Evaluation of Permacol as a cultured skin equivalent.

Skin loss following severe burn requires prompt wound closure to avoid such complications as fluid and electrolyte imbalance, infection, immune suppression, and pain. In clinical situations in which insufficient donor skin is available, the development of cultured skin equivalents (dermal matrices seeded with keratinocytes and fibroblasts) may provide a useful alternative. The aim of this study was to assess the suitability of a porcine-derived dermal collagen matrix (Permacol) to function as a cultured skin equivalent in supporting the growth of keratinocytes in vitro and providing cover to full thickness wounds in the BALB C/nude mouse model. A histological comparison was against Glycerol treated-Ethylene Oxide Sterilised Porcine Dermis (Gly-EO Dermis) which has successfully been used as a cultured skin equivalent in previous studies. Both Gly-EO Dermis and to a lesser extent Permacol were able to support the growth of cultured keratinocytes following a 16-day period of cell culture, however, this study was only able to demonstrate the presence of an epidermal layer on Gly-EO dermis 2 weeks after grafting onto full-thickness wounds in the BALB C/nude mouse model.

Histological evaluation of Permacol as a subcutaneous implant over a 20-week period in the rat model.

This study assessed the suitability of Permacol (a porcine derived, isocyanate cross linked collagen based biomaterial) as an alternative to autologous tissue in soft tissue reconstruction. The Sprague-Dawley rat was used as a model for subcutaneous implantation over a 20 week period and comparison made with two other porcine biomaterials (small intestinal submucosa and glycerol treated-ethylene oxide sterilised porcine dermis). Implants were scored histometrically on the degree of acute inflammation, chronic inflammation, fibrosis and stromal response. The vascularity and percentage composition of collagen within Permacol were assessed by stereology and seescan image analysis, respectively. In general terms, Permacol was well tolerated as a subcutaneous implant, with only a minor chronic inflammatory response remaining after a 20 week period of implantation. There was evidence of collagen degradation during this period and vascular ingrowth into Permacol was limited. Permacol has the potential for a broad range of applications in plastic surgery, but may benefit from modification to promote a more rapid degree of vascularisation.

The diamond CO2 laser as a method of improving the vascularisation of a permanent collagen implant.

For a biomaterial to be considered as anything other than a simple filler material it must allow a certain degree of ingrowth from the surrounding host tissues. Permacol is a novel porcine derived biomaterial that is isocynate crosslinked to prevent its degradation in vivo, but while it appears to be biocompatible in terms of its histological tissue response, our early studies demonstrate only a limited degree of fibrovascular ingrowth. The aim of this study was to assess the effectiveness of both laser perforations and topically applied VEGF as methods of encouraging fibrovascular ingrowth. Treatment of Permacol with the diamond CO(2) laser significantly increased the porosity of the material as assessed by seescan image analysis. Fibrovascular ingrowth as assessed in the Sprague-dawley rat model was found to be limited to the pores themselves and not the surrounding native matrix. Pre-soaking of laser treated Permacol in vascular endothelial growth factor (VEGF) did not further increase fibrovascular ingrowth. Increasing the porosity of Permacol may allow its use as a dermal replacement in the support of an overlying skin graft or keratinocyte sheet; however using the laser may not be the optimal method of achieving this aim.

Evaluation of a porcine origin acellular dermal matrix and small intestinal submucosa as dermal replacements in preventing secondary skin graft contraction.

The degree to which a split thickness skin graft (STSG) contracts after application to its recipient bed is related in part to the proportion of the dermis harvested from the donor site. Harvesting thicker skin grafts may produce better cosmetic results in the recipient bed but result in increased donor site morbidity. The combination of an autologous ultra thin split thickness graft with an underlying non-autologous dermal component may reduce secondary skin graft contraction without further increasing donor site morbidity. This study was aimed at assessing the suitability of two porcine derived biomaterials (Permacol and small intestinal submucosa, SIS) for use in combination with skin grafts in a Sprague-Dawley rat model. Full thickness wounds (1 cm(2)) were created in Sprague-Dawley rats and grafted with skin in combination with Permacol or SIS either as a one-stage operation or following a 2-week-period of vascularisation of these dermal matrices before a second stage operation to cover with skin. Skin graft viability and wound area were assessed at weekly intervals until 4 weeks after graft application. Both Permacol and SIS were able to support an overlying skin graft but had no beneficial effect on skin graft contraction in this model compared to skin grafts alone.

An unusual complication of penile piercing: a report and literature review.

Body piercing has been practiced for many centuries and is thought to have its origins in the Hindu religion. As traditional ear piercings have given way to more adventurous anatomical sites, so too have the associated complications of these practices. We present the case of a 35 year old man who presented 12 months after a penile piercing with an extensive fistula of the glans penis resembling an acquired glanular hypospadius.

