Sarcomas are malignant tumors of mesenchymal origin, and can arise in soft-tissue and in bones. It has been suggested that the abnormal growth regulation in sarcoma cells may be due to an autocrine mechanism, in which the cells are stimulated by an endogenous production of growth factors. Platelet-derived growth factor (PDGF) has been detected in sarcomas, and may be one of the growth factors important for sarcoma growth.

PDGF, originally discovered in platelets, is produced by, and binds to, a variety of cells. PDGF plays several roles both in normal conditions and in disease.

Suramin is a polyanionic drug with antineoplastic activities, and is known to dissociate growth factors from their receptors. Suramin has been shown to inhibit growth in several tumors and tumor cell lines; however, some tumor cells have been unaffected, or even stimulated, by suramin.

The present work was performed in order to a) examine the effects of suramin on sarcoma growth in vivo; b) investigate the kinetics of extravascularly administered PDGF in vivo; c) establish and characterize human sarcoma cells in vitro, including their relation to PDGF; d) evaluate the effects of suramin on sarcoma growth in vitro; e) compare the effects of PDGF on sarcoma growth in vivo and in vitro.

Suramin was shown to inhibit growth of two different human osteosarcoma xenografts grown in nude mice. The action is believed to be mainly cytostatic, as the tumors continued to grow, albeit at a lower pace: the tumors of the suramin treated mice had a volume of one-third or less than the untreated ones. The percentage of cells in S and G₂-M cell cycle phases was increased by suramin treatment, suggesting a selective effect of the drug in the S and G₂ period.

Blood and serum levels of ¹²⁵I, after extravascular administration of ¹²⁵I-PDGF-AB by intraperitoneal, intramuscular or subcutaneous injection in mice, were found to rise to a maximum 2-4 hours after injection. The levels of radioactivity persisted over several hours. Precipitiation of serum with 10% trichloracetic acid revealed that more than 50% of the radioactivity was in a macromolecular form. Gel chromatography of the serum showed that a major portion of the radioactive material in the circulation had the same molecular size as the original ¹²⁵I-PDGF-AB.

Eight cell lines derived from malignant fibrous histiocytomas (MFH) were established and characterized. A heterogeneity in the morphology of the MFH cell lines was noted. This heterogeneity was also reflected in the expression of mRNA for PDGF, transforming growth factor-alpha (TGF-/alpha/) and their receptors, ability to grow in serumfree media and secretion of PDGF into growth media. Two cell lines, able to grow in serum-free medium, coexpressed MRNA for PDGF, TGF-/aplpha/ and their receptors, suggesting that they may be regulated in an autocrine manner. However, other cell lines, unable to grow in a serum-free medium, also displayed this coexpression of mRNA. The simultaneous expression of a growth factor and its receptor is therefore not generally indicative of an autocrine mechanism.

All cell lines, unable to grow in a serum-free medium, were growth inhibited by high-dose suramin (200 ug/ml). The two cell lines, previously noted to grow under serum-free conditions, were not affected by the high-dose suramin treatment. The finding that only serum-dependent human MFH cell lines were inhibited by high doses of suramin indicates that serum dependence in vitro may predict sensitivity of sarcoma cells to suramin.

Two human sarcoma xenografts, one osteosarcoma and one malignant fibrous histiocytoma, were treated with human PDGF-AB when grown in nude mice. No effects on tumor growth were noted, although immunohistochemical studies revealed an expression of PDGF receptors. Furthermore, both sarcomas were markedly stimulated by PDGF-AB in vitro. It is concluded that mechanisms or factors other than available PDGF were limiting the growth of the examined tumors in vivo.

We have investigated the thioredoxin (TRX) levels in severely burned patients and the possible origin of TRX, based on the recent understanding that TRX is a potent antioxidant with cytoprotective functions. Serum and plasma samples from burns patients and healthy blood donors were collected during the first 10 post-bum days and analyzed in a sandwich TRX enzyme-linked immunosorbent assay (ELISA). The TRX levels found were correlated to a panel of blood tests. The presence of TRX in platelets was investigated by immunoelectron microscopy and Western blotting. TRX serum levels of the severely burned patients showed a significant increase, with a mean serum TRX concentration on the day of injury of 76.5 ▒ 19.5 ng/ml (mean ▒ SD) and on post-burn day one 122.6 ▒ 66.9 ng/ml, compared to control blood donor levels of 22.7 ▒ 12.2 ng/ml (p = 0.0041 and 0.0117, respectively). A second peak of increase was found on post-burn days 7 to 9 with a four- to five-fold rise in concentration compared to controls. TRX elevation correlated well with increased platelet (p = 0.007) and leukocyte counts (p = 0.002). We also demonstrated by immunoelectron microscopy and Western blotting the presence of TRX in platelets. In conclusion, our demonstration of TRX release in burn injuries indicates that the TRX system is involved in a rapid antioxidant defense, coagulation processes, cell growth, and control of the extracellular peroxide tone intimately linked to cytoprotection and wound healing in burns. One of the cell types that delivers TRX promptly and efficiently into the blood may be the platelet.

The Os centrale carpi is a relatively rare accessory carpal bone of the wrist that infrequently has been reported to cause symptoms. This report describes 2 cases where an apparently mobile Os centrale carpi caused painful clicking and crepitus and where the symptoms disappeared after arthroscopic removal of the ossicles.

This stndy comprising six separate papers, is concerned with the techniques of, and bevaluation of arthroscopy as a diagnostic and therapeutic tool in different pathologic conditions in the upper extremity.

Arthroscopy proved to increase diagnostic accuracy in 177 examined patients with anterior shoulder instability or shoulder pain. Previously not well described patterns of pathological morphology in the ventral joint capsule after anterior shoulder dislocations were disclosed. A new classification system of rotator cuff pathology to be used for arthroscopy in the subacromial impingement syndrome is suggested. Different stages of rotator cuff pathology were found to influence the clinical results after arthroscopic acromioplasty in 79 patients.

An anatomical stndy on 16 cadaver elbows revealed the close proximity between commonly used arthroscopic portals and important neurovascular structures. The usefulness and potential risks of each portal were demonstrated and a preferred procedure for diagnostic elbow arthroscopy was suggested.

Wrist arthroscopy was performed in 30 patients with long-standing post-traumatic pain and resulted in increased diagnostic accuracy. In patients where clinical examination and radiography had shown no abnormality, arthroscopy demonstrated serious carpal ligament injuries and related instability.

A technique for arthroscopic wrist synovectomy was described in patients with rheumatoid arthritis. In 18 wrists of 16 patients, a reasonably radical synovectomy could be achieved with this atraumatic method. Primary results showed decreased pain and increased grip-strength in all patients and an increased range of motion in some.

