A novel target for the promotion of dermal wound healing: Ryanodine receptors.

Ryanodine receptors have an important role in the regulation of intracellular calcium levels in the nervous system and muscle. It has been described that ryanodine receptors influence keratinocyte differentiation and barrier homeostasis. Our goal was to examine the role of ryanodine receptors in the healing of full-thickness dermal wounds by means of in vitro and in vivo methods. The effect of ryanodine receptors on wound healing, microcirculation and inflammation was assessed in an in vivo mouse wound healing model, using skin fold chambers in the dorsal region, and in HaCaT cell scratch wound assay in vitro. SKH-1 mice were subjected to sterile saline (n = 36) or ryanodine receptor agonist 4-chloro-m-cresol (0.5 mM) (n = 42) or ryanodine receptor antagonist dantrolene (100 µM) (n = 42). Application of ryanodine receptor agonist 4-chloro-m-cresol did not influence the studied parameters significantly, whereas ryanodine receptor antagonist dantrolene accelerated the wound closure. Inhibition of the calcium channel also increased the vessel diameters in the wound edges during the process of healing and increased the blood flow in the capillaries at all times of measurement. Furthermore, application of dantrolene decreased xanthine-oxidoreductase activity during the inflammatory phase of wound healing. Inhibition of ryanodine receptor-mediated effects positively influence wound healing. Thus, dantrolene may be of therapeutic potential in the treatment of wounds.

Effects of Locally Applied Glycerol and Xylitol on the Hydration, Barrier Function and Morphological Parameters of the Skin.

Glycerol and xylitol hydrate the skin and improve its barrier function over a short period. We studied the effects of glycerol and xylitol on the physiological properties and morphology of the skin after longer-term application. Twelve volunteers with dry skin were examined. Three areas on the arms were determined. Area 1 served as untreated control. The vehicle was applied to area 2, while area 3 was treated twice daily with a formulation containing glycerol (5%) and xylitol (5%) for 14 days. Transepidermal water loss (TEWL), hydration and biomechanical properties of the skin were monitored. Biopsies were taken for routine histology and immunohistochemistry for filaggrin and matrix metalloproteinase-1 (MMP-1). The polyols increased the skin hydration and protein quantity of filaggrin, elevated the interdigitation index, decreased the TEWL and improved the biomechanical properties of the skin, but did not change the protein expression of MMP-1. A combination of glycerol and xylitol can be useful additional therapy for dry skin.

Rapid ligations with equimolar reactants in water with the potassium acyltrifluoroborate (KAT) amide formation.

The identification of fast, chemoselective bond-forming reactions is one of the major contemporary challenges in chemistry. We show that chemoselective amide-forming ligations of potassium acyltrifluoroborates (KATs) and O-carbamoylhydroxylamines proceed in the presence of all unprotected functional groups with a second-order rate constant of 20 M(-1) s(-1). PEG chains, lipids, biotin, and dyes were introduced onto an unprotected 31-mer peptide (a GLP-1 analogue) with equimolar ratios of reactants within minutes at 1 mM and within 1 h at 100 µM, even with Mw 20,000 PEG. This conjugation reaction provides a new approach to the synthesis of molecules such as protein-protein and protein-polymer conjugates.

A reagent for the one-step preparation of potassium acyltrifluoroborates (KATs) from aryl- and heteroarylhalides.

Potassium acyltrifluoroborates (KATs) are fascinating functional groups whose further exploration is limited by poor synthetic access. Documented herein is the design and synthesis of a new reagent for their one-step preparation from aryl- and heteroarylhalides. The reagent is a stable, soluble zwitterion prepared by S-alkylation of a novel thioformamide trifluoroboronate. The KATs are prepared by adding one equivalent of nBuLi to a mixture of the aryl halide and the reagent at -78 °C. This protocol is suitable for the preparation of KATs containing pyridines, esters, nitro groups, and halides.

Methane biogenesis during sodium azide-induced chemical hypoxia in rats.

