies revealed cancers: seven basal cell carcinomas (BCCs), one - malignant melanoma, one - Bowen's disease and one - squamous cell carcinomas. Histopathology of six of seven BCCs suggested that non-healing benign wound might have preceded malignancy. We found that leg ulcers which were small (wound area less than 3 cm(2) ), longstanding (duration 24 ≤ weeks), presenting with granulation tissue covering ≥75% of the wound area, with a dull pink appearance of the granulation tissue, or an atypical clinical presentation, can actually be an ulcerating carcinoma. Dull pink granulation tissue or an atypical clinical presentation of ulceration, as a single clinical finding, suggested an underlying malignancy with a statistical significance (71.5% vs. 0%; P = 0.001 and 27.8% vs. 0%; P = 0.0049 respectively). CONCLUSIONS: Prevalence of malignancy, primarily: BCCs in NHWs, may be higher than expected and clinical features suggestive of such a nature of ulcer are an indication for diagnostic biopsy.

Fractal properties of macrophage membrane studied by AFM.

Complexity of cell membrane poses difficulties to quantify corresponding morphology changes during cell proliferation and damage. We suggest using fractal dimension of the cell membrane to quantify its complexity and track changes produced by various treatments. Glutaraldehyde fixed mouse RAW 264.7 macrophage membranes were chosen as model system and imaged in PeakForce QNM (quantitative nanomechanics) mode of AFM (atomic force microscope). The morphology of the membranes was characterized by fractal dimension. The parameter was calculated for set of AFM images by three different methods. The same calculations were done for the AFM images of macrophages treated with colchicine, an inhibitor of the microtubule polymerization, and microtubule stabilizing agent taxol. We conclude that fractal dimension can be additional and useful parameter to characterize the cell membrane complexity and track the morphology changes produced by different treatments.

Pain evaluation and control during and following the treatment of hypertrophic scars and keloids by contact and intralesional cryosurgery--a preliminary study.

BACKGROUND: Intralesional cryosurgery effectively treats hypertrophic scars and keloids (HSK), but pain experienced by the patient during treatment can limit the application of cryosurgery. OBJECTIVES: To characterize the pain response during cryosurgical treatment of HSK, and to evaluate the pain experienced during contact and intralesional cryosurgery that employs a pain-control protocol. METHODS: Twenty-nine patients (17 women, 12 men) aged 17 years and older (mean ages 31.9±12.5 and 38.9±18.6 years, respectively, P=0.24), who were treated for a total of 36 HSKs by intralesional (n=20; 22 cryotreatments) or contact (n=9; 14 cryotreatments) cryosurgery were evaluated. The pain-control protocol involved oral pain-relief tablets (Dipyrone) and translesional local anaesthesia with Bupivacaine hydrochloride 0.5%. Pain evaluation according to the Visual Analogue Scale (VAS) (0-10 cm) was compared between the two groups at three time points: during cryosurgery, immediately after it, and 4 h later. Scores ≤3 cm were considered to define the 'zone of analgesic success'. These results were compared with control data (contact cryosurgery without a pain-control protocol; n=56). RESULTS: Pain in the intralesional group was significantly lower than that in the contact group during and immediately after cryotreatment. During: mean VAS=1.68±2.21 vs. 5.07±4.01 cm; median VAS=0.5 vs. 5.5 cm, respectively; P<0.0001. Immediately after: mean VAS=1.22±1.77 vs. 5.38±3.81 cm; median VAS=0 vs. 6.0 cm, respectively; P=0.001. The control group had more pain during treatment (mean VAS=5.34±2.31, median=6.0) and 4 h later (mean=3.79±2.35, median=4.0) than the intralesional group (P<0.0001 and P=0.988, respectively). The pain level in the control group during the cryotreatment did not differ from that in the contact group (P=0.988). In the intralesional, contact and control groups analgesic success (VAS ≤3 cm) was achieved in 77.3%, 35.7% and 33.9%, respectively, of cases (P=0.002) during cryotreatment, and in 54.5%, 42.9% and 33.9%, respectively, of cases 4 h after treatment (P=0.24). CONCLUSIONS: The pain-control protocol significantly reduced pain severity to tolerable levels (VAS ≤3 cm) during and following intralesional and contact cryosurgery. Intralesional cryosurgery caused the least pain during and immediately after treatment.

Primary monocytes regulate endothelial cell survival through secretion of angiopoietin-1 and activation of endothelial Tie2.

