In vitro labeling of solid tissues with tritiated thymidine for autoradiographic detection of S-phase nuclei.

In vitro measurement of the thymidine labeling index (TLI) of solid tissues requires hyperbaric oxygenation and is potentiated by blockade of thymidylate synthetase by 5-fluoro-2'-deoxyuridine (FUdR) to favor uptake of tritiated thymidine (3H-TdR). Hyperbaric oxygenation can be achieved in a simple system through injection of oxygen into rubber-stoppered test tubes. Incubations are carried out in Hanks' balanced salt solution in a shaker bath at 37 C for 2 hours; an FUdR concentration of approximately 1 micron is optimal. Autoradiographic exposure for 1 week or less is sufficient for TLI measurements on human tissues. With 3 to 4 atmospheres oxygen tension, incorporation of 3H-TdR is sufficient for TLI measurement throughout slices of tissue cut 1 mm thick or less. Mincing of tissue is not necessary, and the anatomic continuity seen in ordinary histological preparations is preserved. A gradient of labeling intensity is present from the surface to the interior of the tissue, but sufficient intensity of labeling for detection of DNA synthesis can be achieved in the interior of the section. The gradient can be reduced only slightly by prior incubation in 3H-TdR with hyperbaric oxygen at 0 C. The method permits TLI measurements on the same specimens, including needle biopsies, that are used for pathologic diagnosis.     

RELATIONSHIP OF BONE MARROW CELLULARITY AND PROLIFERATIVE ACTIVITY: A LOCAL REGULATORY MECHANISM

An analysis has been made of tritiated thymidine (³H-TdR) uptake and cellularity in normal and regenerating marrow. Comparison of right and left femoral marrow in ninety normal rabbits has revealed a significant negative correlation (−0.75) between thymidine uptake per presumptive proliferative cell and the concentration of such cells. Locally irradiated marrow follows the normal marrow regression down to about 40% relative cellularity and then shows an upward displacement. The regenerating mechanically depopulated marrow manifests a similar displacement over the entire cellularity range studied which suggests that recovery was not yet complete as in the case of some of the irradiated marrow samples. The negative regression of ³H-TdR uptake on cellularity is not a consequence of altered thymidine availability, but rather of-a changing DNA synthesis rate and/or a changing fraction of proliferative cells. Since ³H-TdR uptake per presumptive proliferative cell reflects cell production, it follows that cell production is geared to the cellularity of a discrete marrow area. This provides a local self-regulating mechanism.     

TEMPORARY LOCALIZED DEPRESSION OF TRITIATED THYMIDINE INCORPORATION IN MOUSE TISSUES AFTER PULSE LABELLING

Approximately two to six in every 100 mice injected with ³H-TdR appear not to incorporate the labelled precursor into the DNA. The tritium activity appears to be distributed throughout these poor utilizers of thymidine. The lack of incorporation of the precursor is not always general to the whole animal but may be restricted to a given tissue. The effect does not appear to be permanent but varies with time.     

Regional Thymidine Transport and Incorporation in Experimental Brain and Subcutaneous Tumors

Abstract: The regional distribution and local incorporation of [¹⁴C]thymidine into a nonextractable tissue fraction, probably DNA, was measured in normal and neo-plastic tissues. We studied brain tumors induced by avian sarcoma virus and ethylnitrosourea, and transplanted RG-2 intracerebral and subcutaneous gliomas. An incorporation quotient, Q , was calculated for different tumor regions and brain from the methanol nonextractable radioactivity in the tissue and the plasma concentration-time integral of thymidine. The incorporation quotient represents the rate of clearance of thymidine from blood and its incorporation into macromolecules (probably I NA). The values of Q were compared with a labeling index measured in the same tissue regions with conventional autoradiography. The following observations were made: (1) the mean plasma half-life of thymidine was 6.5 min; (2) the regional incorporation quotient in tumors varied from values comparable to normal brain to more than 100 times higher; (3) RG-2 tumors had significantly higher Q s than the other tumor models; (4) Q in subcutaneous tumors varied most widely (>500-fold range); (5) the labeling index reflected the values of Q in some tumor regions but not in others; differences between the two were most frequently related to tumor cell density and the intensity of individual tumor cell labeling. A comparison of these data with previous studies of capillary permeability and blood flow in these tumor models indicates that the incorporation of [¹⁴C]thymidine into a nonextractable tissue fraction can be limited by transcapillary transport in brain tumors and by blood flow in systemic tumors, and that thymidine disposition in these tumors is not always indicative of the rate of DNA synthesis.     

