Effect of total venous occlusion on capillary flow and necrosis in skeletal muscle.

Limb replantation and microvascular transfer of flaps are sometimes complicated by postoperative venous thrombosis. Total venous occlusion can lead to complete shutdown of microvascular perfusion, resulting in failure of the transfer or replantation. Once venous return stops, it must be restored within a critical period of time for tissue survival. The purpose of this experiment was to delineate this critical period of time at which no reflow and irreversible muscle necrosis occurs by the use of a rat gracilis flap microcirculation model. The gracilis muscle of 40 male Wistar rats (135.3 +/- 37.2 g) was elevated on its vascular pedicle and mounted on a raised platform for videomicroscopic analysis. Animals were randomly assigned to one of four groups: (1) sham (no total venous occlusion), (2) 10 minutes of total venous occlusion, (3) 30 minutes of total venous occlusion, and (4) 60 minutes of total venous occlusion. Total venous occlusion was established by placing a microvascular clamp across the femoral vein at the junction of the gracilis pedicle. The number of flowing capillaries in five consecutive high-power fields (832x) were counted at baseline and at 5, 15, 30, 60, 120, 180 minutes, and 24 hours after reperfusion. At 24 hours after reperfusion, the gracilis muscles were harvested and stained with nitroblue tetrazolium. Percentage of muscle necrosis was measured by using computer planimetry. The data were reported as mean +/- standard error of mean and were compared between groups by analysis of variance and appropriate post hoc comparisons. Total venous occlusion for 10, 30, and 60 minutes showed a significant decrease in the number of flowing capillaries through 24-hour postreversal. There was a significant drop (p < 0.01) in the number of flowing capillaries from 30 minutes of total venous occlusion to 60 minutes of total venous occlusion at all times. Muscle necrosis was significantly increased in all three groups of total venous occlusion compared with the sham group (36.1 +/- 1.7 percent, 45.5 +/- 3.4 percent, 74.1 +/- 4.7 percent versus 14.3 +/- 1.7 percent, and p < 0.01). These results indicate that irreversible tissue damage occurs in a very short time interval (60 minutes) in this model, making the early detection of venous occlusion critical to the successful correction of this complication.

Timing of microcirculatory injury from ischemia reperfusion.

The low flow state that results from ischemia and reperfusion injury is a potentially reversible process that is important in numerous clinical situations. However, the point in time during the course of reperfusion where tissue injury becomes irreversible is unknown. This experiment evaluated the continuum of tissue damage in skeletal muscle after ischemic insult by quantifying the number of flowing capillaries and percentage muscle necrosis in a male Wistar rat skeletal muscle model. A gracilis muscle flap was raised on the vascular pedicle of 39 male Wistar rats and examined at 832x using intravital videomicroscopy. The numbers of flowing capillaries in five consecutive high-power fields were counted for baseline values. The flap was then subjected to 4 hours of global ischemia (except in sham animals, n = 7) by placing a microvascular clamp on the pedicle artery and vein. Upon reperfusion, flowing capillaries were counted in the same five high-power fields at intervals of 5, 15, 30, and 60 minutes, then at 2 to 8 (1-hour intervals), 24, and 48 hours. The gracilis muscle was then harvested at these intervals during reperfusion and assessed for viability. Compared with baseline, flowing capillaries from the ischemia and reperfusion group (mean +/- SEM) decreased significantly in the first 8 hours of reperfusion (7.7 +/- 0.2 to 3.2 +/- 0.3, p < 0.001) with minimal change noted from 8 to 48 hours. Percentage muscle necrosis increased progressively in ischemia and reperfusion preparations from 1 to 7 hours of reperfusion (16.5 +/- 2.6 percent to 38.9 +/- 1.2 percent, p < 0.001). No significant change in muscle necrosis in the ischemia and reperfusion group was noted between 7 and 48 hours. Sham preparations showed no change in the number of flowing capillaries through 3 hours of reperfusion, with a slight decrease at 24 hours. This rat gracilis microcirculation skeletal muscle model demonstrates a heterogeneous reperfusion injury. The decrease in flowing capillaries correlated with the increase in percentage necrosis and appeared to stabilize at the 7- to 8-hour interval. This finding may have important implications for the timing of interventions aimed at minimizing tissue damage from ischemia-reperfusion.

