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Biomedical subjects

C J Robinson

Publications and source records attributed to C J Robinson.

At least 37 records · Page 2Linked to original sources
PubMed1997

Murine strain differences in response to mercuric chloride: antinucleolar antibodies production does not correlate with renal immune complex deposition.

Mercuric chloride (HgCl2) induces the production of antinucleolar antibodies (ANucA) in susceptible strains of mice. Responder strains bearing the H-2(5) haplotype as well as several ANucA resistant strains have been shown to develop renal immune complex deposits after HgCl2 treatment. Sera obtained throughout 12 to 16 weeks of HgCl2 treatment from mice of four ANucA responder strains (A.SW/SnJ, A.CA/SnJ, DBA/1J, and P/J) and one ANucA-resistant strain (C57BL/10SnJ) were examined for ANucA production. Terminal sera were also tested for the presence of anti-glomerular basement membrane antibodies, and the kidneys were examined for the deposition of IgG and C3. Only one strain, A.SW, developed significant deposits of IgG in the renal glomeruli, although all four responder strains exhibited similar ANucA induction/production profiles. The differences seen by direct immunofluorescence assay (IFA) in renal immune complex deposition between the A.SW and histocompatibility congenic A.CA mice were corroborated by individually eluting and then quantitating the deposited IgG from renal tissues of Hg-treated A.SW and A.CA mice as well as control A.SW mice. The average amount of IgG eluted from A.SW renal tissue was significantly greater than that eluted from either A.CA or control A.SW renal tissues. All eluates from Hg-treated animals gave only a nucleolar fluorescence pattern when assayed by indirect IFA against a panel of rat organ tissues. In summary, no correlation was found between ANucA production and renal IgG deposition in response to treatment with HgCl2.

Animals↗
PubMed1997

Stress-activated protein kinases: activation, regulation and function.

The response of cells to extracellular stimuli is mediated in part by a number of intracellular kinase and phosphatase enzymes. Within this area of research the activation of the p42 and p44 isoforms of mitogen-activated protein (MAP) kinases have been extensively described and characterised as central components of the signal transduction pathways stimulated by both growth factors and G-protein-coupled receptor agonists. Signaling events mediated by these kinases are fundamental to cellular functions such as proliferation and differentiation. More recently, homologues of the p42 and p44 isoforms of MAP kinase have been described, namely the stress-activated protein kinases (SAPKs) or alternatively the c-jun N-terminal kinases (JNKs) and p38 MAP kinase (the mammalian homologue of yeast HOG1). These MAP kinase homologues are integral components of parallel MAP kinase cascades activated in response to a number of cellular stresses including inflammatory cytokines (e.g., Interleukin-1 (Il-1) and tumour necrosis factor-alpha (TNF-alpha), heat and chemical shock, bacterial endotoxin and ischaemia/cellular ATP depletion. Activation of these MAP kinase homologues mediates the transduction of extracellular signals to the nucleus and are pivotal events in the regulation of the transcription events that determine functional outcome in response to such stresses. In this review we highlight the identification and characterisation of the stress-activated MAP kinase homologues, their role as components of parallel MAP kinase pathways and the regulation of cellular responses following exposure to cellular stress.

Animals↗
PubMed1997

Testing peripheral somatosensory neuroprostheses by recording from raccoon cortex.

A topologically organized representation of the body surface exists within the mammalian somatosensory cortical areas such that stimulation of a part of the body surface will produce a response in a localized region of the contralateral somatosensory cortex. Because of this topography, we propose that the selectivity of a peripheral somatosensory neuroprosthetic electrode array can be tested by noting whether the locus of maximum activation in the cortex moves in a consistent manner when differing portions of the array are stimulated. We further propose that the raccoon might well be the ideal animal in which to test this hypothesis, since the raccoon has a rather unique cortical somatosensory area where each digit is represented in individual subgyri around the tri-radiate sulcus. To demonstrate the feasibility of this concept, a pilot study was carried out in one raccoon under barbiturate anesthesia. The median nerve was stimulated via selective quadrants of a nerve cuff array of four tripolar electrodes implanted around the nerve. Cuff stimulation produced short-latency evoked surface potentials in the digit areas of the raccoon first somatosensory cortex. Response selectivity could be demonstrated, as could a separation between thresholds for producing movement or producing cortical evoked potentials. The sensory and motor responses elicited were consistent with the orientation of the median nerve within the cuff as determined by a postmortem identification of the muscle innervation pattern of the nerve.