Heparan sulphate (HS) represents a heterogeneous class of molecules on cell membranes and extracellular matrices. These molecules are involved in a variety of biological processes, including immune responses, through their binding and functional modulation of proteins. Recently a panel of HS-epitope-specific, human single chain antibodies have been generated by phage display, facilitating analysis of the structural heterogeneity of HS in relation to pathological conditions. In a pilot study a heterogeneous staining pattern in melanoma metastases was observed with one of the clones (EW4G1). Using a double-staining technique, the expression of this epitope was studied in 12 metastatic melanoma lesions in relation to the presence of a CD3 + cell infiltrate. Different staining patterns with EW4G1 were observed in the different lesions. The different staining patterns were associated with the presence and pattern of inflammation with CD3+ cells. A pronounced staining pattern of blood vessels with EW4G1 was associated with a more or less brisk presence of CD3+ cells, while a pronounced staining of tumour cells or tumour cell matrix or absence of staining with EW4G1 was associated with absence of CD3+ cells. These results suggest a dualistic role for HS in the recruitment and intratumoural migration of CD3+ cells, depending on the location of expression of its epitope recognized by EW4G1. Further characterization of the structural diversity of HS and its function in T-cell recruitment and migration is therefore warranted, since detailed understanding of this relation may provide new targets for therapeutic intervention, such that better homing and migration of T cells (in)to tumours might be achieved in immunologically based treatment strategies.

A large number of studies have indicated that specific immune reactivity plays a crucial role in the control of malignant melanoma. In this context, expression of MHC I, MHC II and B7 molecules by melanoma cells is seen as relevant for the immune response against the tumour. For a better understanding of the biological relevance of MHC II and B7 expression by tumour cells in metastatic melanoma, we studied the expression of these molecules in melanoma metastases in relation to the inflammatory response, regression of the tumour and survival from 27 patients treated with biochemotherapy (30 mg m-2 Cisplatin and 250 mg m-2 decarbazine (dimethyl-triazene-imidazole-carboxamide, DTIC) on days 1-3 i.v., and 107 IU IFN-a2b 3 days a week s.c., q. 28d). In 19 out of 27 lesions studied, we found expression of MHC II by the tumour cells, while only in one out of II tumour biopsies obtained from untreated metastatic melanoma patients, MHC II expression was detected. Expression of B7.1 and B7.2 by tumour cells was found in nine out of 24 and 19 out of 24 lesions, respectively. In all cases where B7.1 expression was found, expression of B7.2 by the tumour cells was also seen. In general, no or only few inflammatory cells positive for B7 were found. Expression of MHC II by tumour cells was positively correlated with the presence of tumour-infiltrating lymphocytes, regression of the lesion, and with time to progression (TTP) and overall survival (OS) of the patient. However, no significant correlation between B7.1 or B7.2 expression and regression of the tumour, TTP or OS was found. In light of other recent findings, these data altogether do support a role as biomarker for MHC II expression by tumour cells, however, its exact immunological pathomechanism(s) remain to be established. ⌐ 2003 Cancer Research UK.

Background: There are indications that some features of gastric carcinoma are changing, with a possible impact on prognosis. The aim of this study was to examine any changes in type, location, stage, resection rate, postoperative mortality rate or prognosis for patients with gastric carcinoma in a well defined population. Methods: During 1974-1991, 1161 new cases of gastric adenocarcinoma were diagnosed in Ostergotland County, Sweden. Tumour location, Lauren histological type, tumour node metastasis (TNM) stage, radicality of tumour resection and postoperative complications were recorded after histological re-evaluation of tissue specimens and examination of all patient records. Dates of death were obtained from the Swedish Central Bureau of Statistics. Time trends were studied by comparing the intervals 1974-1982 (period 1) and 1983-1991 (period 2). Results: The proportion of diffuse type of adenocarcinoma increased (from 27 to 35 per cent), while that of mixed type decreased (from 16 to 9 per cent) and that of intestinal type was unchanged. The proportion of tumours located in the proximal two-thirds of the stomach increased (from 32 to 42 per cent) and the proportion of patients with tumours in TNM stage IV decreased (from 32 to 25 per cent). Overall tumour resection rates were unchanged, although the proportion of radical total gastrectomies increased (from 36 to 50 per cent). Excluding tumours of the cardia or gastric remnant after previous ulcer surgery, the 5-year relative survival rate after radical resection increased from 25 to 36 per cent and the postoperative mortality rate decreased for both radical (from 11 to 4 per cent) and palliative (from 18 to 6 per cent) resection. Conclusion: The patterns of tumour histology, location and stage of gastric carcinoma have changed in the authors' region. These changes were paralleled by a significant improvement in survival and postoperative mortality rates.

The vertical rectus abdominis (VRAM) flap has been used for reconstruction of sternal defects, particularly in the interior third, since it was first described 20 years ago. We describe 12 patients with mediastinitis or chronic sternal osteomyelitis after sternotomy treated between 1994 and 1997, nine performed at the Royal Hospitals Trust, London. Sternal osteomyelitis and mediastinitis after median sternotomy is an uncommon (0.4%-8.4%) but often fatal condition. Vascularised pedicles are the treatment of choice, and VRAM flaps were used in all cases. We report good long-term outcome with a follow up of 2-5 years, and no long-term morbidity relating to the VRAM reconstruction. We had only one partial failure of a flap. The operations were largely done in hospitals away from the plastic surgical unit in extremely sick patients, which illustrates the importance of multidisciplinary management to reduce hospital stay, mortality, and morbidity. We argue that early involvement of plastic surgical specialists in the treatment of sternal dehiscence is essential to ensure a successful outcome.

We studied regeneration distance of rat sciatic nerve, with the sensory pinch reflex test and immunocytochemical staining for neurofilaments, four to 21 days after transsection, repair, and enclosure of the repair site in either a non-absorbable silicone tube or an absorbable polyglycolic acid (PGA) tube. The size of both tube-types was carefully selected so that they did not compress the repaired nerve. The opposite nerve was repaired and not inserted in a tube (control). The regeneration distances in repaired nerves enclosed in silicone tube were significantly longer than the control side at all time points, a result not seen when PGA tube was used. The number of proliferating non-neuronal cells (incorporation of 5-bromodeoxyuridine (BrdU)) was studied just proximal to the site of nerve repair after six days. Numerous stained cells were seen, but there where no significant differences between the groups. We conclude that outgrowth of sensory axons after transsection and repair of rat sciatic nerve with sutures can be increased by enclosing the site of repair in a silicone tube but not in a PGA tube. The effect is probably not related to the number of proliferative non-neuronal cells.

