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Introduction: Healing wounds are characterized by high lactate levels. In the past, the shift to an anaerobic energy metabolism was thought to be the main source for increased lactate concentration. Lactate is also generated by leukocytic NADPH-linked oxygenase using oxygen and glucose as one of its substrates, a mechanism called aerobic glycolysis. We hypothesize that hyperbaric oxygen therapy by increasing oxygen availability enhances wound lactate production by aerobic glycolysis. Material and methods: Four wire mesh wound cylinders (n = 142) were implanted underneath the dorsal skin in each of 36 male Sprague-Dawley rats (312 ± 11 g). Animals were randomized to two groups: The first group (n = 18) received 100% oxygen with 2.1 ATA for 90 minutes twice a day for a total of 7 days. The second group (n = 18) was treated with 21% oxygen with 1 ATA for the same time schedule. Hyperbaric treatments were administered in a hyperbaric chamber designed for small animals. Wound fluid from the cylinders was aspirated between the two treatment cycles at day 2 and 5 and analysed for lactate using a lactate analyser. Additionally, at day 10 (after 2 days of treatment with 21% oxygen in 1 ATA in both groups) lactate concentrations in wound fluid were measured again. Data are expressed as mean ± SD. A p-value

Hypothesis: Both hypoxia and lactate independently induce VEGF synthesis. However, an adequate supply of oxygen is required for new vessels to mature. Materials and Methods: We used in vivo matrigel (growth factor reduced) angiogenesis assay and Swiss Webster mice n = 36 (3 groups of 12 with 6 control and 6 experimental in each). Each mouse received a 1 ml subcutaneous matrigel injection. Matrigel solidifies into a gel implant when injected subcutaneously. Control injections/implants were matrigel with no added substance and experimental implants were matrigel supplemented with 30 mgs of crushed lactate polymer or PL (poly DL-Lactide-co-Glycolide 50:50; av. mol.wt. 50–75,000 kilo Daltons). The first group of mice was maintained in 12–14% oxygen, the second group in 21% oxygen, and the third group in 50–52% oxygen continuously for 11 days, at which time the implants were harvested, fixed, sectioned, and stained with Mason’s Trichrome to identify new vessels and collagen. Results: 1. 13% oxygen control: Half showed new blood vessels with thin or incomplete walls and hemorrhages. Few collagen fibers 2. 13% oxygen with PL: same as above 3. 21% oxygen control: none showed new blood vessels/collagen fibers 4. 21% oxygen with PL: five out of six showed new blood vessels. Almost all were mature, with complete walls, collagen fibers, and no hemorrhages 5. 51% oxygen control: None showed new blood vessels/collagen fibers 6. 51% oxygen with PL: all six developed new vessels with some hemorrhage seen in two out of six. Collagen fibers present Note: matrigel contains collagen and stained a pale blue, collagen fibers were seen as dark blue, endothelial cells and RBCs as dark red and red. All blood vessels contained RBCs. Conclusions: Hypoxia and lactate induce VEGF. However, hypoxia limits vascular growth by inhibiting collagen deposition. The largest and most mature new vessels result from a combination of oxygen and lactate.

The significance of the high lactate levels that characterize healing wounds is not fully understood. Lactate has been shown to enhance collagen synthesis by fibroblasts and vascular endothelial growth factor (VEGF) production by macrophages and endothelial cells. VEGF has been shown to induce endothelial cell migration. However, it has not been shown whether accumulated lactate correlates with the biological activity of VEGF. Therefore, we investigated the effect of lactate on migration of endothelial cells. Human umbilical vein endothelial cells and human microvascular endothelial cells were cultured to subconfluent monolayers in standard six-well tissue culture plates. Following a 24-hour serum starvation, cells were treated with the indicated concentrations of l-lactate. Cell migration was assessed using a modified Boyden chamber. VEGF protein in the cell culture supernatant was measured by enzyme-linked immunoassay. Lactate-enhanced VEGF protein synthesis in a time- and dose-dependent manner. Lactate added into the bottom well did not stimulate cellular migration from the upper well. However, lactate when added together with endothelial cells to the bottom well of the Boyden chamber increased cellular migration in a dose-dependent manner. This effect was blocked by anti-VEGF and by cycloheximide. Lactate enhances VEGF production in endothelial cells, although lactate, itself, is not a chemoattractant. We conclude that the lactate-mediated increase in cellular migration is regulated by VEGF.

