Validity and responsiveness of the spinal cord index of function: an instrument on activity level.

OBJECTIVES: To evaluate the validity and responsiveness of the Spinal Cord Index of Function (SIF), a new instrument on activity level, measuring the ability to perform various transfers in non-walking patients with a spinal cord lesion. SETTINGS: Spinal Injuries Unit, Sahlgrenska University Hospital, Gothenburg, Sweden. METHODS: Twenty-nine patients with a spinal cord lesion classified as grade A, B or C according to the American Spinal Injury Association/International Medical Society of paraplegia classification were included. Each patient was evaluated from the acute phase until discharge, every second week, by their physiotherapist, according to SIF and the Swedish physiotherapy clinical outcome variables (S-COVS). To determine validity, Spearman's rho correlation coefficient was calculated between the total scores of SIF and S-COVS, and the determination coefficient was calculated. Responsiveness was determined by computing effect sizes. RESULTS: Spearman's correlation between SIF and S-COVS was 0.933 and the determination coefficient was 0.87. The effect size for SIF was 9.1. CONCLUSION: The results of the study prove that SIF is a valid and sensitive instrument, which will be useful for physiotherapists in goal-planning programs and in evaluating progress during a patient's rehabilitation. SIF could also be used in research and in evaluating the patient's functional ability at follow-ups.

Effects of abdominal binding on breathing patterns during breathing exercises in persons with tetraplegia.

STUDY DESIGN: Cross-sectional, experimental. OBJECTIVES: To investigate and compare static lung volumes and breathing patterns in persons with a cervical spinal cord lesion during breathing at rest, ordinary deep breathing, positive expiratory pressure (PEP) and inspiratory resistance-positive expiratory pressure (IR-PEP) with and without an abdominal binder (AB). SETTING: The outpatient clinic at the Spinal Unit at Sahlgrenska University Hospital, Goteborg, Sweden. METHOD: The study group consisted of 20 persons with complete cervical cord lesion at C5-C8 level. Breathing patterns and static lung volumes with and without an AB were measured using a body plethysmograph. RESULTS: : With an AB, static lung volumes decreased, vital capacity increased, breathing patterns changed only marginally and functional residual capacity remained unchanged during PEP and IR-PEP. CONCLUSION: Evidence supporting the general use of an AB to prevent respiratory complications by means of respiratory training is questionable. However, the interindividual variation in our results indicates that we cannot rule out that some patients may benefit from the treatment.

Regulation of substance P mRNA expression in human dermal microvascular endothelial cells.

Vascular endothelial cells have been identified as a source of substance P (SP) which may act in an autocrine/paracrine fashion to bring about nitric oxide (NO)-dependent vasodilatation and mitogen-induced cell division or immunologic and inflammatory responses. Whilst SP is localised in and released from endothelial cells, an endothelial mRNA expression of SP has not previously been shown. In the present study, mRNA expression of SP in human dermal microvascular endothelial cells is demonstrated using in situ hybridisation techniques with enhancement procedures. Incubation of microvascular endothelial cells with nerve growth factor (NGF) under conditions of increased shear stress increases the mRNA expression and release of SP Endothelin (ET) release is also enhanced. These changes are pertinent to circulatory, events that may occur in Raynaud's phenomenon in systemic sclerosis.

Earthquake nucleation by transient deformations caused by the M = 7.9 Denali, Alaska, earthquake.