Today, in vitro culturing of autologous cells is an established method in the field of tissue reconstruction. It can be applied to urothelial cells and could have many clinical implications in urological reconstructive surgery. This development calls for quality controls concerning cells used for clinical treatment when cells are autotransplanted back to the patient. We have studied cultured cells in order to detect whether genetic or morphologic changes occur. Urothelial cells isolated from bladder lavage were cultured according to different protocols based on the presence or absence of feeder cells. Genetic studies were performed by means of karyotyping with standard G-banding and interphase fluorescent in situ hybridization (FISH) analyses. The morphology of these epithelial cells was judged as well as immunostaining for epithelial cell markers. In addition, to minimize the risk of feeder cell contamination, proliferation studies were performed on cultures including feeder cells that had been pretreated with different doses of mitomycin or radiation. In initial studies, when using feeder cells in each passage according to standard protocols, urothelial cells proliferated unfavourably after the fourth passage with increasing numbers of mouse cells as well as urothelial tetraploid cells. We could also show that urothelial cells from bladder lavage need feeder cells in order to establish primary cultures. Further propagation up to 14 passages was performed without feeder cells and the urothelial cells retained normal karyotypes. We also found that mitomycin treatment had its main effect on feeder cells during the first 2 h. When feeder cells were irradiated, 20 Gy was effective and no feeder cell contamination was seen. In conclusion, we found that a high standard of quality in urothelial cell culturing can be achieved with a careful culturing technique.

Full thickness wounds, such as deep burns, need restoration of both the dermal and epidermal layers of the skin. In normal wound healing, re-epithelialization occurs by migration and proliferation of keratinocytes from the wound edges and by differentiation of stem cells from remaining hair follicles. Restoration of dermis occurs by influx of growth factors secreted by macrophages, platelets, and fibroblasts; by fibroblast proliferation and subsequent synthesis and remodeling of collagenous dermal matrix. In the case of full-thickness acute burn injuries and chronic wounds (e.g. pressure ulcers, venous ulcers and diabetic foot ulcers), these processes are defective. With the principles of tissue engineering in mind (to correct, improve and maintain tissues and their functions), researchers have developed promising materials and methods to make it possible to restore either the dermal (Integra^® DRT, Alloderm^®) or the epidermal layer (split thickness skin grafts (STSG), cultured epithelial autografts (CEA), autologous keratinocytes in single cell suspension). It is now well established that superior results are obtained if both dermal and epidermal components are combined, for example in a bilayered skin equivalent. Apligraf^® is recommended for use on venous ulcers and is the only bilayered living skin equivalent currently approved by the FDA. Studies on different factors affecting the wound healing capacity as well as techniques in use provide valuable information for further development.

In this licentiate thesis, we evaluated different transplantation techniques for delivering cultured human keratinocytes in single cell suspension, a measure becoming more frequently used in addition to STSG and CEA for restoring the epidermal layer of the skin. We found that the pressure device, commonly used to spray cell suspension onto the wound with pressures as high as 200 kPa, killed around 0% of the cells. In comparison, an ordinary syringe with the attachment of a spray nozzle showed almost 90% viable cells post transplantation and provided an equally good distribution of the cell suspension.

We also studied different silver containing dressings regarding silver accumulation in human skin. In addition, we graded the re-epithelialization to evaluate whether the dressings caused any delay in the wound healing process. We found that the silver dressings tested, with few exceptions, caused dermal accumulation of silver, primarily aggregated around blood vessels. We could also show that most of the dressings had negative effect on the re-epithelialization.

For the restoration of the dermal layer of the skin, Integra^® DRT functions as a scaffold for guided tissue regeneration of the dermis. We had the possibility to study a case of necrotizing fasciitis were the treatment consisted of the use of Integra^® DTR together with sub-atmospheric pressure (after initial surgical debridement) and later transplantation of split thickness skin grafts. This measure proved to be safe as well as giving satisfactory pliable and aesthetically acceptable result.

Local blood flow reflexes in the foot skin of healthy controls and in young diabetic patients with or without nerve dysfunction have been studied using laser Doppler perfusion imaging (LDPI). A neurophysiological follow-up study on nerve dysfunction is presented as a complerhent to the work on blood flow regulation in the young diabetics.

An enhanced high-resolution LDPI (EHR-LDPI), intended for visualization and interpretation of flow dynamics in separate microvessels, has been adapted and evaluated in in vitro tube models and in an in vivo tissue model (hamster cheek pouch). By focusing the laser beam to 40 µm in the focal plane and reducing the step length to 25 µm, full format images (4096 measurement sites) of microvascular tissue areas as small as 1,5 x 1,5 mm were created.

The objectives of the work were to study if the vasoconstrictor response seen during change in posture is a mechanism elicited by a rise in venous pressure, but also to investigate if young diabetic patients with nerve dysfunction have an impairment in the local regulation of foot skin blood flow (postural vasoconstriction and hyperemic response) compared to diabetics without nerve dysfunction. An additional aim was to elucidate whether abnormal nerve conduction is retarded or even prevented by tight metabolic control in patients with type 1 diabetes mellitus.

The experimental studies aimed to improve the resolution of the EHR-LDPI system, to evaluate the system flow response in an in vitro model and to evaluate the performance, the limitations and the future potentials by studying flow dynamics in a tissue containing separate microvessels. It was concluded that:

(1) The LDPI recorded skin perfusion during variations in venous stasis and posture, adding information on flow distribution changes. The difference in flow distribution seen suggested an additive regulatory mechanism to a venoarteriolar reflex during change in posture.

(2) Subclinical nerve conduction defects were more common than microvascular abnormalities as measured by LDPI in the present models in young diabetic patients. Although, no signs of established retinopathy or nephropathy in this patient group, resting skin blood flow abnormalities were present, and these findings were related to high HbA1c-levels.

(3) Tight longwterm metabolic control, with HbA1c values less than 6,5%, could retard nerve dysfunction in patients with type 1 diabetes mellitus and a mean disease duration of 12 years.

(4) Using EHR-LDPI a decrease in signal level was obtained as the tube diameter increased, although the algorithm scaled linearly with velocity and was found not to be sensitive to hematocrit variations. Individual microvessel diameters could be estimated, which on average resulted in a difference of 11 µm compared to microscopic measurements.

(5) A dynamic overview of the vascular tree with volumetric flow estimate as well as RBC velocities of separate vessels was obtained. The need for further focusing of the beam and reduction of the step length appeared to be important tasks to solve in order to get a more accurate vessel diameter determination and to refine the volumetric flow estimate.

An enhanced high-resolution laser Doppler imager (EHR-LDI), configured to fit the demands of a measurement area containing separate microvessels, was evaluated for perfusion measurements in hamster cheek pouch preparations during ischemia, reperfusion, and pharmacologically induced vasodilation and vasoconstriction. Measurements in separate microvessels where the laser beam was smaller than the vessel diameter were referred to as red blood cell (RBC) velocity estimates, as previously validated in vitro, whereas a relative flow index, RFI (mean RBC velocity/tissue area), was introduced as a volumetric flow measure. Microvessel diameter and RBC velocity changes during ischemia, reperfusion, as well as during vasoconstriction and vasodilation correlated to the data obtained from the microscope. Correspondingly, during the described provocations anticipated volumetric flow changes were registered as changes in the RFI. When data on intravessel RBC velocity profiles are presented they reflect a parabolic flow profile usually seen in this size microvessel. The EHR-LDI appears a promising tool for investigation of the microvasculature, as it almost simultaneously provides information on relative changes of both in vivo RBC velocity and volumetric flow (RFI), although the latter estimate needs to be further refined.