Previous studies demonstrated methane generation in aerobic cells. Our aims were to investigate the methanogenic features of sodium azide (NaN(3))-induced chemical hypoxia in the whole animal and to study the effects of l-α-glycerylphosphorylcholine (GPC) on endogenous methane production and inflammatory events as indicators of a NaN(3)-elicited mitochondrial dysfunction. Group 1 of Sprague-Dawley rats served as the sham-operated control; in group 2, the animals were treated with NaN(3) (14 mg·kg(-1)·day(-1) sc) for 8 days. In group 3, the chronic NaN(3) administration was supplemented with daily oral GPC treatment. Group 4 served as an oral antibiotic-treated control (rifaximin, 10 mg·kg(-1)·day(-1)) targeting the intestinal bacterial flora, while group 5 received this antibiotic in parallel with NaN(3) treatment. The whole body methane production of the rats was measured by means of a newly developed method based on photoacoustic spectroscopy, the microcirculation of the liver was observed by intravital videomicroscopy, and structural changes were assessed via in vivo fluorescent confocal laser-scanning microscopy. NaN(3) administration induced a significant inflammatory reaction and methane generation independently of the methanogenic flora. After 8 days, the hepatic microcirculation was disturbed and the ATP content was decreased, without major structural damage. Methane generation, the hepatic microcirculatory changes, and the increased tissue myeloperoxidase and xanthine oxidoreductase activities were reduced by GPC treatment. In conclusion, the results suggest that methane production in mammals is connected with hypoxic events associated with a mitochondrial dysfunction. GPC is protective against the inflammatory consequences of a hypoxic reaction that might involve cellular or mitochondrial methane generation.

Catalytic hydrogenation with frustrated Lewis pairs: selectivity achieved by size-exclusion design of Lewis acids.

Catalytic hydrogenation that utilizes frustrated Lewis pair (FLP) catalysts is a subject of growing interest because such catalysts offer a unique opportunity for the development of transition-metal-free hydrogenations. The aim of our recent efforts is to further increase the functional-group tolerance and chemoselectivity of FLP catalysts by means of size-exclusion catalyst design. Given that hydrogen molecule is the smallest molecule, our modified Lewis acids feature a highly shielded boron center that still allows the cleavage of the hydrogen but avoids undesirable FLP reactivity by simple physical constraint. As a result, greater latitude in substrate scope can be achieved, as exemplified by the chemoselective reduction of α,ß-unsaturated imines, ketones, and quinolines. In addition to synthetic aspects, detailed NMR spectroscopic, DFT, and (2)H isotopic labeling studies were performed to gain further mechanistic insight into FLP hydrogenation.

Limb ischemia-reperfusion differentially affects the periosteal and synovial microcirculation.

BACKGROUND: Joints are privileged compartments that enjoy increased protection against the inflammatory reactions affecting the extremities. We hypothesized that the functional characteristics of the microvasculature would contribute to the differential defensive potential of the synovial membrane. METHODS: We investigated the synovial microcirculatory reactions and compared them with those of the tibial periosteum in response to 60 min of total limb ischemia, followed by 180 min of ischemia-reperfusion (IR) in rats. Carrageenan/kaolin-induced knee monoarthritis, a neutrophil-driven synovial inflammation model, served as the positive control. RESULTS: IR brought about a significant reduction in red blood cell velocity in the capillaries and increases in rolling and adherence of the neutrophil leukocytes in the postcapillary venules (intravital microscopy), in adhesion molecule expression (intercellular adhesion molecule-1 immunohistochemistry) and in xanthine oxidoreductase activity in the periosteum. These changes were also pronounced in carrageenan/kaolin-induced monoarthritis but were almost completely absent in the synovium after the IR challenge. Most importantly, even after IR and in carrageenan/kaolin monoarthritis, the synovial microcirculation was characterized by significantly greater red blood cell velocities than that in the periosteum under resting conditions. CONCLUSIONS: The ischemic duration, which significantly affected the functional integrity of the periosteal microcirculation, did not bring about a marked deterioration in that of the synovial membrane, suggesting that the synovial microcirculation is less endangered to the consequences of short-term tourniquet exposure than the periosteum. The greater microcirculatory red blood cell velocities and lower IR-induced endothelial expression of intercellular adhesion molecule-1 in the synovial membrane might explain the greater resistance of this compartment to the inflammatory consequences of IR.

A novel murine model for the in vivo study of transdermal drug penetration.