The Influence of Hyperbaric Oxygenation (HBO) on Proliferation and Differentiation of Human Keratinocyte Cultures In Vitro

A drop in tissue oxygen partial pressure below 30mm Hg as a result of reduced perfusion in an extensive area of acute skin damage, or where a large number of chronic skin defects occur, inhibits collagen synthesis and neoangiogenesis in the various phases of wound healing. Subsequent granulation and epithelialisation are correspondingly impaired.Hyperbaric oxygenation is now recognised as a valuable supplementary method of treatment for problematic wounds. Stimulation of fibroblast and endothelial cell proliferation through Hyperbaric oxygenation has been demonstrated in numerous studies.The aim of this study was to investigate the effect of hyperbaric oxygen treatment on the proliferation and differentiation of human keratinocyte cultures.The influence of hyperbaric oxygenation on the proliferation of human keratinocyte cultures was demonstrated using flow-through cytometry and a fluorescence activated cell sorter, which detects fluorescence intensity following incorporation of 5-bromo-2-deoxyuridine in cell DNA.The degree of cell differentiation was deduced from the expression of various components of the cytoskeleton, such as cytokeratin 10 and involukrin, the production of which was quantified through the determination of monoclonal antibodies against cytokeratin 10 and involukrin from measurements of fluorescence activity in a flow-through cytometer.Hyperbaric oxygenation of cell cultures in vitro did not produce a significantly higher rate of cell proliferation, so that no increase in vitality was observed.An interesting observation following exposure to hyperbaric oxygen was the marked increase in expression of both cytokeratin 10 and involukrin, as an indication of accelerated cell differentiation.     

Expanding the limits of composite grafting: a case report of successful nose replantation assisted by hyperbaric oxygen therapy

A successful nose replantation assisted by hyperbaric oxygen therapy is presented, with a brief discussion of the possible mechanisms and a brief literature review of the use of hyperbaric oxygen in tissue preservation and replantation. Although it is not certain that the hyperbaric oxygenation ensured the survival of the replanted nose in this 2-year-old girl, there was documented change in graft appearance during the initial hyperbaric oxygen treatment. A 1-month, 1-year, and 2-year follow-up is included.     

Retrograde axonal transport specificity in the locus coeruleus neurons after [H]noradrenaline injection into the rat olfactory bulb

Retrograde axonal transport process was investigated in the afferent systems to the rat olfactory bulb, after [³H]noradrenaline ([³H]NA) injection into the olfactory bulb, in order to provide evidence regarding its specificity and the biochemical basis supporting this specificity.

Radioautographs showed that [³H]NA unilaterally injected into the olfactory bulb at a concentration of 10⁻³ M, resulted in labeling of the structures afferent to the olfactory bulb, mainly on the injected side: locus coeruleus (LC), dorsal and central raphes, nucleus of the lateral olfactory tract and piriform cortex. Heavy labeling was observed on the noradrenergic LC cell bodies, whereas the radioautographic reaction was less intense on the other structures. After 10⁻⁴ M injection, the labeling intensity of the LC cell bodies was lower while very rare weakly labeled cell bodies persisted in the dorsal raphe, nucleus of the lateral olfactory tract and piriform cortex. The LC cell bodies were exclusively labeled when the concentration of [³H]NA injection was 10⁻⁵ M. All the other structures were devoid of labeling. It was still possible to detect labeled cell bodies in the LC for a 10⁻⁶ M concentration.

