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Biomedical subjects

C J Green

Publications and source records attributed to C J Green.

At least 109 records · Page 6Linked to original sources
PubMed1995

Increased axonal regeneration over long nerve gaps using autologous nerve-muscle sandwich grafts.

Poor recovery is seen after repair of long defects in peripheral nerves when denatured muscle grafts or regeneration chambers providing physical support alone are used. The presence of Schwann cells and neurotrophic factors is required for axons to migrate significant distances. In this study we have used immunohistochemical techniques and axon counts to quantify the regeneration seen when 5 cm defects in rabbit sciatic nerve were repaired with a composite graft consisting of 2-3 mm lengths of fresh autologous nerve sandwiched between 1 cm frozen-thawed muscle grafts. This technique led to a similar pattern of regeneration as that seen in autologous nerve grafts, used as controls, and a significantly (P < 0.0001) greater axonal and Schwann cell regeneration compared with that seen in frozen-thawed muscle grafts of the same total length. In conclusion, we present a simple technique for incorporating a depot of Schwann cells and other essential components into a nerve conduit which has a marked effect on axonal regeneration across long defects.

Animals↗
PubMed1995

The importance of calcium-related effects on energetics at hypothermia: effects of membrane-channel antagonists on energy metabolism of rat liver.

During normothermic metabolism, the active pumping of Ca2+ across the cell membrane, mitochondria, and specialized sequestration organelles accounts for a large proportion of total energy expenditure in the cell. This study was designed to determine the effects of Ca2+ channel antagonists (chlorpromazine, verapamil, nifedipine, prenylamine, and nisoldipine) on energy metabolism and levels of glycolytic substrate (glucose) and anaerobic endproduct (lactate) during cold ischemia in rat livers. We hypothesized that if the passive channels were blocked during cold ischemia, then the ATP requirement of active ion pumping would be reduced and ATP levels and energy charge ratios would remain higher throughout the ischemic period; thus, viability of the liver would also be increased after prolonged ischemia. The most positive effect on energy metabolism was observed in the chlorpromazine-treated livers, followed by verapamil treatment. In the chlorpromazine treatment, total adenylate (TA) contents were 0.5-1.0 mumol/g (P < 0.05) higher than the sham group for most of the 24-h time course. Energy charge (EC) ratios were 0.05-0.07 higher than the sham values up to 4-10 h ischemia. Verapamil treatment was less effective, but still exhibited positive effects on TA levels at several time points (20 min, 10 h, and 24 h) throughout the entire 24-h period. In both of these groups, TA values by 24 h ischemia were similar to levels at 10 h in the sham group (3.1 mumol/g), thus showing a considerable effect in maintaining adenylate levels. Despite similar pharmacological antagonist activities, ATP levels in the nifedipine, prenylamine, and nisoldipine treatment groups were 1.0-1.5 mumol/g (P < 0.05) less than the corresponding sham group (without Ca2+ antagonists) over the first 1 h ischemia. The decreases in high energy adenylate levels were reflected in lower EC ratios in these three groups; values were 0.06-0.17 (P < 0.05) lower than corresponding sham values. Finally, it was an unexpected finding that the sham injection (0.5 mg/kg ethanol+PEG400) resulted in the sustained elevation of ATP, total adenylates, and EC values over the first h; EC ratios remained at initial (t = 0) values (EC = 0.71 +/- 0.01) up to 1 h.

Adenosine Monophosphate↗
PubMed1995

An underlying mechanism for improved liver preservation with a combined histidine-lactobionate-raffinose flush solution.

