Search PubMed⌕ Search

Biomedical subjects

C J Green

Publications and source records attributed to C J Green.

At least 73 records · Page 4Linked to original sources
PubMed1998

Depression of endothelial and smooth muscle cell oxygen consumption by endotoxin.

An optical method based on the oxygen-dependent quenching of a phosphorescent probe (palladium-porphyrin) was used to investigate the effect of bacterial endotoxin [lipopolysaccharide (LPS)] on oxygen consumption (VO2) by vascular cells. Endothelial (EC) and smooth muscle (SMC) cells from pig aorta were suspended in culture medium in the presence of palladium-porphyrin and transferred to glass capillary tubes that were sealed to create a hypoxic environment. Measured PO2 changed as a function of time in a highly predictable fashion when cell suspensions were exposed to agents or treatment known to affect cellular metabolism. Both EC and SMC showed a significant decrease in VO2 as cell density increased, and SMC VO2 was significantly higher than EC (1.94 +/- 0.09 vs. 1.0 +/- 0.15 nmol . min-1 . 10(6) cells-1). Exposure to LPS (1 microg/ml) caused a decrease in VO2 of 46% and 15% for EC and SMC, respectively. Pretreatment of cells with N-acetyl-L-cysteine, a substrate for glutathione synthesis with antioxidant properties, restored VO2 to normal values after exposure to LPS. These data suggest that endotoxin impairs VO2 in cells derived from the vascular wall and indicate the importance of EC and SMC respiration in maintaining vascular homeostasis under conditions of sepsis.

Acetylcysteine↗
PubMed1997

Involvement of the heme oxygenase-carbon monoxide pathway in keratinocyte proliferation.

It has been suggested that nitric oxide (NO), a small gaseous molecule with a multiplicity of cellular functions, plays an important part in the regulation of cellular proliferation. We have examined the effect of the NO donor sodium nitroprusside (SNP) on heme oxygenase-1 (HO-1) expression in human epidermal keratinocytes and investigated the contribution of the heme oxygenase pathway in the control of keratinocyte proliferation. Incubation of keratinocytes with 0.5 mM SNP resulted in a 2.5-fold increase in heme oxygenase activity which was reflected by a significant increase in HO-1 protein expression, as measured by Western blot. This effect was associated with a 200% increase in keratinocyte proliferation. The proliferative effect of the NO donor was totally abolished by co-incubation of SNP with tin protoporphyrin IX, a potent inhibitor of heme oxygenase, or hydroxocobalamin, a NO scavenger. These results suggest that the heme oxygenase pathway is involved in keratinocyte proliferation mediated by NO.

Carbon Monoxide↗
PubMed1997

Apoptosis after intestinal ischemia-reperfusion injury: a morphological study.

BACKGROUND: Apoptosis has been identified after ischemia-reperfusion (IR) injury to the brain, heart, kidney, retina, and the adrenals. Intestinal IR injury causes villous and crypt damage, which has so far been attributed to cellular necrosis. This study was undertaken to investigate the possible role of apoptosis after reperfusion of cold-stored small bowel grafts in syngeneic rats. METHODS: Small intestinal grafts were stored at 4 degrees C for 24 hr in saline (n=6) or in modified University of Wisconsin solution (n=6), followed by reperfusion for 1 hr in syngeneic Lewis rats. Small bowel samples were obtained before storage, after preservation and after 1 hr of reperfusion. They were processed for light and electron microscopy and analyzed for cell death, with particular emphasis on apoptosis. RESULTS: Less than one apoptotic event was seen per 10 crypts in normal and stored bowels. An occasional normal and some denuded villous epithelial cells of stored bowels exhibited apoptosis. After isotransplantation and 1 hr of reperfusion, marked increase in apoptosis was seen in the crypts and denuded villous epithelial cells of both saline- and modified University of Wisconsin-stored bowels. Secondary necrosis was seen in apoptotic cells, as were dark cells. Only a few cells showed signs of primary ischemic necrosis. CONCLUSIONS: Apoptosis occurs after intestinal IR injury. Modulation of its genetic regulatory and biochemical effector machinery might alleviate or even prevent IR injury in small bowel transplanted after similar periods of storage.

