Biomedical subjects
G D Griffiths
Publications and source records attributed to G D Griffiths.
Management of diabetic midfoot ulcers.
Diabetic midfoot ulcers are caused by bone and joint disruption that occur as a consequence of progressive peripheral neuropathy associated with this disease. This results in osseus deformities and areas of high pressure on the plantar surface of the midfoot, which cause the ulcers. These lesions are difficult to heal and frequently lead to amputation. In a series of 348 patients, 40 developed 54 midfoot ulcers. Limb preservation was achieved in 33 (61%). Wound closure was achieved in 32 (60%). The amputation rate was highest (83%) in the 10 of 12 patients with peripheral vascular disease. Successful therapy for limb preservation most often included an operation combining resection of underlying osseus deformities with debridement of affected soft tissues.
Sub-lethal poisoning by self-injection with ricin.
Ricin, one of the most potent toxins known, has a lethal dose range of 1-10 mg per kilogram (Crompton and Gall, 1980) when injected. There is no effective treatment. We report a case of sub-lethal poisoning by self-injection.
Meticulous attention to foot care improves the prognosis in diabetic ulceration of the foot.
Ulceration of the foot is a major cause of morbidity in patients with diabetes, and its treatment has become a significant part of general surgical practice. It is, therefore, important to develop an efficient and effective approach to the care of this complication. We established a clinic dedicated to the care and prevention of foot ulcers in diabetic patients and since its inception in 1985, 343 patients have been seen. We provide regular prophylactic care and education to patients without ulcers, as well as treating those with ulcers. To assess the effectiveness of the clinic, we compared two groups of patients. Group 1 contained those who had ulcers while attending our prophylactic care program. Group 2 comprised those who were referred to us with lesions already present. There were 21 patients in group 1 and 150 in group 2. There were no statistical differences between the two groups with respect to age, sex, type and duration of diabetes, smoking history, prevalence of peripheral neuropathy, peripheral vascular disease, renal impairment and retinopathy. The sites and sizes of lesions were also no different between the groups. In spite of these similarities, however, patients in group 1 had a significantly better prognosis than those in group 2. The over-all number of lesions per patient was lower (1.52 +/- 0.98, compared with 2.06 +/- 1.33, p less than 0.05), the mean time required for lesions to heal was shorter (111.9 +/- 80.5 days compared with 160.5 +/- 151.3 days, p less than 0.05). The major amputation rate was lower and fewer patients required partial foot amputation. Prior to the opening of the clinic, the mean length of inpatient treatment was 30 days. This now has been reduced to 12.9 +/- 12.8 days. We conclude that the improved prognosis for those in group 1 can be attributed to the earlier detection and treatment of both potential and actual foot lesions. These results support the contention that the establishment of a dedicated diabetic foot care clinic and regular patient review can reduce the morbidity associated with diabetic foot ulceration.
A histochemical study of changes observed in the mouse diaphragm after organophosphate poisoning.
A sublethal dose of sarin (GB, isopropyl methylphosphonofluoridate) was administered to mice. The animals were killed up to 28 d after dosing and frozen sections were made of the excised diaphragms which were stained using haematoxylin and eosin and a modified Gomori trichrome method. Muscle fibre degeneration and mononuclear infiltration were seen, notably at 24 h and 3 d. A number of histochemical procedures were carried out, including the GBHA procedure for ionized calcium. Calcium accumulation, seen at 4 h, was the earliest abnormality observed. All changes were rapidly regressing by 5 d and histological appearances were normal by 14 d. It was concluded that sarin produced myopathic changes preceded by calcium accumulation.
Metatarsal head resection for diabetic foot ulcers.