Enhancement of the transdermal penetration of different active agents is an important research goal. Our aim was to establish a novel in vivo experimental model which provides a possibility for exact measurement of the quantity of penetrated drug. The experiments were performed on SKH-1 hairless mice. A skin fold in the dorsal region was fixed with two fenestrated titanium plates. A circular wound was made on one side of the skin fold. A metal cylinder with phosphate buffer was fixed into the window of the titanium plate. The concentration of penetrated drug was measured in the buffer. The skin fold was morphologically intact and had a healthy microcirculation. The drug appeared in the acceptor buffer after 30 min, and its concentration exhibited a continuous increase. The presence of ibuprofen was also detected in the plasma. In conclusion, this model allows an exact in vivo study of drug penetration and absorption.

Ibuprofen penetration enhance by sucrose ester examined by ATR-FTIR in vivo.

The aim of this work was to investigate the skin penetration enhancer effect of a sucrose ester (SE) in an Ibuprofen (IBU) containing hydrogel and to examine its influence on the special lipid bilayer of the stratum corneum (SC). ATR-FTIR spectroscopic measurements were performed combined with tape stripping method on hairless mice in vivo. A SE containing gel was compared to another gel without SE. It was found that the preparations caused only minimal modifications in the lipid and the protein structure, promoting the skin hydration and therefore also the penetration of IBU. Although the degree of moisturization and penetration were more intense in the case of the SE containing gel treatment, it did not cause greater alterations in the SC structure than the gel without SE. It has been proven that SE acts as an effective and non-irritating hydration and penetration enhancer for IBU through skin.

[Investigation of surgical applicability of acryl-based tissue expanders]. / Akril alapú szöveti expanderek sebészeti felhasználhatóságának vizsgálata.

INTRODUCTION: Hydrogels made of acrylamide (AAm), acrylic acid (AAc) and N-isopropylacrylamide (NIPAAm) display outstanding ability of swelling in vitro and this property can be influenced by the composition of the polymers. Our objective was to study the in vivo behavior of hydrogels made of acrylics and their potential role in surgery. MATERIALS AND METHODS: Cylinder-shaped hydrogels were implanted under the skin of rats in the dorsal region. In group 1 AAm, in group 2 AAc while in group 3 NIPAAm expanders were applied. The observation period lasted 18 days, during this time the length and the diameter of the hydrogels were measured daily. After removal of the polymers the moisture mass and the rheological parameters were determined. Furthermore, biopsies were taken from adjacent tissues. RESULTS: Expanders achieved the peak of swelling in two weeks. During the observation period their mass significantly increased. NIPAAm polymers demonstrated the highest tendency to retain their preformed shape. Histological analysis revealed that implantation of AAc devices was associated with serious tissue damage while healthy skin was harvested with the other two polymers. CONCLUSION: In conclusion, NIPAAm hydrogel seems to be the most promising candidate for surgical utilization.

Reconstruction of Alar-Perialar Defects with a Combined Subcutaneous and Cutaneous Pedicled Rotation-Advancement Nasolabial Flap.

Aim of the Study: During the reconstruction of alar defects involving the upper lip, reconstructive surgeons face the need for various thicknesses of tissues crucial to preserving the facial sulcus which is important for a cosmetically acceptable result. Our aim was to reconstruct the deep perialar and thinner lateral nasal alar defect in a single step procedure with a suitable flap which is reliable, has appropriate blood supply and provides an esthetically good result. Materials and Methods: Extended alar defect was reconstructed with a combined flap in 10 cases. During the procedure, a subcutaneous pedicle was created and the proximal part of the flap was rotated into the defect as a rotational flap. The procedure and the follow-up have been photo-documented in all cases. Furthermore, the perfusion of the flaps was monitored by means of laser Doppler flowmetry. Postoperative complications were evaluated with a semi-quantitative score and the patients completed a patient satisfaction questionnaire, too. Results: An optimal esthetic result was obtained in all cases after the operation. The lateral nasal alar part of the defect was reconstructed with the thinner proximal part of the flap while the deeper perialar region involving the upper lip was covered with the thicker distal part. The flaps have shown sufficient blood flow after the operation. There was no significant pin cushioning or "trap-door" effect in any case. Mild erythema and edema was found in few cases. The patients were satisfied with the cosmetic result of the intervention. Conclusions: The flap is suitable for the reconstruction of alar defects involving the perialar region. It has the advantage of covering the deeper perialar and the thinner alar defects, whilst eliminating the pin cushioning effect of the conventional subcutaneous island pedicle flaps.

