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Biomedical subjects

R A Clark

Publications and source records attributed to R A Clark.

At least 253 records · Page 14Linked to original sources
PubMed1989

Optimization of immunohistochemical techniques to detect extracellular matrix proteins in fixed skin specimens.

Complete antigen visualization in the context of well-preserved tissue architecture is the goal of all immunohistochemical techniques. Frozen tissue section techniques achieve optimal antigen visualization but preserve tissue architecture poorly. On the other hand, formalin-fixed tissue section techniques preserve tissue architecture very well but result in antigen masking. Enzymatic digestion or salt extraction of formalin-fixed sections has been used to reestablish antigen expression. Recently acid-alcohol-fixed tissue has been used as a successful compromise between tissue architecture preservation and the visualization of cytoskeletal antigens. In an attempt to find an improved immunohistochemical process for non-cytoskeletal antigens, we compared avidin-biotin immunofluorescence staining in frozen, formalin-fixed, and acid-alcohol-fixed tissues. The fixed tissues were either untreated or treated with enzyme digestion or salt extraction. For this study, we examined healing cutaneous wounds in Yorkshire pigs with antibodies to fibronectin, laminin, von Willebrand factor VIII, and keratin. Although tissue architecture was poor, frozen sections provided the best antigen visualization and were therefore used as the standard for complete antigen expression. Formalin-fixed tissues had excellent tissue architecture, but most antigens were completely masked. Pre-treatment technique only partially overcame the antigen masking caused by formalin. In contrast, acid-alcohol fixation preserved tissue architecture almost as well as formalin and sometimes allowed complete antigen visualization; however, laminin and fibronectin were partially masked. Total recovery of the expression of these antigens could be obtained by pre-treating the acid-alcohol-fixed tissue with either hyaluronidase or 1 M NaCl. Therefore, acid-alcohol-fixed tissue appears best for extracellular matrix (ECM) protein immunostaining as well as for cytoskeletal staining. However, certain ECM antigens require hyaluronidase or 1 M NaCl treatment for optimal visualization.

Acetates↗
PubMed1989

Controlling delayed vomiting: double-blind, randomized trial comparing placebo, dexamethasone alone, and metoclopramide plus dexamethasone in patients receiving cisplatin.

The majority of patients receiving cisplatin at a dose of 120 mg/m2 experience delayed nausea and vomiting occurring between 24 and 120 hours after chemotherapy administration. Ninety-one patients who were receiving cisplatin (120 mg/m2) as initial chemotherapy were entered into this double-blind trial. All patients received intravenous (IV) metoclopramide, dexamethasone, and lorazepam for the control of acute emesis during the period from 0 to 24 hours after cisplatin. Patients were then randomized to one of three treatment regimens: placebo; oral dexamethasone, 8 mg twice daily for two days, then 4 mg twice daily for two days; or the combination of oral metoclopramide, 0.5 mg/kg four times daily for four days, plus oral dexamethasone administered as above. Forty-eight percent of individuals who received the two-drug combination of metoclopramide plus dexamethasone experienced delayed vomiting as opposed to 65% who were administered dexamethasone alone and 89% who received placebo (P = .006). Scores assessing the severity of delayed nausea and vomiting were consistently worse in individuals receiving placebo. The incidences of sleepiness, restlessness, heartburn, hiccoughs, loose bowel movements, insomnia, and acute dystonic reactions did not differ significantly among the three regimens and were mild and self-limited. The two-drug combination of oral metoclopramide plus dexamethasone is well tolerated, safe, and more effective than dexamethasone alone or placebo in controlling delayed vomiting following cisplatin.

Adult↗
PubMed1989

Screening mammography in the Tampa Bay area: current status and implications for the next decade.