For many years, lactate has been known to accelerate collagen deposition in cultured fibroblasts and, without detailed explanation, has been presumed to stimulate angiogenesis. Similarly, hypoxia has been linked to angiogenic effects and collagen deposition from cultured cells. Paradoxically, however, wound angiogenesis and collagen deposition are increased by breathing oxygen and decreased by hypoxia. Lactate accumulates to 4-12 mM in wounds for several reasons, only one of which is the result of hypoxia. Oxygen in wounds is usually low but can be increased by breathing oxygen (without change in lactate). We have reported that lactate elicits vascular endothelial growth factor (VECF) from macrophages, as well as collagen, some heat shock proteins, and VECF from endothelial cells, and collagen from fibroblasts, even in the presence of normal amounts of oxygen. Hypoxia exerts many of these same effects in cultured cells. In this study, we elevated extracellular lactate in wounds by implanting purified solid-state, hydrolysable polyglycolide. A steady-state 2-3 mM additional elevation of lactate resulted. With it, there was a significant short-term elevation of interleukin-1beta, a long-term elevation of VECF (2x) and transforming growth factor-beta1 (2-3x), a 50% elevation in collagen deposition, and a large reduction of insulin-like growth factor-1 (- 90%). We propose that lactate induces a biochemical "perception" of hypoxia and instigates several signals that activate growth factor/cytokine signals while the continued presence of molecular oxygen allows endothelial cells and fibroblasts to reproduce and deposit collagen. The data are consistent with ADP-ribosylation effects and oxidant signaling. (WOUND REP REG 2003;11:504-509)

Macrophages respond to various stimuli to produce angiogenic factors but few mechanistic details are known. We examined the effects of hypoxia, lactate and nicotinamide on the expression of vascular endothelial growth factor by cultured macrophages. These agents were chosen because they down-regulate polyadenosine diphosphoribose levels. Following exposure, conditioned media were analyzed for vascular endothelial growth factor protein. Nicotinamide adenine dinucleotide, polyadenosine diphosphoribose, and vascular endothelial growth factor mRNA were measured in the cellular fraction. Angiogenic capacity of the conditioned media was tested in rabbit corneas and Matrigel implants. All three agents, hypoxia, lactate and nicotinamide, elicited significantly increased levels of vascular endothelial growth factor mRNA and vascular endothelial growth factor in the conditioned media, and these levels were paralleled by their angiogenic activity. Polyadenosine diphosphoribose in the cellular fraction was correspondingly depressed. Anti-vascular endothelial growth factor antibody inhibited most of the angiogenic response whereas anti-basic fibroblast growth factor antibody had little effect. We propose that redox changes associated with the alteration of cellular nicotinamide adenine dinucleotide and polyadenosine diphosphoribose are involved in lactate-mediated VEGF expression.

Targeted disruption of the stromelysin-1 gene in mice causes a delay in excisional wound healing due to a failure in wound contraction. Therefore, we postulated that stromelysin-1 activity is responsible for initiating contraction. To test this hypothesis, we compared the contractile capacity of fibroblasts from stromelysin-1 knockout mice (strom-1 KO) with that of normal fibroblasts using a collagen gel contraction model. Fibroblast cultures were established from explants of skin and lung parenchyma from strom-1 KO and wild-type mice, then transferred to the surface of collagen gels. The extent of contraction was determined by measuring greatest gel diameter. Results demonstrated that (1) all fibroblasts contracted collagen gels in a uniform concentric fashion, (2) skin fibroblasts from both sets of mice exhibited greater gel contraction than did lung fibroblasts, and (3) strom-1 KO fibroblasts demonstrated significantly less contraction (21-23%) than wild-type fibroblasts. These data support the hypothesis that absence of stromelysin-1 results in defective fibroblast contraction that may contribute to delayed wound healing.