A Randomized Controlled Trial with a Medical Device Containing Sodium Hyaluronate and Nicotinic Acid to Increase the Efficacy of Ultraviolet Phototherapy in Psoriasis.

INTRODUCTION: The dry and scaly skin of psoriatic patients decreases the efficacy of ultraviolet B (UVB) phototherapy. Different agents are used to facilitate the transmission of light, but most of these preparations are cosmetically unfavorable. We have tested a novel preparation containing sodium hyaluronate and nicotinic acid (UV Fotogel®; Pernix Ltd.) with the double aim to improve the efficacy of UVB phototherapy and assess the cosmetic acceptability of the preparation. METHODS: Ninety patients with plaque psoriasis were enrolled in the study, of whom 44 received narrow-band UVB (NB-UVB) phototherapy. Prior to phototherapy, one side of the patient's body was treated with UV Fotogel while the other side served as a control. The other 46 patients used the preparation at their homes before regular sunbathing. The Local Psoriasis Severity Index (L-PSI), cosmetic acceptability and tolerability were recorded. The median values with the 25th and 75th percentiles (25p and 75p, respectively) were determined for the UV Fotogel-treated and control sites and then compared. RESULTS: The sides of the body to which UV Fotogel was applied prior to NB-UVB phototherapy had a significantly lower median L-PSI score than the non-treated control sides at the end of the treatment (1.0 [25p-75p: 0.0-2.0] vs. 2.0 [1.0-3.0], respectively). The application of UV Fotogel prior to sunbathing also led to a significant decrease in L-PSI score. There was a significant reduction in the median L-PSI score of patients at the final visit compared to baseline (2.5 [25p-75p: 1.5-3.5] vs. 6.0 [6.0-7.0], respectively). Use of the preparation was not accompanied by considerable adverse effects, and the patients found it cosmetically acceptable. Application of UV Fotogel prior to sunbathing was well tolerated by the patients, and the cosmetic acceptability was also good. CONCLUSION: UV Fotogel is potentially a useful device for enhancement of the efficacy of phototherapy in patients with psoriasis.

Efficacy and safety of long pulse 1064 and 2940 nm lasers in noninvasive lipolysis and skin tightening.

Noninvasive body shaping is becoming a growing demand. The aim of this study was to investigate the efficacy and safety of the combined treatments of 1064 nm Nd:YAG and 2940 nm Er:YAG in noninvasive lipolysis and skin tightening. Ten females were enrolled, and all women's side of the waist or the lower part of the abdomen were treated. In the first step, the 1064 nm Nd:YAG was used. As a second step, the 2940 nm Er:YAG laser was applied. Each woman was treated four times, once every 2 weeks. The effects were determined by comparative photo documentation, waist circumference measurement, two-dimensional B-mode ultrasonography and low-dose native computer tomography (CT), whereas body fat was monitored with bioelectric impedance. The tissue firmness was measured by ultrasound shear wave elastography. Combined laser treatment significantly reduced waist circumference and total body fat. Ultrasonography has revealed that the treatment considerably decreased fat thickness and improved skin stiffness in the treated region. Subcutaneous fat volume, measured by low-dose CT, displayed a moderate decrease in the waist region. The combined 1064 nm Nd:YAG and 2940 nm Er:YAG laser treatment results in the reduction of fat tissue and tightens the skin as confirmed by objective measurements.

A Possible Technique for the Complex Reconstruction of Exposed Breast Implant: Applicability and Microcirculation of the Capsule Flap.

Aim of the Study: Immediate breast reconstruction is often applied after mastectomy. However, inappropriate surgical technique, postoperative radiotherapy and infection may lead to tissue necrosis and implant protrusion. Traditional therapies frequently fail. However, previous data suggested that capsule flaps may be appropriate for the salvage of implants. Our goal was to investigate the usefulness of capsuloplasty in patients with exposed breast implant and to monitor the blood supply of capsule flaps during the operation. Materials and Methods: Capsuloplasty was performed in 19 patients with exposed implant. After removal of necrotic tissue, capsulotomy was performed, the planned flap was dissected free, the implant was covered with the flap and the wound was then closed. During operation, the blood flow of the flap was determined by means of laser Doppler flowmetry. Moreover, tissue samples were taken for histology and immunostaining for CD34. Results: The postoperative follow-up showed that capsule flaps survived in each case: no complications were found. The blood flow of the flaps did not change significantly during the intervention as compared with the baseline values. The histology and the immunohistochemistry revealed considerable vascularization and angiogenesis in the flap. Conclusions: Capsule flaps seem to be appropriate for the salvage of exposed implants and for enhancement of implant cover in the case of thin and injured tissue.