The exact nature of the decrease in foot skin blood flow seen after a change in posture remains unsettled. This mechanism has previously been examined by non-invasive techniques such as the laser Doppler perfusion monitor (laser Doppler flowmetry). Taking into account the shortcomings of laser Doppler perfusion monitoring when applied to the determination of skin blood flow, which normally shows substantial heterogeneity, we have applied an emerging technology, the laser Doppler perfusion imager (LDPI). This technique provides a more comprehensive picture of the blood flow distribution in the skin, as it maps skin blood flow over a surface area (120×120 mm, 4096 measurement sites). It was used to examine if the reduction in tissue perfusion or the alterations in flow distributions seen after a change in posture (supine to dependency) could be fully explained by an increase in venous pressure (venous stasis of 50 mmHg) or if the data suggest a complementary mechanism.

Skin blood flow of the forefoot decreased from 0.60 V (volt) (median) during rest to 0.40 and 0.38 V during venous stasis and dependency, respectively. Although almost identical median values were obtained during stasis and dependency, the flow distributions were different, with a loss of high flow values during venous stasis. Biological zero was 0.24 V.

As the LDPI technique readily records skin perfusion during variations in venous stasis and posture, as well as information on flow distribution changes, it appears promising for future application in stimuli-response studies of skin blood flow. The difference in flow distribution seen between increased venous pressure and dependency suggests an additive regulatory mechanism to the veni-vasomotor reflex during a change in posture.

Providing cutaneous wounds with sufficient epidermis to prevent infections and fluid loss is one of the most challenging tasks associated with surgical treatment of burns. Recently, application of cultured keratinocytes in this context has allowed this challenge to be met without several of the limitations connected with the use of split-thickness skin grafts. The continuous development of this novel approach has now revealed that transplantation of cultured autologous keratinocytes as single-cell suspensions exhibits several advantages over the use of cultured epidermal grafts. However, a number of methodological problems remain to be solved, primarily with regards to the complexity of culturing these cells, loss of viability and other negative effects during their preparation and transportation, the relatively long period of time required following transplantation to obtain a sufficiently protective epidermis. In the present investigation we attempted to eliminate these limitations by culturing the keratinocytes on macroporous gelatin spheres. Accordingly, the efficacies of normal human keratinocytes in single-cell suspension or growing on macroporous gelatin spheres, as well as of split-thickness skin grafts in healing wounds on athymic rats were compared. Human keratinocytes were found to adhere and proliferate efficiently both on the surface and within the pores of such spheres. Transplantation of such cells adherent to the spheres resulted in significantly more rapid formation of a stratified epidermis than did transplantation of single-cell suspensions or spheres alone. Twenty-three days after transplantation, the epidermis formed from the cells bound to the spheres was not as thick as the epidermis on wounds covered with split-thickness skin grafts, but significantly thicker than on wounds to which single-cell suspensions, spheres alone or no transplant at all was applied. Furthermore, fluorescence in situ hybridisation revealed that the transplanted keratinocytes, both those adherent to gelatin spheres and those in single-cell suspension, were components of the newly formed epidermis. These findings indicate that application of biodegradable macroporous spheres may prove to be of considerable value in designing cell-based therapies for the treatment of acute and persistent wounds. © 2006 Elsevier Ltd and ISBI.

The vascular bed in skeletal muscle plays an important role in the regulation of the systemic circulation. The present study was undertaken to investigate how capillary blood flow and oxygenation in skeletal muscle are affected by: a) a reduction in the levels of respiratory gases in blood, hypocapnia or hypoxia, and b) the interaction of anesthetics (pentobarbital, propofol, ketamine) during hemorrhage, and c) hypotension induced pharmacologically by adenosine, sodium nitroprusside or acetylcholine.

The experiments were performed on the vastus medialis muscle in mechanically ventilated anesthetized rabbits. Skeletal muscle microvascular perfusion was investigated with a local hydrogen clearance technique (LHC) (using a multi wire microelectrode) and laser-Doppler flowmetry (LDF). Skeletal muscle oxygen pressures (Pt02) and pH wereassessed using a multiwire Clark-type oxygen or an antimony (pH) microelectrode.

Hypocapnia (arterial PC02 2.3 kPa) decreased LDF flow and Pt02, whereasmuscle tissue pH remained unchanged. This was interpreted as being due to a reduction in microvascular perfllsion induced by vasoconstriction. This led to a decline in both tissue oxygenation and in the removal of acid metabolites, which counteracted a developing tissue alkalosis. Apart form the vasoregulatory role of carbon dioxide, it appears that muscle tissue pH is an important factor in the control of skeletal muscle perfusion. Hypoxia (arterial P02 4.0 kPa) reduced LHC flow and Pt02. This was reversed by the administration of ritanserin (serotonin antagonist), despite a further reduction in blood pressure. This supports the concept that the decline in capillary perfusion during systemic hypoxia is at least partly mediated by serotonin.

Hemorrhage was induced by withdrawal of blood to a mean arterial blood pressure of 40 mmHg, and measurements (LHC, LDF) were carried out during the spontaneous recovery period. When ketarnine wa<> used as anesthetic a higher capillary perfusion was found as compared to pentobarbital or propofol. These differences may be mediated, as suggested by the results from other studies, by the cardiovascular stimulating properties of ketamine, the effects on the renin-angiotensin system by pentobarbital and venomotor tone by propofol.

Hypotension, 20-25% reduction in mean arterial blood pressure, induced by adenosine, was associated with a decrease in skeletal muscle capillary blood flow and an increase in PtOz. This may be explained by a redistribution of capillary flow and/or a reduction in local oxygen demand. Hypotension induced by sodiuin nitroprusside to a similar level, on the other hand, increased capillary blood flow, mostly in the high flow range, whereas the oxygen pressure distributions were reduced. This may be caused by an increased local oxygen demand. Acetylcholine-induced hypotension decreased capillary blood flow which was most pronounced in the high flow range, while oxygen pressure distributions decreased homogeneously in a manner similar to that seen during hemorrhage.

The findings presented rnay be of importance in clinical situations with compromised skeletal muscle circulation.