Following bilateral injections of [³H]NA (10⁻³ M) the total radioactivity retrogradely transported to the LC represented about 4 times the total radioactivity measured in the periaqueductal gray substance (as control tissue of the tracer diffusion). Fractional study by ethanol of LC tissue homogenate and liquid scintillation counting of each fraction showed that 60% of the total radioactivity (about 2.5 times the control value) was in the supernatant and 40% (about 20 times the control value) was associated with the precipitate. In the other regions such as the dorsal and central raphes and periaqueductal gray substance, radioactivity was mainly found in the supernatant, except for the dorsal raphe whose precipitate contained a low amount of radioactivity (about 4 times the control value).

Colchicine (an axonal transport inhibitor) bilaterally injected into the medial forebrain bundle and systemic administration of desipramine (a noradrenaline uptake inhibitor) decreased the radioactivity associated with the LC precipitate by 90 and 85% and the LC supernatant radioactivity by 55 and 35%, respectively. These pretreatments did not significantly affect the radioactivity amounts measured in the different fractions of dorsal and central raphes and periaqueductal gray substance. Radioautographic study after colchicine treatment showed a large decrease in the labeling intensity of the LC cell bodies as compared to the non-treated side.

Therefore, we suggest that low concentrations (10⁻⁵ M) of [³H]NA injected in the olfactory bulb determine specific conditions of noradrenergic pathway labeling. This specific labeling after migration could be supported by the radioactive ethanol precipitate which would appear to contain [³H]NA- and/or ³H-derivatives-binding protein. Such a ³H-macromolecular complex, which could represent the specific carrier, may well undergo retrograde transport from the nerve terminals towards the cell bodies.     

Inhibitory effect of high oxygen pressure on potassiuminduced activation of pyruvate dehydrogenase and glucose metabolism in rat brain slices

The effects of high oxygen pressure on pyruvate dehydrogenase (pyruvate: lipoate oxidoreductase (decarboxylating and acceptor-acylating), EC 1.2.4.1) activity, tissue concentration of ATP, and CO₂ production from glucose were studied in rat brain cortical slices. The increase in pyruvate dehydrogenase activity and the lowering of cellular ATP, occurring during potassium-induced depolarization at 1 atm of oxygen, were reversed by increasing the oxygen pressure to 5 atm. When brain slices were incubated at 1 atm oxygen with [U-¹⁴C]glucose, a high potassium medium approximately doubled the production of ¹⁴CO₂. Oxygen at 5 atm abolished this potassium-dependent increase in ¹⁴CO₂ production with no significant effect on glucose oxidation in normal Krebs-Ringer phosphate medium. Adding 4 atm helium to 1 atm oxygen did not interfere with the ability of potassium ions to activate pyruvate dehydrogenase, lower ATP, or increase glucose oxidation. The results show that toxic effects of hyperbaric oxygen, not manifest in “resting” tissue, may be revealed during stress such as potassium depolarization. The site of the toxic effects of oxygen is probably the cell membrane where excess oxygen appears to interfere with the action of the sodium pump, calcium transport or other processes stimulated by increased concentrations of extracellular potassium.     

Effect of hyperbaric oxygenation on the Na+, K+-ATPase and membrane fluidity of cerebrocortical membranes after experimental subarachnoid hemorrhage

It is reported that CNS hemorrage causes membrane dysfunction and may exacerbate this damage as a result of secondary ischemia or hypoxia. Since hyperbaric oxygenation improves oxygen metabolism, it may reduce this membrane damage. The present study was conducted to reveal whether hyperbaric oxygenation influences membrane alteration after hemorrhage. Thirty minutes after subarachnoid hemorrhage induction, rats were treated with hyperbaric oxygenation 2 ATA for 1 hour. Rats were decapitated 2 hours after subarachnoid hemorrhage induction. Na⁺, K⁺-ATPase activity measurement, and spin-label studies were performed on crude synpatosomal membranes. Subarachnoid hemorrhage decreased Na⁺, K⁺-ATPase activity. Spin label studies showed that hydrophobic portions of near the membrane surface became more rigid and the mobility of the membrane protein labeled sulfhydryl groups decreased after subarachnoid hemorrhage. Hyperbaric oxygenation significantly ameliorated most of the subarachnoid hemorrhage induced alterations. We conclude that hyperbaric oxygenation may be a beneficial treatment for acute subarachnoid hemorrhage.     