OBJECTIVE: Monocyte recruitment and interaction with the endothelium is imperative to vascular recovery. Tie2 plays a key role in endothelial health and vascular remodeling. We studied monocyte-mediated Tie2/angiopoietin signaling following interaction of primary monocytes with endothelial cells and its role in endothelial cell survival. METHODS AND RESULTS: The direct interaction of primary monocytes with subconfluent endothelial cells resulted in transient secretion of angiopoietin-1 from monocytes and the activation of endothelial Tie2. This effect was abolished by preactivation of monocytes with tumor necrosis factor-α. Although primary monocytes contained high levels of both angiopoietin 1 and 2, endothelial cells contained primarily angiopoietin 2. Seeding of monocytes on serum-starved endothelial cells reduced caspase-3 activity by 46 ± 5.1%, and 52 ± 5.8% after tumor necrosis factor-α treatment and decreased detected single-stranded DNA levels by 41 ± 4.2% and 40 ± 3.5%, respectively. This protective effect of monocytes on endothelial cells was reversed by Tie2 silencing with specific short interfering RNA. The antiapoptotic effect of monocytes was further supported by the activation of cell survival signaling pathways involving phosphatidylinositol 3-kinase, STAT3, and AKT. CONCLUSIONS: Monocytes and endothelial cells form a unique Tie2/angiopoietin-1 signaling system that affects endothelial cell survival and may play critical a role in vascular remodeling and homeostasis.

Keloid histopathology after intralesional cryosurgery treatment.

BACKGROUND: Keloid presents a great healthcare challenge. The patients suffer from aesthetic disfiguration and occasionally from pruritus, pain and discomfort. Although various treatments are recommended, a single, highly effective treatment represents a great clinical need. OBJECTIVE: The cellular events and histopathology that follow intralesional cryosurgery were evaluated including cell proliferation, the number of cells expressing fibroblast markers, collagen synthesis and organization and mast cell infiltration. METHODS: Biopsies were collected before and after intralesional cryoneedle procedure. Collagen structure was evaluated with confocal microscopy. Mast cells, blood vessels and cell proliferation were evaluated using immunohistochemistry. RESULTS: Keloids contain abnormally thick collagen bundles, organized in swirls comprising closely bound fibrils. After intralesional cryosurgery, the collagen bundles lost their swirl structure, the thickness of the collagen layer decreased, and the bundles became more compact with less space between the fibres. A clear distinct transition zone separated the treated from the unaffected area. The frozen tissue was devoid of proliferating cells and mast cells whereas the number of blood vessels remained unaltered. Most of the fibroblasts expressed all tested myofibroblast markers although some exclusively expressed one and not the other. Few nuclei were observed in the affected area after treatment and very few of them expressed any fibroblast markers. CONCLUSIONS: Intralesional cryosurgery resulted in major changes in collagen structure and organization. The treatment reduced the number of proliferating cells, of myofibroblasts and of mast cells. These results may explain the reduction in no-response rate and the amelioration of the clinical symptoms after intralesional cryosurgery treatment.

Atypical fibroxanthoma of the scalp following hair transplantation in a 35-year-old male.

Atypical fibroxanthoma (AFX) is an uncommon spindle cell neoplasm of the elderly. This case report presents an atypical case of AFX of the scalp 8 years after hair transplantation in a 35-year-old male patient. Possible synergistic effects of previous sun exposure radiation to the scalp, together with the thermal and radiation injury of carbon dioxide (CO(2)) laser, might explain the mechanisms of the development of AFX at such an early age. To the best of our knowledge, this case report is the first description in the medical literature of development of skin malignancy on a hair-transplanted scalp.

Enhancement of ischemic wound healing by inducement of local angiogenesis.

OBJECTIVE: To determine if monocytes activated toward an angiogenic phenotype can be used to improve ischemic tissue healing in a rat skin flap model. STUDY DESIGN: Prospective experimental study on Wistar rats. METHODS: A caudally based 9 x 3 cm dorsal skin/panniculus carnosus flap was raised in 15 rats. The animals were divided into three groups: the monocyte group (N = 5) received subcutaneous topical application of 0.1-0.2 cc of i-Monogrid, a collagen gel containing M2 angiogenic monocytes; control group 1 (N = 5) received application of cell-free collagen; and control group 2 (N = 5) received no treatment. Skin flaps were stapled in place and observed for wound ischemia and necrosis of the skin flap. One week postoperatively, skin and underlying muscle were harvested for histologic analyses. RESULTS: No macroscopic differences in wound healing or microscopic differences in skin viability were observed. However, the monocyte group showed significantly greater vascular improvement than C1 (P = .047, chi = 3.96), and a trend toward greater vascular improvement than C2 (P = .103, chi = 2.67). CONCLUSIONS: Delivery of activated pro-angiogenic monocytes to an ischemic skin flap tended to improve histologic evidence of vascularity without corresponding microscopic or gross evidence of improved flap survival. These results are encouraging regarding the use of monocytes as a potential method of improving vascularization of ischemic tissue.