We studied the effect of hyperbaric oxygen treatment on axonal outgrowth in grafts of sciatic nerves in 40 rats. The sciatic nerve was transsected and a 10 mm long segment from the opposite side was immediately sutured in as a nerve graft. Postoperatively 17 animals were treated with 100% oxygen at 3.2 atmospheres absolute pressure for 45 minutes and the treatment was repeated at four and eight hours postoperatively and then every eight hours until evaluation. At seven days the axonal outgrowth was evaluated by immunohistochemical staining of neuro-filaments in the nerve grafts. The axonal outgrowth was significantly longer in animals treated with hyperbaric oxygen. We conclude that hyperbaric oxygen can improve nerve regeneration in sciatic nerve grafts in rats.

Previous experimental studies have shown positive effects of hyperbaric oxygen treatment in the early regeneration phase in the first few days following a nerve injury. In this study, functional effects of hyperbaric oxygen treatment were studied in 2 series of rats after an injury to the sciatic nerve - a standardized crush injury and nerve transection and repair, respectively. Postoperatively the animals were treated with 100% oxygen at 2.5 atmospheres absolute pressure for 90 minutes and the treatment was employed twice daily for 7 days. The animals were evaluated with walking track analysis up to twice weekly. The experiments were terminated after 90 days when the tetanic force was measured in the tibial anterior and gastrocnemius muscles. No statistically significant differences were found in either of these tests. It is concluded that hyperbaric oxygen treatment, given in accordance with clinical protocols used in limb crush injuries and other peripheral conditions, was not effective in the restoration of gait or the muscular strength after 90 days in rats after these nerve injuries. This study does not support nerve crush injury or nerve transection and repair as indications for hyperbaric oxygen treatment.

A nerve injury in the hand often results in profound abnmmalities in sensory perception despite careful alignment and microsurgical repair in order to facilitate accurate nerve regeneration. The present experimental and clinical studies were undertaken in an attempt to functionally evaluate peripheral and central effects of nerve regeneration.

In the experimental studies the rat foot was used as a model for the human hand. Three months after neurotomy and repair a flUlctional evaluation was performed of regenerated and misrouted polymodal nociceptor C-fibers and low-threshold mechanoreceptive axons by mechanical stimulation on the foot and electrical recordings proximal to the lesion and by Evans blue albumin extravasation.

It was found that 1) functional regeneration of misrouted axons related to polymodal nociceptive units and low-threshold mechanoreceptive units is more efficient in hairy skin of the rat foot whereas only misrouted polymodal nociceptor C-fibers recover function in glabrous skin, 2) following epineural suture and repair with a silicone tube after sciatic neurotomy there is similar effect on the regeneration of polymodal C-fibers after three months, and 3) functional regeneration of C-fibers is more successful in 3-month-old than in new-born rats.

In the clinical studies the functional effects of median nerve injury and regeneration were evaluated by two-point discrimination test, electroneurography, somatosensory evoked potentials and functional magnetic resonance imaging (IMRI). A method oftMRI during tactile stimulation was developed, and the normal cortical activation during stimulation was studied in 12 healthy volunteers.

The effects of nerve injury were studied in 6 injured adult men 15-55 months after median nerve injury and repair. It was found that 1) the 2-point discrimination was > 15 mm, 2) the nerve conduction velocity and signal amplitude were decreased in the severed nerve segment, but the cortical evoked response was normal at nerve stimulation proximal to the lesion, 3) tactile stimulation of the glabrous skin of the hand in healthy volunteers caused bilateral cortical activation (fMRI) in the primary somatosensory cortex, 4) a loss of sensory discrimination in the hand after median nerve injury was associated with a normal or even elevated activation (fMRI) in the somatosensory cortex during tactile stimulation of the digit IT-m.

Object. The aim of this study was to assess the effects of median nerve injury and regeneration on neuronal activation in the somatosensory cortex by means of functional magnetic resonance (fMR) imaging and somatosensory evoked potentials (SSEPs). Methods. Ten injured male patients (mean age 26 years) were examined 15 to 58 months after a total transection of the median nerve at the wrist that was repaired with epineural sutures. Two-point discrimination was lost in Digit II-III and sensory nerve conduction displayed decreased velocity (-29%) and amplitude (-84%) in the median nerve at the wrist. The fMR images were obtained during tactile stimulation (gentle strokes) performed separately on the volar surface of either Digit II-III or Digit IV-V (eight patients: two were excluded because of movement artifacts). The SSEPs were obtained using electrical stimulation proximal to the median nerve lesion. Conclusions. Patients with loss of sensory discrimination after median nerve damage and regeneration had larger areas of activation in fMR imaging near the contralateral central sulcus during tactile stimulation of the injured compared with the noninjured hand. The increase relative to the unaffected hand was 43% (p < 0.02) for Digit II-III stimulation and 46% (p < 0.02) for Digit IV-V stimulation. The SSEP data showed normal latency and amplitude. The enlarged area of cortical activation may be the result of reorganization, and it may indicate that larger cortical areas are involved in the discriminatory task after a derangement of the peripheral input.

Peripheral nerve lesions are often complicated by difficulties in approximating the nerve tumps without tension. The aim of the present study was to evaluate C-fïbre function after nerve regeneration in rats in which the nerve had been lengthened by leaving a 5 mm gap inside a silicone tube (n = 5). The outcome was compared with nerve regeneration after epineural end-to-end suture (n = 5). The innervated skin territory was defined by Evans blue extravasation after antidromic nerve stimulation. Five rats acted as controls. After three months, there was similar functional reinnervation in both experimental groups, which indicates that silicone tubes may reduce tension over a nerve repair with no adverse effects.

It is generally believed that nerve injuries in children regenerate better than those which occur in adults. However, there are no functional experimental studies that support this belief. This study evaluates the functional regeneration of polymodal C-fibres after nerve regeneration in newborn and mature rats 3 months after unilateral sciatic nerve neurotomy and suture. The distribution of polymodal C-fibres was tested by measuring the Evans blue-stained area in the skin after antidromic nerve stimulation. In the newborn group of regenerated animals showed that functional C-fibres were present in a significantly (P<0.05) smaller area than found in the adult group. We conclude that the functional regeneration of C-fibres is superior in mature rats compared with newborns, 3 months after regeneration.