The secretion of growth hormone, an important anabolic agent, declines with aging. We hypothesize that growth hormone levels (measured as insulin-like growth factor-1) correlate with postoperative tissue repair in otherwise healthy, elderly persons. The goal was to determine whether growth hormone supplementation can improve wound healing in this circumstance. We conducted a randomized controlled double-blind trial of 6 months of growth hormone replacement or placebo in 28 healthy older men (>69 years of age) with low baseline plasma insulin-like growth factor-1. Growth hormone doses were adjusted to elevate insulin-like growth factor-1 to levels expected in younger adults. Wound healing was tested by implanting 10 cm expanded polytetrafluoroethylene porous tubes for 10 days, then measuring the content of collagen (as hydroxyproline), DNA, and total protein. Hydroxyproline content was 15% greater in the wounds of the growth hormone group (n = 13) compared with the placebo group (n = 15), (4.52 +/- 0.94 versus 3.92 +/- 0.78 microg/cm; p = 0.04). Therefore, healthy older men who took growth hormone had enhanced reparative collagen deposition during the wound healing process. This action may be clinically useful after selected surgery or trauma in the elderly.

Complications of irradiated tissue include infections and impaired healing. Although fibrosis and hypovascularity contribute, a cellular mechanism has not been identified. This study examines the effect of radiation (10 to 30 Gy) on neutrophil function in a rabbit wound cylinder model. At 3 to 12 weeks after radiation, subcutaneous wound cylinders were implanted in both irradiated and control fields in 19 rabbits. Wound neutrophils were subsequently assayed for phagocytosis ( 3 H-labeled Staphylococcus aureus assay), superoxide production (cytochrome c reduction assay), and surface Mac-1 expression (flow cytometric assay using MHM 23 monoclonal antibody). Phagocytosis of 3 H-labeled S. aureus was significantly lower in neutrophils from irradiated fields compared with controls at 6 and 12 weeks after radiation (6.5 versus 18.9 bacteria per neutrophil at 12 weeks ; p = 0.027). Stimulated neutrophils from irradiated tissue could not increase superoxide production or Mac-1 expression as much as controls, with differences increasing as postirradiation time increased. The diminished phagocytosis, superoxide production, and Mac-1 expression provide a cellular mechanism that may account for susceptibility to infection and poor healing in irradiated tissues.

Background and Purpose. Microamperage direct current and microamperage electromagnetic stimulation are used to accelerate healing in bone. Although many clinicians are using microamperage stimulation to relieve pain and facilitate wound healing, there is significant question regarding whether this low-intensity direct current significantly accelerates soft tissue wound repair. The purpose of this study was to determine whether low-voltage pulsed microamperage current (100 μA, 60 V, 0.1 Hz) enhances the healing of superficial, full-thickness, or incisional wounds created to simulate acute abrasions, ulcers, and lacerations. Subjects. Ten adult Yucatan mini pigs served as the subjects for this study. Methods. Variables associated with healing were measured in 60 matched pairs of surgically induced partial-thickness, full-thickness, and incisional wounds after either sham or anodal (positive-polarity) stimulation with pulsed microamperage current (100 μA) was administered for 1 hour per day for 5 days. Sterile, disposable electrodes (2×4 cm) were placed over each wound, which was kept wrapped and protected throughout the study. Results. At 7 days postinjury, all wounds were healing well with no signs of infection. There were no differences in tensile strength, collagen density, maturity, or deposition (hydroxyproline), wound size, or visual appearance between the sham treatment and treatment lesions. No changes in local subcutaneous oxygen or temperature were found in the swine during or after microamperage stimulation. Conclusion and Discussion. This study did not provide any evidence to support the use of microamperage stimulation to accelerate wound healing. No negative effects, however, were found. Further research is needed to determine whether there is a critical interaction between the size of the electrode relative to the wound, the density of the current, the duration of the treatment, the polarity of the treatment electrode, and the acuity or chronicity of wounding and the effectiveness of microcurrent stimulation for wound healing.