In previous experimental liver transplant studies, it was possible to extend cold ischaemic time (CIT) by using a flush/storage solution combining histidine, lactobionate and raffinose (HLR). In this study, energy metabolism, glycolytic substrate (glucose) and anaerobic end-product (lactate) were examined in rat liver over 24 h of cold storage to determine the mechanism of action of the HLR solution. In livers subjected to simple flush and storage with the HLR solution, levels of ATP and ADP were considerably higher than livers stored with modified UW throughout 24 h of storage; at 4 h of storage, ATP and ADP levels were 1.1 and 3.1 mumol/g for HLR solution versus 0.18 and 0.81 mumol/g for UW solution. Total adenylate contents (TA = ATP + ADP + AMP) also remained 1-2 mumol/g higher in HLR-treated livers than those preserved in UW; TA values ranged from 3.8 to 5.7 mumol/g. Glucose increased to 20-35 mumol/g by 10-24 h of storage (similar to the UW group). Lactate rose to almost twice that in livers stored in UW; total lactate accumulation was approximately 10.0 mumol/g. This study demonstrated that the combined HLR solution is able to prolong the maximum 'safe' CIT by increasing anaerobic metabolism and consequently preserving liver energetics. The second part of the experiment examined the effect of continuous perfusion (with/without O2) over the 1st h of cold ischaemia. Under current methods of liver flushing and excision, the 1st h of cold storage may be the critical time of metabolic 'adjustment' since most of the pH and ATP changes occur during this period.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine↗
PubMed1995

Near infra-red spectroscopy: a non-invasive monitor of perfusion and oxygenation within the microcirculation of limbs and flaps.

Reliable early detection of adverse circulatory changes within a flap following free tissue transfer and early re-exploration are vital to minimise flap failure. Most surgeons rely on clinical assessment to monitor these changes but techniques such as plethysmography and laser Doppler have their advocates. These methods are limited however to measuring changes close to the surface. Near infra-red spectroscopy (NIRS) is a relatively new, non-invasive technique which allows continuous monitoring of concentration changes in oxy-, deoxy- and total haemoglobin (HbO2, Hb and HbT), as well as oxidised cytochrome aa3, through tissue up to 10 cm in depth. Information is provided on tissue oxygen supply, cellular oxygen utilisation, blood volume and perfusion status. A study has been performed in 10 rabbit hind limbs to assess the ability of NIRS to detect and distinguish between venous, arterial and total vascular occlusion. Clear patterns of change have been identified which allow rapid detection of vascular occlusion with accurate prediction of site. Arterial occlusions were characterised by an increase in Hb with a corresponding decrease in HbO2 and HbT. Venous occlusions resulted in an increase in HbT with relatively minor fluctuations in Hb and HbO2. Simultaneous occlusion of both artery and vein produced similar changes to those of arterial occlusion except that HbT decreased only minimally. These findings suggest that NIRS has a potentially useful role in the monitoring of free flaps, with the great advantage that perfusion can be measured to a considerable depth and information provided on the oxygenation profiles both accurately and non-invasively.

Animals↗
PubMed1995

Oxidative processes and free radical scavengers in ischaemia-reperfusion injury in adipocutaneous flaps: in vitro lipid peroxidation assessment.

An in vitro lipid peroxidation study measuring Schiff base and TBAR formation in homogenates of the fat and skin fractions of epigastric free flaps of DA-rats was performed to determine the role of oxygen-derived free radicals (ODFRs) in the aetiology of ischaemic injury and of ischaemia and reperfusion injury following cold (0-1 degree C) storage. The storage intervals were 0, 48, 72, 96 and 120 h in the study on ischaemic injury and 72 and 96 h in the study on combined ischaemia and reperfusion injury. Reperfusion was accomplished by anastomosing the pedicle vessels of the flap to the femoral vessels of a recipient; the reperfusion period was 15 min. In addition to a control group (C), three experimental groups were created to test the benefit of a preischaemic single passage perfusion with a hypertonic citrate solution (HCA), a pre- and postischaemic treatment with desferrioxamine (DFX) and a pre- and postischaemic treatment with lipoic acid (LA). The susceptibility of homogenates of skin and fat fractions of epigastric free flaps in DA-rats for lipid peroxidation increased significantly whenever the cold (0-1 degree C) ischaemic interval was prolonged from 0 to 72 h. These findings offer circumstantial evidence for the role of ODFRs in the aetiology of ischaemic injury. Following a short reperfusion period after extended periods of cold (0-1 degree C) ischaemia, no significant increase in susceptibility for lipid peroxidation could be found. Furthermore, no unequivocal role in the prevention of ischaemia and/or reperfusion injury by the use of either HCA, or DFX or LA could be found. The exact role of ODFRs in the aetiology of ischaemia-reperfusion injury in this setting remains unclear.