Twenty-five diabetic patients underwent 34 metatarsal head resections for chronic neuropathic ulceration. All ulcers were located on the plantar surface beneath the metatarsophalangeal joints. The ulcers had been present for a mean of 9.0 +/- 7.8 months before operation, yet they healed in a mean of 2.4 +/- 1.6 months postoperatively. None recurred during the mean follow-up time of 13.8 +/- 11.0 months. Moderate peripheral vascular disease, impaired renal function, and retinopathy did not affect the time required for ulcer healing. There were two complications: one wound infection and one hematoma. No extremities were lost, and none of the patients suffered any long-term sequelae. We recommend metatarsal head resection to achieve the healing of chronic diabetic foot ulcers under the metatarsophalangeal joints.
The influence of renal function on diabetic foot ulceration.
We examined the effect of renal function on the formation, severity, and outcome of diabetic foot lesions. Information was collected from a retrospective hospital chart survey and analyzed by univariate and multivariate linear regression analysis. Creatinine clearance, peripheral neuropathy, and peripheral vascular disease were all found to be independently associated with formation of foot lesions, indicating that each of these acts by distinct biologic mechanisms. Renal function had no bearing on the severity of lesions or on their eventual healing. We conclude that foot ulcers are more likely to develop in diabetic patients who also suffer from renal impairment, but they are no less likely to heal than are those in patients with normal renal function. We further conclude that attempts to preserve functional limbs in these patients are justified.
Diphtheria with a difference--a rare Corynebacterium fatality with associated apoptotic cell death.
A case of infection with Corynebacterium ulcerans (C. ulcerans), resulted in the sudden death of a previously healthy 73-year-old woman. Death from Corynebacterium diphtheriae (C. diphtheriae) infection is well-documented. Fatalities following infection with C. ulcerans are unreported; this would appear to be the first documented death due to such infection.
Pathological aspects of ricin toxicity in mammalian lymph node and spleen.
In an earlier study (Griffiths et al., 1987) we demonstrated that two toxic plant proteins, ricin and abrin bring about apoptotic rather than necrotic cell death in mammalian lymphoid tissues and intestine. Here we expand upon the previous study, and report further observations relating purely to ricin and its pathology in lymph node and spleen. Rats were injected with ricin and killed at time intervals, tissue being excised and examined by light or electron microscopy. As well as previously reported apoptotic changes in the lymph node, we observed sinusoidal haemorrhage accompanied by erythrophagocytosis and loss of normal structure. With the aid of lymphocyte typing, we noticed a complete relocation of T and B lymphocyte populations, with destruction of B cells. Follicular centres of the spleen exhibited similar pathology to that seen in lymph nodes, along with loss of lymphocytes in areas adjacent to the central artery of the peri-arteriolar lymphoid sheath (PALS).
Intestinal pathology following intramuscular ricin poisoning.
Within the gut, intramuscular ricin poisoning results in major pathological changes. Immunocytochemistry and electron microscopy illustrate that these changes appear confined to the small intestine, the stomach and large intestine remaining virtually unaffected. These changes include apoptosis crypt and mucosal epithelial cells, hydropic change in enterocytes, infiltration of large numbers of plasma cells into the lamina propria, subsequent apoptosis of many of the plasma cells, and the appearance within the lamina propria of highly active macrophages. Of these changes, greatest significance is attached to the infiltration and apoptosis of large numbers of plasma cells. This is not a common pathological feature within the gastro-intestinal tract, and may prove to be specifically diagnostic for ricin or a group of toxins having structurally similar characteristics. In this study, the toxin was administered by intramuscular injection and not orally; considering this mode of administration, damage to the small intestine was far more severe than might perhaps be expected. The intensity of cellular infiltration within the lamina propria is especially interesting, as it appears to be more analogous to a local immune response triggered by an oral toxin rather than one administered by an intramuscular route.
Demonstration of ricin within the mammalian para-aortic lymph node. I. Comparison of the localization, after intramuscular injection, with three immunocytochemical methods.