Animals↗
PubMed1997

Neurotrophin-3-enhanced nerve regeneration selectively improves recovery of muscle fibers expressing myosin heavy chains 2b.

The purpose of this study was to evaluate the effect of neurotrophin 3 (NT-3) enhanced nerve regeneration on the reinnervation of a target muscle. Muscle fibers can be classified according to their mechanical properties and myosin heavy chain (MHC) isoform composition. MHC1 containing slow-type and MHC2a or 2b fast-type fibers are normally distributed in a mosaic pattern, their phenotype dictated by motor innervation. After denervation, all fibers switch to fast-type MHC2b expression and also undergo atrophy resulting in loss of muscle mass. After regeneration, discrimination between fast and slow fibers returns, but the distribution and fiber size change according to the level of reinnervation. In this study, rat gastrocnemius muscles (ipsilateral and contralateral to the side of nerve injury) were collected up to 8 mo after nerve repair, with or without local delivery of NT-3. The phenotype changes of MHC1, 2a, and 2b were analyzed by immunohistochemistry, and fiber type proportion, diameter, and grouping were assessed by computerized image analysis. At 8 mo, the local delivery of NT-3 resulted in significant improvement in gastrocnemius muscle weight compared with controls (NT-3 group 47%, controls 39% weight of contralateral normal muscle; P < 0.05). NT-3 delivery resulted in a significant increase in the proportion (NT-3 43.3%, controls 35.7%; P < 0.05) and diameter (NT-3 87.8 micron, controls 70.8 micron; P < 0.05) of fast type 2b fibers after reinnervation. This effect was specific to type 2b fibers; no normalization was seen in other fiber types. This study indicates that NT-3-enhanced axonal regeneration has a beneficial effect on the motor target organ. Also, NT-3 may be specifically affecting a subset of motoneurons that determine type 2b muscle fiber phenotype. As NT-3 was topically applied to cut nerves, our data suggest a discriminating effect of the neurotrophin on neuro-muscular interaction. These results would imply that muscle fibers may be differentially responsive to other neurotrophic factors and indicate the potential clinical role of NT-3 in the prevention of muscle atrophy after nerve injury.

Animals↗
PubMed1997

Cyclophilins are induced by hypoxia and heat stress in myogenic cells.

This is a novel study demonstrating that cyclophilins are heat and stress inducible proteins in eukaryotic myogenic cells. We investigated the expression of cyclophilins in embryonal rat heart derived H9c2 myocytes following heat stress and chronic hypoxia. We report here that cyclophilins, the proteins capable of catalysing the interconversion of cis and trans isomers (PPIses) in proteins and peptides, are heat and stress inducible, and are involved in the complex stress response, as their level is significantly elevated after heat stress and hypoxia. A time course analysis showed the gradual increase in expressed levels of cyclophilin after heat stress of cells, with maximal expression as measured by Western blot at 48 hours after the actual treatment. Rat myogenic cells exposed to chronic hypoxia followed by 5 hours reoxygenation resulted in approximately threefold expression of PPI-ases. The results showing that cyclophilins are heat and stress inducible suggest a multiple role for cyclophilins in ischemia: a potential functional association with the different heat shock proteins, with the established protective role in ischaemic injury, as well as the possible involvement of cyclophilins in the protein folding in cooperation with molecular chaperones.

Amino Acid Isomerases↗
PubMed1997

Thiol compounds interact with nitric oxide in regulating heme oxygenase-1 induction in endothelial cells. Involvement of superoxide and peroxynitrite anions.