Effect of hypothermia on radiosensitization

Hypothermia reduces metabolism and oxygen utilization by tissues. If the blood supply to a solid tumour can be maintained at a sufficient level, the hypoxic fraction of tumour cells may be reduced and radiosensitivity increased. This may be achieved if hyperbaric oxygen is used in combination with the hypothermia. The blood supply and oxygen tension have been measured in C3H mouse mammary tumours under hypothermia and hyperbaric oxygen, and the enhancement of radiosensitivity by hyperbaric oxygen has been estimated in mice irradiated at different temperatures with and without anaesthesia. Measurement of xenon-133 clearance showed that the blood supply of a tumour tended to increase when anaesthetized mice became hypothermic. Oxygen cathode data showed that the oxygen tension tended to be relatively higher in tumours and lower in subcutaneous tissue when mice exposed to hyperbaric oxygen became hypothermic under anaesthesia. Hyperbaric oxygen enhanced the radiation response of the tumour in terms of an increase in regrowth delay by a factor of 1.7 when the mice had been anaesthetized, whether or not they became hypothermic. A lower factor of 1.4 was obtained without anaesthesia although induced hypothermia increased the response to a small extent. We conclude that anaesthesia and hypothermia affect oxygen metabolism in tumours by different mechanisms.     

Thymidine incorporation in Lake Cisó: Problems in estimating bacterial secondary production across oxic-anoxic interfaces

Abstract Heterotrophic bacterial activity was measured by means of the ³H-thymidine (³H-TdR) incorporation technique in Lake Cisó, a small holomictic lake with anoxic hypolimnion. We tested several methodological questions across the vertical profile: TdR concentration at which maximal incorporation is reached, linearity of incorporation and isotope dilution, during holomixis and stratification periods. The TdR concentration at which maximal incorporation is reached changed seasonally and vertically. During holomixis, maximal incorporation was not always reached at concentrations up to 40 nM. Uptake was always linear in short incubation times and decreased from epi- to hypolimnion. The isotope dilution technique indicated a degree of participation in DNA synthesis higher than 50%, although a linear relationship between the inverse of ³H-TdR incorporation and increasing ‘cold’ thymidine concentration was not always observed. Autoradiographic experiments showed a low percentage of bacteria taking up ³H-TdR in both aerobic and anaerobic samples. The percentage of total labeled bacteria seemed to be generally higher in the metalimnion (11% maximal value) than in the hypolimnion. Labeled Amoebobacter and Chromatium cells were detected in field samples. Amoebobacter cells photoassimilated TdR in culture. Therefore, our results show that ³H-TdR incorporation is not an appropriate technique to estimate bacterial secondary production in anaerobic systems and in oxic-anoxic interfaces.     

The role of oxygen monitoring during photodynamic therapy and its potential for treatment dosimetry.

Understanding of the biology of photodynamic therapy (PDT) has expanded tremendously over the past few years. However, in the clinical situation, it is still a challenge to match the extent of PDT effects to the extent of the disease process being treated. PDT requires drug, light and oxygen, any of which can be the limiting factor in determining efficacy at each point in a target organ. This article reviews techniques available for monitoring tissue oxygenation during PDT. Point measurements can be made using oxygen electrodes or luminescence-based optodes for direct measurements of tissue pO2, or using optical spectroscopy for measuring the oxygen saturation of haemoglobin. Imaging is considerably more complex, but may become feasible with techniques like BOLD MRI. Pre-clinical studies have shown dramatic changes in oxygenation during PDT, which vary with the photosensitizer used and the light delivery regimen. Better oxygenation throughout treatment is achieved if the light fluence rate is kept low as this reduces the rate of oxygen consumption. The relationship between tissue oxygenation and PDT effect is complex and remarkably few studies have directly correlated oxygenation changes during PDT with the final biological effect, although those that have confirm the value of maintaining good oxygenation. Real time monitoring to ensure adequate oxygenation at strategic points in target tissues during PDT is likely to be important, particularly in the image guided treatment of tumours of solid organs.     