The permanent and dynamic (transient) stress changes inferred to trigger earthquakes are usually orders of magnitude smaller than the stresses relaxed by the earthquakes themselves, implying that triggering occurs on critically stressed faults. Triggered seismicity rate increases may therefore be most likely to occur in areas where loading rates are highest and elevated pore pressures, perhaps facilitated by high-temperature fluids, reduce frictional stresses and promote failure. Here we show that the 2002 magnitude M = 7.9 Denali, Alaska, earthquake triggered widespread seismicity rate increases throughout British Columbia and into the western United States. Dynamic triggering by seismic waves should be enhanced in directions where rupture directivity focuses radiated energy, and we verify this using seismic and new high-sample GPS recordings of the Denali mainshock. These observations are comparable in scale only to the triggering caused by the 1992 M = 7.4 Landers, California, earthquake, and demonstrate that Landers triggering did not reflect some peculiarity of the region or the earthquake. However, the rate increases triggered by the Denali earthquake occurred in areas not obviously tectonically active, implying that even in areas of low ambient stressing rates, faults may still be critically stressed and that dynamic triggering may be ubiquitous and unpredictable.

Breathing patterns during breathing exercises in persons with tetraplegia.

STUDY DESIGN: Cross-sectional, observational, controlled study. OBJECTIVES: To survey breathing patterns during breathing at rest, ordinary deep breathing (DB), positive expiratory pressure (PEP) and inspiratory resistance-positive expiratory pressure (IR-PEP) among individuals with a cervical spinal cord lesion (SCL) compared with able-bodied controls. SETTING: Sahlgrenska University Hospital, Göteborg, Sweden. METHOD: Participants consisted of 20 persons with a complete SCL at the C5-C8 level (at least 1 year postinjury) and 20 matched, able-bodied controls. Breathing patterns and static lung volumes were measured using a body plethysmograph. RESULTS: Compared to the controls, breathing patterns at rest among the people with tetraplegia were characterised by a decreased tidal volume, stable respiratory rate and total cycle duration resulting in decreased mean inspiratory and expiratory flow, and alveolar ventilation. All volume and flow parameters increased except respiratory rate, which decreased during DB and PEP. During IR-PEP, tidal volume increased less compared to PEP, and combined with a decreased respiratory rate the alveolar ventilation was lower than during breathing at rest. The functional residual capacity increased during PEP and IR-PEP in people with tetraplegia. CONCLUSION: DB exercises with or without resistance during expiration or the whole breathing cycle affect the breathing pattern in persons with tetraplegia. DB was superior in increasing volumes and flow. PEP and IR-PEP increased FRC but IR-PEP decreased volumes and flows. However, large interindividual differences in the SCL group indicate the need for caution in generalising the results. SPONSORSHIP: This work was supported in part by grants from the Memorial Foundation of the Swedish Association of registered Physiotherapists and the Association of Cancer and Road Accident Victims.

P2X4 and P2X6 receptors associate with VE-cadherin in human endothelial cells.

We investigated the expression of P2X4 and P2X6 receptors on human umbilical vein endothelial cells (HUVECs) and found that both P2X receptor subtypes on plasma membranes are largely restricted to areas of cell-cell contact. Co-labelling experiments at the confocal and electron microscopy levels revealed that P2X4 and P2X6 receptors are strongly co-localised with the cell adhesion molecule VE-cadherin. The P2X4 and P2X6 receptors on plasma membranes at cellular junctions are rapidly (within 5 min) internalised specifically after decreasing extracellular [Ca2+]. Disruption of microfilaments, microtubules and integrin-mediated adhesion or stimulation of P2 receptors with ATP did not alter P2X4 and P2X6 receptor expression on HUVEC plasma membranes. Membraneous P2X4 and P2X6 receptors resisted extraction with Triton-X 100, whereas cytoplasmic P2X receptors were Triton-X 100 soluble. P2X4 receptors, but not P2X6 receptors, could be co-immunoprecipitated with VE-cadherin and vice versa. We conclude that P2X4 and P2X6 receptors are associated with VE-cadherin at HUVEC adherens junctions.

ATP is released from guinea pig ureter epithelium on distension.