Effect of hyperbaric oxygen on neutrophil CD18 expression.

Previous work has shown that treatment with hyperbaric oxygen significantly reduces neutrophil adhesion to postcapillary venules in a rat microcirculation model of ischemia-reperfusion injury. The mechanism of this process is unknown. The purpose of this study was to evaluate the effect of hyperbaric oxygen on neutrophil CD18 adhesion sites by flow cytometry in an animal model of ischemia-reperfusion injury. The gracilis muscle flap was raised in three groups of male Wistar rats: (1) a sham group (n = 25), (2) a group that underwent 4 hours of ischemia (n = 25), and (3) a group that underwent 4 hours of ischemia and received hyperbaric oxygen (100% 02, 2.5 atmospheres absolute, during the last 90 minutes of ischemia) (n = 25). Samples from one subgroup of each group (n = 5) were divided into two portions, and one portion was stimulated with phorbol-12 myristate 13-acetate (PMA). Samples from another subgroup of each group (n = 5) were treated in the same manner, and a flap flush was added at the end of reperfusion to determine the number of CD18 adhesion sites on adherent neutrophils remaining in the flap. Venous blood was drawn 10 minutes after the operation, at 5 minutes of reperfusion, and at 90 minutes of reperfusion. Hematocrit and white blood cell count were measured. Samples were analyzed by flow cytometry, and the antibody binding capacity was assessed using microbead standards and linear regression (antibody binding capacity was expressed as the mean number of sites per cell +/- SEM). Microbeads were used to align the flow cytometer and to provide external and internal standards. Ischemia-reperfusion injury increased the expression of CD18 by neutrophils (p < 0.05). Expression of CD18 was not decreased by hyperbaric oxygen treatment. Stimulation with PMA increased the expression of CD18 in all groups (p < 0.01). These results suggest that ischemia-reperfusion injury does increase the expression of CD18 by neutrophils. Hyperbaric oxygen, as administered in this experiment, did not prevent the increase in CD18 expression.

L-selectin and leukocyte function in skeletal muscle reperfusion injury.

HYPOTHESIS: Treatment with anti-L-selectin monoclonal antibody will reduce venular neutrophil-endothelial rolling (flux and velocity) and adhesion associated with ischemia reperfusion injury in rat skeletal muscle. DESIGN: Prospective, randomized experimental trials. SETTING: Basic science research laboratory. MATERIALS: Male Wistar rats weighing 109 +/- 5 g (mean +/- SEM). INTERVENTIONS: Gracilis pedicle muscle flaps were elevated and microcirculation was observed by intravital microscopy. Two groups were evaluated: (1) the control group, which received 4 hours of global ischemia, and (2) the experimental group, which received 4 hours of global ischemia, plus treatment with anti-L-selectin monoclonal antibody 30 minutes before reperfusion. MAIN OUTCOME MEASURES: The number of rolling and adherent leukocytes in postcapillary venules were counted in the 2 groups at baseline and at 1 through 5, 10, 15, 20, 30, 45, and 60 minutes of reperfusion. RESULTS: Treatment with the monoclonal antibody to L-selectin significantly reduced the number of rolling leukocytes (flux) at 2 through 5, 20, 30, 45, and 60 minutes of reperfusion compared with controls (P<.05). Use of the monoclonal antibody significantly reduced the number of adherent neutrophils at 5, 10, 15, 20, 30, 45, and 60 minutes of reperfusion (P<.05). There was no significant difference in leukocyte velocity. CONCLUSION: L-Selectin plays a significant role in leukocyte rolling and adherence to venular endothelium in rat skeletal muscle ischemia reperfusion injury.

Role of the thromboxane A2 receptor in the vasoactive response to ischemia-reperfusion injury.