If a biodegradable scaffold is applied, the dermis can be regenerated by guided tissue regeneration. Scaffolds can stimulate in-growth of cells from the surroundings that migrate into them and start to produce autologous extracellular matrix as the scaffold is degraded. Several materials are available, but most of them are in the form of sheets and need to be laid on an open wound surface. A number of injectable fillers have been developed to correct soft-tissue defects. However, none of these has been used for guided tissue regeneration. We present a new technique that could possibly be used to correct dermal defects by using macroporous gelatine spheres as a biodegradable scaffold for guided tissue regeneration. In eight healthy volunteers, intradermal injections of macroporous gelatine spheres were compared with injections of saline and hyaluronic acid (Restylane (R)). Full-thickness skin biopsy specimens of the implants and surrounding tissue were removed 2, 8, 12 and 26 weeks after injection, and the (immuno) histological results were analysed. The Restylane (R) merely occupied space. It shattered the dermal tissue and compressed collagen fibres and cells at the interface between the implant and the dermis. No regeneration of tissue was found with this material at any time. The macroporous gelatine spheres were populated with fibroblasts already after 2 weeks. After 8 weeks the spheres were completely populated by fibroblasts producing dermal tissue. After 12 and 26 weeks, the gelatine spheres had been more or less completely resorbed and replaced by vascularised neodermis. There were no signs of capsular formation, rejection or adverse events in any subject. Further in vivo studies in humans are needed to evaluate the effect of the macroporous spheres fully as a matrix for guided tissue regeneration with and without cellular pre-seeding. However, the results of this study indicate the possibility of using macroporous gelatine spheres as an injectable, three-dimensional, degradable matrix for guided tissue regeneration.

For the majority of patients with metastatic malignant melanoma the prognosis is poor. Immunotherapy and biochemotherapy have shown promise with a subset of durable responses, but there is still a great need for a better understanding of the mechanisms of action during treatment to optimize future treatment schedules. In the present study Bcl-2 expression was studied in biopsies from ten patients with metastatic malignant melanoma (five with regional disease and five with systemic disease) treated with biochemotherapy, (cisplatinum 30 mg/m2 days 1-3, DTIC 250 mg/m2 days 1-3 i.v. and Interferon-a2b 10 MIU s.c. 3 days a week, on a 28-day cycle). The expression of Bcl-2 by the tumour cells was separately recorded in areas of histopathological regressive changes and in areas of unaffected tumour growth. Comparisons were made with biopsies from 14 untreated patients. In 10 of 10 treated patients a high expression of Bcl-2 by the tumour cells was found in areas of unaffected tumour growth. In contrast, only in 5 of 13 untreated patients was a high expression of Bcl-2 by the tumour cells found in these areas (P = 0.008). A significant difference was also found in the expression of Bcl-2 by the tumour cells between areas of unaffected tumour growth and areas of histopathological regressive changes (P=0.03). The significantly higher expression of Bcl-2 by the tumour cells in areas of unaffected tumour growth in treated patients compared to untreated patients indicates that clones with a high expression of Bcl-2 may be present after therapy, preventing apoptosis and eventually in many patients resulting in progressive disease. Supporting this concept, a difference was also found between the expression of Bcl-2 in areas of unaffected tumour growth, i.e. in areas of treatment failure, and the expression in areas of histopathological regressive changes. Thus immunohistochemical analysis of tumour biopsies shortly after therapy seems to be a good surrogate endpoint that allows a detailed analysis of Bcl-2 expression. The high expression of Bcl-2 shown in unaffected tumour areas after therapy suggests the need for additional treatment, e.g. Bcl-2 antisense therapy.

The therapeutic efficacy of biochemotherapy in metastatic malignant melanoma still carries a low remission rate, but with some durable responses. It would therefore be of considerable importance if patients with a high probability of responding could be identified using predictive tests. The response to interferon-alpha (IFN-a) correlates with the occurrence of CD4+ lymphocytes identified by fine-needle aspirates from melanoma metastases (Hσkansson et al, 1996). The present investigation studies a possible correlation between tumour-infiltrating CD4+ lymphocytes in malignant melanoma metastases and the therapeutic effect of biochemotherapy. A total of 25 patients with systemic and 16 with regional metastatic melanoma were analysed before initiation of biochemotherapy (cis-platinum 30 mg/m2 d.1-3, DTIC 250 mg/m2 d.1-3 i.v. and IFN-a2b 10 million IU s.c. 3 days a week, q. 28d.). A monoclonal antibody, anti-CD4, was used to identify tumour-infiltrating lymphocytes in fine-needle aspirates before start of treatment. The presence of these lymphocytes was correlated to response, time to progression and overall survival. A statistically significant correlation (P = 0.01) was found between the occurrence of CD4+ lymphocytes and tumour regression during biochemotherapy in patients with systemic disease. Out of 14 patients with moderate to high numbers of infiltrating CD4+ lymphocytes, 12 achieved tumour regression. In contrast, among patients with low numbers of these cells in metastatic lesions, 8 out of 11 had progressive disease. We also found a significantly longer time to progression (P < 0.003) and overall survival (P < 0.01) among patients with moderate to high numbers of these cells compared to patients with low numbers of these cells before initiation of biochemotherapy. Furthermore, in patients with regional disease, we found a significantly longer time to progression (P = 0.01) and a trend toward a longer overall survival time (P = 0.09). Based on these results and as previously shown with IFN-a therapy alone, there seems to be a need for CD4+ lymphocytes infiltrating the tumours before the start of biochemotherapy to make the treatment successful. Determination of these cells in fine-needle aspirates seems to be a method to predict responders to biochemotherapy, thus increasing the cost-benefit of this treatment strategy considerably, both in terms of patient adverse reactions and health care costs. ⌐ 2001 Cancer Research Campaign.

Immunotherapy and combination treatments such as biochemotherapy have shown promise, with higher response rates and a subset of durable responses, however, as the majority of responses are still of short duration, they do not provide any survival benefit. There is therefore a great need to better understand the mechanisms whereby tumours escape immune surveillance. The present study examines the expression of CD28 in patients with untreated and treated melanoma metastases. Twenty-eight patients with metastatic malignant melanoma were treated by biochemotherapy (cisplatinum 30 mg/m2 days 1-3, DTIC 250 mg/m2 days1-3 i.v., and IFN-a2b 10 million IU s.c. three days a week for 28 days treatment cycle). Tumours were resected post-biochemotherapy and analysed for the expression of CD28 in CD4+ and CD8+ lymphocytes in areas where histopathological regressive changes had occurred, and close to tumour cells in areas of unaffected tumour growth using a double-staining technique. A high percentage of the lymphocytes in areas with regressive changes were found to be CD4+ CD28-. In contrast, the vast majority of CD4+ lymphocytes migrating close to the tumour cells were found to be CD28+ (P<0.001). A similar difference in the expression of CD28 was also found for the CD8+ subset (P=0.004). A difference in down-regulation of the expression of CD28 was found between CD4+ and CD8+ lymphocytes both in the areas of regressive changes and in the unaffected tumour areas. The present study demonstrates that extensive down-regulation of the co-stimulatory factor CD28 is found in metastases following biochemotherapy. These results indicate that parameters of importance for the immune function have already undergone modification after one or two treatment cycles and that this down-regulation occurs in particular in areas with regressive tumour changes.