Animals↗
PubMed1997

Resistance to mineralocorticoids in Wistar-Furth rats.

Wistar-Furth rats (WF) do not develop hypertension when treated with salt and mineralocorticoids and therefore may be useful for investigating the mechanisms of mineralocorticoid action and hypertension. In the present studies, we determined vascular and renal responses of WF to mineralocorticoids. Control Wistar rats (W) developed deoxycorticosterone acetate (DOCA)-NaCl and dexamethasone hypertension, whereas WF rats developed dexamethasone hypertension only. Aldosterone treatment of vascular smooth muscle cells cultured from WF resulted in 82% less upregulation of angiotensin II radioligand binding, 50% less induction of angiotensin II AT1a receptor mRNA, and 76% less potentiation of angiotensin II-stimulated inositol phosphates than did aldosterone treatment of cells from W. Similarly, DOCA-NaCl potentiated angiotensin II- and phenylephrine-stimulated contractions in aortic rings from W but not from WF. Although DOCA-NaCl treatment affected hypokalemia to an equal degree in WF and W, increases in renal citrate synthase activity (a specific renal mineralocorticoid response) were greater in W than in WF. WF manifest a partial defect in mineralocorticoid responsiveness in vascular smooth muscle and, possibly, in the kidney.

Aldosterone↗
PubMed1996

Treatment of vascular smooth muscle cells with antisense phosphorothioate oligodeoxynucleotides directed against p42 and p44 mitogen-activated protein kinases abolishes DNA synthesis in response to platelet-derived growth factor.

We have investigated the requirement for mitogen-activated protein (MAP) kinase in the stimulation of DNA synthesis by platelet-derived growth factor (PDGF) in rat aortic smooth muscle cells using a phosphorothioate-modified oligodeoxy-nucleotide (ODN) to deplete MAP kinase. Treatment for 72 h with MAP kinase antisense ODN directed against both the p42 and p44 isoforms of MAP kinase abolished the expression of MAP kinase and reduced agonist-stimulated MAP kinase activity by approx. 95%. The scrambled control ODN was without effect, but the sense control ODN slightly enhanced the expression of both isoforms. Abolition of MAP kinase activity by antisense ODN treatment prevented angiotensin II- and PDGF-stimulated activation of p90 ribosomal S6 kinase activity, but did not affect activation of MAP kinase kinase. In addition, antisense ODN pretreatment reduced PDGF-stimulated [3H]thymidine incorporation to < 5% of control, and decreased basal incorporation by approx. 90%. In contrast, basal [3H]thymidine incorporation was enhanced approx. 60% by control sense ODN treatment. These results indicate an obligatory role for MAP kinase in the activation of a number of early events in mitogenesis, including DNA synthesis, in vascular smooth muscle cells.

Animals↗
PubMed1996

Efficacy of agonist-stimulated MEK activation determines the susceptibility of mitogen-activated protein (MAP) kinase to inhibition in rat aortic smooth muscle cells.