Thiols are very important antioxidants that protect cells against oxidative insults. Recently, a different and new physiological role has been defined for these compounds because of their involvement in nitric oxide (NO) binding and transport in biological systems. In view of these characteristics, we examined the effect of thiols and NO on the expression of the inducible form of heme oxygenase (HO-1), a stress protein that degrades heme to carbon monoxide and biliverdin. Cultured bovine aortic endothelial cells exposed to the NO donors sodium nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine (SNAP) resulted in increased heme oxygenase activity and HO-1 expression. Co-incubation with N-acetylcysteine, a precursor of glutathione synthesis, significantly attenuated heme oxygenase induction by SNP and SNAP, and a reduction in heme oxygenase activity was also observed when cells were preincubated with N-acetylcysteine for 16 h prior to exposure to NO donors. This effect appears to be associated with NO stabilization by thiols through the formation of S-nitrosothiols. Hydroxocobalamin, a specific NO scavenger, significantly decreased endothelial heme oxygenase activity, indicating a direct involvement of NO released by NO donors to regulate the expression of this stress protein. Moreover, superoxide anion (O-2) and its reaction product with NO, peroxynitrite (ONOO-), were found to partially contribute to the observed NO-mediated activation of endothelial heme oxygenase. Thus, we suggest the existence of a dynamic equilibrium among free NO, O-2, and endogenous glutathione, which might constitute an interactive signaling mechanism modulating stress and adaptive responses in tissues.

Acetylcysteine↗
PubMed1997

Measurements of tissue viability in transplantation.

Near-infrared spectroscopy has primarily been used in monitoring changes in cerebral haemoglobin oxygenation and haemodynamics. However its use as a method for the assessment of tissue viability following transplantation has recently been explored experimentally in our laboratory. The ability to measure changes in oxygenation and perfusion during harvesting and following transplantation of organs or transfer of free and pedicled flaps potentially important in reconstructive surgery. We have found that near-infrared spectroscopy is extremely useful in detecting vaso-occlusive events and can accurately and reliably distinguish between arterial, venous or total occlusions. Venous congestion indicated by raised levels of deoxygenated haemoglobin with a concomitant increase in blood volume and the presence and magnitude of reactive hyperaemia are both easily recognizable features by near-infrared spectroscopy. We have shown that near-infrared spectroscopy measurements of venous congestion in kidneys (and other tissues) following prolonged storage correlate with medullary vascular congestion confirmed by angiographical and histological analysis of intrarenal perfusion. Clinically we have shown that flap perfusion can be improved by altering fluid replacement regimes and the addition of ionotropes. Cerebral near-infrared spectroscopy measurements in a liver transplant model showed statistically significant differences within minutes after the anhepatic phase in cerebral perfusion and oxygenation, between animals transplanted with ischaemically damaged livers compared to those isografted with minimally stored livers. Similarly we have found that near-infrared spectroscopy can be used as a monitor to assess the adequacy of fluid or blood replacement in haemorrhagic and hypovolaemic models. We believe that near-infrared spectroscopy provides a sensitive and reliable postoperative method for the assessment of tissue viability following the transfer of free and pedicled flaps and organs.

Animals↗
PubMed1997

The development of neovascularisation in flap perfabrication with vascular implantation: an experimental study.

Femoral arteriovenous pedicles were implanted beneath the abdominal skin in rats (n = 30) to investigate the development of neovascularisation over time. New vessel formation was assessed by micro-angiography and quantified under light microscopy. Flap viability was assessed by dye injection studies at different time intervals. Two weeks after implantation of the vascular pedicles, neovascularisation was confined to the vicinity of the arteriovenous pedicle only and none of the flaps were viable. At 4 and 6 weeks, 4 out of 6 flaps were viable, with new vessel formation being widespread throughout the subcutaneous tissue. All flaps were viable at 8 and 12 weeks. Neovascularisation in the dermis up to the epidermis was observed after 8 weeks. Neovascularisation in the panniculus carnosus showed a steady increase after 6 weeks. There was a decrease in the total number of vessels at 12 weeks compared to 8 weeks but at 12 weeks the diameters of the vessels were considerably larger. Flap survival was best predicted by the amount of neovascularisation in the panniculus carnosus. In this experimental model, it is concluded that prefabricated flaps should be raised at 8 weeks instead of 4 or 6 weeks after the implantation of an arteriovenous pedicle, to reduce the risk of flap failure.

