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Biomedical subjects

R Cross

Publications and source records attributed to R Cross.

At least 19 recordsLinked to original sources
PubMed2003

Player sensitivity to changes in string tension in a tennis racket.

Forty-one advanced recreational tennis players were tested to determine their ability to detect differences in string tension in a tennis racket. Subjects were given pairs of rackets that varied in tension by up to 98 N (10 kg) and were asked whether they noticed a difference in tension and if so, which racket was strung at a higher tension. Only 11 (27%) of those tested could correctly identify a tension difference of 5 kg (11 lb) or less. Fifteen (37%) could not pick a difference of 10 kg (22 lb). To examine the importance of sound as a means of discrimination, an additional test was undertaken where participants wore earplugs. Of the 26 subjects undertaking this additional test, only 6 (23%) were successful. It was concluded that advanced recreational tennis players demonstrated limited ability to correctly identify differences in string tension and that impact sound was an important factor for those participants who were successful at various levels of discrimination.

Adult↗
PubMed2001

Unusual properties of the fungal conventional kinesin neck domain from Neurospora crassa.

Fungal conventional kinesins are unusually fast microtubule motor proteins. To compare the functional organization of fungal and animal conventional kinesins, a set of C-terminal deletion mutants of the Neurospora crassa conventional kinesin, NcKin, was investigated for its biochemical and biophysical properties. While the shortest, monomeric construct comprising the catalytic core and the neck-linker (NcKin343) displays very high steady-state ATPase (k(cat) = 260/s), constructs including both the full neck and adjacent hinge domains (NcKin400, NcKin433 and NcKin480) show wild-type behaviour: they are dimeric, show fast gliding and slower ATP turnover rates (k(cat) = 60-84/s), and are chemically processive. Unexpectedly, a construct (NcKin378, corresponding to Drosophila KHC381) that includes just the entire coiled-coil neck is a monomer. Its ATPase activity is slow (k(cat) = 27/s), and chemical processivity is abolished. Together with a structural analysis of synthetic neck peptides, our data demonstrate that the NcKin neck domain behaves differently from that of animal conventional kinesins and may be tuned to drive fast, processive motility.

Adenosine Diphosphate↗
PubMed2001

KIF1D is a fast non-processive kinesin that demonstrates novel K-loop-dependent mechanochemistry.

The KIF1 subfamily members are monomeric and contain a number of amino acid inserts in surface loops. A particularly striking insertion of several lysine/arginine residues occurs in L12 and is called the K-loop. Two recent studies have employed both kinetic and single-molecule methods to investigate KIF1 motor properties and have produced very different conclusions about how these motors generate motility. Here we show that a hitherto unstudied member of this group, KIF1D, is not chemically processive and drives fast motility despite demonstrating a slow ATPase. The K-loop of KIF1D was analysed by deletion and insertion mutagenesis coupled with characterization by steady state and transient kinetics. Together, the results indicate that the K-loop not only increases the affinity of the motor for the MT, but crucially also inhibits its subsequent isomerization from weak to strong binding, with coupled ADP release. By stabilizing the weak binding, the K-loop establishes a pool of motors primed to undergo their power stroke.

Adenosine Diphosphate↗
PubMed2001

Concurrent naive and memory CD8(+) T cell responses to an influenza A virus.