In rat aortic smooth muscle cells, platelet-derived growth factor (PDGF) stimulated a sustained activation of mitogen-activated protein kinase (MAP kinase) while the response to angiotensin II (AII) was transient. This was due to a relatively greater initial activation of MAP kinase kinase (MEK) and a correspondingly greater residual MEK activity at later time points. Pretreatment of cells with the novel MEK inhibitor PD 098059 reduced MEK activation at 5 min in response to each agonist by a similar proportion (70%); however, at this time point MAP kinase activation in response to PDGF was only marginally affected while the response to AII was substantially reduced. PD 098059 did, however, reduce PDGF-stimulated MEK activity after 30 min and this correlated with a loss in MAP kinase activity and DNA synthesis. Pretreatment with forskolin also caused a similar pattern of inhibition of agonist-stimulated MEK and MAP kinase activity. Only following protein kinase C down-regulation were both AII- and PDGF-stimulated MAP kinase activation substantially reduced and this correlated with the virtual loss of both MEK and c-Raf-1 activity in response to both agents. The differential inhibition of MAP kinase activation by forskolin was not due to specific activation of A-Raf by PDGF; both PDGF and AII stimulated A-Raf kinase and this activity was strongly inhibited by forskolin. These results suggest that the efficacy of MEK activation determines the duration of MAP kinase activation and the susceptibility of MAP kinase activation to inhibition by different agents. The results also argue against the selective activation of A-Raf by PDGF as a mechanism to explain the differences in the kinetics of MAP kinase activity stimulated by AII and PDGF.

Angiotensin II↗
PubMed1996

The raccoon as an animal model for upper limb neural prosthetics.

The raccoon was evaluated as an animal model for upper limb neural prosthetics. This animal was selected primarily because the functional use of its forelimb mimics in many ways the usage in humans and because of its optimal size and commercial availability. Eight cadaver and fresh specimen forearms were dissected. Important characteristics of the raccoon forearm were: 1) large muscles in the volar forearm, 2) large digits in the paw that appear more similar to humans than to other species such as cat or dog, 3) persistence of two median nerve cords into the forearm, 4) no separation of individual tendons of flexor digitorum superficialis and flexor digitorum profundus in the carpal tunnel, 5) a small thumb digit with little function and 6) a primary origin of flexor policis longus on the proximal ulna with a secondary origin on the radius. Four animals were anesthetized and responses of the forearm and paw to stimulation of the volar forearm muscles with percutaneous electrodes were evaluated. A pair of stimulating electrodes was placed in each of four muscles or muscle groups. Recording electrodes were placed in two muscles which showed the greatest separation of muscle movements to stimulation. Stimulation currents just above threshold produced discrete motion as well as recordable EMG M-waves. Incremental increases in stimulation current produced an increase in M-wave amplitude up to a maximal stimulating current. Torque recordings for pronation, wrist flexion and finger flexion showed graded and selective responses. These results including anatomical descriptions indicate both the limitations of this animal model and its potential use in the development of upper limb neural prosthetics. We conclude that the raccoon model may be superior to other nonprimate animal models such as the cat because of its extensive forearm and paw movements.

Animals↗
PubMed1996

The international standard for epidermal growth factor (EGF): comparison of candidate preparations by in vitro bioassays and immunoassays. National Institute for Biological Standards and Control.

Four preparations of recombinant human sequence epidermal growth factor (EGF), two of the full length 53-amino acid chain, and two of the 52-amino acid chain, lacking the carboxyl-terminal arginine, were evaluated, using a variety of in vitro bioassays and immunoassays, by 12 laboratories in seven countries, for their suitability to serve as the international standard for EGF. The study shows that some assay systems appear to discriminate between the full length and the 52-amino acid forms of EGF and that use of a common standard in place of the various in-house standards leads to a substantial decrease in between-laboratory variances in estimates of potency of EGF samples. On the basis of the results reported here, the World Health Organization (WHO) established one of the preparations of the full-length EGF molecule (coded 91/530) as the first international standard (IS) for EGF, with an assigned unitage of 2000 International Units/ampoule, and one of the preparations of EGF(1-52) (coded 91/550) as the first international reference reagent (IRR) for EGF (1-52) with a nominal mass content of 1.75 microg EGF(1-52)/ampoule. The IS for EGF and the IRR for EGF(1-52) may be obtained by writing to NIBSC, PO Box 1193, Potters Bar, EN6 3QH, UK.

