Functional in vitro tension measurements of fascial tissue - a novel modified superfusion approach.

INTRODUCTION: While two laboratory techniques are commonly used to assess the tensile properties of muscle tissue, emerging evidence suggests that the fascial components of these tissues also serve an active role in force generation. Hence, we investigated whether these techniques are sensitive for assessment of fascial micromechanics. METHODS: Force measurements on dissected fascial tissue were performed either using the classical immersion organ bath or using an improved superfusion approach simulating pulsed pharmacological triggers. Rat deep dorsal fascial strips as well as rat testicular capsule were pharmacologically challenged either with mepyramine or oxytocin. RESULTS: The classical immersion technique yielded a lower force response to mepyramine than the superfusion method (median: 367.4 vs. 555.4µN/mm(2)). Pause in irrigation before application reduced irregularities during bolus application. The superfusion approach was improved further by the following points: The high sensitivity of the superfusion method to bolus addition was voided by deviation of fluid supply during bolus addition. CONCLUSION: Although both methods demonstrated pharmacologically induced contractile responses in lumbar fascia samples, the modified superfusion method may improve force registrations of slow contracting fascial tissue and minimize artefacts of fluid application.

Remifentanil in combination with ketamine versus remifentanil in spinal fusion surgery--a double blind study.

AIM: This study is aimed at conducting a program for two different anesthetic methods used during a spinal fusion surgery to ensure better intra-operative hemodynamic stability and post-operative pain control. METHODS: A prospective, randomized, double blind study in patients scheduled for spinal fusion surgery, who were randomly allocated to two groups, G1 and G2, (n = 15 per group), class I-II ASA, was carried out. Both groups received pre-operatively midazolam, followed intra-operatively by propofol, sevoflurane, atracurium, and either remifentanil infusion 0.2 microg/kg/min (G1), or the same dose of remifentanil infusion and low doses of ketamine infusion 1 microg/kg/min (G2) anesthetics, antidote medication and post-operative morphine doses. HR, MAP, vital signs, surgical bleeding, urine output, duration of surgery and duration of anesthesia were recorded. In a 24-h recovery period in a post-anesthesia care unit (PACU) the recovery time, the first pain score and analgesic requirements were measured. RESULTS: Intra-operative HR and arterial BP were significantly less (p < 0.05) in G1 as compared to G2. In the PACU the first pain scores were significantly less (p < 0.05) in G2 than in G1. The time for the first patient analgesia demand dose was greater in G2, as also morphine consumption which was greater in G1 than G2 (p < 0.05). Other results were the same. None of the patients had any adverse drug reaction. CONCLUSIONS: Adding low doses of ketamine hydrochloride could be a routine therapy to improve the hemodynamic stability and reduce the post-operative morphine consumption during spinal fusion surgery.

Re-visiting Luck's classification: a histological analysis of Dupuytren's disease.

Luck (1959) described a histological staging system for Dupuytren's disease, classifying the disease into three stages. Previous biochemical and immunochemical studies have detailed the decrease in type III/I collagen ratio with disease progression. Herovici (1963) described a histological stain that produced a differential red/purple and blue colour for type I and III collagen respectively. We stained 15 specimens of Dupuytren's disease and quantified the different collagen types in each using computer analysis. We found a corresponding decrease in the amount of type III collagen as a percentage of the total collagen with disease progression: stage I range 35-49% (mean 38%); stage 2 range 21-33% (mean 27%) and stage 3 range 11-19% (mean 14%). We propose a new staging system based on the relative amount of type III collagen, where stage 1: >35%, stage 2: >20% and <35%, and stage 3: <20%.

Pentoxifylline inhibits mature burn scar fibroblasts in culture.

Fibroblasts are thought to be (in part) responsible for the persisting contractile forces that result in burn contractures. Using monolayer and fibroblast populated collagen lattice (FPCL) models we subjected burn scar fibroblasts to the anti-fibrinolytic agent Pentoxifylline (PFX) in an attempt to reduce proliferation and contraction of these cells. Fibroblasts were isolated from mature burn scars at reconstructive surgery. Fibroblasts were grown in monolayer or incorporated into FPCL's and exposed to PFX. Fibroblast numbers and FPCL surface areas were calculated using digital photography and image analysis. PFX showed a dose-dependent inhibition of contraction and reduced proliferation of burn scar fibroblasts. In monolayer, cell number proliferation was markedly reduced. FPCL's containing 0, 0.25, 0.5, 1, and 2 mg/ml of PFX had relative surface areas of 31, 40, 43, 59, and 85%, respectively. One and 2 mg/ml FPCL's contracted significantly less than controls (p < 0.0001). This is the first study to show the dose-dependent effects of Pentoxifylline on the proliferation and contraction of burn scar fibroblasts. This study suggests that Pentoxifylline has a direct effect on inhibiting burn scar fibroblasts. Further study of PFX on burn scars will provide opportunities to reduce burn scar contractures in vivo.