Occurrence of singlet oxygen oxygenation of oleic acid and linoleic acid in the skin of live mice

To assess the contribution of singlet molecular oxygen [O₂ (¹Δ_g)] to lipid peroxidation in vivo, this study combined gas chromatography-mass spectrometry with thin layer chromatography to analyse peroxidized lipids in the skin of hairless mice. Hydroxyoctadecenoate isomers and unconjugated hydroxyoctadecadienoate isomers derived from peroxidized oleic acid and linoleic acid, respectively, which are specific to O₂ (¹Δ_g)-dependent oxygenation, were detected in the skin of live mice under ordinary feeding conditions. Short-term ultraviolet A (UVA)-irradiation of the skin in vivo elevated levels of the unconjugated hydroxyoctadecadienoate isomers significantly, whereas the irradiation of skin homogenate in vitro increased levels of all isomers derived from both O₂ (¹Δ_g) and free radical-dependent oxygenation to a much greater extent. This is the first report to demonstrate the occurrence of O₂ (¹Δ_g)-specific oxygenation of unsaturated fatty acids in living animals.     

The effect of hyperbaric oxygenation on the viability of human fat injected into nude mice

Autologous free-fat injection for the correction of soft-tissue defects has become a common procedure in plastic surgery. The main shortcoming of this method for achieving permanent soft-tissue augmentation is the partial absorption of the injected fat, an occurrence that leads to the need for both overcorrection and repeated fat reinjection. Improving the oxygenation of the injected fat has been suggested as a means of helping to overcome the initial critical phase that occurs postinjection (when the fat cells are nourished by osmosis), increasing phagocyte activity, accelerating fibroblast activity and collagen formation, and enhancing angiogenesis. In addition, the hyperbaric oxygen-mediated decrement in endothelial leukocyte adhesion will decrease cytokine release, thereby reducing edema and inflammatory responses. The purpose of the present study was to examine the effect of hyperbaric oxygenation on improving the viability of injected fat. Adipose tissue obtained from human breasts by suction-assisted lipectomy was injected into the subcuticular nuchal region in nude mice. The mice were then exposed to daily hyperbaric oxygen treatments, breathing 100% oxygen at 2 atmospheres absolute (ATA) for 90 minutes. The duration of the administered hyperbaric oxygen therapy was 5, 10, or 15 days, according to the study group. Mice exposed to normobaric air alone served as the control group, and each group included 10 animals. The rats were killed 15 weeks after fat injection. The grafts were dissected out, weight and volume were measured, and histologic evaluation was performed. In all of the study groups, at least part of the injected fat survived, giving the desired clinical outcome. No significant differences could be found between the groups regarding fat weight and volume. Histopathologic examination of the dissected grafts demonstrated a significantly better integrity of the fat tissue in the group that received hyperbaric oxygen for 5 days (p = 0.047). This finding was manifested by the presence of well-organized, intact fat cells, along with a normal appearance of the fibrous septa and blood vessels. The worst results were found in animals treated by hyperbaric oxygenation for 15 consecutive days. An inverse correlation was found between an increased dose of the high-pressure oxygen and fat tissue integrity (r = -0.87, p = 0.076). The toxic effects of highly reactive oxygen species on fat cells might explain the failure of an excessively high dose of hyperbaric oxygen to provide any beneficial outcome. The clinical relevance of these results should be further investigated.     