Adipose Tissue↗
PubMed1995

Different muscle graft denaturing methods and their use for nerve repair.

In an attempt to enhance recovery through denatured muscle grafts we have used different methods to denature muscle, either using dry heat in a microwave oven or by warming in sterile distilled water. Axonal regeneration was studied at intervals from day 5 to day 60 after insertion of the muscle grafts and compared to results found in frozen-thawed muscle grafts and in autologous nerve grafts used as controls. Axonal regeneration, Schwann cell behaviour and the degree of inflammation were quantified using immunohistochemical techniques and computerised image analysis. Autologous nerve grafts supported the highest rates and volumes of axonal regeneration until day 30 when the microwaved muscle grafts had the highest values for axonal immunostaining and at 60 days the highest level of Schwann cell immunoreactivity. We conclude that microwave heating is suitable as an alternative denaturing method for successful muscle grafts and has a potential clinical use.

Animals↗
PubMed1995

Protective properties of amino acids in liver preservation: effects of glycine and a combination of amino acids on anaerobic metabolism and energetics.

BACKGROUND/AIMS/METHODS: In this study, we investigated the hepatoprotective effects of three storage solutions containing glycine (180 mM), glycylglycine (180 mM), and a mixture of 20 amino acids (combined concentration of 180 mM) on energy metabolism and levels of glucose and lactate (as an index of glycolytic flux) in rat livers. All effects were compared to those of livers flushed/stored with a modified University of Wisconsin solution. RESULTS: Glycine-treatment showed no improvement in liver energetics (ATP, ADP, AMP) and lactate accumulation; this solution had the lowest buffering capacity of the four tested (approximately 30% of the University of Wisconsin solution). The glycylglycine solution had the highest buffering capacity of the four solutions tested (including University of Wisconsin solution). Complete titration of the glycine-, combined amino acids-, and University of Wisconsin solutions (from 8.0 to pH = 6.0) resulted in a minor decrease in glycylglycine buffer pH; pH dropped by 0.2 pH units. In glycylglycine-treated livers, energetics showed an improvement over the first 1 h cold storage; ATP and 'energy charge' values remained high and ADP levels (and consequently total adenylate contents) were 0.7-2.4 micro mol/g greater than livers stored in University of Wisconsin solution. A 2-fold increase in lactate accumulation suggested that the improvement in liver energetics for the glycylglycine buffer was due to maintained flux through glycolysis brought about by enhanced buffering capacity. The solution containing a combination of amino acids exhibited maximum maintenance of liver energetics via increased glycolytic flux, despite its slightly inferior buffering capacity (85% of University of Wisconsin solution). ATP levels were maintained over the first 2 h storage and ADP levels (and consequently, total adenylate contents) were 1.2-2.1 micro mol/g greater than University of Wisconsin solution-treated livers during the entire 24 h storage period. Energy charge values for livers treated with the combination of amino acids were also significantly higher than with glycine-, glycylglycine- and University of Wisconsin solution-treatment; even at 24 h, energy charge was 0.36 (comparable to only 4 h storage in University of Wisconsin solution). CONCLUSIONS: Our data suggest that a combination of amino acids may be required for maximum protection of the liver, and furthermore there may be several independent mechanisms, including buffering capacity, responsible for cytoprotection of the liver during cold storage.

Adenosine↗
PubMed1995

Nutritional support in HIV infection and AIDS.