Angiography↗
PubMed1997

Inter-relationships between angiogenesis and nerve regeneration: a histochemical study.

Whilst increases in capillary number and permeability occurring during nerve regeneration suggest an interaction between regenerating axons and blood vessels, clinical attempts to improve nerve regeneration by augmenting nerve graft vascularisation have produced conflicting results and the nature of their relationship remains obscure. A better understanding of the process might be exploited in the development of a synthetic alternative to the autologous nerve graft and bring an improvement in the clinical results of nerve repair. To clarify this relationship the growth of axons and blood vessels through mats of orientated fibronectin grafted in rat sciatic nerves was assessed morphologically. Fibronectin, which supports axonal regeneration, is initially acellular, ensuring all vascular and neural elements within the graft are newly formed. To follow the progression of the elements, grafts were harvested between 3 and 30 days and stained with antibodies against endothelial cells (RECA-1), Schwann cells (S-100) and axons (a polyclonal or monoclonal panaxonal marker). Dual fluorescence staining combined with double exposure photography allowed the simultaneous visualisation of these elements and the demonstration of their true relative positions. Graft vascularisation came initially from the adjacent muscle bed. A neovascularisation front preceded axonal regeneration, although vessel and axonal orientation appeared similar. Schwann cells and axons extended together, never exceeding the area of vascularisation and appeared most numerous in well vascularised areas containing longitudinally orientated vessels. These results suggest that provision of a well vascularised, longitudinally orientated conduit may enhance nerve regeneration.

Animals↗
PubMed1997

Cold intolerance following peripheral nerve injury. Natural history and factors predicting severity of symptoms.

Cold intolerance can be severe and debilitating following injury to the hand. Little is known about its natural history and factors predicting symptom severity. We looked retrospectively at upper limb peripheral nerve injuries over a 12-year period. Information was obtained using a patient questionnaire and patient records. The incidence of cold intolerance was 83%. In 48% the onset of symptoms was within 1 month of the initial injury. At a mean follow-up of 51 months improved symptoms were reported by 21%, while 18% deteriorated. Patients were more likely to develop cold intolerance if they smoked and less likely if they suffered a sharp injury. A score defining the severity of cold induced symptoms, based on the information collected, was calculated for each patient. Significantly increased severity was associated with complete nerve division, median and ulnar nerve division and an associated vessel injury. Symptom improvement was significantly more likely in non-smokers and a deterioration most likely with a high severity score.

Adolescent↗
PubMed1997

Measurements of oxygenation and perfusion in skeletal muscle using multiple microelectrodes.

This paper describes an apparatus to measure tissue oxygenation and perfusion (as measured by the wash-in rate of gaseous hydrogen) simultaneously at multiple points in muscle using needle microelectrodes. The development of suitable electrodes and apparatus is described, as well as the development of the method and its validation. In particular, the potential for tissue damage secondary to electrode insertion, the need for in vivo voltammetric determination of the operating potential and the extent of any electrode-tissue and of electrode-electrode interactions are explored, and are shown to be insufficient in magnitude to affect the technique. Its subsequent use to characterise oxygenation and perfusion in rabbit skeletal muscle at rest is also described. In resting tibialis anterior muscle of the rabbit the mean pO2 was 18 +/- 13.3 mm Hg and the mean perfusion was 4.4 +/- 1.3 ml s-1 100 g-1. There was a heterogeneity in simultaneously-measured values of pO2 and perfusion at different points within muscle, and also a temporal variation at the same site. The spans between the highest and lowest simultaneously-measured values of pO2 in muscle ranged from 14 to 80 mm Hg, and for perfusion, from 1 to 12 mls-1 100 g-1. No significant correlation was evident from histological examination between either pO2 or perfusion and surrounding fibre type or capillary density.