Neutrophil-endothelial adhesion in venules and progressive vasoconstriction in arterioles seem to be important microcirculatory events contributing to the low flow state associated with ischemia-reperfusion injury of skeletal muscle. Although the neutrophil CD-18 adherence function has been shown to be a prerequisite to the vasoconstrictive response, the vasoactive substances involved remain unknown. The purpose of this study was to evaluate the role of thromboxane A2 receptor in the arteriole vasoactive response to ischemia-reperfusion injury. An in vivo microscopy preparation of transilluminated gracilis muscle in male Wistar rats (175 +/- 9 g) (n = 12) was used for this experiment. Three experimental groups were evaluated in this study: (1) sham, flap raised, no ischemia (20 venules, 20 arterioles), (2) 4 hours of global ischemia only (19 venules, 22 arterioles), and (3) 4 hours of global ischemia + thromboxane A2 receptor antagonist (ONO-3708) (17 venules, 20 arterioles). ONO-3708 (5 mg/kg), a specific competitive antagonist of thromboxane A2 receptor, was infused at a rate of 0.04 ml/minute into the contralateral femoral vein 30 minutes before reperfusion. Mean arterial blood pressure was not changed at this dose of ONO-3708 (88 +/- 6 mmHg before infusion, 81 +/- 4 mmHg after infusion, n = 3). The number of leukocytes rolling and adherent to endothelium (15-sec observation) were counted in 100-microm venular segments, and arteriole diameters were measured at 5, 15, 30, 60, and 120 minutes of reperfusion. Leukocyte counts and arteriole diameters were analyzed with two-way factorial analysis of variance for repeated measures and Duncan's post hoc mean comparison. Statistical significance was indicated by a p < or = 0.05. The ischemia-reperfusion-induced vasoconstriction was significantly reduced by the thromboxane A2 receptor antagonist (ONO-3708). The mean arteriole diameters at 30, 60, and 120 minutes reperfusion were significantly greater in the treated animals than in the ischemia-reperfusion controls. Despite a significant increase in treated mean arteriole diameters, 30 percent of arterioles still demonstrated vasoconstriction. Neutrophil-endothelial adherence was not reduced by ONO-3708. Thromboxane A2 receptor blockade significantly reduces but does not eliminate ischemia-reperfusion-induced vasoconstriction in this model. This finding suggests that additional and perhaps more important vasoactive mediators contribute to vasoconstriction. Furthermore, thromboxane A2 receptor blockade has no effect on polymorphonuclear endothelial adherence.

Effects of L-NAME and L-arginine on ischemia-reperfusion injury in rat skeletal muscle.

The involvement of nitric oxide in ischemia-reperfusion injury remains controversial and has been reported to be both beneficial and deleterious, depending on the tissue and model used. This study evaluated the effects of the nitric oxide synthase inhibitor N(G)-nitro-L-arginine-methyl ester (L-NAME) and the substrate for nitric oxide synthase, L-arginine on skeletal muscle necrosis in a rat model of ischemia-reperfusion injury. The rectus femoris muscle in male Wistar rats (250 to 500 g) was isolated on its vascular pedicle and subjected to 4 hours of complete arteriovenous occlusion. The animals were divided into five groups: (1) sham-raised control, no ischemia, no treatment (n = 6); (2) 4 hours of ischemia (n = 6); (3) vehicle control, 4 hours of ischemia + saline (n = 6); (4) 4 hours of ischemia + L-arginine infusion (n = 6); and (5) 4 hours of ischemia + L-NAME infusion (n = 6). The infusions (10 mg/kg) were administered into the contralateral femoral vein beginning 5 minutes before reperfusion and during the following 30 to 45 minutes. Upon reperfusion, the muscle was sutured in its anatomic position and all wounds were closed. The percentage of muscle necrosis was assessed after 24 hours of reperfusion by serial transections, nitroblue tetrazolium staining, digital photography, and computerized planimetry. Sham (group 1) animals sustained baseline necrosis of 11.9 +/- 3.0 (percentage necrosis +/- SEM). Four hours of ischemia (group 2) significantly increased necrosis to 79.2 +/- 1.4 (p < 0.01). Vehicle control (group 3) had no significant difference in necrosis (81.17 +/- 5.0) versus untreated animals subjected to 4 hours of ischemia (group 2). Animals treated with L-arginine (group 4) had significantly reduced necrosis to 34.6 +/- 7.5 versus untreated (group 2) animals (p < 0.01). Animals infused with L-NAME (group 5) had no significant difference in necrosis (68.2 +/- 6.7) versus untreated (group 2) animals. L-Arginine (nitric oxide donor) significantly decreased the severity of muscle necrosis in this rat model of ischemia-reperfusion injury. L-arginine is known to increase the amount of nitric oxide through the action of nitric oxide synthase, whereas L-NAME, known to inhibit nitric oxide synthase and decrease nitric oxide production, had comparable results to the untreated 4-hour ischemia group. These results suggest that L-arginine, presumably through nitric oxide mediation, appears beneficial to rat skeletal muscle subjected to ischemia-reperfusion injury.

