Biomedical subjects

L Gerrard

Publications and source records attributed to L Gerrard.

9 recordsLinked to original sources

PubMed2003

Tachykinin expression in cartilage and function in human articular chondrocyte mechanotransduction.

OBJECTIVE: To assess whether substance P and the corresponding neurokinin 1 (NK1) receptor are expressed in human articular cartilage, and whether these molecules have a role in chondrocyte mechanotransduction. METHODS: Transgenic studies, immunohistochemistry, Western blotting, and reverse transcriptase-polymerase chain reaction were used to assess the expression of the preprotachykinin (PPT) gene, substance P, and NK1 in developing mice, in adult human articular cartilage, and in human chondrocytes in culture. Chondrocytes obtained from PPT knockout mice and human articular chondrocytes were mechanically stimulated in the presence or absence of inhibitors of substance P signaling, and cell membrane potentials or relative levels of aggrecan messenger RNA (mRNA) were measured. RESULTS: Replacing a region of the PPT gene transcriptional site that contains a dominant repressor of the proximal promoter activity with the constitutive minimal promoter of the human beta-globin promoter allowed expression of a marker gene in areas of chondrogenesis during mouse development and in adult chondrocytes grown in culture. Adult human articular chondrocytes expressed endogenous PPT mRNA, substance P, and the corresponding NK1 receptor in vivo and in vitro. Blockade of substance P signaling by a chemical antagonist to the NK1 receptor inhibited chondrocyte responses to mechanical stimulation. CONCLUSION: Substance P is expressed in human articular cartilage and is involved in chondrocyte mechanotransduction via the NK1 receptor in an autocrine and paracrine manner. This suggests that substance P and the NK1 receptor have roles in the maintenance of articular cartilage structure and function that were previously unrecognized.

Age Factors↗

PubMed2000

A role for Octamer binding protein motifs in the regulation of the proximal preprotachykinin-A promoter.

We have recently developed cell line models that express the endogenous rat preprotachykinin-A (rPPT) gene and support reporter gene expression directed by the rPPT promoter. These are the neuronal derived cell line NF2C and the pancreatic cell lines RINm5F and a derivative RIN-1027-B2. Reporter gene activity in these cell lines is similar to that observed in primary cultures of rat dorsal root ganglion neurons. Analysis of reporter gene expression supported by various fragments of the rPPT promoter demonstrated that although -865 to +92 supported expression, addition of fragments between +92 and +500 led to repression of expression. Two previously defined octamer binding motifs, present both 5' and 3' of the major transcriptional start site, have been postulated to be potential enhancers of rPPT activity and we have now used these cell lines to determine the role of these regions in the rPPT promoter. Site specific mutagenesis of these elements has shown not only that both sites are potential enhancers of gene expression but also that the 3' element binds multiple factors, of which at least one has repressor function. Binding of this repressor protein over or adjacent to the 3' octamer binding protein site leads to the observed repression of promoter activity in the -865 to +500 construct relative to the to -865 to +92 the fragment.

Amino Acid Motifs↗

PubMed2000

Molecular models to analyse preprotachykinin-A expression and function.

Towards an understanding of the mechanisms controlling Preprotachykinin A (PPT) expression we have generated a variety of molecular models to determine the mechanisms regulating both the tissue-specific and stimulus-inducible expression of the PPT gene. The approaches used include transgenic and virus vector models complementing biochemical analysis of promoter interactions with transcription factors. We have identified and characterised a yeast artificial chromosome (YAC) containing the human PPT gene and generated transgenic mouse lines containing multiple copies of this chromosome on a normal mouse genetic background. This resulted in a pattern of expression in the nervous system remarkably similar to that reported for PPT mRNA in rodents. In addition, this transgenic model has been constructed in such a manner to allow for over expression of tachykinins based on the number of extra alleles in the transgenic mouse. These animals allow us to further examine the function of the tachykinins and acts as a useful complement to existing PPT ablated mice. In vitro we have introduced the proximal PPT promoter in reporter gene constructs into adult neurones in both DRG and the CNS by an adenoassociated virus (AAV) vector or by biolistic transfection respectively. Using the AAV vector we have demonstrated that the proximal promoter can mediate the effects of NGF in adult rat DRG. These models allow us to delineate transcriptional domains involved in the physiological and pathological expression of the PPT gene.

Animals↗

PubMed1999

Novel cell lines for the analysis of preprotachykinin A gene expression identify a repressor domain 3' of the major transcriptional start site.

Until now, no clonal cells have been identified that support the expression of a marker gene expressed from the rat preprotachykinin A (rPPT) promoter. We have analysed recently available cell lines that are candidates for supporting reporter gene expression directed by the rPPT promoter. These are the neuronal-derived cell line NF2C and the pancreatic cell lines RINm5F and a derivative RIN-1027-B2. The NF2C line was derived from the brain homogenate of a transgenic animal in which a temperature-sensitive simian virus 40 large T antigen was expressed from a neurofilament promoter. All three lines are able to support expression of a reporter gene directed by a fragment of the 5' rPPT promoter. Analysis of reporter gene expression supported by various fragments of the rPPT promoter demonstrated that although -865 to +92 bp supported expression, the addition of fragments between +92 and +447 bp led to repression of expression. Subsequent analysis of reporter gene constructs microinjected into primary cultures of dorsal root ganglion neurons (DRG) confirmed the existence of this repressor domain. This repression could be relieved totally in both RIN cell lines and partly in NF2C cells by mutating residues between +373 and +396 bp. This indicates that these cell lines support PPT promoter activity similar to that observed in DRG and determines a novel repressor domain within the promoter.