Animals↗
PubMed1997

Characterization of apoptosis in intestinal ischaemia-reperfusion injury--a light and electron microscopic study.

Intestinal ischaemia-reperfusion (IR) injury has largely been attributed to cellular necrosis. Apoptosis, a distinct form of cell death has been observed following IR to the brain, heart, adrenals and the kidneys. In order to characterize the role of apoptosis in intestinal IR, small bowel grafts were stored in saline (n = 6) or modified University of Wisconsin solution (n = 6) at 4 degrees C for 12 h and reperfused for 6 h in syngeneic rats. Samples of normal, stored and reperfused intestines at 1, 3 and 6 h were analysed by light and electron microscopy. Following reperfusion, there was crypt and villous epithelial apoptosis, loss of crypt and villous structures, and an increase in mucosal inflammatory cell infiltration. Ongoing apoptosis was maximum at 1 h, its degree decreasing with increasing reperfusion intervals. Large numbers of apoptotic bodies dominated the picture from 3 h of reperfusion. This study has demonstrated the induction of apoptosis by intestinal IR injury, which begins within an hour of reperfusion and is probably responsible for the observed crypt and villous loss. This has potential therapeutic implications as, opposed to necrosis, apoptosis is an active process with genetic regulators and biochemical effectors, which can be specifically targeted to prevent or alleviate IR injury.

Adenosine↗
PubMed1997

Neurotrophin-3 delivered locally via fibronectin mats enhances peripheral nerve regeneration.

A better understanding of the mechanisms of nerve regeneration could improve the outcome of surgical nerve repair. We have previously shown that axonal regeneration is increased by nerve growth factor. Neurotrophin-3 (NT-3) belongs to the same family as nerve growth factor but acts on a distinct neuron subpopulation. As little is known about its role following nerve injury, we have investigated the effect of NT-3 delivered via fibronectin mats, previously shown to support nerve regeneration comparable to nerve grafts. NT-3 stimulation (0.1-1000 ng/ml) of neurite extension from embryonic chick dorsal root ganglia in vitro has shown that fibronectin can bind and release bioactive NT-3. Fibronectin mats impregnated with NT-3 (500 ng/ml) were grafted into 1 cm sciatic nerve defects in adult Lewis rats. Plain mats were used as controls. Computerized quantification of penetration distance, volume of axonal regeneration and myelinated fibre counts was undertaken using immunostaining for axonal markers (growth-associated protein 43, calcitonin gene-related peptide, substance P, vasoactive intestinal peptide and neuropeptide tyrosine), or S100 or thionine blue staining up to 8 months postoperatively. The maximal effect of NT-3 occurred at day 15, when for GAP43-immunostained axons both penetration distance (NT-3, 6.10 +/- 0.42 mm; control, 4.11 +/- 0.41 mm; P < 0.01) and staining area (NT-3, 0.137 +/- 0.012 mm2; control, 0.077 +/- 0.018 mm2; P < 0.05) were significantly increased. Similar results were found for each neuronal subpopulation investigated. By 8 months after repair, the NT-3 group supported a significantly greater number of myelinated axons (NT-3, 7003 +/- 402; control, 4932 +/- 725; P < 0.05) of similar diameter and g-ratio to controls. These results demonstrate the contribution of NT-3 to the increase of nerve regeneration promoted by growth factors.

Administration, Topical↗
PubMed1997

Neuropathy in non-freezing cold injury (trench foot).