The current status of screening mammography in the Tampa Bay area is evaluated by three parameters: accessibility, cost and quality, and the screening resources necessary to reach the National Cancer Institute's (NCI) cancer control goals for the year 2000 in this area are projected. Fifty-two mammographic facilities were surveyed comprising 64 units in Pinellas, Hillsborough, Pasco and Manatee Counties. The number of units is adequate to serve the current needs of the population eligible for screening; however, the number of facilities offering low-cost examinations is not adequate. Only a few facilities have documented quality assurance through the American College of Radiology. These results and those projected for the year 2000 suggest that new efforts will be necessary to meet the NCI-2000 goal for screening mammography. We have made recommendations to clinicians and radiologists for strategies to meet this challenge.

Adult↗
PubMed1989

Development of persistent late onset asthma following treatment with captopril.

We describe the first case of de novo asthma following treatment with the angiotensin converting enzyme (ACE) inhibitor captopril. Despite drug withdrawal there was evidence of persistent airways obstruction and bronchial hyperreactivity. This suggests the possibility that ACE inhibitors may uncover an asthmatic tendency in patients with pre-existing bronchial hyperreactivity.

Aged↗
PubMed1989

Obstructive airways disease in surgical practice.

Asthma and chronic obstructive pulmonary disease (COPD), i.e. chronic bronchitis and emphysema are common. The pathological and clinical features of these diseases are described. Ventilatory function, lung volumes and 6-min walking tests are used to assess respiratory function while blood gas estimations are essential when managing respiratory failure. The causal mechanisms of respiratory failure are described. In COPD careful assessment with maximization of respiratory function is essential preoperatively. Continued smoking increases postoperative complications sixfold. Bronchial irritability, common in asthma and COPD, increases the anaesthetic risks while many anaesthetic agents adversely affect respiratory function. Postoperative pulmonary complications remain common particularly in COPD. The hypoxia occurring in the first 2 hours post-surgery is usually more pronounced in COPD. Careful postoperative monitoring with aggressive treatment of any complications is essential in these patients. Surgical treatment may significantly improve respiratory function in some patients with bullous emphysema.

Humans↗
PubMed1988

Two cytosolic neutrophil oxidase components absent in autosomal chronic granulomatous disease.

Neutrophils kill microorganisms with oxygen radicals generated by an oxidase that uses the reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) as substrate. This system requires both membrane and cytosolic components and is defective in patients with chronic granulomatous disease. A cytosolic complex capable of activating latent membrane oxidase was eluted from guanosine triphosphate-agarose and was used to raise polyclonal antiserum that recognized 47- and 67-kilodalton proteins. These proteins were restricted to the cytosol of myeloid cells. Both proteins were associated with NADPH oxidase-activating capacity when neutrophil cytosol was purified on nucleotide affinity matrices or molecular sizing columns. Neutrophils from patients with two different forms of autosomal chronic granulomatous disease lacked either the 47- or 67-kilodalton protein.

Blotting, Western↗
PubMed1988

Cryptic chemotactic activity of fibronectin for human monocytes resides in the 120-kDa fibroblastic cell-binding fragment.

Monocytes and lymphocytes form a second wave of infiltrating blood leukocytes in areas of tissue injury. The mechanisms for monocyte accumulation at these sites are not completely understood. Recently, however, fragments from extracellular matrix proteins including collagen, elastin, and fibronectin have been shown to induce monocyte chemotaxis. In this report we demonstrate that chemotactic activity for human monocytes is expressed when a 120-kDa fragment containing the RGDS cell-binding peptide is released from intact fibronectin or from larger fibronectin fragments. Monocytes, either from mononuclear cell Ficoll-Hypaque preparations (10-20% monocytes, 89-90% lymphocytes) or from elutriation preparations (95% monocytes, 5% lymphocytes), but not lymphocytes, migrated toward 120-kDa fragment preparations (10(-7) M) in blind-end chambers when the cells were separated from the chemoattractant by a 5-micron pore polycarbonate filter either alone or overlying a 0.45-micron pore nitrocellulose filter. Neutrophils migrated toward zymosan-activated serum but not toward 10(-5)-10(-8) M concentrations of the 120-kDa fragment. Intact fibronectin had no chemotactic activity for human monocytes. Fibronectin was isolated from citrated human plasma by sequential gelatin-Sepharose affinity and DEAE ion-exchange chromatography in the presence of buffers containing 1 mM phenylmethylsulfonyl fluoride to prevent fragmentation. Controlled enzymatic digestion with thermolysin cleaved fibronectin into 30 kDa fibrin, 45 kDa collagen, and 150/160-kDa cell and heparin domains. Upon prolonged digestion, purified 150/160-kDa fragments were cleaved into 120-kDa cell and 30/40-kDa heparin-binding fragments. Even though the intact fibronectin molecule, the 150/160-kDa fragments, and the 120-kDa fragment, have cell binding activity for Chinese hamster ovary fibroblasts, only the 120-kDa fragment expressed chemotactic activity for human monocytes. Thus, the 120-kDa fibroblastic cell-binding fragment contains a cryptic site for monocyte chemotaxis which is expressed upon enzymatic cleavage of fibronectin.