Oxygen tension and collagen deposition were measured in standardized, subcutaneous wounds in 33 postoperative surgical patients. Pertinent clinical and wound parameters were analyzed by Pearson's correlation test and sequential linear regression analysis. Collagen deposition was directly and significantly proportional to wound oxygen tension and measures of perfusion. There were no significant correlations with hematocrit, estimated blood loss, length of operation, smoking, age, weight, sex, or urine output. This study in humans confirms animal experiments showing that collagen deposition and tensile strength in wounds are limited by perfusion and tissue oxygen tension. It appears unnecessary to maintain hemoglobin at normal levels to support repair, provided that peripheral perfusion can be maintained at a high level in compensation for anemia. These circumstances reflect the fact that although oxygen is essential to many aspects of healing, and must be delivered at adequate partial pressures, reparative tissue consumes relatively little of it.

The significance of the high lactate levels that characterize healing wounds is not fully understood. Lactate has been shown to enhance collagen synthesis by fibroblasts and vascular endothelial growth factor (VEGF) production by macrophages and endothelial cells. VEGF has been shown to induce endothelial cell migration. However, it has not been shown whether accumulated lactate correlates with the biological activity of VEGF. Therefore, we investigated the effect of lactate on migration of endothelial cells. Human umbilical vein endothelial cells and human microvascular endothelial cells were cultured to subconfluent monolayers in standard six-well tissue culture plates. Following a 24-hour serum starvation, cells were treated with the indicated concentrations of l-lactate. Cell migration was assessed using a modified Boyden chamber. VEGF protein in the cell culture supernatant was measured by enzyme-linked immunoassay. Lactate-enhanced VEGF protein synthesis in a time- and dose-dependent manner. Lactate added into the bottom well did not stimulate cellular migration from the upper well. However, lactate when added together with endothelial cells to the bottom well of the Boyden chamber increased cellular migration in a dose-dependent manner. This effect was blocked by anti-VEGF and by cycloheximide. Lactate enhances VEGF production in endothelial cells, although lactate, itself, is not a chemoattractant. We conclude that the lactate-mediated increase in cellular migration is regulated by VEGF.

Insulin-like growth factor-I (IGF-I) has been shown to promote angiogenesis by enhancing vascular endothelial growth factor (VEGF) expression. However, how IGF-I-induces VEGF expression is not yet fully understood. With this investigation, we propose a new possible mechanism involving downregulation of poly(ADP-ribosyl)ation (pADPR). We first demonstrated that IGF-I increased VEGF protein expression in endothelial cells. Inhibitors of mitogen activated kinase (PD 98059), phosphatidyl-3-inositol-kinase (LY 294002), and protein kinase C (staurosporine) diminished the IGF-I effect suggesting the involvement of signal transduction. Since there is an established link between pADPR and transcriptional activity, we focused on a possible role of poly(ADP-ribose)polymerase (PARP). The inhibition of PARP by 3-aminobenzamide or nicotinamide enhanced VEGF expression. Additionally, IGF-I markedly decreased PARP activity. Furthermore, the IGF-I-mediated inhibition of PARP could be demonstrated as a result of protein phosphorylation since phosphorylation of PARP decreased its activity in vitro and IGF-I treatment of endothelial cells induced PARP phosphorylation. The IGF-I-mediated phosphorylation and inhibition of PARP represent a novel mechanism of VEGF protein expression.

Introduction: Healing wounds are characterized by high lactate levels. Lactate has been shown to enhance collagen synthesis by fibroblasts and VEGF production by macrophages. However, little is known whether the increased VEGF level correlates with enhanced biological activity. Therefore, we investigated the migration of endothelial cells treated with lactate. Material and methods: Human umbilical vein endothelial cells (HUVEC) were cultured to subconfluent monolayers in standard 6 well tissue culture plates. Following a 16 hour serum starvation period, cells were treated with indicated concentrations of lactate for 20 hours in a standard incubator at 95% air and 10% CO2. Cellular migration was assessed using a modified Boyden-chamber. VEGF protein in the cell culture supernatant was measured by ELISA. All experiments were performed in triplicates and repeated at least twice. Results are given as mean ± SD; a p-value less than 0.05 calculated by Student’s t-test was considered significant. Results: Lactate enhanced VEGF protein synthesis in a time- and dose-dependent manner. Lactate added to the bottom well did not stimulate cellular migration from the upper well. However, lactate together with endothelial cells in the bottom well of the Boyden-Chamber increased cellular migration in a dose-dependent manner. This effect could be blocked both by anti-VEGF or cyclohexamide as well. Conclusion: Lactate enhances VEGF production in human endothelial cells. However, lactate is not a chemoattractant. Lactate mediated increase in cellular migration is regulated by VEGF.