Memory Thy-1(+)CD8(+) T cells specific for the influenza A virus nucleoprotein (NP(366-374)) peptide were sorted after staining with the D(b)NP(366) tetramer, labeled with CFSE, and transferred into normal Thy-1.2(+) recipients. The donor D(b)NP(366)(+) T cells recovered 2 days later from the spleens of the Thy-1.2(+) hosts showed the CD62L(low)CD44(high)CD69(low) phenotype, characteristic of the population analyzed before transfer, and were present at frequencies equivalent to those detected previously in mice primed once by a single exposure to an influenza A virus. Analysis of CFSE-staining profiles established that resting tetramer(+) T cells divided slowly over the next 30 days, while the numbers in the spleen decreased about 3-fold. Intranasal infection shortly after cell transfer with a noncross-reactive influenza B virus induced some of the donor D(b)NP(366)(+) T cells to cycle, but there was no increase in the total number of transferred cells. By contrast, comparable challenge with an influenza A virus caused substantial clonal expansion, and loss of the CFSE label. Unexpectedly, the recruitment of naive Thy-1.2(+)CD8(+)D(b)NP(366)(+) host D(b)NP(366)(+) T cells following influenza A challenge was not obviously diminished by the presence of the memory Thy-1.1(+)CD8(+)D(b)NP(366)(+) donor D(b)NP(366)(+) set. Furthermore, the splenic response to an epitope (D(b)PA(224)) derived from the influenza acid polymerase (PA(224-233)) was significantly enhanced in the mice given the donor D(b)NP(366)(+) memory population. These experiments indicate that an apparent recall response may be comprised of both naive and memory CD8(+) T cells.

Animals↗
PubMed2001

Soman-induced seizures: limbic activity, oxidative stress and neuroprotective proteins.

Soman, a potent acetylcholinesterase inhibitor, induces status epilepticus in rats followed by conspicuous neuropathology, most prominent in piriform cortex and the CA3 region of the hippocampus. Cholinergic seizures originate in striatal-nigral pathways and with fast-acting agents (soman) rapidly spread to limbic related areas and finally culminate in a full-blown status epilepticus. This leads to neurochemical changes, some of which may be neuroprotective whereas others may cause brain damage. Pretreatment with lithium sensitizes the brain to cholinergic seizures. Likewise, other agents that increase limbic hyperactivity may sensitize the brain to cholinergic agents. The hyperactivity associated with the seizure state leads to an increase in intracellular calcium, cellular edema and metal delocalization producing an oxidative stress. These changes induce the synthesis of stress-related proteins such as heat shock proteins, metallothioneins and heme oxygenases. We show that soman-induced seizures cause a depletion in tissue glutathione and an increase in tissue 'catalytic' iron, metallothioneins and heme oxygenase-1. The oxidative stress induces the synthesis of stress-related proteins, which are indicators of 'stress' and possibly provide neuroprotection. These findings suggest that delocalization of iron may catalyze Fenton-like reactions, causing progressive cellular damage via free radical products.

Animals↗
PubMed2001

Enzymatic removal of nitric oxide catalyzed by cytochrome c' in Rhodobacter capsulatus.

Cytochrome c' from Rhodobacter capsulatus has been shown to confer resistance to nitric oxide (NO). In this study, we demonstrated that the amount of cytochrome c' synthesized for buffering of NO is insufficient to account for the resistance to NO but that the cytochrome-dependent resistance mechanism involves the catalytic breakdown of NO, under aerobic and anaerobic conditions. Even under aerobic conditions, the NO removal is independent of molecular oxygen, suggesting cytochrome c' is a NO reductase. Indeed, we have measured the product of NO breakdown to be nitrous oxide (N(2)O), thus showing that cytochrome c' is behaving as a NO reductase. The increased resistance to NO conferred by cytochrome c' is distinct from the NO reductase pathway that is involved in denitrification. Cytochrome c' is not required for denitrification, but it has a role in the removal of externally supplied NO. Cytochrome c' synthesis occurs aerobically and anaerobically but is partly repressed under denitrifying growth conditions when other NO removal systems are operative. The inhibition of respiratory oxidase activity of R. capsulatus by NO suggests that one role for cytochrome c' is to maintain oxidase activity when both NO and O(2) are present.

Cytochrome c Group↗
PubMed2000

Contact dynamics during keratocyte motility.