The effect of basic fibroblast growth factor on scarring.

Granulation-tissue myofibroblasts are important in wound contraction, disappearing in normal scars but persisting in hypertrophic scars. While transforming growth factor beta 1 (TGF-beta 1) is implicated in excessive scarring and induces the accumulation of myofibroblasts, the role of basic fibroblast growth factor (FGF-2) on scarring remains unreported. Four linear full-thickness incisions, each 20 mm long, were made dorsally on 25 adult male Hooded Lister rats. In each animal, one wound was unmanipulated (control), one was injected with TGF-beta 1 (positive control), one with vehicle alone and one with FGF-2 (test). The wounds were injected daily for 5 days. Animals were sacrificed in groups of five on days 5, 10, 15, 20 and 30 after wounding. Wounds were subjected to tensiometry. Sections were stained for collagen fibres, immunostained for myofibroblasts and studied under light microscopy and electron microscopy. Myofibroblasts were present in the granulation tissue on day 5, reached their maximum number on day 10 and disappeared from all wounds by day 30. Treatment with FGF-2 inhibited this transient phenotypic change of granulation-tissue fibroblasts into myofibroblasts, relative to controls (15.23% versus 58.71%, 15.23% versus 54.71% and 15.23% versus 53.15%; P<0.003 on day 10, paired t-test). Test collagen-fibre orientation resembled that of the normal dermis, in contrast to that of the control wounds. Test wound breaking strength was unreduced. These results suggest a possible anti-scarring effect of FGF-2 during wound healing.

Dupuytren's disease. The way forward?

Histologically, Dupuytren's disease has been compared to the process of neoplasia because of fibroblast proliferation, recurrence, chromosomal abnormalities and antigenic profiles. However, a comparison of Dupuytren's tissue with the granulation tissue formed in wound healing could be more valid. Histology reveals similarities in cell types, proliferation, vascularity and collagen morphology. Pharmacologically, both tissues have a similar range of agonist and antagonist responses. Biochemical analysis reveals new collagen synthesis, an increased ratio of type III to type I collagen, and similar changes of the ground substance in both processes. Considering such similarities perhaps it is possible to regard some of the models used for the investigation of wound healing and granulation tissue as the missing experimental "model" for the study of Dupuytren's disease. Recently great strides have been made in the basic understanding of wound biology, and such a comparison might well provide novel therapeutic options for Dupuytren's disease.

The origin and mode of fibroblast migration and proliferation in granulation tissue.

The factors which regulate the properties of granulation tissue remain uncertain. The key cells in granulation tissue are fibroblasts and their role was investigated in rat skin excisional wounds. Full thickness wounds in 28 Hooded Lister rats were made and traced. On days 1 to 7, animals in groups of 4 received colchicine 1 mg/kg subcutaneously, 4 h before being killed. Wound sections were stained to locate and count mitotic cells. All wounds healed with a normal coefficient of contraction. The fibroblasts' mitotic indices (% of mitotic cells) were calculated. Dermis: 0.1% vs. 2.9% (control vs. test P < 0.001, paired t-test); fascia; 0.07% vs. 5.6% (control vs. test P < 0.001); granulation tissue: 9.7%. Granulation tissue fibroblasts come mainly from the adjacent fascia although the dermis may have an initial role. Once they are in the wound replication is extremely rapid. This study highlights for the first time the rapidity of fibroblastic migration and proliferation in excisional wounds, and suggests that future therapeutic interventions to control granulation tissue should consider these kinetics.

The in vitro enhancement of rat myofibroblast contractility by alterations to the pH of the physiological solution.

Wound contraction achieved by myofibroblast contraction is vital for the repair of cutaneous wounds. Many changes occur during tissue repair one of which is a lowering of pH. This study was designed to determine if myofibroblast contractility, as mimicked by using in vitro preparations, was sensitive to alterations of the pH. The responses of strips of rat superficial fascia when stimulated in vitro by adenosine, calcium and potassium ions, and mepyramine in physiological solutions at pH 5.5, 6.1, 7.3 and 8.1 were clearly pH dependent with acidic media producing an enhanced in vitro contractility. Perhaps modifying the pH of the wound environment could enhance wound contraction.