Copper, endoproteolytic processing of the prion protein and cell signalling.

Recently, understanding of many molecular interactions has progressed appreciably and cellular events once thought to be by-products of more important reactions or to be detrimental to cellular function are now known to be part of complex interactions of the cell with its environment. Numerous proteins can elicit differing effects depending upon post-translational modification events such as complex glycosylation and endoproteolytic cleavage or through binding co-factors including metal ions; the prion protein (PrP) is likely one such example. Its absolute requirement for pathogenesis has made the function of PrP an area of intense study but with apparently inconsistent results. This may, in part, stem from the ability of PrP to undergo different modifications to varying extents depending upon precise cellular circumstances. Specific modifications may promote altered association with binding partners resulting in apparent promiscuity of PrP interactions and activation of different signalling pathways, producing the diversity of functions suggested for this protein. This review discusses how modification of PrP by internal cleavage and metal ion co-ordination might influence, or be influenced by, signal transduction cascades. 2.

Monocyte activation state regulates monocyte-induced endothelial proliferation through Met signaling.

Direct interaction of unactivated primary monocytes with endothelial cells induces a mitogenic effect in subconfluent, injured endothelial monolayers through activation of endothelial Met. We now report that monocytes' contact-dependent mitogenicity is controlled by activation-mediated regulation of hepatocyte growth factor. Direct interaction of unactivated monocytes with subconfluent endothelial cells for 12 hours resulted in 9- and 120-fold increase in monocyte tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta) mRNA levels and bitemporal spike in hepatocyte growth factor that closely correlates with endothelial Met and extracellular signal-related kinase (ERK) phosphorylation. Once activated, monocytes cannot induce a second wave of endothelial cell proliferation and endothelial Met phosphorylation and soluble hepatocyte growth factor levels fall off. Monocyte-induced proliferation is dose dependent and limited to the induction of a single cell cycle. Monocytes retain their ability to activate other endothelial cells for up to 8 hours after initial interaction, after which they are committed to the specific cell. There is therefore a profoundly sophisticated mode of vascular repair. Confluent endothelial cells ensure vascular quiescence, whereas subconfluence promotes vessel activation. Simultaneously, circulating monocytes stimulate endothelial cell proliferation, but lose this potential once activated. Such a system provides for the fine balance that can restore vascular and endothelial homeostasis with minimal overcompensation.

Sclerosing mucoepidermoid carcinoma of the parotid gland: case report.

Mucoepidermoid carcinoma is the most common malignant salivary gland neoplasm. Its sclerosing morphologic variant, however, is an extremely rare entity. Only 7 cases have been reported in the literature, and only 2 of those reports have discussed this unusual malignancy in detail. This report describes a case involving a 57-year-old woman with an intermediate-grade sclerosing mucoepidermoid carcinoma of the parotid gland. She underwent superficial parotidectomy and radiation therapy with excellent results. Diagnosis and options for treatment are discussed, as well as a review of the literature, which demonstrates that this is an extremely rare malignancy with no strict protocol for treatment.

Photon-exposure-dependent photon-stimulated desorption for obtaining photolysis cross section of molecules adsorbed on surface by monochromatic soft x-ray photons.