Biological Assay↗
PubMed1995

Circulating human factor IX produced in keratin-promoter transgenic mice: a feasibility study for gene therapy of haemophilia B.

It has previously been suggested that keratinocytes might provide a suitable target cell for delivery of factor IX to the systemic circulation for gene therapy of haemophilia B. Here, an investigation of the use of cellular gene promoters specific for keratinocytes was undertaken to examine whether factor IX could be passed from the epidermis to the systemic circulation. Utilizing two bovine cytokeratin gene promoters, BKIII and BKVI, three lines of transgenic mice were generated with targeted expression of human factor IX in the epidermis. All three transgenic mouse lines secreted epidermally derived human factor IX into the blood system. Most effective factor IX expression (46 ng/ml steady-state levels of circulating human factor IX) was obtained utilizing the BKVI gene promoter, the human homologue of K10, which is expressed exclusively in differentiated keratinocytes, localized distal to the basement membrane. This report demonstrates, for the first time, that human factor IX can be efficiently synthesized and secreted from keratinocytes in situ, and can cross the epidermal basement membrane to reach the systemic circulation. The transgenic mouse model will provide a good in vivo system with which to optimize the efficiency of different keratin gene promoter constructs for delivery of therapeutic gene products to the serum, especially for those promoters, such as K10, which are not effectively expressed in vitro.

Animals↗
PubMed1994

Biomechanical and reflex responses to joint perturbations during electrical stimulation of muscle: instrumentation and measurement techniques.

A test device is developed to measure ankle joint compliance and muscle activity when the ankle is subjected to perturbations in angular position (or torque) from bias positions achieved volitionally or via electrical stimulation. The ankle measurement system uses a pivoting footplate and is operable with the subject sitting or supine. A companion platform for the knee is developed that uses a rotary arm and attached leg brace and is operable with the subject's leg in the horizontal or vertical plane. The knee fixture's pivoting arm can slide to account for the cam-like movement of the knee during rotation. The devices use similar hardware and share common instrumentation and control. Precise torque or position perturbations are delivered by a computer-controlled torque motor to the ankle or knee. Angular displacement, torque, acceleration, knee fixture moment arm and electromyographic data are collected on analogue tape and simultaneously digitised and stored. A special stimulator/recording amplifier permits the recording of electromyographic signals from the stimulated muscle. Experimental data indicate that the ankle and knee devices, operated horizontally, are purely inertial systems. Sample ankle and knee joint responses to perturbations are presented.

Ankle Joint↗
PubMed1994

An in vitro bioassay for nerve growth factor based on 24-hour survival of PC-12 cells.

We have developed an in vitro bioassay for nerve growth factor (NGF) which offers several advantages over currently used alternatives. The assay uses the commercially available PC-12 cell line, and requires only 24 hr incubation with NGF. The cells do not require priming, and are dosed as they are originally seeded in serum-free medium in 96-well cell culture plates coated with collagen. Concentrations between one and 20 ng/ml NGF can be measured, and only standard cell culture equipment and a microplate reader are required.

Animals↗
PubMed1994

The international standard for basic fibroblast growth factor (FGF-2); comparison of candidate preparations by in vitro bioassays and immunoassays.

Three preparations of recombinant basic fibroblast growth factor (bFGF, FGF-2) were evaluated, using a variety of in vitro bioassays and immunoassays, by 11 laboratories in six countries for their suitability to serve as international standards (IS) for bFGF. Two preparations of human sequence bFGF showed broadly similar behavior in different assays, but the relative activity of the third preparation, a modification of the human sequence, showed more variability. All three preparations were predicted to be sufficiently stable to serve as an IS. On the basis of the results reported here, the World Health Organization (WHO) established one of the preparations (coded 90/712) as the international standard for basic fibroblast growth factor, and assigned it a content of 1600 International Units (IU) per ampoule. The IS may be obtained for use in the calibration of local standards by writing to NIBSC, PO Box 1193, Potters Bar, EN6 3QH, UK.