Chemotaxis, Leukocyte↗
PubMed1988

Depolarization blunts the oxidative burst of human neutrophils. Parallel effects of monoclonal antibodies, depolarizing buffers, and glycolytic inhibitors.

The anti-neutrophil mAb PMN 7C3 and IIC4 inhibited the respiratory burst of neutrophils as measured by the generation of superoxide anion or hydrogen peroxide in response to PMA, serum-treated zymosan, and FMLP. To examine the effect of these mAb on neutrophil transmembrane potential, a fluorescent probe was used in a continuous assay. Compared with control cells, antibody-treated neutrophils were partially depolarized at rest and had a blunted response when stimulated. The F(ab)2 fragment of PMN 7C3 had similar effects on both the respiratory burst and transmembrane potential, whereas the Fab fragment did not. The unrelated antineutrophil mAb 31D8 had no effect on either the respiratory burst or on transmembrane potential. Neutrophils suspended in high potassium buffers also exhibited partial depolarization of the resting cell membrane and a blunted depolarization response to stimuli and produced less superoxide anion and hydrogen peroxide in response to stimuli than did control cells in physiologic buffer. Exposure of neutrophils to 2-deoxy-D-glucose resulted in dose- and time-dependent depression of the respiratory burst. 2-Deoxy-D-glucose also caused depolarization of the resting membrane and impaired subsequent stimulus-induced depolarization. Similar effects were seen with addition of iodoacetamide or depletion of glucose. The parallel effects of anti-neutrophil mAb, depolarizing buffers, and glycolytic inhibitors on both neutrophil membrane depolarization and activation of the respiratory burst indicate a close association between these two events. The evidence suggests that the inhibitory effects of these antibodies are mediated through partial membrane depolarization which interferes with signal transduction on subsequent stimulation of the cells. The impairment in oxidative responses to phorbol esters as well as to receptor-dependent activating agents points to interruption at a distal step, e.g., subsequent to Ca2+ mobilization.

Antibodies, Monoclonal↗
PubMed1988

Problems with asthma following treatment of thyrotoxicosis.

We describe two cases of asthma associated with concomitant thyrotoxicosis where initial improvement followed antithyroid treatment. Relapse of asthma on thyroxine replacement was accompanied by subclinical hyperthyroidism with elevated levels of triiodothyronine. This emphasizes the need to follow up asthmatic patients closely for biochemical relapse following treatment of thyrotoxicosis.

Adult↗
PubMed1988

Atopic dermatitis and aeroallergen contact sensitivity.