Weight loss and wasting are common features of HIV infection and AIDS. Patterns of weight loss can be acute or chronic which appear to be related primarily to systemic infections and gastrointestinal pathology, respectively. However, weight loss is not inevitable. and periods of weight stability and weight gain have been documented. Reduced food intake appears to be a major cause of weight loss in HIV infection. Since time of death has been associated with degree of wasting, it seems reasonable to suggest that nutritional support may contribute to enhancing survival and quality of life. All patients should have early access to a qualified dietitian such that assessment of individual situations can be made and appropriate dietary advice given, within a multi-disciplinary approach. Choice of nutritional therapy should be made based on an assessment of the causes of weight loss and an assessment of gut function. Treating infections and alleviating symptoms is vital for ensuring effective nutritional support. Enhancing the energy and protein density of foods and use of oral supplements should be considered if a normal diet alone cannot meet nutritional requirements. Unnecessary dietary restrictions should be avoided. Enteral feeding is indicated for patients unable to meet their needs via the oral route, and in cases of inadequate gut function, parenteral nutrition may be necessary. There is currently insufficient clinical evidence to justify the need for special enteral formulae specifically for patients with HIV infection.

Journal Article↗
PubMed1995

Nerve-muscle sandwich grafts: the importance of Schwann cells in peripheral nerve regeneration through muscle basal lamina conduits.

An interposed segment of nerve was used to enhance the distance over which freeze-thawed muscle autografts will support effective peripheral nerve regeneration. Gaps were created in the sciatic nerves of adult Lewis rats. Regeneration through 1 and 1.5 cm freeze-thawed muscle grafts was compared to regeneration through nerve-muscle sandwich grafts in which muscle grafts of equivalent length were divided and a 2 mm segment of the distal nerve sutured between the two halves of the muscle, providing an intermediate depot of Schwann cells. Electrophysiological and morphological evaluation was carried out 40 weeks after operation. Despite lengthening the graft, and having four anastomoses instead of two, this manoeuvre enhanced nerve regeneration over each gap studied and for the 1.5 cm gaps compared favourably with perfect match nerve autografts. In addition, a number of grafts were examined at 7 and 14 days by S100 immunohistochemistry. Schwann cell migration was seen to proceed both proximally and distally from the intermediate segment at a rate similar to that from the distal stump. It is concluded that sandwich grafts may prove to be effective alternatives to cutaneous nerve grafts for peripheral nerve reconstruction.

Analysis of Variance↗
PubMed1995

Orientated mats of fibronectin as a conduit material for use in peripheral nerve repair.

This study introduces a new nerve conduit material consisting of orientated strands of the cell adhesive fibronectin. Axonal regeneration, Schwann cell behaviour and the degree of inflammation were quantified using immunohistochemical techniques and computerized image analysis. The results when fibronectin was used to bridge a 1 cm defect in rat sciatic nerve were compared to those with autologous nerve grafts and freeze-thawed muscle grafts used as controls. The nerve grafts supported the highest rate and amount of axonal regeneration in the first 10 days; however, the fibronectin supported a significantly faster rate of growth (P = 0.0008) and amount (P = 0.0001) of axons than the freeze-thawed muscle grafts. From day 15 onwards, the fibronectin and nerve grafts had comparable amounts of regenerating axons and Schwann cells. We conclude that the orientated form of fibronectin is a suitable material for successful nerve repair and has potential clinical use.

Analysis of Variance↗
PubMed1995

Septic patients in multiple organ failure can oxidize infused glucose, but non-oxidative disposal (storage) is impaired.