Distension of the perfused guinea pig ureter at pressures from 20 to 700 cmH(2)O increased the amount of ATP released from the epithelium in a pressure-dependent manner. During basal perfusion (40 microl/min), the perfusate contained 10 pmol/ml ATP; this increased 10- to 50-fold at various distending pressures. ATP was released from epithelial cells during distension as mechanical removal of the urothelium blocked release. No lactate dehydrogenase was detected in the perfusate, and scanning electron microscopy confirmed an intact urothelium after distension. ATP was not released due to the activation of stretch-activated channels, as gadolinium (10 microM) failed to affect ATP release. Glibenclamide (10 microM), known to inhibit two members of the ATP-binding cassette (ABC) protein family, did not affect ATP release after distension; nor did verapamil (10 microM). In contrast, both monensin (100 microM) and brefeldin A (10 microM), which interfere with vesicular formation or trafficking, inhibited distension-evoked ATP release, which was Ca(2+)-dependent. This suggests that ATP release from the ureter epithelium might be mediated by vesicular exocytosis. The role of ATP released by distension of hollow visceral organs is discussed in relation to the concept of purinergic mechanosensory transductions, with special reference to nociception and the activation of P2X(3) receptors on the subepithelial sensory nerves.

Purinergic signalling: ATP release.

Adenosine triphosphate (ATP) has a fundamental intracellular role as the universal source of energy for all living cells. The demonstration of its release into the extracellular space and the identification and localisation of specific receptors on target cells have been essential in establishing, after considerable resistance, its extracellular physiological roles. It is now generally accepted that ATP is a genuine neurotransmitter both in the central and peripheral nervous systems. As such, there are numerous arguments which prove that the release of ATP by nerve terminals is by exocytosis. In some non-neuronal cells, however, recent evidence suggests that ATP release could also be carrier-mediated and would involve ATP-binding cassette proteins (ABC), an ubiquitous family of transport ATPases.

Evidence that release of adenosine triphosphate from endothelial cells during increased shear stress is vesicular.

In response to increased shear stress, vascular endothelial cells release adenosine triphosphate (ATP) by an unknown mechanism. We have investigated this mechanism using different approaches. First, we discovered that quinacrine, used to locate intracellular stores of ATP bound to peptides, displayed a granular fluorescence, typical of vesicular storage. Second, we found that two inhibitors of vesicular transport (monensin and N-ethylmaleimide) produced a highly significant reduction in the release of ATP from vascular endothelial cells in response to increased shear stress. Preliminary experiments using inhibitors of the cystic fibrosis transmembrane regulator, the sulfonylurea receptor, and the multidrug resistance protein showed no involvement of these ATP-binding cassette transporter proteins (previously characterized in endothelial cells) in the mechanism of release of ATP. We suggest, therefore, that the release of ATP from vascular endothelial cells, like that of nerve cells, is probably by vesicular exocytosis.

P2X3 knock-out mice reveal a major sensory role for urothelially released ATP.

The present study explores the possible involvement of a purinergic mechanism in mechanosensory transduction in the bladder using P2X(3) receptor knock-out (P2X(3)-/-) and wild-type control (P2X(3)+/+) mice. Immunohistochemistry revealed abundant nerve fibers in a suburothelial plexus in the mouse bladder that are immunoreactive to anti-P2X(3). P2X(3)-positive staining was completely absent in the subepithelial plexus of the P2X(3)-/- mice, whereas staining for calcitonin gene-related peptide and vanilloid receptor 1 receptors remained. Using a novel superfused mouse bladder-pelvic nerve preparation, we detected a release of ATP proportional to the extent of bladder distension in both P2X(3)+/+ and P2X(3)-/- mice, although P2X(3)-/- bladder had an increased capacity compared with that of the P2X(3)+/+ bladder. The activity of multifiber pelvic nerve afferents increased progressively during gradual bladder distension (at a rate of 0.1 ml/min). However, the bladder afferents from P2X(3)-/- mice showed an attenuated response to bladder distension. Mouse bladder afferents of P2X(3)+/+, but not P2X(3)-/-, were rapidly activated by intravesical injections of P2X agonists (ATP or alpha,beta-methylene ATP) and subsequently showed an augmented response to bladder distension. By contrast, P2X antagonists [2',3'-O-(2,4,6-trinitrophenyl)-ATP and pyridoxal 5-phosphate 6-azophenyl-2',4'-disulfonic acid] and capsaicin attenuated distension-induced discharges in bladder afferents. These data strongly suggest a major sensory role for urothelially released ATP acting via P2X(3) receptors on a subpopulation of pelvic afferent fibers.