Atopic dermatitis (AD) may be worsened by ingested foods or contact with irritants. We have identified 10 patients (six male and four female subjects, aged 1 to 54 years) with AD and contact sensitivity to a variety of aeroallergens. Marked improvement in skin symptomatology was noted when these patients were removed from their usual environment. The patients had markedly positive immediate wheal-and-flare reactions to a variety of aeroallergen extracts in response to prick tests and were subsequently patch tested on uninvolved skin with aeroallergen extracts (1:20 wt/vol, 50% glycerine) that elicited positive prick tests. Patch tests were applied for 48 hours, removed, and then were interpreted 24 hours later. Fifty percent glycerine was used as a negative control. Significant delayed cutaneous responses to a variety of aeroallergens were noted: house dust mite, tree, grass and weed pollens, animal danders, and various molds. Positive delayed cutaneous responses correlated strongly with aeroallergens identified in the patient's environment and/or suspected by the patients as provocateurs of their AD. Delayed cutaneous reactions were negative to aeroallergens not historically relevant to their AD. We conclude that aeroallergen contact may play an important role in selected patients with AD. The demonstration of immediate and delayed cutaneous responses in AD suggests both IgE and cell-mediated hypersensitivity as contributory mechanisms.

Adolescent↗
PubMed1988

The influence of heparin on the wound healing response to collagen implants in vivo.

The biologic response to fibrillar collagen (collagen) and fibrillar collagen plus heparin (collagen/heparin) implants have been compared in the rat subcutaneous and guinea pig dermal wound models. The reconstituted bovine dermal collagen implants were injected subcutaneously in rats at concentrations ranging from 18 to 30 mg/ml and in volumes ranging from 0.5 to 1.0 ml. The biologic response to the collagen implants alone was characterized by a transient invasion of a modest number of inflammatory cells within the first three days of implantation that was followed by limited fibroblast invasion into the peripheral 1/3 of the implant during the course of the next three to four weeks. Occasionally, blood vessels were observed to invade the peripheral regions of the implant. The degree (number) and extent (depth) of cell invasion were inversely related to initial collagen implant concentration. Addition of heparin (0.3-20 micrograms/mg collagen) to these implants resulted in a significant dose-dependent increase in the degree and extent of fibroblast invasion. Radiolabeling studies showed that the collagen and collagen/heparin implants were cleared from the subcutis at identical rates. Implantation of these formulations in a guinea pig dermal wound model was also performed, using a semi-occlusive wound dressing (Opsite) to maintain the implant in the wound site. The fibrillar collagen implant alone was pushed upward by developing granulation tissue at the base of the wound and served as a support for epidermal cell migration, proliferation, and differentiation as wound closure proceeded. The implant was slowly invaded and turned over as granulation tissue developed from the base and margins of the wound bed. The inclusion of heparin in these implants resulted in a significantly different pattern of wound healing. The collagen/heparin implants histologically presented a more broken-up or porous appearance following implantation, which was associated with a greater degree of penetration of developing granulation tissue into the implant itself as compared to the collagen implants. Radiolabeling studies revealed that clearance rates of implants with and without heparin from wound sites were similar, as noted in the rat subcutis. Laser doppler flowmetry studies suggested that the heparin--containing implants were more vascular than control wound sites or sites treated with collagen alone.

Animals↗
PubMed1988

Fibronectin extracellular matrix assembly by human epidermal cells implanted into athymic mice.

Human epidermal keratinocytes reorganize into epidermal inclusion cysts when implanted subcutaneously into athymic mice. During the organization and maturation of these cysts, fibronectin accumulates in the surrounding extracellular matrix and the basement membrane proteins bullous pemphigoid antigen and laminin appear at the epithelial-stromal interface. The sequence in which these proteins appear parallels that seen during reepithelialization of a skin wound in vivo. Fibronectin appears during aggregation of the epidermal cells and persists in the area surrounding the cysts for at least 7 days. Bullous pemphigoid antigen and laminin appear later (by 4 and 7 days, respectively) and ultimately become organized into a continuous band at the periphery of the cyst. This distribution of bullous pemphigoid antigen and laminin at the stromal-epithelial interface persists at least 5 weeks, suggesting that the implanted epidermal cells are capable of developing and maintaining a stable basement membrane zone. Fibronectin, which is abundant in the matrix adjacent to the epidermal cysts and in the surrounding stroma during cyst organization and maturation, diminishes to undetectable levels by 5 weeks. While much of the fibronectin derives from the host tissues, species-specific antibodies to human fibronectin reveal that at least a portion of this protein is synthesized and deposited by the implanted epidermal cells.

Animals↗