BACKGROUND: Keratocytes are specialised, rapidly moving cells that generate substantial contractile force perpendicular to their direction of locomotion. Potential roles for contractile force in cell motility include cell-body transport, regulation of adhesion, and retraction of the cell's trailing edge. RESULTS: To investigate contact dynamics, we used simultaneous confocal fluorescence and interference reflection microscopy to image keratocytes injected with fluorescent vinculin. We found that contacts formed behind the leading edge and grew beneath both the lamellipodium and the cell body. Contacts in the middle of the cell remained stationary relative to the substrate and began to disassemble as the cell body passed over them. In contrast, contacts in the lobes of the cell grew continuously and more rapidly, incorporated more vinculin, and slid inwards towards the sides of the cell body. Contact sliding often led to merging of contacts before their removal from the substrate. CONCLUSIONS: We suggest a synthesis of two existing, apparently conflicting models for keratocyte motility, in which network contraction progressively reorients actin filaments using the contacts as pivots, forming bundles that then generate lateral tension by a sliding-filament mechanism. Contact dynamics vary between the middle of the cell and the lobes. We propose that laterally opposed contractile forces first enhance contact growth and stability, but escalating force eventually pulls contacts from the substrate at the back of the cell, without interfering with the cell's forward progress.

Actins↗
PubMed2000

Protein profiled features patterned via confocal microscopy.

Protein patterns were printed using conventional microlithographic materials in a bilayer arrangement and unconventional exposure tools. The bilayer resist stack consisted of a lower poly(tert-butyl methacrylate) layer and an upper diazonaphtoquinone/novolak layer. The protein features were printed in either 'contact printing', or 'step and repeat' mode. The latter printing mode can be managed in a flow-cell consisting of a standard microscope slide and cover slip, spaced apart by about 20 microm, as follows: (i) the exposure step is carried out in the cell using focused 488 nm beam of a confocal laser scanning microscope; (ii) the development step is performed by flowing the photoresist developer through the cell; (iii) the selective deposition of the protein (FITC-labelled avidin) is achieved via the flow of the protein solution through the cell until a desired contrast has been reached; (iv) the control of the process is assured using on-line monitoring of the photo-activated red fluorescence of the developing resist layer, and of the green fluorescence of the FITC-protein patterns, respectively. The protein printing technique uses equipment routinely available in biological laboratory. The 'step and repeat' patterning yields high and controllable resolution. The process can be applied in the fabrication of medical microanalysis devices.

Biosensing Techniques↗
PubMed2000

Advanced life support: retention of registered nurses' knowledge 18 months after initial training.

In 1996, The Wesley Hospital introduced a 2 day Advanced Life Support (ALS) course, targeted at all critical care registered nurses and medical officers. The purpose of this study was to explore the retention of theoretical knowledge and clinical skills of registered nurses who had successfully completed the 2 day ALS course 18 months previously and to establish effective retesting timeframes. The study utilised a repeated post-test measure design. Forty registered nurses participated in the study. Data were collected during ALS retesting using scores from a theoretical examination and from the results of four practical skill assessments (basic life support, airway management, defibrillation and code management). Using Wilcoxon test, data were analysed with and compared to the participant's original scores from the training program 18 months previously. The findings demonstrate that the participant's theoretical knowledge remained at an equivalent level over the 18 month timeframe. However, 18 months after successfully completing an ALS course, only 75 per cent (n = 30) of participants passed the practical skill assessment components, with the 25 per cent (n = 10) requiring a second attempt to pass. The implications from this study focus on the model of assessment utilised and the dichotomy between theoretical and practical skill assessment results. Additional study is required to determine the optimal timeframe for ALS retesting and educational strategies to help retain skills over time.

Adult↗
PubMed2000

Cytochrome c' from Rhodobacter capsulatus confers increased resistance to nitric oxide.

We report the cloning and sequencing of the gene containing cytochrome c' (cycP) from the photosynthetic purple bacterium Rhodobacter capsulatus and the regions flanking that gene. Mutant strains unable to synthesize cytochrome c' had increased sensitivity to nitrosothiols and to nitric oxide (which binds to the heme moiety of cytochrome c').

Cell Division↗
PubMed1999

Coupled chemical and mechanical reaction steps in a processive Neurospora kinesin.