Earthquake triggering by seismic waves following the Landers and Hector Mine earthquakes.

The proximity and similarity of the 1992, magnitude 7.3 Landers and 1999, magnitude 7.1 Hector Mine earthquakes in California permit testing of earthquake triggering hypotheses not previously possible. The Hector Mine earthquake confirmed inferences that transient, oscillatory 'dynamic' deformations radiated as seismic waves can trigger seismicity rate increases, as proposed for the Landers earthquake. Here we quantify the spatial and temporal patterns of the seismicity rate changes. The seismicity rate increase was to the north for the Landers earthquake and primarily to the south for the Hector Mine earthquake. We suggest that rupture directivity results in elevated dynamic deformations north and south of the Landers and Hector Mine faults, respectively, as evident in the asymmetry of the recorded seismic velocity fields. Both dynamic and static stress changes seem important for triggering in the near field with dynamic stress changes dominating at greater distances. Peak seismic velocities recorded for each earthquake suggest the existence of, and place bounds on, dynamic triggering thresholds. These thresholds vary from a few tenths to a few MPa in most places, depend on local conditions, and exceed inferred static thresholds by more than an order of magnitude. At some sites, the onset of triggering was delayed until after the dynamic deformations subsided. Physical mechanisms consistent with all these observations may be similar to those that give rise to liquefaction or cyclic fatigue.

Triggering of earthquake aftershocks by dynamic stresses.

It is thought that small 'static' stress changes due to permanent fault displacement can alter the likelihood of, or trigger, earthquakes on nearby faults. Many studies of triggering in the near-field, particularly of aftershocks, rely on these static changes as the triggering agent and consider them only in terms of equivalent changes in the applied load on the fault. Here we report a comparison of the aftershock pattern of the moment magnitude Mw = 7.3 Landers earthquake, not only with static stress changes but also with transient, oscillatory stress changes transmitted as seismic waves (that is, 'dynamic' stresses). Dynamic stresses do not permanently change the applied load and thus can trigger earthquakes only by altering the mechanical state or properties of the fault zone. These dynamically weakened faults may fail after the seismic waves have passed by, and might even cause earthquakes that would not otherwise have occurred. We find similar asymmetries in the aftershock and dynamic stress patterns, the latter being due to rupture propagation, whereas the static stress changes lack this asymmetry. Previous studies have shown that dynamic stresses can promote failure at remote distances, but here we show that they can also do so nearby.

Sitting position - posture and performance in C5 - C6 tetraplegia.

OBJECTIVES: To investigate how sitting position and seating affect posture and performance (balance, transfers, wheelchair skills, physical strain during wheelchair propulsion, spasticity and respiration) in persons with C5 and C6 tetraplegia. SETTING: Outpatient clinic 'Spinalhälsan', Göteborg, Sweden. METHOD: Baseline measurements of sitting position and performance were performed followed by an intervention period. The intervention was individually adapted to each person with emphasis on reduction of kyphotic posture and pelvic obliquity. Furthermore, a functional requirement was that the new sitting position was used in everyday life and did not impair balance, transfers, wheelchair skills, physical strain during wheelchair propulsion, spasticity and respiration. RESULTS: Four persons with complete C5 - C6 tetraplegia who reported dissatisfaction with posture and seating took part in the study. A comparison of photographs before and after the intervention showed a reduction of kyphotic posture and pelvic obliquity. Balance, transfers, wheelchair skills, physical strain during wheelchair propulsion, spasticity and respiration were affected by the sitting position in an individual manner. CONCLUSION: Solution of problems concerning sitting and posture for persons with C5 - C6 tetraplegia requires good knowledge of the physical impairment, wheelchair adaptation, seating systems and cushions as well as an understanding of the individual's demands and wishes. Due to the complexity of the issue, standard solutions are not applicable. Thus, an analytical working method is required and co-operation between professionals - occupational therapists and physiotherapists - is important.