Single-vessel arteriovenous revascularization of the amputated ear.

Successful single-vessel arteriovenous replantation of a completely amputated human ear is described. This result was followed by an experimental study using a rabbit model, to confirm that an amputated ear could survive replantation with only a single arteriovenous anastomosis. Fifteen animals were placed in one of two experimental groups: Group 1-arteriovenous replantation, no treatment (n = 6); and Group 2-arteriovenous replantation with medicinal leeching (n = 9). All ears demonstrated initial reperfusion of the replantation immediately following microanastomosis. Laser Doppler flow readings in the non-leeched replanted ears fell to zero by 8 hr and, by 12 hr, the non-leeched ears demonstrated signs of necrosis. All the leeched, replanted ears demonstrated perfusion and complete viability at the time of sacrifice. The case report, combined with the results from the experimental study, confirm that single-vessel arteriovenous replantation of an amputated ear is feasible.

Effect of hyperbaric oxygen and medicinal leeching on survival of axial skin flaps subjected to total venous occlusion.

This study evaluates the effect of hyperbaric oxygen and medicinal leeching on axial skin flaps subjected to total venous occlusion. Axial epigastric skin flaps (3 x 6 cm) were elevated on their vascular pedicles in 40 male Wistar rats. Total venous occlusion was achieved by division of all veins draining the skin flap. Arterial inflow was left intact. Animals were randomly assigned to one of five groups: sham (n = 8); control, total venous occlusion only (n = 8); occlusion with hyperbaric oxygen (n = 8); occlusion with leeching (n = 8); occlusion with leeching and hyperbaric oxygen (n = 8). The hyperbaric oxygen protocol consisted of 90-minute treatments, twice daily, with 100% O2 at 2.5 atmospheres absolute for 4 days. The leeching protocol consisted of placing medicinal leeches on the congested flaps for 15 minutes, once daily, for 4 days. Laser Doppler measurements of flap perfusion were recorded preoperatively, postoperatively, and on postoperative days 1 and 3. The percentage of flap necrosis was evaluated on postoperative day 3. Mean percentage necrosis and mean laser Doppler readings were compared between both groups. The flaps in the sham group demonstrated 99 percent survival, whereas the flaps in the occlusion-only group demonstrated 100 percent necrosis. The flaps in the occlusion with oxygen, the occlusion with leeching, and the occlusion with oxygen and leeching groups demonstrated 1, 25, and 67 percent survival, respectively. Sham laser Doppler readings remained within normal limits. Laser Doppler readings in the occlusion-only and the occlusion with oxygen groups decreased to negligible levels on postoperative day 1, and on postoperative day 3 no perfusion was demonstrated. In both the occlusion with leeching and the occlusion with leeching and oxygen groups, there was also a significant decrease in laser Doppler measurements after surgery, but perfusion remained stable throughout the remainder of the study. This study demonstrates that hyperbaric oxygen alone is not an effective treatment for skin flaps compromised by total venous occlusion. The combination of leeching and hyperbaric oxygen treatment of total venous occlusion results in a significant increase in flap survival above that found with leeching alone. It appears that hyperbaric oxygen is effective because of the venous outflow provided by leeching as demonstrated by laser Doppler flow readings.

Assessment of intraneural oxygen tension and blood flow in a mobilized peripheral-nerve model.