Publisher Summary The most important and commonly used methods for oxygen measurement, range from microelectrode measurements of wound oxygen gradients to clinically useful measurements in humans. Studies of the role of oxygen in wound healing turn on the ability to measure oxygen: in cell and tissue culture, in animal models, and in human wounds. The development of methods for the accurate measurement of wound oxygen tension has led to studies demonstrating that activation of the sympathetic nervous system by such common perioperative stressors as hypothermia, pain, and hypovolemia decreases wound oxygen tension and impairs wound healing. Tissue oxygen measurements are relatively simple to make, particularly with current highly stable probes. Concepts related to subcutaneous wound tissue oximetry and measurement of transcutaneous oxygen in patients with ulcers of the lower extremity are also discussed.

Hypothesis Anti-inflammatory corticosteroids significantly impair wound healing. Retinoids partially, but significantly, reverse this effect. Little is known about the mechanism of steroid retardation or retinoid reversal. We hypothesized that corticosteroids lower transforming growth factor-β (TGF-β) and insulin-like growth factor-I (IGF-I) levels and tissue deposition in wounds and that retinoids stimulate corticosteroid-impaired TGF-β and IGF-I release and collagen production. Design Randomized controlled trial. Setting Wound healing research laboratory. Participants Animal study. Interventions Four wire mesh wound cylinders were implanted subcutaneously into the backs of 72 male Sprague-Dawley rats. Wound healing was impaired by a single subcutaneous injection of 6 mg of methylprednisolone acetate (Depo-Medrol). Two preparations of retinoids were used in separate experiments: all- trans -retinoic acid and 9- cis -retinoic acid that were fed orally. Main Outcome Measures Hydroxyproline content was measured in the healing tissue and TGF-β and IGF-I levels were analyzed in the wound fluid. Results Methylprednisolone treatment significantly decreased TGF-β and IGF-I levels in the wound fluid and hydroxyproline content in the tissue ( P trans - and 9- cis -retinoic acid partially reversed the TGF-β and IGF-I decrease and significantly increased hydroxyproline content toward normal levels ( P trans -retinoic acid enhanced collagen deposition, TGF-β and IGF-I levels over normal chow fed control animals ( P Conclusions Steroids and retinoids have antagonistic effects on growth factors and collagen deposition in wound healing. These effects can be relevant for treatment options in a clinical setting.

Premature infants and neonates are vulnerable to bacterial sepsis. This susceptibility may be due to the relative immaturity of their immune systems. To determine if neonates and, in particular, premature infants have decreased polymorphonuclear leukocyte (PMN) phagocytosis, we tested PMN phagocytosis of Staphylococcus aureus as a function of gestational age in the fetal lamb model. Because phagocytosis is made more efficient by the presence of opsonins in plasma, fetal and postnatal PMN phagocytosis were also measured after exposure to fetal and adult plasma. PMNs were isolated from fetal lambs at 104, 114, 124, and 141 days' gestation (term gestation for the fetal lamb is 145 days), as well as from 10-day-old neonatal sheep and adult sheep. Labeled S aureus were opsonized by incubation in either fetal or adult plasma, or left unopsonized for baseline values. Phagocytosis was measured as a percent of adult PMN phagocytosis after adult plasma opsonization. It was found that fetal PMN function is limited by two factors during the early third trimester: a primary defect in the ability of the PMN to phagocytose S aureaus despite adequate opsonization, and the diminished ability of autologous fetal plasma to opsonize bacteria. The defect in PMN phagocytosis disappears late in the third trimester, but the inability of the fetal plasma to opsonize effectively continues until after birth.

