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Biomedical subjects

Keith Harding

Publications and source records attributed to Keith Harding.

At least 19 recordsLinked to original sources
PubMed2006

Optimisation of the azinobis-3-ethyl-benzothiazoline-6-sulphonic acid radical scavenging assay for physiological studies of total antioxidant activity in woody plant germplasm.

A robust spectroscopic method for determining total antioxidant activity in aqueous extractions has been applied to tissues from diverse woody plant species, including seeds of Coffea arabica and in vitro shoots from Ribes nigrum, Picea sitchensis and Shorea leprosula. The assay involves scavenging of an ABTS [2,2'-azinobis-(3-ethyl-benzothiazoline-6-sulphonic acid)] radical generated by the reaction of potassium persulphate with ABTS to produce an ABTS*(+) chromophore (lambda=734 nm). Antioxidants reduce ABTS*(+) back to ABTS with a concomitant decrease in absorbance. Aqueous extractions from C. arabica and S. leprosula had considerably higher (110-205 micromol Trolox eq. g(-1) FW) total antioxidant activities than P. sitchensis and R. nigrum (6-11 micromol Trolox eq. g(-1) FW). Further studies in two of these species showed that the inclusion of water-insoluble polyvinylpyrrolidone during aqueous tissue extraction enabled the combined phenolic and alkaloid antioxidant activity to be determined. These fractions accounted for 85% and 60% of total antioxidant activity for C. arabica seeds and R. nigrum shoots, respectively. The ABTS radical scavenging assay is presented herein as a robust method for determining total antioxidant activity in germplasm from diverse woody plant tissues and species. Its applicability to study oxidative stress in tissue cultures and germplasm employed in plant biotechnology, breeding and stress physiology programmes is discussed.

Antioxidants↗
PubMed2006

The management of deep sternal wound infections using vacuum assisted closure (V.A.C.) therapy.

A group of international experts met in May 2006 to develop clinical guidelines on the practical application of vacuum assisted closure (V.A.C.)+ therapy in deep sternal wound infections. Group discussion and an anonymous interactive voting system were used to develop content. The recommendations are based on current evidence or, where this was not available, the majority consensus of the international group. The principles of treatment for deep sternal wound infections include early recognition and treatment of infection. V.A.C. therapy should be instigated early, following thorough wound irrigation and surgical debridement. V.A.C. therapy in deep sternal wound infections requires specialist surgical supervision and should only be undertaken by clinicians with adequate experience and training in the use of the technique.

Humans↗
PubMed2005

Risk of plantar ulceration in diabetic patients with single-leg amputation.

BACKGROUND: There is a disconcerting rate of bilateral limb loss in patients with diabetes. Therefore, this study aimed to explore plantar loading of the surviving foot following unilateral trans-tibial amputation within a wider context of daily walking activity to investigate the precise risk to the surviving limb. METHODS: Twenty-one subjects with diabetic neuropathy and trans-tibial amputation were matched for weight; height; age and gender with 21 control subjects with diabetic neuropathy without history of plantar ulceration. Gait parameters, in-shoe plantar pressure distribution and daily walking (using the step activity monitor) were recorded. Student's t-tests were used to compare groups (alpha-level: 0.05). FINDINGS: The trans-tibial amputations group walked almost 30% slower compared to controls (P < 0.01), with reduced cadence (P < 0.01), and shorter strides (P < 0.01). Despite walking slower, the surviving foot showed higher mean peak plantar pressures in the trans-tibial amputations group over the heel (P < 0.001) however there was no significant difference over the I-II and lateral III-IV-V metatarso-phalangeal regions. Pressure time integral was higher over the heel (P < 0.00), I-II (P < 0.01) and III-IV-V metatarso-phalangeal (P < 0.05) in the trans-tibial amputations group. The amputee group walked less steps per day (P < 0.01). INTERPRETATION: Adaptations in gait and level of walking activity affect plantar pressure distribution and ultimately the risk of ulceration to the surviving foot. Therefore rehabilitation measures should consider implications for plantar loading and the potential risk of ulceration to the surviving foot.