Analysis of Variance↗
PubMed1993

Inhibiting the hyperreflexic bladder with electrical stimulation in a spinal animal model.

Uninhibited bladder contractions are a problem in spinal cord injured patients. Accordingly, methods using electrical stimulation to inhibit the bladder were investigated in chronic spinal cord injured (C6-T1) male cats. In unanesthetized, restrained animals, spontaneous bladder contractions were observed after the bladder was filled above the micturition threshold. In 3 of the 5 cats studied, this bladder activity could be inhibited with stimulation of either sacral nerves or pudendal nerves. Pudendal nerve stimulation, however, was more selective than sacral nerve stimulation for inhibition with fewer side effects such as leg spasms. Tibial nerve stimulation was ineffective and caused leg spasms and increased bladder activity. Finally, high-frequency stimulation (1,000 Hz) of the sacral nerves was shown to block bladder contractions in 2 of 3 cats investigated. However, this method had adverse side effects such as leg flexion and secondary bladder contractions. We conclude that pudendal nerve/pelvic floor stimulation at low frequency is a relatively effective method in this model.

Animals↗
PubMed1993

Growth factors: therapeutic advances in wound healing.

Polypeptide growth factors regulate cellular processes involved in wound healing. Application of exogenous growth factors can modify the healing process and, with recombinant DNA technology, growth factors can now be made in sufficient quantity to be used therapeutically. Several growth factors are showing promising results in clinical trials, especially in cases of impaired healing, such as chronic ulcers. Preclinical studies indicate that further growth factors may have therapeutic potential in a wide range of wound-healing applications. The use of specifically designed and modified growth factors, growth-factor inhibitors, and sequential and combinatorial dosing regimes offer further possibilities for enhancing wound healing.

Animals↗
PubMed1992

Comparison of direct bladder and sacral nerve stimulation in spinal cats.

Neuroprosthetic techniques have been used to facilitate voiding via electrical stimulation for bladder management following spinal cord injury (SCI), but high urethral resistance has been a problem. This problem was investigated here in the chronic, spinal, male cat (C6-T1) using direct bladder and sacral nerve stimulation. Direct bladder stimulation was only conducted during terminal procedures with an open abdomen and with four hook electrodes inserted into the bladder wall. Sacral stimulation was conducted daily during the 10 weeks post-SCI and during terminal procedures. Stimulation was conducted with both implanted epidural electrode and surface electrodes over the sacral bone. Both of these sacral methods stimulated anterior and posterior roots. However, these sacral methods were generally ineffective for inducing voiding during the study. In three of the five animals investigated, stimulation did not empty the bladder. In the remaining two animals, the bladder was emptied with sacral stimulation, but only after return of bladder reflex activity, 2 to 4 weeks post-injury. When poor voiding occurred in spite of high bladder pressures, it indicates high urethral resistance. This was confirmed using video cystourethrography where the membranous urethra was observed to remain closed following stimulation. Direct bladder stimulation was then compared to sacral nerve stimulation during terminal procedures. Direct bladder stimulation induced voiding at a high rate both during and after stimulation, whereas sacral nerve stimulation with implanted electrodes induced voiding at a lower rate and only after stimulation. A simple urethral resistance measure, the ratio of bladder pressure to voiding rate, was lower with direct bladder stimulation than sacral nerve stimulation. Stimulation-facilitated voiding has also been associated with the development of bladder wall hypertrophy. This problem was investigated by evaluating bladder wall thickness postmortem in three groups of animals: the first group was the spinal-stimulated animals detailed above; the additional two groups were a spinal-nonstimulated but instrumented group maintained for 10 weeks following injury, and an intact group of animals. The stimulated spinal cats tended to have the thickest bladder wall followed by the nonstimulated spinal cats. The wall thickness of intact animals served as a control.

Animals↗