Prefabricated skin flaps in a rat model based on a dermal replacement matrix Permacol.

The reconstruction of three-dimensional structures is one of the most challenging problems in plastic surgery. While shaped frameworks incorporated within soft tissue envelopes often lose their fine detail, the integration of a blood supply to a stable biomaterial scaffold allows skin graft cover and emphasis of fine detail. The aim of this study was to evaluate whether a porcine derived isocyanate crosslinked collagen tissue matrix (Permacol) could be vascularised from an isolated vascular pedicle and subsequently sustain an overlying skin graft as a pedicled prefabricated flap. A vascular induction technique was employed using the epigastric pedicles in Sprague-Dawley rats to vascularise 2 x 2 cm(2) pieces of Permacol of standard 0.75 and 1.5 mm thickness Permacol and laser treated Permacol of 0.4 and 0.75 mm thickness. Flaps were able to support overlying skin grafts as early as 2 weeks following vessel implantation and laser treated Permacol had an increased vascularity compared to standard Permacol with no evidence of degradation. The study showed that Permacol could be a useful matrix for use in tissue engineered flaps.

Bath PUVA--an investigation of the distribution of trioxsalen (TMP) and 8-methoxypsoralen (8-MOP) in bathwater.

Trioxsalen (TMP) bath PUVA avoids the side effects of nausea and headache associated with oral 8-methoxypsoralen (8-MOP) treatment and allows shorter irradiation times that can be advantageous in some patients. However we noted that a number of patients developed unusual patterns of phototoxic burning. We thought that this was related to an uneven distribution of the TMP in the bathwater and for this reason, a study of bath water TMP concentrations achieved using different TMP preparations was undertaken. The distribution of 8-MOP in an 8-MOP bath was also measured for comparison. Our results confirm that an uneven distribution of TMP is achieved using TMP capsules or suspension and would explain our observed patterns of burning. With an ethanolic solution of TMP, or the commercial equivalent Tripsor, or with Puvasoralen-8 (an 8-MOP preparation), a homogeneous psoralen distribution is achieved, and they are therefore preferable for use in bath PUVA.

Radiation from ultraviolet phototherapy sources results in photodegradation of 12(R) and 12(S)-hydroxy-eicosatetraenoic acid: high-performance liquid chromatography and polymorph migration studies.

The effect of irradiation from Sylvania PUVA lamps (emitting predominantly in the ultraviolet (UVA) region) and broadband Philips TL-12 lamps (peaking in the UVB region) on two inflammatory mediators, 12(R)- and 12(S)-hydroxy-eicosatetraenoic acid (HETE) was studied. A high-performance liquid chromatography study showed significant photodegradation of both enantiomers at a concentration of 5 micrograms/ml in phosphate-buffered saline following irradiation with 10 J.cm-2 UVA or 0.375 J.cm-2 UVB. The in vitro chemokinetic microdroplet migration response of human peripheral polymorphonuclear leukocytes from normal and psoriatic subjects was significantly reduced following irradiation of 12(R)-HETE at a concentration of 1 micrograms/ml in medium with 40 J.cm-2 UVA and 1.5 J.cm-2 UVB respectively. No such effect was seen with 12(S)-HETE. The effect of ultraviolet on skin physiology, and in particular in the successful phototherapy of a range of inflammatory skin disorders, is not fully understood. The photodegradation of inflammatory mediators such as 12-HETE, as shown in this study, provides another factor of possible therapeutic significance.

Filtration as part of the validation and routine testing of a purified water system.

A distillation system, which includes a steam condensate supply, pre-treated before distillation by passage through carbon and mixed resin barrels, is described. Filtration of 5-litre aliquots from a storage tank/ring mains system through a 0.45-micron cellulose filter identified contaminants (rust from the ring mains and carbon from the barrel) which were not detected by the British Pharmacopoeia tests for purified water.

HPLC studies on the degradation profiles of glucose 5% solutions subjected to heat sterilization in a microprocessor-controlled autoclave.

A practically useful relationship between degradation and F0 at various temperatures is given. This may be of value for identifying the most suitable sterilization conditions for a number of glucose products and other pharmaceuticals. It was found that autoclaving at a high temperature to a low final F0 value gave the maximum product integrity.

Reversed-phase high-performance liquid chromatographic method for the quantification of 5-hydroxymethylfurfural as the major degradation product of glucose in infusion fluids.

Analysis of 5-hydroxymethylfurfural (5-HMF) is an important indicator of degradation in glucose infusion fluids. Current pharmacopoeial methods for analysis are insensitive and non-specific. A method is described here, incorporating an internal standard, to give a sensitive, accurate and precise method, suitable for use in the quality control of glucose infusion fluids, and with possible applications for the quantitative determination of 5-HMF in food and drink, bacterial cultures and hydrothermolysed wood.