Adult↗
PubMed2005

HPLC analysis of plant DNA methylation: a study of critical methodological factors.

HPLC analysis of nucleosides is important for determining total DNA methylation in plants and can be used to help characterise epigenetic changes during stress, growth and development. This is of particular interest for in vitro plant cultures as they are highly susceptible to genetic change. HPLC methodologies have been optimised for mammalian and microbial DNA, but not for plants. This study examines critical methodological factors in the HPLC analysis of plant DNA methylation using in vitro cultures of Ribes ciliatum. HPLC revealed that complete removal of RNA from plant DNA extractions is difficult using RNase (A and T1) digestions and LiCl precipitation. This suggests that base analysis should be avoided when using these RNA removal techniques, as bases from residual RNA fragments will inflate peak areas for DNA-derived bases. Nucleoside or nucleotide analysis is therefore recommended as a more suitable option as RNA and DNA constituents can be readily separated. DNA digestion was also a critical factor as methylation was under-estimated following incomplete nuclease digestion and over-estimated following incomplete phosphatase digestion. The units of enzyme required for complete DNA digestion was optimised and found to be 20-200 times less for nuclease P1 and 15 times less for alkaline phosphatase as compared with previous protocols. Digestion performance was conveniently monitored using marker peaks that indicate incomplete digestion products. This study identifies critical components of HPLC analysis and offers a comprehensive guide for the stringent analysis of DNA methylation in plants.

Chromatography, High Pressure Liquid↗
PubMed2004

Cardiff Wound Impact Schedule: the development of a condition-specific questionnaire to assess health-related quality of life in patients with chronic wounds of the lower limb.

The purpose of this study was to develop and validate a questionnaire to measure the impact of chronic wounds (leg ulcers and diabetic foot ulcers) on patient health-related quality of life (HRQoL) and identify areas of patient concern. The Cardiff Wound Impact Schedule (CWIS) was created following a three-stage process. Stage 1 included a focus group (n=10) and a series of semi-structured interviews (n=13) to generate items for the questionnaire. Stage 2 involved piloting the 28-item questionnaire on 124 patients (64.5% with leg ulceration and 35.5% with diabetic foot ulceration): data from this stage were analysed using factor analysis. The reliability, validity and reproducibility of the resulting scale were assessed in Stage 3, a 3-month follow-up study of 135 patients in which patients completed the CWIS and the SF-36. Factor analysis identified three domains of HRQoL: physical symptoms and daily living, social life and well-being. There were no significant differences in scores across the wound types. Internal consistency was good (alpha=0.77-0.96) as was reproducibility (P<0.001). The CWIS was able to discriminate between those with healed ulcers and active ulcers (P<0.01). Construct validity was demonstrated by strong correlations between related items on CWIS and SF-36 (P<0.01-P<0.0001). The data suggest that CWIS has high internal consistency and the ability to discriminate between health states and good reproducibility. CWIS is a valid tool for studying the impact of chronic wounds of the lower leg on HRQoL. CWIS allows clinicians to identify items of patient concern, which can then be used to negotiate options of care most suited to individual patients.

Adult↗
PubMed2003

Wound bed preparation: a systematic approach to wound management.