Extensive mobilization of injured peripheral nerves is often required to allow a tension-free repair. The purpose of this study was to determine the effect of surgical mobilization and division on intraneural blood flow and oxygen tension in a rat sciatic nerve model. The right and left sciatic nerves were exposed in male Lewis rats Four experimental groups were studied: 1) nerve in situ/intact (n=7); 2) nerve mobilized and intact (n=7); 3) nerve in situ/divided (n=7); and 4) nerve mobilized and divided (n=7). Intraneural oxygen tension and blood flow were measured postoperatively. Mean oxygen tension (mmHg) in mobilized/intact nerves (17.47+/-4.79) was significantly lower than in in situ/intact nerves (38.32+/-5.16) [p < 0.05]. Mean oxygen tension in mobilized/divided nerves (3.10+/-1.25) was significantly lower than in in situ/divided nerves (30.30+/-6.36) [p < 0.005]. Laser Doppler flow (ml/min/100 g) in mobilized divided nerves (11.31+/-1.81) was significantly lower than in in situ/divided nerves (25.60+/-3.31) [p < 0.05]. Laser Doppler flow in mobilized/intact nerves was not statistically significantly different from in situ/intact nerves. Mobilization of the extrinsic blood supply was shown to significantly lower intraneura) oxygen tension and laser Doppler flow in mobilized intact and divided rat peripheral nerves.

Effect of hyperbaric oxygen therapy on testicular ischemia-reperfusion injury.

PURPOSE: Testicular torsion is a urologic emergency representing a form of ischemia-reperfusion (IR) injury that requires prompt care to achieve tissue salvage and a reduction in post-torsion morbidity. Hyperbaric oxygen (HBO) has shown benefits in previous musculoskeletal models of IR. We evaluated the efficacy of HBO treatment in a rat testicular torsion model. MATERIALS AND METHODS: Four groups of male Wistar rats were included in this study: 1) Sham (n=16), spermatic cords exposed but not occluded; 2) Control (n=16), 4 hours of bilateral spermatic cord occlusion; 3) HBO during ischemia (n=18), 4 hours of occlusion and administration of HBO during the last 90 minutes of ischemia; and 4) HBO on reperfusion (n=8), HBO administered immediately upon reperfusion of the testes. The animals were sacrificed at two weeks and architecture and germinal epithelial cell thickness were determined by histological examination on each testicle. Average thickness (in cell layers) of each group was compared with control using Student's t test. RESULTS: Control testicles showed a significant reduction in germinal cell thickness compared with sham (1.7 versus 6.3, p <0.05). The animals treated with HBO during ischemia showed a significant increase in epithelial cell thickness compared with control (2.8 versus 1.7, p <0.05). Hyperbaric oxygen treatment during reperfusion had the greatest beneficial effect compared with control (5.1 versus 1.7, p <0.05). CONCLUSIONS: Adjunctive HBO therapy administered during ischemia or reperfusion significantly reduced injury to the testicle in this animal model. These results suggest a potential benefit of HBO treatment in clinical situations of testicular torsion.

Effect of ketorolac tromethamine (Toradol) on ischemia-reperfusion injury in skeletal muscle.

The eicosanoids, leukotriene B4 (LTB4) and thromboxane A2 (TXA2), contribute to neutrophil adhesion and arteriole vasoconstriction, important microcirculatory events in ischemia-reperfusion (I-R) injury. The purpose of this study was to evaluate the effect of ketorolac on I-R injury of skeletal muscle. A videomicroscopic preparation of gracilis muscle in male Wistar rats (n=7) in two experimental groups was evaluated: Group 1-4 hr global ischemia only (19 arterioles, 19 venules), and Group 2-4 hr ischemia plus ketorolac (13 arterioles, 14 venules). Ketorolac (0.86 mg/kg, i.m.) was given 30 min prior to reperfusion. The number of neutrophils, rolling and adherent, was counted in 100-micron venular segments, and arteriole diameters were measured at 5, 15, 30, 60 and 120 min of reperfusion. The I-R-induced increase in neutrophil adhesion was significantly reduced by ketorolac, which significantly increased arteriolar vasodilation in the first 30 min of reperfusion and significantly reduced the I-R-induced vasoconstriction in arterioles at 30 min; this effect was lost at 1 hr of reperfusion. Although ketorolac augments immediate arteriole vasodilation and blocks subsequent vasoconstriction, this effect appears to be transient. These findings suggest that ketorolac may have potential as a treatment for I-R injury.

Evaluation of hyperbaric oxygen for diabetic wounds: a prospective study.