Effect of shear stress on the release of soluble ecto-enzymes ATPase and 5'-nucleotidase along with endogenous ATP from vascular endothelial cells.

Stimulation of endothelial cells from human umbilical vein by shear stress induced release of endogenous ATP which was accompanied by an extracellular increase in the activity of enzymes degrading both ATP (ATPases) and AMP (5'-nucleotidases). The activity of soluble ATPase was progressively increased from 1.62+/-0.27 to 12.7+/-1.0 pmoles ml(-1) h(-1) after 60 min of stimulation by shear stress. The rate of [(3)H]-ATP hydrolysis in the medium was inhibited by the purinergic agents suramin, Reactive blue 2 and pyridoxalphosphate-6-azophenyl-2'4'-disulphonic acid, and remained insensitive to the classic inhibitors of ion-pumping and intracellular ATPases. Shear stress also increased the activity of 5'-nucleotidase in the medium from 2.0+/-0.5 to 27.2+/-2.8 pmoles ml(-1) h(-1). When shear stress was applied after removal of ecto-5'-nucleotidase by phosphatidylinositol-specific phospholipase C, the release of 5'-nucleotidase was drastically reduced. These results show that soluble ATPase and 5'-nucleotidase which are released during shear stress are not released from an intracellular compartment together with ATP but have an extracellular origin.

Free radical involvement in endothelium-dependent responses of the rat thoracic aorta in moderate hypoxic conditions.

This study investigates the effects of agents which act on the production or efficacy of free radicals on the hypoxic responses of rat aorta rings. Under moderate hypoxic conditions, the resting tension of the rings was not changed but in rings precontracted with 5-hydroxytryptamine, there was a relaxation followed by a contraction. Removal of the endothelium with saponin suppressed relaxation to acetylcholine and abolished the contractions produced by hypoxia. In rings with a functional endothelium, hypoxic vasoconstriction was strongly inhibited by mannitol and exifone, but was not reduced by N(G)-nitro-L-arginine methyl ester, superoxide dismutase + catalase, or deferoxamine. Hypoxic vasodilatation was only partially inhibited by mannitol. To conclude, hypoxic constriction of the rat thoracic aorta is largely endothelium-dependent and involves free radicals whereas hypoxic dilatation is partially endothelium-dependent and partially involves free radicals. There is also indirect evidence for lack of direct involvement of nitric oxide/endothelium-derived relaxing factor (NO*/EDRF), hydroxyl radical (OH*) and superoxide anion in the hypoxic constriction and relaxation of the rat aorta.

Lack of uptake, release and action of UTP at sympathetic perivascular nerve terminals in rabbit ear artery.

A possible role of uridine 5'-triphosphate (UTP) and uridine at sympathetic nerve terminals was studied in the rabbit ear artery after incubation of isolated vessels with [3H]uridine or [3H]noradrenaline. It was found that [3H]uridine was taken up by rabbit ear artery. This uptake was largely suppressed after the removal of endothelium and was inhibited by ethidium bromide and dipyridamole. Chemical denervation of the vessels with 6-hydroxydopamine did not reduce the uptake. Following pre-incubation of the isolated vessels with [3H]uridine, there was a release of radioactivity from the superfused rabbit ear artery. UTP, UDP, UMP and uridine were detected by thin layer chromatography both in the superfusate and inside the vessels. Transmural electric stimulation (30 V, 5 Hz) induced a contraction of the vessels but did not increase the release of uridine nucleotides into the superfusate. [3H]Noradrenaline was released during electric stimulation and the addition of UTP (100 microM) had no effects on this release. To conclude, this study shows that in contrast to endothelial cells, the sympathetic nerve terminals of the rabbit ear artery do not take up uridine and do not release uridine-derived nucleotides. UTP at 100 microM is also unable to modulate the evoked release of noradrenaline. These results mainly confine the role of UTP in endothelium-derived vasodilatation via P2Y2 and/or P2Y4 receptors.