Protective effects of glycerol and xylitol in keratinocytes exposed to hyperosmotic stress.

Purpose: Our goal was to study whether glycerol and xylitol provide protection against osmotic stress in keratinocytes. Methods: The experiments were performed on HaCaT keratinocytes. Hyperosmotic stress was induced by the addition of sorbitol (450, 500 and 600 mOsm). Both polyols were applied at two different concentrations (glycerol: 0.027% and 0.27%, xylitol: 0.045% and 0.45%). Cellular viability and cytotoxicity were assessed, intracellular Ca2+ concentration was measured, and the RNA expression of inflammatory cytokines was determined by means of PCR. Differences among groups were analyzed with one-way ANOVA and Holm-Sidak post-hoc test. When the normality test failed, Kruskal-Wallis one-way analysis of variance on ranks, followed by Dunn's method for pairwise multiple comparison was performed. Results: The higher concentrations of the polyols were effective. Glycerol ameliorated the cellular viability while xylitol prevented the rapid Ca2+ signal. Both polyols suppressed the expression of IL-1α but only glycerol decreased the expression of IL-1ß and NFAT5. Conclusions: Glycerol and xylitol protect keratinocytes against osmotic stress. Despite their similar chemical structure, the effect of these polyols displayed differences. Hence, joint application of glycerol and xylitol may be a useful therapeutic approach for different skin disorders.

A Review of Electroporation-based Antitumor Skin Therapies and Investigation of Betulinic Acid-loaded Ointment.

BACKGROUND: Electrochemotherapy is a novel treatment for cutaneous and subcutaneous tumors utilizing the combination of electroporation and chemotherapeutic agents. Since tumors have an increasing incidence nowadays as a result of environmental and genetic factors, electrochemotherapy could be a promising treatment for cancer patients. OBJECTIVE: The aim of this article is to summarize the novel knowledge about the use of electroporation for antitumor treatments and to present a new application of electrochemotherapy with a well-known plant derived antitumor drug betulinic acid. For the review we have searched the databases of scientific and medical research to collect the available publications about the use of electrochemotherapy in the treatment of various types of cancer. METHOD: By the utilization of the available knowledge, we investigated the effect of electroporation on the penetration of a topically applied betulinic acid formulation into the skin by ex vivo Raman spectroscopy on hairless mouse skin. RESULTS: Raman measurements have demonstrated that the penetration depth of betulinic acid can be remarkably ameliorated by the use of electroporation, so this protocol can be a possibility for the treatment of deeper localized cancer nodules. Furthermore, it proved the influence of various treatment times, since they caused different spatial distributions of the drug in the skin. CONCLUSION: The review demonstrates that electrochemotherapy is a promising tool to treat different kinds of tumors with high efficiency and with only a few moderate adverse effects. Moreover, it presents a non-invasive method to enhance the penetration of antitumor agents, which can offer novel prospects for antitumor therapies.

Electroporation-enhanced transdermal diclofenac sodium delivery into the knee joint in a rat model of acute arthritis.