Quantitative multi‐spectral oxygen saturation measurements independent of tissue optical properties

Imaging of tissue oxygenation is important in several applications associated with patient care. Optical sensing is commonly applied for assessing oxygen saturation but is often restricted to local measurements or else it requires spectral and spatial information at the expense of time. Many methods proposed so far require assumptions on the properties of measured tissue. In this study we investigated a computational method that uses only multispectral information and quantitatively computes tissue oxygen saturation independently of tissue optical properties. The method is based on linear transformations of measurements in three isosbestic points. We investigated the ideal isosbestic point combination out of six isosbestic points available for measurement in the visible and near‐infrared region that enable accurate oxygen saturation computation. We demonstrate this method on controlled tissue mimicking phantoms having different optical properties and validated the measurements using a gas analyzer. A mean error of 2.9 ± 2.8% O₂Sat was achieved. Finally, we performed pilot studies in tissues in‐vivo by measuring dynamic changes in fingers subjected to vascular occlusion, the vasculature of mouse ears and exposed mouse organs.

Selected steps of spectral transformations applied to oxygenation spectra. The original reflectance spectrum M(λ) is transformed in step 1 to overlap with reference spectra (grey) in three isosbestic points, resulting in M″(λ). In step 2, the gradient of M″(λ) is computed resulting in M″_grad(λ), which can be used for quantitative oxygenation computation.     

Differential spermatogonial stem-cell survival and mutation frequency

One group of adult C3H×101 hybrid male mice was given 3 injections of 12.5 μCi of [³H]thymidine at 9-h intervals and irradiated 24 h after the last injection with X-ray doses of 100, 300, 500, 600, 1000 R or the first fraction of a split 1000-R dose given as two 500-R exposures 24 h apart. Mice were killed 207 and 414 h after irradiation. A second group of mice was given a single injection of 12.5 μCi of [³H]thymidine 1 h before irradiation with single exposures of 300, 500, 600, 1000 R, or the first fraction of a 1000-R exposure given as two 500-R fractions 24 h apart. Mice were killed 120 and 207 h after irradiation. In both experiments, parallel groups of mice were given X-ray only as a control for the effect of [³H]thymidine. Two sets of slides were prepared for each mouse receiving [³H]thymidine: one set was not autoradiographed and was used for scoring cell survival; the second set was coated with emulsion and used for scoring percentage of labeled cells. The dose-response curves for survival at 120 and 207 h were curvilinear, with no evidence of discontinuity over the 100–1000-R range. After multiple injections of [³H]thymidine and irradiation 24 h later, percentage of labeled cells at 207 h was comparable for controls, 100, 300, and 600 R; significantly lower than controls for 1000 R; and significantly above controls after 500 + 500 R. Thus the surviving stem-cell population was qualitatively the same for that portion of the dose-response curve giving a linear increase in mutation rate but was different for both 1000-R and 500 + 500-R exposures, and the single and fractionated 1000-R exposures differed from each other. This parallelism between survival of labeled cells and mutation frequency in spermatogonial stem cells suggests that a stage in the cell cycle 24–42 h after DNA synthesis is resistant to cell killing but sensitive to mutation induction. The mutation rate after a single 1000-R exposure is low because labeled, mutation-sensitive cells have been selectively killed. Mutation frequency after the 500 + 500-R dose is increased because of synchronization induced by the first dose combined with selective killing of unlabeled cells by the second fraction. Irradiation 1 h after labeling with [³H]-thymidine demonstrated that the S phase of the spermatogonial stem-cell cycle is sensitive to radiation-induced cell killing.     

Nitrofurazone-induced DNA damage to tissues of mice

Cytotoxicity and DNA damage by nitrofurans has previously been correlated with metabolic reduction of these drugs in vitro. In the present study, nitrofurazone increased the rate of disappearance of stable [³H]thymidine labelled DNA from tissues of mice fed 0.1% nitrofurazone in the diet. Significant loss of labelled DNA occurred within 25 days after the start of the diet in all tissue observed, and loss was in relation to the rate of metabolic reduction of nitrofurazone. A similar correlation was found when another endpoint for DNA damage was used; nitrofurazone reduced by mouse tissue slices caused DNA single-strand breaks in cultured mouse L cells incubated in vitro with the tissues. Again, the ability of each tissue to produce toxic nitrofurazone metabolites determined the amount of DNA damage to the L cells.     