Photon-exposure-dependent positive- and negative-ion photon-stimulated desorption (PSD) was proposed to study the photoreactions and obtain the photolysis cross sections of molecules adsorbed on a single-crystal surface by monochromatic soft x-ray photons with energy near the core level of adsorbate. The changes in the F(+) and F(-) PSD ion yields were measured from CF(3)Cl molecules adsorbed on Si(111)-7x7 at 30 K (CF(3)Cl dose=0.3x10(15) molecules/cm(2), approximately 0.75 monolayer) during irradiation of monochromatic soft x-ray photons near the F(1s) edge. The PSD ion yield data show the following characteristics: (a) The dissociation of adsorbed CF(3)Cl molecules is due to a combination of direct photodissociation via excitation of F(1s) core level and substrate-mediated dissociation [dissociative attachment and dipolar dissociation induced by the photoelectrons emitting from the silicon substrate]. (b) the F(+) ion desorption is associated with the bond breaking of the surface CF(3)Cl, CF(2)Cl, CFCl, and SiF species. (c) the F(-) yield is mainly due to DA and DD of the adsorbed CF(3)Cl molecules. (d) The surface SiF is formed by reaction of the surface Si atom with the neutral fluorine atom, F(+), or F(-) ion produced by scission of C-F bond of CF(3)Cl, CF(2)Cl, or CFCl species. A kinetic model was proposed for the explanation of the photolysis of this submonolayer CF(3)Cl-covered surface. Based on this model and the variation rates of the F(+)F(-) signals during fixed-energy monochromatic photon bombardment at 690.2 and 692.6 eV [near the F(1s) edge], the photolysis cross section was deduced as a function of energy.

Analysis of in vitro activities and modes of action of synthetic antimicrobial peptides derived from an alpha-helical 'sequence template'.

OBJECTIVES: Cationic antimicrobial peptides (AMPs) are indispensable components of innate immune systems and promising candidates for novel anti-infective strategies. We rationally designed a series of peptides based on a template derived from known alpha-helical AMPs, which were then analysed regarding efficacy against clinical isolates and antibiotic mechanisms. METHODS: Efficacy tests included standard MIC and synergy assays. Whole cell assays with staphylococcal strains included killing kinetics, efflux experiments and determination of membrane depolarization. The transcriptional response of AMP-treated Staphylococcus aureus SG511 was analysed using a Scienion genomic microarray covering (approximately 90% of) the S. aureus N315 genome and AMP P16(6|E). RESULTS: The AMPs showed remarkable broad-spectrum activity against bacteria and fungi regardless of any pre-existing antibiotic resistance mechanism. Whole cell assays indicated that the AMPs target the cytoplasmic membrane; however, significant membrane leakage and depolarization was only observed with a standard laboratory test strain. Transcriptional profiling identified up-regulation of putative efflux pumps and of aerobic energy generation mechanisms as major counter activities. Important components of the staphylococcal cell wall stress stimulon were up-regulated and the lipid metabolism was also affected. CONCLUSIONS: The broad spectrum activity of amphiphilic helical AMPs is based on multiple stresses resulting from interactions with microbial membranes; however, rather than killing through formation of pores, the AMPs appear to interfere with the coordinated and highly dynamic functioning of membrane bound multienzyme complexes such as electron transport chains and cell wall or lipid biosynthesis machineries.

Regulation of endothelial cell proliferation by primary monocytes.

OBJECTIVE: Endothelial cell-monocyte cross talk is essential for vascular repair. Monocytes colocalize with endothelial cells forming a complex set of interactions distinct from the growth promoting cytokines secreted by differentiated macrophages. In the present work we examined the growth regulation and in vitro wound repair early after binding of monocytes to endothelial cells. METHODS AND RESULTS: After direct contact with primary unactivated monocytes, endothelial cells enter S-phase through a mechanism mediated in part by contact-dependent activation of endothelial Met as demonstrated by siRNA silencing of Met, neutralizing antibodies for hepatocyte growth factor and Met as well as by specific inhibition of Met by the Met kinase inhibitor SU11274. Monocytes robustly promote endothelial cell proliferation and migration into a wounded endothelial monolayer. Monocyte-induced endothelial cell proliferation is accompanied by prolonged extracellular signal-regulated kinase (ERK) activation and is inhibited by the specific ERK inhibitor PD98059. The contact-mediated effect of monocytes is specific to endothelial cells and does not occur with vascular smooth muscle cells. Interestingly, although Flk1 is activated by monocytes, the proliferative effect of monocytes reported here is minimally mediated by Flk1 signaling. CONCLUSIONS: These results suggest that the early interaction between endothelial cells and monocytes is critical for the regulation of endothelial cell proliferation. This complex regulation is mediated in part by contact-dependent Met and ERK phosphorylation. These findings add to a broader set of leukocyte-endothelial contact mediated signals that together regulate endothelial function in health and disease.

Soft x-ray photoreactions of CF3Cl adsorbed on Si(111)-7x7 studied by continuous-time photon-stimulated desorption spectroscopy near F(1s) edge.