The purpose of this study was to prospectively evaluate the effect of hyperbaric oxygen (HBO2) on the healing of diabetic lower extremity wounds. Ten consecutive insulin-dependent diabetic patients with chronic lower extremity wounds were referred for HBO2 treatment. The control group consisted of five patients, two claustrophobic and three rural. The latter refused HBO2 treatments because of logistic reasons. Five patients underwent 30 HBO2 treatments in the problem wound protocol (100% oxygen, 2 atm abs, 2 h/day, 5 days/wk). All patients were evaluated with transcutaneous oxygen measurements and had an initial surgical debridement of the wound. Weekly tracings of the wound surface area were made by a nurse or resident who was blinded to the group assignment. At the end of 7 wk, the mean wound area expressed as a percentage of pretreatment baseline area was compared between groups (analysis of variance, Duncan's post hoc). No significant differences were noted between groups with respect to age, gender, baseline wound area, wound site O2 tension, or presence of osteomyelitis. At the completion of each of the 7-wk treatment periods, a significantly greater reduction in wound surface area was noted in the HBO2 vs. the control group (P < 0.05). HBO2 treatment significantly reduced wound size compared to controls in this small, non-randomized prospective study. These results should serve as a basis for larger multicenter prospective, randomized, double-blind controlled studies to definitively evaluate the effect of HBO2 on the healing of diabetic foot wounds.

Ischemia-reperfusion injury in skeletal muscle: CD 18-dependent neutrophil-endothelial adhesion and arteriolar vasoconstriction.

The purpose of this study was to evaluate if the venular neutrophil-endothelial adhesion associated with ischemia-reperfusion of skeletal muscle is dependent on leukocyte adhesion glycoprotein CD18 function and to determine if this interaction influences the vasoactive response in nearby arterioles. An in vivo microscopy preparation of transilluminated gracilis muscle in 13 male Wistar rats was used for this experiment. Observations of nonischemic muscle (sham) demonstrated this preparation to be stable for 8 hours with negligible change in neutrophil adherence or arteriole diameter. Three groups were evaluated in this study: (1) sham, no ischemia, no treatment (n = 5, 20 arterioles. 20 venules), (2) 4 hours of global ischemia only (n = 4, 19 venules, 22 arterioles), and (3) 4 hours of ischemia plus monoclonal antibody against CD18 (n = 4, 12 venules, 9 arterioles). The murine monoclonal antibody (WT.3, Seikagaku America, Inc.), which binds the rat leukocyte function antigen I CD18 chain, was infused into the contralateral femoral vein 30 minutes prior to reperfusion. The number of leukocytes rolling and adherent to endothelium (15 seconds of observation) was counted in 100-microns venular segments, and arteriole diameters were measured at various times during reperfusion. All counts and measurements were normalized to baseline preischemic readings for each animal. Mean changes from baseline were compared between groups. The increase in ischemia-reperfusion-induced neutrophil-endothelial adherence in venules was blocked by monoclonal antibody, but rolling behavior was not changed. The ischemia-reperfusion-induced progressive vasoconstriction in arterioles was blocked by monoclonal antibody. These results suggest that (1) neutrophil-endothelial adherence function associated with ischemia-reperfusion in this model is CD18-dependent, (2) neutrophil rolling function does not appear to be dependent on CD18, and (3) neutrophil CD18 function is a prerequisite for ischemia-reperfusion-induced arteriolar vasoconstriction. These findings provide important mechanistic information that may help explain the deleterious microcirculatory events associated with ischemia-reperfusion injury skeletal muscle.

Effect of hyperbaric oxygen on neutrophil concentration and pulmonary sequestration in reperfusion injury.