Animals↗

PubMed1999

An intronic domain within the rat preprotachykinin-A gene containing a CCCT repetitive motif acts as an enhancer in differentiating embryonic stem cells.

Previous attempts by several groups to clone fragments containing intron 2 of the rat preprotachykinin-A gene have generated deletions of various sizes. We have determined that these deletions occur within a specific region of the intron spanning a CCCT tandem repeat domain. We show that this intronic domain is able to support reporter gene expression in mouse embryonic stem (ES) cells that have been induced to differentiate but not in undifferentiated ES cells. No significant expression was observed in the HeLa clonal cell line. This demonstrates that this intron 2 domain is a highly restrictive enhancer and may be associated with differentiation.

Animals↗

PubMed1999

Factors that may influence the implementation of nurse-centred telemedicine services.

A qualitative study of four UK telemedicine sites was carried out to explore reactions to nurse-centred telemedicine services and their possible implications. Using semistructured schedules, 36 people directly involved in these services were interviewed--25 nurses, four general practitioners, two advising medical consultants, three service managers and two researchers. Factors identified which may influence successful implementation included: early involvement of all groups affected, prior consideration of the practical implications, thorough initial equipment testing, good technical support, imaginative training and clarity of the purpose of the service. The study showed that all those involved must be prepared for the rapid learning that is implicit in the implementation of change. The challenge is to maximize the potential for the primary operator to use new techniques. In respect of nurse-centred services, the nurse must not be viewed as a technician, nor should telemedicine be seen as a substitute for an available doctor.

Clinical Competence↗

PubMed1997

The Dundee out-of-hours cooperative: preliminary outcomes for the first year of operation.

The Dundee out-of-hours cooperative (DDOC) was the first of its kind to be developed in a city in Scotland. In its first year of operation, the key features of the cooperative were that (a) two doctors could deal with the large majority of out-of-hours calls for a population of just under 100,000 patients, (b) only one third of calls required a home visit, (c) most calls were handled within a period of time acceptable to patients, (d) seven out of 10 patients rated the service equal to or better than previous services, and (e) there were extended opportunities for general practitioners (GPs) involved in the scheme to meet with other colleagues during out-of-hours work.

Family Practice↗

PubMed1995

Effect of combined atenolol and nifedipine administration on psychomotor performance in normal subjects.

The aim of the present study was to evaluate the central effects of single doses of the beta-adrenoceptor antagonist atenolol and the calcium antagonist nifedipine retard, alone and in combination, in normal subjects. Twelve normal males received single oral doses of atenolol 100 mg, nifedipine retard 20 mg, atenolol 100 mg and nifedipine retard 20 mg in combination, diazepam 5 mg (active control), and each of two matching placebos in a double-blind, randomised fashion. Psychomotor performance was assessed using digit symbol substitution, letter cancellation (LCT), continuous attention, choice reaction time, finger tapping, immediate recall and short-term memory. Two flash fusion and critical flicker fusion thresholds were measured and subjective assessments made using visual analogue scales (VAS). Diazepam 5 mg significantly worsened LCT scores at 4h, significantly impaired alertness at 2 h and 4 h, and tended to increase reaction time and impair continuous attention and physiological measurements. Atenolol 100 mg alone significantly reduced alertness at 2 h and 4 h, and also tended to impair physiological measurements. Nifedipine retard 20 mg produced no significant psychomotor effects. Combined atenolol and nifedipine retard administration produced a small but significant improvement in continuous attention and a reduction in body sway, with no adverse effects being evident on performance or subjective awareness. The results suggest that no significant adverse effects on psychomotor performance are produced by single doses of atenolol 100 mg and nifedipine retard 20 mg when given together in normal subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗

PubMed1993

Central effects of combined bendrofluazide and atenolol administration. A single dose study in normal subjects.

Twelve normal male subjects received single oral doses of atenolol 100 mg (AT), bendrofluazide 5 mg (BFZ), combined atenolol 100 mg and bendrofluazide 5 mg (AT/BFZ), diazepam 5 mg (Dz), or one of two matching placebos, on each of 6 study days. Tests of psychomotor performance [digit symbol substitution (DSST), letter cancellation (LCT), continuous attention, choice reaction time (CRT), finger tapping, short-term memory, body sway], physiological measurements [critical flicker fusion (CFF), two-flash fusion (2FF)] and subjective assessments using visual analogue scales (VAS), were performed at 2 and 4 hours post-ingestion. Dz (active control) significantly worsened VAS scores at 2 h (+0.68) and reduced DSST scores at both 2 h (-15.0) and 4 h (-11.0). AT and BFZ given alone, each produced significant worsening of VAS at 2 h [AT +1.0; BFZ +1.38], but had no significant effects on performance. In combination however, AT/BFZ at 4 h produced significant impairment of DSST scores (-10.4), reduced finger tapping (-16.5) and increased involuntary rest pauses (+16.5). Despite these effects, no change in VAS scores occurred. In summary, we have demonstrated significant impairment of psychomotor performance in normal subjects with the AT/BFZ combination, which was not evident with the single agents and which occurred in the absence of a change in subjective awareness. These central effects may have important clinical implications for patients taking combined antihypertensive medication.

Adult↗