The continuous-time core-level photon-stimulated desorption (PSD) spectroscopy was employed to monitor the monochromatic soft x-ray-induced reactions of CF3Cl adsorbed on Si(111)-7x7 near the F(1s) edge (681-704 eV). Sequential F+ PSD spectra were measured as a function of photon exposure at the CF3Cl-covered surface (dose=0.3x10(15) molecules/cm2, approximately 0.75 ML). The F+ PSD and total electron yield (TEY) spectra of molecular solid CF3Cl near the F(1s) edge were also measured. Both F+ PSD and TEY spectra show two features at the energy positions of 690.2 and 692.6 eV, and are attributed to the excitations of F(1s) to 11a1[(C-Cl)*] and (8e+12a1)[(C-F)*] antibonding orbitals, respectively. Following Auger decay, two holes are created in the F(2p) lone pair and/or C-F bonding orbitals forming the 2h1e final state which leads to the F+ desorption. This PSD mechanism, which is responsible for the F+ PSD of solid CF3Cl, is employed to interpret the first F+ PSD spectrum in the sequential F+ PSD spectra. The variation of spectrum shapes in the sequential F+ PSD spectra indicates the dissipation of adsorbed CF3Cl molecules and the formation of surface SiF species as a function of photon exposure. From the sequential F+ PSD spectra the photolysis cross section of the adsorbed CF3Cl molecules by photons with varying energy (681-704 eV) is determined to be approximately 1.0x10(-17) cm2.

Antibacterial substances of low molecular weight isolated from the blowfly, Lucilia sericata.

Low molecular weight compounds were isolated by high-performance liquid chromatography from the maggot or haemolymph extracts of Lucilia sericata (Meigen) (Diptera: Calliphoridae). Using gas chromatography-mass spectrometry analysis, three compounds were obtained: p-hydroxybenzoic acid (molecular weight 138 Da), p-hydroxyphenylacetic acid (molecular weight 152 Da) and octahydro-dipyrrolo[1,2-a;1',2'-d] pyrazine-5,10-dione (molecular weight 194 Da), also known as the cyclic dimer of proline (or proline diketopiperazine or cyclo[Pro,Pro]). All three molecules revealed antibacterial activity when tested against Micrococcus luteus and/or Pseudomonas aeruginosa, and the effect was even more pronounced when these molecules were tested in combination and caused lysis of these bacteria.

Effect of skin surface temperature on skin pigmentation during contact and intralesional cryosurgery of keloids.

BACKGROUND: This 15-month study was designed to compare the effect of skin surface temperature on skin pigmentation following a single intralesional or contact cryosurgical treatment of keloids. PATIENTS/METHODS: Thirty Caucasian patients with 45 keloids present for more than 6 months were included in this study. Twenty-one keloids were treated by the contact method while the remaining 24 scars were managed using an intralesional cryosurgery technique. The skin surface temperature at the keloids was measured and recorded using a Ni/Cd thermocouple. Four variables of the thermal history were evaluated with the contact and the intralesional methods, namely cooling rate, hold time, end temperature and thawing rate. Assessment of the local hypopigmentation was performed 6 months after the treatment using a pigmentation scale. RESULTS: Significantly slower cooling (6.09 +/- 4.56 degrees C/min) and thawing rates (54.52 +/- 32.17 degrees C/min) were recorded with the intralesional cryosurgery method when compared with the cooling rates (13.47 +/- 9.04 degrees C/min) and thawing rates (89.00 +/- 86.42 degrees C/min) of the contact method (P < 0.000001). The end temperature of the contact technique was significantly cooler (-46.77 +/- 14.74 degrees C) when compared with that of the intralesional method (-15.55 +/- 6.77 degrees C) (P < 0.000001). There was a trend for the hold time of intralesional cryosurgery to be longer (82.67 +/- 138.03 s) than that of the contact method (16.86 +/- 23.49 s) (P < 0.059). A significant difference in skin pigmentation was demonstrated between the two cryosurgical methods. In 91.7% of the keloids treated by the contact technique a significant hypopigmentation was noticed, while no marked hypopigmentation was detected in the skin surface of the keloids treated by the intralesional method (P < 0.0001). CONCLUSION: We hypothesize that the thermal history of the skin surface during the intralesional cryosurgery technique provides a better survival environment for the melanocytes than the contact method, thus producing a lower rate of permanent hypopigmentation and disfiguring.