Non-freezing cold injury (trench foot) is characterized, in severe cases, by peripheral nerve damage and tissue necrosis. Controversy exists regarding the susceptibility of nerve fibre populations to injury as well as the mechanism of injury. Clinical and histological studies (n = 2) were conducted in a 40-year-old man with severe non-freezing cold injury in both feet. Clinical sensory tests, including two-point discrimination and pressure, vibration and thermal thresholds, indicated damage to large and small diameter nerves. On immunohistochemical assessment, terminal cutaneous nerve fibres within the plantar skin stained much less than in a normal control whereas staining to von Willebrand factor pointed to increased vascularity in all areas. The results indicate that all nerve populations (myelinated and unmyelinated) were damaged, possibly in a cycle of ischaemia and reperfusion.

Adult↗
PubMed1997

Anticoagulation in chronic nonvalvular atrial fibrillation: a critical appraisal and meta-analysis.

OBJECTIVE: To assess the outcomes associated with warfarin treatment of patients with chronic nonvalvular atrial fibrillation (CNVAF) for prevention of primary stroke. DATA SOURCES: MEDLINE was searched for literature published from 1987 to August 1996. Search terms used were 'atrial fibrillation' and 'anticoagulants'. STUDY SELECTION: Five published randomized controlled trials concerning primary stroke prevention. DATA EXTRACTION: Data were pooled across trials to estimate the magnitude of the effect for each of nine reported end-points. The annual probability of occurrence of each outcome was calculated, including standard errors and Mantel-Haenszel significance tests with 95% CIs. DATA SYNTHESIS: In view of the lack of blinded assessment and documented low inter-rater reliability of soft neurological end-points, the analysis was limited to the relatively objective end-points of major strokes, fatal strokes, major bleeding and fatal bleeding. Warfarin did not reduce the incidence of fatal strokes to a statistically significant extent, nor was incidence of fatal bleeding increased significantly. Warfarin reduced the absolute annual incidence of major strokes in patients with CNVAF by 0.89%, while at the same time it increased the absolute annual risk of major bleeding incidents by 1.8%. Though small, these differences were statistically significant. CONCLUSIONS: On balance, the margin between expected benefit and harm for warfarin prophylaxis in patients with CNVAF is uncomfortably thin. These results and conclusions differ from those of a previously published meta-analysis of these same studies.

Anticoagulants↗
PubMed1996

Effect of mannitol and polyethylene glycol on the action of frusemide during renal storage and transplantation.

Hypoxic injury is a major cause of tubular necrosis in the corticomedullary junction of isolated perfused kidneys, and is ameliorated by inhibitors of active reabsorption, such as frusemide. Our objective was to determine whether frusemide has a similar effect on hypothermically stored transplanted kidneys and whether this effect is modulated by impermeant solutes included in the preservation solution. The effect of frusemide on cytochrome oxidase (cyt aa3) oxidation, renal hemodynamics, and morphology was investigated in the New Zealand White rabbit renal autograft model using near-infrared spectroscopy and light microscopy. A total of 30 kidneys were autografted in six groups. Kidneys were transplanted with or without frusemide either (1) without storage (groups 1 and 2) or after 72 hr of storage in: (2) hypertonic citrate containing mannitol (groups 3 and 4); and (3) hypertonic citrate containing polyethylene glycol (PEG) (groups 5 and 6). In unstored transplanted kidneys, frusemide infusion stimulated a significant (P < 0.05) increase in hemoglobin oxygenation, compared with untreated controls. There was a tendency for cyt aa3 to become reduced, but there were no significant differences between groups 1 and 2. After 72 hr of storage, frusemide infusion stimulated a significant increase in hemoglobin oxygenation in kidneys stored in mannitol (P < 0.005), but there was no significant change in the kidneys stored in PEG. There was a corresponding reduction in cyt aa3 in kidneys stored in mannitol (P < 0.05) but no change in those stored in PEG. These results suggest that frusemide has a significant effect on cortical hemoglobin oxygenation in transplanted kidneys and on active reabsorption in the corticomedullary junction. The selection of impermeant is important and mannitol is significantly superior to PEG in this model.

Animals↗