Although immunotherapy and biochemotherapy have shown promise, producing a subset of durable responses, for the majority of patients with metastatic melanoma the prognosis is still poor. Therefore there is a great need for predictive tests to identify patients with a high probability of responding. Furthermore, there is also a need for a better understanding of the mechanisms of action during treatment in order to be able to monitor the relevant antitumour reactivity during treatment and to optimize the efficacy of future immunotherapy and biochemotherapy. In the present study histopathological regression criteria were used to study the efficacy of biochemotherapy. Thirty-two patients with metastatic malignant melanoma (18 with regional disease and 14 with systemic disease) were treated with biochemotherapy (cisplatin 30 mg/m2 intravenously on days 1-3, dacarbazine 250 mg/m2 intravenously on days 1-3 and interferon-a2b 10 million IU subcutaneously 3 days a week, every 28 days). Pre-treatment fine needle aspirates were obtained from metastases to analyse the number of tumour-infiltrating CD4+ lymphocytes. Therapeutic efficacy was evaluated in metastases resected after treatment using histopathological criteria of tumour regression. Comparisons were also made with metastases from 17 untreated patients, all with regional disease. Regressive changes of 25% or more (of the section area) were found in two of the 17 untreated patients with regional disease compared with 13 of the 18 patients with regional disease and 10 of the 14 patients with systemic disease after biochemotherapy. Fifty per cent of the patients with regional disease showed a high degree of regressive changes (75-100% of the section area) after biochemotherapy. These results demonstrate the occurrence of an antitumour reactivity in the majority of patients. Patients with extensive regressive changes in 75-100% of the analysed biopsies were also found to have a longer overall survival (P = 0.019). In patients with regional disease there was a close correlation between a larger number of CD4+ lymphocytes pre-treatment and a higher degree of regressive changes post-treatment (P < 0.05). Thus, immunohistochemical analysis of tumour biopsies shortly after treatment seems to be a good surrogate endpoint This technique also allows detailed analysis of antitumour reactivity and escape mechanisms. ⌐ 2003 Lippincott Williams & Wilkins.

Facial palsy is a relatively common clinical condition with a variety of causes. Irrespective of its etiology, facial palsy always represents a very serious problem for the patient. This underlines the need for more effective treatment procedures. Retrospective evaluation of a clinical material of 16 patients with facial palsy treated at the University Hospital of Linköping during the period 1990-2000 showed that to improve the results of microsurgical nerve repair experimental research - controlled studies on homogeneous materials - is imperative.

To produce relevant experimental data we used a rat model. Dissections showed that the mandibular branch (MB) of the rat facial nerve is suitable for experimental studies. Electron rnicroscopy revealed that the normal rat MB contains some 2,200 axons, 1,825 of which are myelinated and show a unimodal size distribution with a mode at 4.5 µm. It was also found that the normal rat MB contains myelinated and unmyelinated sympathetic axons and that about half the C-fibers in the normal rat MB belong to capsaicin-sensitive putative polymodally nociceptive sensory neurons. Importantly, repair of the MB through transmedian grafting in one stage and in two stages, respectively, had largely similar outcomes in terms of anatomy.

These data evoked questions concerning the functional outcome of the two types of repair. Electrophysiological analysis (force recordings andelectromyography) revealed that repair of the rat MB through transmedian grafting in one stage gives a somewhat better functional restoration than repair in two stages.

The observations on the rat MB called for experimental studies on the effects of denervation and repair on mimic muscle. We found that the rat dilator naris muscle (DNM) is suitable for that purpose. The normal DNM contains 1,200 fast MyHC fibers with MyHC IIB fibers predominating. It is a very fast contracting muscle without static functions. A brief denervation of the DNM followed by spontaneous reinnervation by the MB did not influence fiber number, had long-term effects on fiber diameter, and had little effect on fiber types. Fiber number, fiber diameter, and occurrence of fiber types in the DNM remained abnormal both after immediate and delayed surgical repair of the MB. Long-term denervation of the DNM had severe effects on qualitative histology, fiber number, fiber diameter and fiber type distribution. Hence, a long delay between facial nerve injury and repair reduces the chances of restoring normal facial muscle function.

Conventional cross facial grafting includes division of a facial nerve branch (the donor nerve) on the intact side of the face. This non-optimal situation prompted us to test cross-facial grafting with a less traumatic end-to-side procedure in the rat. After coaptation of a sural nerve graft to the rat MB in an end-to-side fashion, including opening of a perineurial window, axons in the donor nerve emitted myelinated and umnyelinated sprouts into the graft. A predegenerated graft did not stimulate sprouting more than a fresh graft. Retrograde tracing with fast blue and fluoro-ruby showed that many sprouts originate from facial motor axons. Of all traced facial motoneurons 50% projected exclusively into the graft, 45% projected into the graft and the donor MB and 5% projected exclusively into the donor MB. This shows that the end-to- side procedure can be used for cross-facial repair of the rat MB.

Altogether these results provide new experimental data, which hopefully will contribute to improvements of the microsurgical treatment of patients with facial palsy.

Burn surgery has gone through revolutionary changes during the last century, even the most severe burns can now be safely treated. Modern surgical treatments have been followed by improvements in intensive burn care and anaesthesiology. These developments have allowed immediate surgery within 24 hours of the injury to take place in our centre, therefore obtaining advantageous results in shortening hospital stay, improving patient’s functional results and simplifying the treatment itself. This treatment is economically beneficial as well. In future we count on rapid development of a new discipline, tissue engineering, which should take a greater role in burn care.

The aim of this study is to analyse retrospectively the outcome of microsurgical treatment of patients with facial palsy at the University Hospital of Linköping during the period 1990-2000. Ten patients with facial palsy during 1 year or more completed the evaluation.

Patients subjected to cross facial nerve grafting (CFNG) only included 3 females. The CFNGs appeared to be fimctional in all patients. The measurements revealed good symmetry of the eye rim at rest and at eye closure. All three patients had an asymmetric angle of the mouth with a mean dropping of 2.3 mm at rest and 5.5 mm when smiling. The House-Brackmann grade was 4-5. The average Facial Grading System score was 33. The mean decrease in quality of life was 34%. The mean improvement after treatment was 48%. The degree of post-lesional impairment and the degree of postoperative satisfaction were inversely related. All patients were aware of their disability but they had not completely accepted the current facial status.

The group of patients with CFNG and a free muscle flap transfer included six females and one male. The transferred muscle flaps were activated by the CFNGs. Symmetry of the eye rims at rest was observed in 4 cases and with closure in 3 cases. In asymmetric cases, the mean difference was 1. 8 mm at rest. With eyes closed the mean difference was 4.4 mm. The angle of the mouth was asymmetric in all patients, the mean lowering being 3 mm at rest and 7.5 mm at smile. The House Brackmann grade was 4-5 and the average Facial Grading System score was 31. Four patients with early or congenital facial palsy could not provide data on impairment of quality of life. In the others, the post-lesional impairment was 63%. For all patients in this group the average postoperative improvement was 76%. The patients expressed a rather good acceptance of the achieved status after reconstruction.