Background: "Respiratory burst" activity, ie, O 2 −production, is dependent on Po 2 , temperature, pH, and glucose concentrations within the physiologic range. Objectives: To determine whether environmental conditions characteristic of wounds may limit human neutrophil respiratory burst metabolism and to clarify the degree to which bactericidal oxidant production depends on local Po 2 . Methods: Human blood and wound neutrophils were stimulated with phorbol myristate acetate. Oxygen consumption and superoxide production were measured over a range of 30 to 300 mm Hg Po 2 , 0 to 40 mmol/L glucose, pH 6.0 to 8.0, and 30°C to 37°C. The apparent Michaelis Menten constant for oxidant production with respect to Po 2 was calculated. Results: Oxygen consumption and O 2 −production were dependent on Po 2 throughout the range tested. Half-maximal oxidant production occurred in the range of 45 to 80 mm Hg Po 2 and maximal at Po 2 higher than 300 mm Hg. These data agree with the highest previous estimates. Oxidant generation was also dependent on pH, temperature, and glucose concentration, but to a lesser extent. Conclusions: Leukocyte bacterial killing capacity as measured by oxygen consumption and superoxide production are substantially impaired at the low oxygen tensions often found in wounds. Changes in pH, temperature, and glucose concentration have lesser but nonetheless significant consequences. The data provide a plausible mechanism for the vulnerability of some wounds to infection and for the previous finding that increasing oxygen tension at wound sites enhances bactericidal function. Thus, the data serve as a basis for future studies on prevention of wound infection. Arch Surg. 1997; 132:991-996

Multiple System Organ Failure develops in 15% of surgical intensive care unit patients despite being adequately resuscitated by conventional criteria. Of these, greater than 50% will eventually die1. Some current theories implicate the persistence of inadequate gut oxygenation as the principle inciting event2,3. It is proposed that gut hypoxia compromises both mucosal barrier integrity and transmural oxidative bacterial killing. The net effect is an increased rate of bacterial translocation, which if unchecked can rapidly progress to sepsis and death. The early detection of gut hypoxia offers an opportunity to intervene and possibly prevent this scenario.

The object of this animal study was to examine and further develop the expanded polytetrafluoroethylene wound healing model. The goal was to increase its potential for assessing wound healing by increasing yield, reducing variability, establishing the elements of a standard technique, and further testing its ability to detect variations of healing which have clinical significance. Expanded polytetrafluoroethylene implants of various dimensions and fabrications and several implantation and sterilization techniques were compared in rats. Hydroxyproline, DNA, and protein deposition into the expanded polytetrafluoroethylene implants as parameters for wound healing were assessed. Additionally, a 4 cm skin incision for tensile strength assessment was created. Wound healing was assessed under normal and corticosteroid-impaired healing conditions. The highest yield of collagen was found in the stiffer fabrication of expanded polytetrafluoroethylene with the larger pore size and after the more traumatic implantation technique of incisional placement. Variability was unaffected by fabrication, implantation technique, indexing by various geometric dimensions of the implant, sterilization, or sampling techniques. Variability was the same in the individual animals as in groups of animals. The expanded polytetrafluoroethylene method also detects the influence of antiinflammatory corticosteroids and reflects the tensile strength of incisional wounds made in other sites in the same animal.

Gossypol is present in antiinflammatory poultices made from the medicinal tree Thespesia populnea. Isolated human neutrophils exposed to 3-20 microM gossypol for 15-90 min were assayed in vitro for superoxide production and surface expression of Mac-1 (CD11b/CD18). Gossypol increased superoxide production in a time- and concentration-dependent fashion consistent with a moderate, delayed respiratory burst. Surface Mac-1 expression was increased within 15 min by 3-5 microM gossypol, resulting in a 14-fold increase over controls and a threefold greater increase over that produced by PMA. Staurosporine failed to block gossypol induction of superoxide and Mac-1, while EDTA inhibited induction of Mac-1 only, implicating a calcium-dependent mechanism. Gossypol increased intracellular calcium to peak levels, but in a delayed fashion as compared to FMLP. These findings demonstrate that gossypol is a highly potent stimulant of Mac-1 expression and suggest at least two protein kinase C-independent pathways of neutrophil activation. The resultant exhaustion of neutrophils may account for the antiinflammatory properties of plants containing gossypol.