PURPOSE: Since electroporation (EP) can increase the permeability of biological membranes, we hypothesized that it offers an opportunity to enhance the transdermal delivery of drugs for intra-articular indications. Our aim was to compare the anti-inflammatory and analgesic efficacy of EP-combined topical administration of diclofenac sodium hydrogel (50 mg mL-1 in 230 µL volume) with that of an equivalent dose of oral (75 mg kg-1) and simple topical administration. METHODS: Arthritis was induced with the injection of 2% λ-carrageenan and 4% kaolin into the right knee joints of male Sprague Dawley rats. EP was applied for 8 min with 900 V high-voltage pulses for 5 ms followed by a 20 ms break. Drug penetration into the synovial fluid and plasma was detected by high-performance liquid chromatography. Leukocyte-endothelial interactions were visualized by intravital videomicroscopy on the internal surface of the synovium. Inflammation-induced thermal and mechanical hyperalgesia reactions, knee joint edema, and inflammatory enzyme activities were assessed at 24 and 48 h after arthritis induction. RESULTS: EP significantly increased the plasma level of diclofenac as compared with the topical controls 10 min after the 2% λ-carrageenan and 4% kaolin injection. Increased leukocyte-endothelial interactions were accompanied by joint inflammation, which was significantly reduced by oral and EP diclofenac (by 45% and by 30%, respectively) and only slightly ameliorated by simple topical diclofenac treatment (by 18%). The arthritis-related secondary hyperalgesic reactions were significantly ameliorated by oral and EP-enhanced topical diclofenac treatments. The knee cross-section area (which increased by 35%) was also reduced with both approaches. However, simple topical application did not influence the development of joint edema and secondary hyperalgesia. CONCLUSION: The study provides evidence for the first time of the potent anti-inflammatory and analgesic effects of EP-enhanced topical diclofenac during arthritis. The therapeutic benefit provided by EP is comparable with that of oral diclofenac; EP is a useful alternative to conventional routes of administration.

Systemic phosphatidylcholine pretreatment protects canine esophageal mucosa during acute experimental biliary reflux.

AIM: To characterize the consequences of short-term exposure to luminal bile on mucosal mast cell reactions in a canine model, and to determine the effects of systemic phosphatidylcholine pretreatment in this condition. METHODS: Twenty mongrel dogs were used for experiments. Group 1 (n = 5) served as a saline-treated control, while in group 2 (n = 5) the esophagus was exposed to bile for 3 h. In group 3 (n = 5) the animals were pretreated with 7-nitroindazole to inhibit the neuronal isoform of nitric oxide synthase. In group 4 (n = 5) phosphatidylcholine solution (50 mg/kg) was administered iv before the biliary challenge. Mucosal microcirculation was observed by intravital videomicroscopy. Myeloperoxidase and nitric oxide synthase activities, the degrees of mast cell degranulation and mucosal damage were evaluated via tissue biopsies. RESULTS: Exposure to bile evoked significant mast cell degranulation and leukocyte accumulation. The red blood cell velocity and the diameter of the postcapillary venules increased significantly. The tissue ATP content and constitutive nitric oxide synthase activity decreased, while the inducible nitric oxide synthase activity increased significantly as compared to the control values. 7-nitroindazole treatment significantly exacerbated the mucosal mast cell degranulation and tissue damage. In contrast, phosphatidylcholine pretreatment prevented the bile-induced ATP depletion, the inducible nitric oxide synthase and myeloperoxidase activity and the mast cell degranulation increased. CONCLUSION: The neuronal nitric oxide synthase--mast cell axis plays an important role in the esophageal mucosal defense system. Systemic phosphatidylcholine pretreatment affords effective protection through ameliorating the bile-induced ATP depletion and secondary inflammatory reaction.

Pharmacological Targeting of the Epidermal Barrier.

The most important function of the skin is to form a barrier between the body and the external environment. The epidermal barrier prevents transepidermal water loss from the skin, but also serves as a barrier to the entry of harmful environmental allergic, toxic or infectious substances. Inherited defects in the genes encoding the components of the epidermal barrier result in the development of rare genetic disorders, whereas polymorphisms in these genes together with environmental factors cause frequent inflammatory skin diseases, such as atopic dermatitis. In this review, components of the skin-barrier function will be reviewed with special emphasis on how the altered epidermal barrier might be repaired. The different strategies to increase the transdermal penetration of drugs is also discussed.

ATR-FTIR and Raman spectroscopic investigation of the electroporation-mediated transdermal delivery of a nanocarrier system containing an antitumour drug.

The aim of the present work was the optimization of the transdermal delivery of a lyotropic liquid crystal genistein-based formulation (LLC-GEN). LLC was chosen as medium in view of the poor solubility of GEN in water. Membrane diffusion and penetration studies were carried out with a Franz diffusion cell, through a synthetic membrane in vitro, a chick chorioallantoic membrane ex ovo, and ex vivo excised human epidermis. Thereafter, LLC-GEN was combined with electroporation (EP) to enhance the transdermal drug delivery. The synergistic effect of EP was verified by in vivo ATR-FTIR and ex vivo Raman spectroscopy on hairless mouse skin.