The healing process in acute wounds has been extensively studied and the knowledge derived from these studies has often been extrapolated to the care of chronic wounds, on the assumption that nonhealing chronic wounds were simply aberrations of the normal tissue repair process. However, this approach is less than satisfactory, as the chronic wound healing process differs in many important respects from that seen in acute wounds. In chronic wounds, the orderly sequence of events seen in acute wounds becomes disrupted or "stuck" at one or more of the different stages of wound healing. For the normal repair process to resume, the barrier to healing must be identified and removed through application of the correct techniques. It is important, therefore, to understand the molecular events that are involved in the wound healing process in order to select the most appropriate intervention. Wound bed preparation is the management of a wound in order to accelerate endogenous healing or to facilitate the effectiveness of other therapeutic measures. Experts in wound management consider that wound bed preparation is an important concept with significant potential as an educational tool in wound management. This article was developed after a meeting of wound healing experts in June 2002 and is intended to provide an overview of the current status, role, and key elements of wound bed preparation. Readers will be able to examine the following issues; the current status of wound bed preparation; an analysis of the acute and chronic wound environments; how wound healing can take place in these environments; the role of wound bed preparation in the clinic; the clinical and cellular components of the wound bed preparation concept; a detailed analysis of the components of wound bed preparation.

Cell Proliferation↗
PubMed2003

Healing responses of skin and muscle in critical illness.

OBJECTIVE: Many aspects of the care and underlying pathologies in patients suffering critical illness can detrimentally influence the normal healing processes of skin and soft tissues. Although a great diversity of pathologies exists, some aspects of the diseases and their treatments are common in critically ill patients. We aimed to identify some features, both common and specific, that could influence wound healing and the mechanisms by which they may do so. DESIGN: In this review, we first outline the biology of normal skin and muscle healing and then explore how critical illness may influence the normal healing cascade. FINDINGS: The healing of skin and skeletal muscle in critical illness is influenced by both underlying disease processes and the intensive care environment. Local and systemic factors can contribute to impaired healing, with the potential to prolong functional disability and increase the likelihood of wound complications. The frequency and number of soft tissue injuries derived from accidental injury, surgical intervention, and the need for invasive monitoring and therapies in the intensive care unit setting are likely to compromise the innate immunity and potentially further jeopardize the patient's ability to heal. Alterations in coagulation, tissue perfusion, inflammation, immune functioning, metabolism, nutrition, and drug therapies will influence healing responses by modifying the biological responses to tissue disruption. Locally, wound contamination, sepsis, tissue hypoxia, edema, and excessive or prolonged local pressure all have the potential to compromise soft tissue healing. One or more of these factors may be present at any time. CONCLUSION: The skin and soft tissues are vulnerable to both injury and compromised healing when a patient is critically ill and exposed to a critical care environment. The identification of risk factors may aid in forming and modifying treatment strategies when caring for the critically ill patient with soft tissue injuries.

Critical Care↗
PubMed2003

Deciphering the impaired cytokine cascades in chronic leg ulcers (review).

Chronic leg ulcers are typically wounds that do not heal at a normal rate. Impaired healing appears to be due to primary microvascular changes and it is aggravated by ongoing bacteria-driven vasculitis. The various cytokines identified in experimental wounds are also present in leg ulcers. VEGF is strongly implicated as a promoter of blood vessel growth in patients with venous disease. In addition, there is good evidence of increased expression of bFGF, TGF-beta1, and PDGF in lipodermatosclerosis. All of these growth factors are involved in wound healing. Upregulated TGF-beta1 is probably one of the main causes of the fibrosis observed in lipodermatosclerosis. In leg ulcers, cytokines appear to be trapped in the perivascular fibrinoid deposits. It is not the nature and amount of cytokines that are inadequate in leg ulcers, but rather their spatial distribution. Dermal dendrocytes (DD) are resident factor XIIIa-enriched macrophages. They likely play a role in tissue repair when boosted adequately. New therapies aiming at helping the release of cytokines by DD apparently promote and improve the healing phase.

Animals↗
PubMed2002

Evaluating the effectiveness of a dressing for use in malodorous, exuding wounds.