Surgical methods to correct facial paresis should be directed towards achieving a symmetric resting tone on the injured side as well as symmetric movements. At present, our methods, when utilised on proper indications and with a precise timing, can give the patient a reasonably good but not perfect result. Further refinements of the technique are needed and should be sought through experimental studies.

This study examined by electron microscopy the normal fibre composition of the mandibular branch (MB) of the rat facial nerve and the outcome of axon regeneration in the MB after transmedian grafting in one or two stages. The average normal MB contained 2,185 axons, 17 % of which were unmyelinated. The myelinated axons had a unimodal diameter distribution (range 1.5-9.5 μm, mode 4.5 μm). After superior cervical ganglionectomy, the MB lost 1/3 of the C-fibres and 10% of the myelinated axons. In neonatally capsaicin-treated rats the occurrence of unmyelinated axons was reduced by about 50%. After repair in one or two stages the MB contained more myelinated and unmyelinated axons than normal. The myelinated axons showed a unimodal size distribution with a subnormal diameter range. Statistical comparisons showed that MBs from both experimental groups were significantly abnormal with respect to total axon number as well as numbers of unmyelinated and myelinated axons. In these respects the grafted MBs did not differ significantly from each other. However, the myelinated axons in MBs from one-stage cases showed larger mean and maximum diameters compared to MBs from two-stage cases. These data suggest that the normal MB of the rat contains myelinated and unmyelinated sympathetic axons and that about half the C-fibres in the normal MB come from capsaicin-sensitive sensory neurons. The comparison of the two reparative procedures used provides evidence in favor of the one-stage alternative.

A previous study examined the morphological outcome of axonal regeneration in the mandibular branch (ramus marginalis mandibulae) of the rat facial nerve after transmedian nerve grafting in one or two stages. The present study supplements the morphological data with a functional evaluation. Recordings of the force of tetanic muscle contractions elicited through stimulation of the mandibular branch showed that upper and lower lip data obtained from animals grafted in one stage did not differ significantly from control data. However, animals grafted in two stages exhibited significantly lower muscle forces compared to one-stage data and to control data. Electromyographic recordings of the M-response showed multiple prolonged potential fluctuations with subnormal amplitudes in grafted cases. In both groups of grafted rats, the mean voltage amplitudes recorded from the upper lip were weaker than the amplitudes seen at the angle of the mouth or the lower lip. The two-stage cases exhibited the most obvious deficit. In conclusion, the present results show that, with respect to the functional restoration achieved three months after nerve injury, repair through transmedian grafting in one stage gives better results than repair in two stages. This finding, which conforms with previous morphological data, suggests that the one-stage procedure should be considered for clinical use.

In spite of a continuous technical refinement the results of microsurgical facial nerve repair remain unsatisfactory. This situation prompted the present study, which evaluates axonal regeneration into a rat facial nerve branch after crossfacial end-to-side neurorrhaphy in adult rats. A fresh (group A, 12 rats) or predegenerated (group B, 12 rats) sural nerve graft (SNG) was sutured to a perineurial window in the mandibular branch (MB) of the right facial nerve. It was then tunneled under the chin and sutured to the distal stump of the cut left MB. Twelve weeks after grafting some rats were prepared for electron microscopy (EM). EM examination showed that distal to the window the right MB contained mainly myelinated axons with relatively thick sheaths. Aberrant small-diameter axons with thin sheaths were also observed in most specimens. The counts revealed a total average of about 1 800 axons (14 % unmyelinated) in both groups. In the SNG thin myelinated and unmyelinated axons predominated. The counts showed 900-1 000 axons (63-68% unmyelinated) with no significant difference between the groups. The left MB contained some thin myelinated axons and many unmyelinated axons. The number of axons was 1 200-1300 (81-83% unmyelinated). Again, the two groups did not differ. Other rats were subjected to retrograde tracing. The SNG-right MB coaptation was exposed and each nerve was cut off. Two or 3 days after application of fast blue (FB) to the right MB and fluoro-ruby (FR) to the SNG the animals were perfused with paraformaldehyde and the brain stem was transversely cryosectioned. Counts of labeled motoneurons were used to calculate the percentage of motoneurons with different tracer combinations. The results showed FBlabeling only (motoneurons with axons in the right MB distal to the coaptation) in 5-11% of the counted profiles, FR-labeling only (motoneurons with axons in the SNG) in 43-54% of the profiles, and a combined FB- and FR-labeling (motoneurons with axons in the right MB as well as in the SNG) in 41-46% of the profiles. The two groups did not differ statistically in this respect. We conclude that (i) coaptation of a SNG to the MB of the facial nerve in an end-toside fashion is followed by growth of myelinated and unmyelinated axons from the donor nerve into the graft; (ii) these axons originate from facial motor axons; (iii) some 50% of the traced facial neurons project into the SNG only, 45% project into the SNG and into the MB and 5% project into the MB only; (iv) predegeneration of the SNG does not enhance the number of axons in the graft.

This study describes changes in a rat facial muscle innervated by the mandibular and buccal facial nerve branches 4 months after nerve injury and repair. The following groups were studied: (A) normal controls; (B) spontaneous reinnervation by collateral or terminal sprouting; (C) reinnervation after surgical repair of the mandibular branch; and (D) chronic denervation. The normal muscle contained 1200 exclusively fast fibers, mainly myosin heavy chain (MyHC) IIB fibers. In group B, fiber number and fiber type proportions were normal. In group C, fiber number was subnormal. Diameters and proportions of MyHC IIA and hybrid fibers were above normal. The proportion of MyHC IIB fibers was subnormal. Immediate and delayed repair gave similar results with respect to the parameters examined. Group D rats underwent severe atrophic and degenerative changes. Hybrid fibers prevailed. These data suggest that spontaneous regeneration of the rat facial nerve is superior to regeneration after surgical repair and that immediacy does not give better results than moderate delay with respect to surgical repair. Long delays are shown to be detrimental.

Background: Severe contusion of an artery often presents clinical problems in that it affects flow distal to the injury. However, the effect of a contusion on the microvascular flow regulation in the distal part of the limb is still largely unknown. Methods: A multipoint microelectrode technique was used to assess both tissue oxygenation (P(t)O2) and microflow (hydrogen clearance) on the skeletal muscle surface in a standard contusion injury to the femoral vessels in rats. Results: A significant increase in and an altered distribution of (P(t)O2) as well as a reduction in and altered distribution of microflow on the muscle surface distal to the injury was found in all animals (n = 27) compared with the uninjured control leg. These findings could not be reproduced experimentally by sympathectomy or when the adjacent skeletal muscle alone was injured. Conclusion: The results suggest that the changes observed distal to the injury are of vascular origin, possibly as a result of endothelial damage at the site of the contusion.