Because various immune functions are impaired at temperatures only 1 degrees to 3 degrees C less than normal, we tested the hypothesis that mild hypothermia during anesthesia impairs resistance to dermal infections. Guinea pigs were anesthetized for 6 hours with 1% inspired halothane. Their core temperatures were maintained at either 39 degrees C (normal for guinea pigs, n = 12) or 36 degrees C (n = 12). Two hours after induction of anesthesia, three doses each of Staphylococcus aureus (10(8), 10(7), and 10(6) organisms) were injected intradermally at nine sites on each animal's back. Core temperatures were not controlled after recovery from the anesthetic, and animals in each group were maintained in the same environment. Four days after anesthesia, each injection site was excised to obtain a count of viable bacteria. Subcutaneous oxygen partial pressure values, averaged over time, were 53 +/- 3 mm Hg (mean +/- SEM) in the hypothermic group and 62 +/- 4 mm Hg in the normothermic group (p = 0.06). Capillary perfusion, as assessed by laser Doppler flowmetry, was comparable in the two groups. One day after injection of 10(8) bacteria, the area of induration was 89 +/- 11 mm(2) in the hypothermic group but only 61 +/- 6 mm(2) in the normothermic group (p0.05). On postanesthetic day 4, the area of induration was 72 +/- 6 and 59 +/- 6 mm(2) in the hypothermic and normothermic groups, respectively (p0.05). After inoculation with 10(8) bacteria, the fraction recovered was 1.0 +/- 0.2 in the hypothermic groups and 0.6 +/- 0.2 in the normothermic group (p0.05). After inoculation with 10(7) and 10(6) bacteria, the fraction recovered was less than 0.2, and no difference was found between the hypothermic and normothermic animals. Thus mild hypothermia during halothane-induced anesthesia slightly impairs resistance to dermal infection.

Until recently, there has been little interest in measuring tissue oxygen tension because there has been no theoretical basis on which to interpret the results and therefore no practical value. Now, however, strong theoretical and practical bases are emerging, and it is time for re-examination.

INTRODUCTION: High concentrations of lactate approx. 10 mM are characteristic of wound healing. We have previously shown that lactate instigates release of VEGF from macrophages and endothelial cells. Our present study uses an in vivo murine Matrigel model to study how lactate enhances vascular development and repair. METHODS: Matrigel is a reconstituted basement membrane complex, a liquid at 4 degree C. When injected subcutaneously, it reconstitutes as a gel. 30 mgs of finely divided polylactate polymer (DL-lactide-co-glycolide) was mixed with 1cc of Matrigel, and 2 injections were made into the dorsum of each of 60 (average 7/group), 6-month-old Swiss Webster mice. The gel implants were harvested at 3, 6, 9, 11 and 16 days post injection, and were fixed in buffered formalin or frozen. Cell migration and vascular development were assessed using H&E, anti-CD31 and anti-MAC3 antibody stainings for leukocytes, fibroblasts, endothelial cells, and macrophages. Cells and vessels were quantified microscopically. RESULTS: Control implants (without polylactate) developed little or no inflammation or angiogenesis. Polylactated implants developed highly significant number of endothelial cells and recognizable vessels at 6 and 9 days. At 11 and 16 days, sizeable vessels with surrounding connective tissue were found. Though a moderate inflammation occurred in the lactate groups, no foreign body reaction was seen. Larger implants of high molecular weight instigated little inflammation and few vessels. CONCLUSION: Slowly released lactate leads to the formation of new vessels. Our results provide support for further clinical applications of lactate polymer in wounds. NIH/NIGMS Trauma Training Grant