Malodorous, exuding wounds can cause problems for patients in terms of discomfort and social isolation. A prospective, noncomparative, multicenter study was conducted to evaluate the effectiveness of an odor-controlling dressing in the management of patients with chronic and acute exuding, malodorous wounds. Forty-six patients with various malodorous wounds (mean duration prior to study entry of 231.6 weeks) participated in the study. The majority of the patients were > 65 years of age (mean 70.7 years). Wound characteristics, odor, and dressing exudate absorption were assessed at baseline and at every dressing change (total: 348 dressing changes). Overall assessment of the dressing's ability was rated as good or excellent for odor control (91% of patients), exudate management (82%), and overall comfort (86%). Patient and nurse perception of odor were similar. While appropriate treatment to address the cause of exudate and wound odor remains a priority, the results of this study suggest that discomfort and isolation may be reduced through the use of odor-control dressings.

Aged↗
PubMed2002

Improved body-outline imaging technique for localization of sentinel lymph nodes in breast surgery.

UNLABELLED: Many centers use techniques for localizing the sentinel lymph node (SLN) associated with a breast tumor. Protocols involve the use of an intraoperative probe and blue dye but may or may not include scintigraphy. METHODS: Three methods of body-outline imaging were investigated to aid SLN localization. Body outlines were acquired using a handheld source; a transmission image was acquired from a (57)Co-flood source; and, finally, a new method is described using a (153)Gd-line source attenuation correction for body outline. RESULTS: Method 1 images were of low quality, although 83% of SLNs were visualized. Body outline was unsatisfactory for surgical localization. Method 2 was unsatisfactory because of the lack of a lateral image. Method 3 enabled good presurgical visualization of the SLN (73%) and speeded surgical localization. CONCLUSION: The (153)Gd-line source consistently gives optimal-quality body-outline images. This source is simple, quick, and clearly locates the SLN on multiple projections.

Breast Neoplasms↗
PubMedDate not supplied

Effect of human fibroblast-derived dermis on expansion of tissue from venous leg ulcers.

Novel approaches to healing of chronic wounds, such as venous leg ulcers, include the use of tissue-engineered skin substitutes, e.g., human fibroblast-derived dermis. The exact mechanisms of action of these products and their effects on wound healing at a cellular level are yet to be fully defined. The aim of our study was to evaluate the potential effects of human fibroblast-derived dermis on the healing of chronic wounds using an experimental model. We used a tissue expansion model to examine the effect of human fibroblast-derived dermis on the growth of human tissue biopsied from venous leg ulcers. Further characterization of the cytokine profile produced by human fibroblast-derived dermis in culture was performed using enzyme-linked immunosorbent assay techniques. Addition of medium conditioned with human fibroblast-derived dermis significantly increased the outgrowth of cells from venous leg ulcer biopsies (p = 0.001). We detected bioactive levels of hepatocyte growth factor/scatter factor and interleukin-8 in media conditioned with human fibroblast-derived dermis. Therefore, conditioned media from human fibroblast-derived dermis enhances ex vivo expansion of tissue taken from chronic venous leg ulcers, and contains potent angiogenic factors. These experimental findings may explain the enhanced healing seen with clinical applications of human fibroblast-derived dermis on chronic wounds.

Cell Culture Techniques↗
PubMedDate not supplied

Genetic integrity of cryopreserved plant cells: a review.

Cryopreservation techniques exist for some 100-plant species. Cryopreservation for the long-term conservation of in vitro germplasm results in the exposure of tissues to physical, chemical and physiological stresses causing cryoinjury. Although, the effects of cryoinjury upon the genome are often unknown, any accumulative DNA polymorphisms may not be induced by cryopreservation per se but are the result of the whole culture-cryoprotection-regeneration process. It is desirable to assess the genetic integrity of plants surviving cryogenic storage to determine if they are true to type after cryopreservation. This can be done at the phenotypic, histological, cytological, biochemical and molecular levels. The relevance of these approaches to stability investigations is discussed with their limitations. This review provides a definition for Cryobionomics - a novel term describing the re-modelled concept of genetic stability and the re-introduction of cryopreserved plants into the environment.

Biochemistry↗