A study of the photodegradation of leukotriene B4 by ultraviolet irradiation (UVB, UVA).

In view of the presence of the polymorphonuclear leukocyte (PMN) chemoattractant Leukotriene B4 (LTB4) in the surface scale of the psoriatic lesion and the known therapeutic effect of phototherapy in psoriasis, the photostability of LTB4 was investigated. LTB4 was irradiated with dosages of UVB (290-320 nm) ranging from 100-1500 mJ cm-2 and with dosages of UVA (320-400 nm) ranging from 5-40 J cm-2. Topical application of UVB irradiated LTB4 to the forearm skin of normal volunteers showed a marked reduction in cutaneous erythema, paralleled histologically by reduced transepidermal PMN migration when compared with sites of application of unirradiated and UVA irradiated LTB4. High performance liquid chromatography (HPLC) demonstrated a dose-dependent photodegradation of LTB4 by UVB irradiation. UVA irradiation produced no such effect. The wavelengths responsible lie within the absorption spectrum of LTB4. In vitro chemotaxis studies, using an under agarose technique, showed a statistically significant reduction in the migration of PMN from both psoriatic and non-psoriatic subjects to the UVB irradiated LTB4 compared with the unirradiated LTB4, whilst UVA irradiated LTB4 produced a normal PMN chemotactic response.

Polymorphonuclear leucocyte chemotaxis in response to leukotriene B4 in treated and untreated psoriatics.

Various chemoattractants have been implicated in the aetiology of the polymorphonuclear leucocyte (PMN) migration into the epidermis seen in early lesions of psoriasis. Using an under-agarose technique, the in vitro chemotactic responses of PMN to the arachidonic acid lipoxygenase product leukotriene B4 (LTB4) were assayed in five groups of subjects: normal healthy volunteers (n = 12), untreated psoriatics (n = 11) and psoriatics treated with topical tar (n = 12), PUVA (n = 11) and UVB phototherapy (n = 10). No significant difference was observed between the responses of control subjects and of untreated psoriatics, nor between the untreated psoriatic group and the PUVA- and UVB-treated groups, respectively. However, comparison of the tar-treated and untreated groups revealed a significantly increased chemotactic response to LTB4 in the tar-treated group (p less than 0.01).

Drug dilution in anaesthetic practice.

The accuracy of dilution by anaesthetists of commonly used drug preparations was assessed by ultra violet spectrophotometric assay of the diluted drugs. Percentage deviation from target concentrations was then calculated. Of a total of 358 drug dilutions carried out by 28 anaesthetists only 58% were within 10% of the target concentration while 6% lay outside 50% of the target concentration.

Dapsone and human polymorphonuclear leucocyte chemotaxis in dermatitis herpetiformis.

Polymorphonuclear leucocyte chemotaxis was investigated in 6 patients with active dermatitis herpetiformis in the untreated state and when under control with Dapsone. Control studies were undertaken in 7 healthy volunteers. No significant difference in chemotaxis was demonstrated between the active, treated and control groups. Furthermore, when Dapsone in physiological concentrations was added separately in all three groups there was no additional effect on chemotaxis. These results clearly show that polymorphonuclear leucocytes from active untreated dermatitis herpetiformis have normal chemotactic activity compared with those from the same patients in the treated state and controls. Additional Dapsone did not alter these findings.

Cell-mediated immune response to basal cell carcinoma.

The role of cell-mediated immunity in controlling Basal Cell Carcinoma (BCC) growth was studied by measuring the transformation of lymphocytes when challenged in vitro with aqueous extracts of the patients' BCCs and of control skin. Tests were carried out in the presence of autologous and heat-treated plasma. Three patients out of 8 showed significantly raised thymidine uptake ratio (TUR). Heat-treated plasma produced higher TURs, indicating the presence of an inhibitory factor.

Electrophoretic mobility of lymphocyte subpopulations in infectious mononucleosis.

The electrophoretic mobilities (EPM) of lymphocytes from seventeen patients with infectious mononucleosis (IM) and fifteen normal controls were compared. The IM patients were divided into three groups, two in the first week of illness and one in the second week. The lymphocyte EPM distribution was separated by histogram analysis into fast and slow populations and further division of the fast cell population was possible in all but one of the two groups of IM patients during their first week of illness. The percentage of cells of the faster subpopulation in the first week of illness was lower than that of the controls (P less than 0.05) but the absolute lymphocyte count was higher (P less than 0.005). However, the percentage of slow moving cells of both groups in the first week of illness was significantly higher than in either the second week or the normal controls (P less than 0.01-0.001). Absolute counts were higher in all lymphocyte subpopulations of IM (P less than 0.01-0.001).