Introduction: Vascular Endothelial Growth Factor (VEGF) mediated angiogenesis plays a key role in wound healing. Insulin-like Growth Factor I (IGF-I) has been reported to be angiogenic. However, the mechanism is not known. Recently, a link between transcriptional activity and inhibition of poly(ADP-Ribose)polymerase (PARP) has been reported. We investigated whether IGF-I increases VEGF expression and whether this effect is regulated by the inhibition of PARP. Material and methods: Subconfluent monolayers of human umbilical vein cells were cultured and serum starved. Cultures were treated with Long-R3-IGF-I for 20 h. VEGF in the supernatant was measured by ELISA and lactate by a lactate analyser. PARP activity was assessed by measuring the incorporation of 14C-radiolabeled NAD+. All experiments were performed in triplicate. Results are given as percent change compared to control ± SD; p

Hypothesis Women deposit more collagen after major abdominal surgery than men. Design A post hoc analysis of data obtained from 2 prospective, randomized, double-blind clinical trials. Setting University hospital general surgical service. Patients One hundred sixteen patients undergoing colon resection. Main Outcome Measures Protein and hydroxyproline (collagen) deposition during the first 7 postoperative days in expanded polytetrafluoroethylene implants positioned subcutaneously. Results On univariate analysis, men and women deposited comparable amounts of collagen (257 ± 120 vs 281 ± 117 ng/mm, respectively). When potential confounding factors were entered into a generalized mixed-effects model, only the interaction between age and sex was a significant factor ( P = .047). Collagen deposition decreased with age in men, being 317 ± 133 ng/mm in men younger than 45 years, but only 238 ± 113 ng/mm in those older than 45 years ( P = .03). In contrast, collagen deposition was virtually identical in women younger than 45 years (280 ± 133 ng/mm) and in those older than 45 years (281 ± 110 ng/mm). Only 3 of these women were receiving hormone replacement therapy. Conclusions Collagen deposition after surgery decreased significantly with age in men, while remaining unchanged in women. Younger men and women deposited similar amounts of collagen. Therefore, older men made less collagen after surgery than older women, perhaps explaining the consistent observation that wound dehiscence is twice as common in men than in women. Our results differ from previous studies conducted in healthy, nonsurgical volunteers, which showed that (1) young women made significantly more collagen than young men and (2) collagen deposition was reduced in postmenopausal women, but deposition returned to premenopausal values with hormone replacement therapy. Differences between our results and those reported previously likely stem from the populations studied. In particular, multiple perioperative factors decrease collagen deposition, which apparently obscures the differences observed previously in healthy, unstressed volunteers.

Chemoattractant activity for irradiated and nonirradiated rabbit skin fibroblast and bovine aortic arch endothelial cells was assayed in rabbit wound fluid and sera using a modification of the agarose well method originally described for polymorphonuclear leukocytes. Both serum and wound fluid contained chemoattractants for fibroblasts and endothelial cells. Fibroblast migration was decreased by 70 to 80% when the serum or wound fluid was heated to 56 degrees C for 30 min while endothelial cell migration was reduced by 50 to 60%. Platelet-poor plasma-derived serum had no directive effect on the migration of either cell type.

We compared simultaneous healing processes in fetal, newborn, and maternal rabbits using a miniaturized wound cylinder of expanded Gore-Tex tubing. The tubing was placed subcutaneously in fetal and maternal rabbits on day 23 of pregnancy (term = 31 to 32 days), and in 7-day-old newborn rabbits. At specific intervals, the tubing was removed and analyzed for hydroxyproline accumulation, histology, and cellular enzyme-histochemistry. Granulation tissue ingrowth and accumulation of hydroxyproline were each inversely related to age (fetus greater than newborn greater than maternal). The fetus showed an impressive infiltration of macrophages by day 4, fibroblasts by day 7, and a conspicuous lack of neutrophils in all specimens. Newborns and mothers had few cells until day 7, when a mixture of macrophages, neutrophils, and some fibroblasts appeared. We conclude that the fetus heals wounds rapidly by both mesenchymal cell proliferation and collagen deposition, and that these processes are more rapid in fetuses than in newborn or adult animals despite relative fetal hypoxemia.