The qualities and conduct of an English surgeon in 1446: as described in a manuscript attributed to Thomas Morstede.

The year 1996 marks the 550th Anniversary of an anonymous manuscript which represents one of the earliest surgical works written in English. Generally attributed to Thomas Morstede, Serjeant-Surgeon to King Henry V, the book was for many centuries considered to have been lost and has escaped detailed examination by the surgeons of today. We present a modern translation of its first chapter in which the author outlines the range of equipment a fifteenth-century surgeon would use, the personal qualities all surgeons should possess, and the manner in which surgical practice should be conducted.

The contractility of knuckle pads. An in vitro study.

Strips of tissue from knuckle pads contract in response to the myofibroblast stimulant mepyramine in a reversible, repeatable and dose-dependent manner. The significance of this finding is discussed.

Back to the future for wound care?

The European Renaissance was a time of enormous change and rapid progress in the arts, sciences, and medicine. A glimpse of wound care in the last phase of the European Renaissance is provided by the analysis of work by Wilhelm Fabry, the "father of German surgery," as provided in his book De Combustionibus ("Burns") which details his range of treatments for the burn wound, as well as his approach to the later problems of scarring and contracture. We describe some of the historic events which may have stimulated Fabry's writings, in particular, the influences passed down from the medical school of Padua which thereby advanced the cause of wound care and surgery. Finally, we briefly explore the potential of such an approach to the works of our medical forefathers.

Human skin histology as demonstrated by Herovici's stain: a guide for the improvement of dermal substitutes for use with cultured keratinocytes?

The potential for the widespread use of cultured keratinocytes for burns treatment is handicapped by practical problems such as fragility, poor take and, often, unsatisfactory cosmesis. Although dermal equivalents reduce these problems there remains a lack of consensus on what is the best structure of such equivalents. At present the commonest support is type I collagen. This histological study, however, using Herovici's stain, clearly shows that in human skin from a variety of anatomical sites the epidermis is not in direct contact with type I collagen but rather with a distinct layer of type III collagen. We suggest that dermal equivalents may have to be constructed so as to include a layer of type III collagen at the interface between the keratinocytes and a type I collagen neo-dermis, so mimicking normal skin structure more closely.

A modern translation and interpretation of the treatise on burns of Fabricius Hildanus (1560-1634).

At the beginning of the seventeenth century, some surgeons were as interested in the classification, prognosis and treatment of burns as their successors are at the close of the twentieth century. We present extracts from our translation of the visionary treatise on Burns by Fabricius Hildanus (Wilhelm Fabry), the "Father of German surgery", and show how some of the present day approaches to burns have their origins almost 400 years ago.

A picropolychrome staining technique applied to Dupuytren's tissue.

Although the histology of Duputyren's tissue is well-documented, conventional stains do not distinguish between the different types of collagen which biochemistry and immunochemistry suggest are present. Duputyren's specimens [nodules (n = 26), cords (n = 15) and dermofasciectomies (n = 6)] were stained with haematoxylin and eosin, Van Gieson's Mallory's, Masson's, and Herovici's picropolychrome stain, and examined for both cellularity and collagen staining characteristics. All stains illustrated the marked cellularity of the nodules, contrasting with a paucity of cells within the cords. The first four stains demonstrated uniformity of the collagen staining within the tissues. Herovici's picropolychrome, however, showed distinct staining patterns for the dermis, nodules and cords, with both purple/red and blue areas. Other studies suggest that those fibres stained purple/red and blue are types I and III collagens respectively. These findings may shed further light on the tissue of origin of Duputyren's disease.

An experimental model to investigate the dynamics of wound contraction.

An excisional wound model in the Hooded Lister rat is described. The methods used to inflict, measure and characterise the process of wound contraction are comprehensively discussed. The model is shown to be reliable, reproducible and capable of detecting the effect of systemically administered prednisolone.

Modifications to the rate of wound contraction by allopurinol.

The effect of allopurinol (50 mg/kg) on the rate of full thickness excisional wound contraction in the Hooded Lister rat was assessed by planimetric and histological examinations. Compared with control animals, those treated with allopurinol showed a significantly lower coefficient of wound area contraction for days 0-7 (p < 0.05) than those of control animals. Histologically, in the allopurinol treated wounds the granulation tissue was less cellular but appeared to contain more collagen. This inhibition by allopurinol of the contraction phase of wound healing associated with an increased quantity of granulation tissue suggests that mediation of the process may involve a complex interaction between the fibroblasts/myofibroblasts and free radicals.