1. Patients suffering trauma and sepsis are insulin resistant, but no studies have specifically been made of patients suffering multiple organ failure. 2. We have studied exogenous glucose utilization in multiple organ failure using a combination of the hyperglycaemic glucose clamp and indirect calorimetry to quantify glucose utilization in multiple organ failure, partitioning it into oxidative and nonoxidative disposal (storage). 3. Fourteen septic patients with multiple organ failure were studied. APACHE II (Acute Physiological and Chronic Health Evaluation Mark II) scores on the day of the study ranged from 11 to 31 (median 16). Twenty percent D-glucose was infused and blood glucose was clamped at 12 mmol/l for 3 h. The results were compared with those obtained on seven healthy control subjects. 4. Glucose utilization and energy expenditure were similar in the two groups for the first 90 min of the clamp, after which glucose utilization and energy expenditure increased steadily in the control subjects but did not change in the patients. Respiratory exchange ratio rose in both groups; considered over the whole of the clamp period, respiratory exchange ratio was slightly lower in the patients than in the control subjects (P < 0.05) but not at any specific time point. Glucose oxidation rose in both groups but non-oxidative glucose disposal (storage) rose only in the control subjects. Glucose oxidation was slightly lower in the patients (P < 0.05) but not at any specific time point and there was no difference between the groups in the amount by which glucose oxidation increased. Non-oxidative disposal in the patients fell significantly (P < 0.01) over the course of the clamp and was significantly lower than in the control subjects (P < 0.01). 5. Growth hormone increased in response to glucose infusion in the patients but not in the control subjects. 6. Like patients suffering uncomplicated sepsis or trauma, patients with multiple organ failure are also insulin resistant. The defect appears to lie in an impairment of the ability to store glucose rather than oxidize it, and this may be due in part to the increase in growth hormone in patients with multiple organ failure.

Adolescent↗
PubMed1995

Regeneration pattern of blood vessels and nerves in cultured keratinocyte grafts assessed by confocal laser scanning microscopy.

The aim of this study was to investigate the pattern of both neovascularization and reinnervation, and the relationship between the two processes, in keratodermal grafts, using confocal laser scanning microscopy, at different time points during the healing process. Keratodermal grafts were prepared in pigs by combining autologous dermis with cultured autologous keratinocytes. Immunohistochemistry was carried out on thick cryostat sections (100-150 microns), using antisera to the endothelial marker von Willebrand factor (vWf) and the pan-neuronal marker protein gene product 9.5 (PGP9.5). The results suggest that the neovascularization and reinnervation in the cultured keratodermal graft is almost complete at 6 weeks. Neovascularization precedes innervation, reaching the surface covered by the keratinocytes at 2 weeks, initially with a linear vascular pattern. From 3 weeks, there is a gradual arborization of the vessels to form a typical vascular plexus. The process of reinnervation is similar in pattern to that of neovascularization, although slower in developing a full network of fibres. In conclusion, the use of confocal microscopy allows the precise definition of complex patterns of neovascularization and nerve growth, which are not fully apparent when using conventional microscopy. Because angiogenesis occurs first, it probably plays a leading role in the survival of keratodermal grafts during wound healing. Indeed, new blood vessels form a pathway for the subsequent innervation process, and quickly reach the epidermal layer which, in turn, may play a key role in the tropism of both blood vessels and nerves.

Animals↗
PubMed1995

High-level expression of soluble recombinant RNase P protein from Escherichia coli.

We have expressed recombinant RNase P protein from Escherichia coli in high yield. A hexahistidine sequence at the amino terminus allowed protein purification in a single step. Mass spectrometry confirmed the molecular weight of the purified protein and indicated a purity of > 95%. Protein functionality was demonstrated by reconstitution of active holoenzyme.

Bacterial Proteins↗
PubMed1995

Dantrolene sodium protects against experimental ischemia and reperfusion damage to skeletal muscle.

The effect of 4 hours of ischemia followed by reperfusion for 1 hour has been studied in fully anesthetized rabbits. Muscles from the limb subjected to ischemia and reperfusion showed considerable ultrastructural damage, although the distribution of damage between muscles was not uniform (anterior tibialis > soleus > quadriceps). Damage to the muscle was associated with a significant increase in the concentration of some indicators of free radical-mediated processes (thiobarbituric acid-reactive substances and diene conjugates), although others (glutathione and protein sulfhydryl groups) were unchanged. Reperfused muscles also showed considerable changes in their calcium and sodium contents. Treatment of animals with dantrolene sodium (4 mg/hr) throughout the periods of ischemia and reperfusion was found to preserve the ultrastructural appearance of quadriceps, soleus and anterior tibialis muscles. No effect of dantrolene sodium on indicators of free radical activity or muscle cation content was seen.

Animals↗