Increased release of ATP from endothelial cells during acute inflammation.

OBJECTIVE AND DESIGN: The effects of lipopolysaccharide (LPS), a potent inflammatory mediator, on the shear stress stimulated release of adenosine triphosphate (ATP) were investigated on endothelial cells from human umbilical vein in primary culture. METHODS: Human umbilical vein endothelial cells (HUVEC) in primary cultures were subjected to shear stress using a cone and plate apparatus. ATP released by the cells was measured by luminometry, using a luciferin-luciferase assay. RESULTS: Under conditions of shear stress alone (25dyn/cm2), ATP accumulates into the culture medium and reaches a maximum after 3 to 5 min of stimulation (121.7+/-13.2 pmol/ml). The shear stress-stimulated release of ATP was significantly increased after a 4 h pre-incubation of endothelial cells with 50 microg/ml (314.4+/-26.7 pmol/ml) and 10microg/ml lipopolysaccharide (207.7+/-22.2 pmol/ml). Dexamethasone, an anti-inflammatory glucocorticoid, inhibited the effects of lipopolysaccharide. CONCLUSIONS: These results show that non-damaged endothelial cells release ATP under experimental inflammatory conditions and support an early role of extracellular ATP in the inflammatory process.

Free radical generators cause changes in endothelial and inducible nitric oxide synthases and endothelin-1 immunoreactivity in endothelial cells from hyperlipidemic rabbits.

Reactive oxygen species (ROS) play an important role in the damage of vascular endothelium during atherogenesis and impaired endothelium-dependent vasorelaxation. We have studied the effect of two ROS generators (H2O2 and menadione) and one of the most potent antioxidants (morin) on the double immunofluorescent staining of endothelial cells (EC) from both Watanabe Heritable Hyperlipidemic (WHHL) and New Zealand White (NZW) rabbits in primary cultures using antibodies against endothelin-1 (ET-1), endothelial (eNOS), and inducible nitric oxide synthase (iNOS). In aortic EC from normal rabbits, ROS decreased the immunoreactivity of eNOS and ET-1 and this effect was significantly reversed by morin. In atherosclerotic rabbits, ROS had the same effect on the immunoreactivity of eNOS and ET-1 but also induced the expression of iNOS immunoreactivity. In general, the cells from WHHL rabbits were less sensitive to the protective effects of morin and more sensitive to the effects of ROS. It thus appears that the protective effect of morin may be due to neutralization of ROS and may be considered for the treatment of early stages of atherosclerosis, before macroscopic lesions have occurred.

ATP-stimulated release of ATP by human endothelial cells.

We investigated the effects of several concentrations of extracellular ATP on the release of intracellular ATP by human umbilical vein endothelial cells (HUVEC) in primary cultures. When ATP is added to the medium of cultured EC at a concentration of 1 microM, it is readily degraded by extracellular enzymes; 10 microM ATP added to the culture medium provokes a transient but significant increase, followed by a decrease in the concentration of extracellular ATP. At a concentration of 100 microM, there was a significant release of ATP and its level was maintained in the culture medium throughout the experiment. Our results show that extracellular ATP leads to a sustained release of intracellular ATP by HUVEC. Such sustained self-perpetuating release of ATP is likely to play an important part in physiological and pathological local vascular control mechanisms.