We show using single molecule optical trapping and transient kinetics that the unusually fast Neurospora kinesin is mechanically processive, and we investigate the coupling between ATP turnover and the mechanical actions of the motor. Beads carrying single two-headed Neurospora kinesin molecules move in discrete 8 nm steps, and stall at approximately 5 pN of retroactive force. Using microtubule-activated release of the fluorescent analogue 2'-(3')-O-(N-methylanthraniloyl) adenosine 5'-diphosphate (mantADP) to report microtubule binding, we found that initially only one of the two motor heads binds, and that the binding of the other requires a nucleotide 'chase'. mantADP was released from the second head at 4 s(-1) by an ADP chase, 5 s(-1) by 5'-adenylylimidodiphosphate (AMPPNP), 27 s(-1) by ATPgammaS and 60 s(-1) by ATP. We infer a coordination mechanism for molecular walking, in which ATP hydrolysis on the trailing head accelerates leading head binding at least 15-fold, and leading head binding then accelerates trailing head unbinding at least 6-fold.

Adenosine Diphosphate↗
PubMed1999

Comparison of two 3-day Helicobacter pylori eradication regimens with a standard 1-week regimen.

BACKGROUND: The duration of Helicobacter pylori eradication regimens has decreased to 1 week with cure rates of over 90%. This can be attributed to the use of triple drug regimens including potent inhibitors of gastric acid secretion and clarithromycin. There is no theoretical reason why shorter regimens should not be possible. AIM: To compare two 3-day, low-dose, twice daily regimens with 1 week of omeprazole 20 mg b.d., clarithromycin 250 mg b.d., and metronidazole 400 mg b.d. (OCM) METHODS: Outpatients referred for gastroscopy were screened by biopsy urease test. H. pylori-positive patients were randomized to receive either lansoprazole 30 mg b.d., tri-potassium dicitrato bismuthate one tablet b.d., clarithromycin 250 mg b.d., and amoxycillin 1 g b.d. for 3 days (LTdbCA), or ranitidine bismuth citrate 400 mg b.d., clarithromycin 250 mg b.d. and amoxycillin 1 g b.d. for 3 days (RbcCA) or omeprazole 20 mg b.d., clarithromycin 250 mg b.d. and metronidazole 400 mg b.d. for 1 week (OCM). They were not pre-treated with a gastric acid inhibitor. After 8 weeks, H. pylori status was assessed by 13C urea breath test. RESULTS: 974 out of 1114 patients referred for gastroscopy were screened by biopsy urease test. 140 patients were not screened either because they were anticoagulated or for technical reasons. 334 patients were H. pylori-positive: 154 were excluded mostly because of allergy to penicillin and personal reasons but 180 were randomized to treatment All regimens were well tolerated. For LTdbCA (n=60), RbcCA (n=59), and OCM (n=61) the H. pylori cure rates (95% CI) were 23% (12-34), 14% (5-23) and 87% (79-95), respectively, using intention-to-treat analysis and 25% (14-36), 15% (6-24) and 88% (80-96), respectively, if analysed per protocol. OCM was significantly superior to LTdbCA and RbcCA (P < 0.001) but there was no significant difference between regimens LTdbCA and RbcCA. CONCLUSIONS: OCM is an extremely effective H. pylori eradication regimen. The 3-day regimens tested both have poor cure rates. Pre-treatment with a proton pump inhibitor, higher doses or more frequent dosing may be necessary to increase the cure rate of short duration regimens. However, this could make them less acceptable than the H. pylori eradication regimens currently available.

2-Pyridinylmethylsulfinylbenzimidazoles↗
PubMed1998

Lymphocyte subsets in cord blood of preterm infants: effect of antenatal steroids.