Hyperbaric oxygen induces apoptosis via a mitochondrial mechanism

During therapeutic hyperbaric oxygenation lymphocytes are exposed to high partial pressures of oxygen. This study aimed to analyze the mechanism of apoptosis induction by hyperbaric oxygen. For intervals of 0.5–4 h Jurkat-T-cells were exposed to ambient air or oxygen atmospheres at 1–3 absolute atmospheres. Apoptosis was analyzed by phosphatidylserine externalization, caspase-3 activation and DNA-fragmentation using flow cytometry. Apoptosis was already induced after 30 min of hyperbaric oxygenation (HBO, P < 0.05). The death receptor Fas was downregulated. Inhibition of caspase-9 but not caspase-8 blocked apoptosis induction by HBO. Hyperbaric oxygen caused a loss of mitochondrial membrane potential and caspase-9 induction. The mitochondrial pro-survival protein Bcl-2 was upregulated, and antagonizing Bcl-2 function potentiated apoptosis induction by HBO. In conclusion, a single exposure to hyperbaric oxygenation induces lymphocyte apoptosis by a mitochondrial and not a Fas-related mechanism. Regulation of Fas and Bcl-2 may be regarded as protective measures of the cell in response to hyperbaric oxygen.     

Dermal excisional wound healing in pigs following treatment with topically applied pure oxygen

Hypoxia, caused by disrupted vasculature and peripheral vasculopathies, is a key factor that limits dermal wound healing. Factors that can increase oxygen delivery to the regional tissue, such as supplemental oxygen, warmth, and sympathetic blockade, can accelerate healing. Clinical experience with adjunctive hyperbaric oxygen therapy (HBOT) in the treatment of chronic wounds have shown that wound hyperoxia may increase granulation tissue formation and accelerate wound contraction and secondary closure. However, HBOT is not applicable to all wound patients and may pose the risk of oxygen toxicity. Thus, the efficacy of topical oxygen treatment in an experimental setting using the pre-clinical model involving excisional dermal wound in pigs was assessed. Exposure of open dermal wounds to topical oxygen treatment increased tissue pO₂ of superficial wound tissue. Repeated treatment accelerated wound closure. Histological studies revealed that the wounds benefited from the treatment. The oxygen treated wounds showed signs of improved angiogenesis and tissue oxygenation. Topically applied pure oxygen has the potential of benefiting some wound types. Further studies testing the potential of topical oxygen in pre-clinical and clinical settings are warranted.     

Biochemical and autoradiographic investigations of the retrograde axonal transport of labeled material following [H]norepinephrine injection in the olfactory bulb

Biochemical and autoradiographic methods were used to investigate the retrograde transport of labeled material after injection of [³H]norepinephrine ([³H]NE) in the olfactory bulb (OB) of rat. Mechanical obstruction of the ventricular recess prevented intraventricular diffusion. At different time intervals following bilateral [³H]NE injections, total radioactivity was measured in the OB, locus caeruleus (LC), raphe dorsalis and periaqueductal gray. Preferential accumulation occurred in the LC, and an approximate rate of retrograde transport of 1–6 mm/h could be calculated. Previous administration of 6-hydroxydopamine in the OB reduced this accumulation by 60%. Sixteen hours after [³H]NE injection, the radioactivity in LC was equally distributed between an ethanol-soluble and -insoluble fraction. A small proportion of the soluble material was recovered as [³H]NE and/or [³H]normetanephrine. Following unilateral injections of [³H]NE, light microscopic autoradiograms demonstrated nerve cell body labeling mainly in the ipsilateral LC and of greater intensity after 16 than 4 and 8 h. These data lead to the conclusion that the movement of radioactive material was indeed representative of retrograde axonal transport rather than of other mechanisms such as diffusion. The observation of a preferential labeling of noradrenergic perikarya in LC supports the hypothesis of a process mediated by specific binding and/or uptake of [³H]NE into noradrenergic axon terminals.