OBJECTIVE: To evaluate the effect of hyperbaric oxygen (HBO) on muscle flap arterial neutrophil concentration and pulmonary neutrophil sequestration following ischemia-reperfusion injury in a rat gracilis muscle microcirculation model. DESIGN: Prospective randomized experimental trials. SETTING: Laboratory. MATERIALS: Male Wistar rats weighing 150 to 300 g. INTERVENTIONS: Gracilis muscle flaps were raised on isolated vascular pedicles. Three groups were evaluated: (1) sham (flap raised, 4-hour observation, no ischemia, no HBO), (2) ischemia control (4-hour flap is ischemia, no HBO); and (3) ischemia (4-hour flap is ischemia) plus HBO (last 90 minutes of ischemia, 100% oxygen, 2.5 atm absolute). MAIN OUTCOME MEASURES: Flap pedicle arterial leukocyte and neutrophil concentrations were assessed at 5 and 90 minutes of reperfusion in the 3 groups. Pulmonary neutrophil sequestration was measured in a blinded fashion, using histologic examination and myeloperoxidase assay in the 3 groups at 5 and 90 minutes of reperfusion. RESULTS: Ischemia produced an increase in pedicle arterial leukocyte and neutrophil concentrations in blood flowing to the gracilis muscle flap; administration of HBO significantly reduced concentration to sham levels (P < .05). There were no significant differences in pulmonary neutrophil sequestration among the experimental groups. CONCLUSIONS: In this rat gracilis muscle microcirculation model, the increase in pedicle arterial leukocyte and neutrophil concentrations following ischemia-reperfusion injury was significantly reduced to sham levels by HBO treatment. This observed reduction was not attributable to HBO-induced pulmonary sequestration, which did not significantly change with HBO administration. Further investigation is required to elucidate the mechanisms of action of HBO in ameliorating ischemia-reperfusion injury in this model.

Functional evaluation of peripheral-nerve repair and the effect of hyperbaric oxygen.

The effect of hyperbaric oxygen (HBO) on peripheral-nerve recovery following devascularization and repair was studied, using the rat sciatic-nerve model. The right sciatic nerve was mobilized, stripped of the extrinsic blood supply, transected, and repaired in an epineurial fashion, using microsurgical technique. Following repair, animals were randomized into one of two groups: 1) control--no HBO (n = 20); 2) HBO treatment--twice daily for one week (1.75 hr dives, 100 percent O2, 2.5 ATA) (n = 16). Nerve recovery was assessed weekly (total of 10 weeks) by walking-track analysis, from which the sciatic function index (SFI) was calculated for each animal. Mean SFI scores were improved in the HBO-treatment group over controls, becoming statistically significant at weeks 7 through 10. These results suggest that functional recovery in transected, devascularized, peripheral nerves may be improved by 1 week of HBO treatment following microsurgical repair.

Effect of artificial airway on ear complications from hyperbaric oxygen.

Hyperbaric oxygen treatment is associated with an increased risk of barotrauma to the tympanic membrane and middle ear. An artificial airway may compromise normal eustachian tube function and equilibration of middle ear pressures. This retrospective study was designed to evaluate the risk of middle ear complications in 267 patients receiving hyperbaric oxygen (HBO) therapy and to compare those with and without artificial airways. Charts of all patients were reviewed for middle ear and tympanic membrane complications and myringotomy tube placement. Eighteen of the 267 patients had artificial airways. Seventeen (94%) of these 18 patients developed middle ear or tympanic membrane complications, and 11 (61%) required tympanostomy tubes for pain, hemotympanum, or serous otitis. In contrast, 114 (45.8%) of the 249 patients without airways developed ear complications, and 53 (21.3%) required tympanostomy tubes. These results suggest that patients with an artificial airway who are receiving HBO therapy are at greater risk for developing tympanic membrane and middle ear complications than nonintubated patients. Similarly, patients with artificial airways receiving HBO frequently require placement of tympanostomy tubes.

Evidence for the existence of a single enzyme catalyzing the phosphorylation of choline and ethanolamine in primate lung.

Choline kinase (ATP:choline phosphotransferase, EC 2.7.1.32) has been isolated and purified 1000-fold from adult African Green monkey lung with a yield of 10%. The purified enzyme also phosphorylated ethanolamine (ratio of ethanolamine kinase to choline kinase = 0.30). This ratio remained constant throughout the purification procedure. The Km for choline (3.0 - 10(-5) M) was lower than that of ethanolamine (1.2 - 10(-3) M.) Choline was also found to inhibit ethanolamine kinase activity by 50% at a concentration of 0.005 mM, while ethanolamine inhibited choline only at very high concentrations (100--150 mM). When the enzyme was subjected to inactivation by heat, hemicholinium-3, trypsin digestion, and p-hydroxymercuribenzoate, both ethanolamine kinase and choline kinase activities were destroyed at the same rate. Freezing and thawing in the absence of glycerol also destroyed both activities at the same rate. Based on these findings, we conclude that in adult African Green monkey lung tissue, there is only one enzyme for the phosphorylation of ethanolamine and choline, and that choline phosphorylation predominates.