The objective of this study was to evaluate prospectively the influence of gestational age (GA) and short-term antenatal steroids on total lymphocyte count and lymphocyte subsets in cord blood from preterm infants. Two-color flow cytometric analyses of lymphocyte subsets were performed on cord blood collected from 67 infants. These infants were grouped according to GA: group I (term, n = 19); group II (GA 33-37 weeks, n = 25); group III (GA <33 weeks, n = 23). The mean absolute lymphocyte counts (ALC) in groups I, II and III were 5.6 +/- 2.5 x 10(3)/ microl, 4.3 +/- 1.5 x 10(3)/ microl and 3. 5 +/- 1.8 x 10(3)/ microl respectively. The mean values for CD4+ lymphocytes in groups I, II and III were 2.7 +/- 0.8 x 10(3)/ microl, 2.0 +/- 0.8 x 10(3)/ microl and 1.6 +/- 0.9 x 10(3)/ microl respectively. Mean values for CD8+ lymphocytes were 0.9 +/- 0.3 x 10(3)/ microl, 0.6 +/- 0.3 x 10(3)/ microl and 0.5 +/- 0.3 x 10(3)/ microl respectively. With decreasing GA, there was a statistically significant decrease in ALC (p = 0.0035), CD4+ lymphocytes (p = 0. 0013) and CD8+ lymphocytes (p = 0.0064). We then evaluated the effect of antenatal steroids, now routinely administered to women with preterm onset of labor to facilitate fetal lung maturation, and found that after adjusting for GA, infants of women on antenatal steroids had significantly fewer ALC (p = 0.0001), CD4+ lymphocytes (p = 0.02) and CD25+ lymphocytes (p = 0.03). In this population of infants, the decreased number of lymphocytes seen at younger GAs is associated with antenatal steroid use.

Betamethasone↗
PubMed1998

Recovery of metabolic activity in retinofugal targets after traumatic optic nerve injury is independent of retinofugal input.

Traumatic injury of the adult optic nerve causes a progressive degeneration of retinal ganglion cells. Despite this ongoing degeneration, a partial recovery of visual behavioral function and of local cerebral glucose use (LCGU) has been observed. To evaluate whether this partial recovery of LCGU is due to a recovery of visual conductance (extrinsic) or intrinsic neuronal activity, visual stimulation alone and combined with physostigmine,an acetylcholinesterase inhibitor, were used to activate the retinofugal pathway. LCGU was determined in 30 male adult rats with or without physostigmine treatment 2 or 9 days after crush or 8 days after cut of the right optic nerve. Analysis of LCGU in contralateral first-order projection areas revealed no differences 8 days after cut and 9 days after optic nerve crush. Furthermore, LCGU in the contralateral areas could not be stimulated by the treatment with physostigmine. We therefore conclude that the increase in LCGU from 2 to 9 days after crush is not due to a recovery in the conductance of visual input. We hypothesize a relief of an injury-dependent active suppression (diaschisis) of LCGU. This reversal of diaschisis may, in part, account for the return of visual functions after mild optic nerve injury.

Journal Article↗
PubMed1997

Apoptosis occurs in endothelial cells during hypertension-induced microvascular rarefaction.

Disappearance of microvessels (microvascular rarefaction) during hypertension is a process that exacerbates the hypertensive condition. The cellular process by which the vessels disappear is not known. In the present study, we investigate the pathogenic role of cell death, specifically apoptosis, in hypertension-induced microvascular rarefaction. An established rodent one kidney/one clip (1K1C) Goldblatt model of hypertension was used. Histological and ultrastructural characteristics of apoptosis and necrosis were used to define incidence of the two types of cell death. The new method of in situ end-labeling DNA fragmentation known to occur in apoptosis was analyzed, and expression of an apoptosis-related gene, clusterin, identified using Northern blots and in situ hybridization. Microvessels in skeletal muscle were compared in 1K1C animals (n = 3 per time point) and control animals (n = 6) at experimental times after surgery up to established hypertension (1, 2, and 4 days and 1, 2, and 6 weeks). Loss of microvessels in hypertensive animals was verified. Endothelial cell apoptosis, not necrosis, was identified and was more frequent in hypertensive animals than in controls. Apoptosis of endothelial cells was found most often within 1 week after 1K1C surgery. Clusterin mRNA transcripts were increased above control levels in all 1K1C treatments, but expression was not localized specifically above endothelial cells. In this instance, increased expression of clusterin in hypertensive animals may be an epiphenomenon, not directly related to the presence of apoptosis. The results demonstrate a role for apoptosis in the development of microvascular rarefaction in hypertension. The significance of this novel finding is that these results may now be used to direct site-specific anti-apoptosis therapy for treatment of structural rarefaction, at present unaffected by conventional anti-hypertensive therapies.

Animals↗