Following a supralethal injection of ricin into thigh muscle of the adult rat, the toxin was demonstrated post-mortem in the para-aortic lymph node, ipsilateral to the side of injection. The relative merits of two immunoenzyme methods, peroxidase anti-peroxidase (PAP) and avidin-biotin-peroxidase complex (ABC) and a silver-enhanced immunogold method (IGSS) were assessed in the detection of ricin in the lymph node tissue. The toxin was clearly seen to be located in association with histiocytes found both within and lining the sinuses of the nodes and also, in some cases, in the subcapsular sinus of the node; the toxin was not demonstrable within lymphoid follicles by light microscopy. However, using electron microscopy and the IGSS technique, cells carrying discrete particles of gold could be visualized within follicular areas. The IGSS and ABC-peroxidase methods were both found to give excellent results without background staining at the light microscopy level. However, when these techniques were used prior to embedding and viewing by electron microscopy, the IGSS technique proved to be far superior.
Distribution of ricin within the mammalian para-aortic lymph node. II. Comparison of the localization, after intramuscular dosage of colloidal gold-labelled ricin in vivo, with in vitro binding characteristics of the native toxin.
Previous work has shown that, following an intramuscular injection of ricin, the toxin becomes localized within histiocytes in the sinuses of lymph nodes draining the 'wound' site. When ricin labelled with colloidal gold was similarly injected, it was found within the same lymphoid cells as seen with native ricin. Biologically inert Indian ink apparently follows a similar fate, as demonstrated by the appearance of carbon particles within sinus histiocytes, as soon as 1 h after intramuscular injection. When the binding in vitro of Indian ink or ricin toxin to sections of lymph node was examined, ricin was seen to bind to the surfaces of the same sinusoidal cells and also, with a much lower frequency, to follicular lymphocytes, whereas Indian ink failed to bind. This indicated an interaction between ricin and cell membrane components. Moreover, this binding was inhibited markedly by the galactose-containing disaccharide, lactose, a target sugar specified by the lectin binding site of ricin and to a much lesser extent by the monosaccharide mannose.
Burns management and junior staff--what do they know?
This study examines the ability of junior doctors to initiate the management of burned patients. One hundred and twenty-four junior doctors were assessed using a questionnaire. Eighty per cent of the sample had had undergraduate lectures on the subject and 43 per cent had experience of managing patients with major burns. Despite this only 3 per cent could correctly carry out all the steps necessary to estimate the fluid requirements of a burned patient. Theoretical knowledge of the 'Rule of Nines' was adequate but 10 per cent of the sample made mathematical errors when supplied with a burns formula and the appropriate values. We suggest that postgraduate instruction be given to junior staff and that burns charts include details of a burns formula and an illustrative example of the calculation required.
Electrophoretic determination of ricin using immunogold silver staining--comparison with simple "protein dot" method.
Studies were performed to establish a sensitive electrophoretic immunodetection system for the highly toxic plant protein, ricin. This has potential criminal application as an agent for causing a delayed death following parenteral administration. The immunodetection system could be used to demonstrate residual traces of the toxin left in certain tissues of the victim's body. Following polyacrylamide gel electrophoresis of ricin added to rat muscle tissue extracts, the gels were electro-blotted onto nitrocellulose paper and ricin bands probed for visualisation by immunostaining. Several immunostaining procedures were investigated in order to select the most sensitive. These included indirect immunoperoxidase, peroxidase-anti-peroxidase (PAP), avidin-biotin complex (ABC) and the immunogold silver staining (IGSS) procedures. The sensitivity of PAP and indirect immunoperoxidase methods were similar at around 50 ng while the ABC technique gave visible staining of 10 ng of electro-blotted ricin. The method with greatest sensitivity was undoubtedly IGSS, which resulted in unequivocal demonstration of 4 ng of ricin. The IGSS-immunoblotting system was considered to readily demonstrate the presence of ricin in muscle tissue from the injection site of dead victims. We compared this system with the very simple method of sample dot staining. Here, samples of ricin were spotted directly onto nitrocellulose. The dots were stained using the IGSS method which was found able to demonstrate less than 10 pg of ricin.
The toxic plant proteins ricin and abrin induce apoptotic changes in mammalian lymphoid tissues and intestine.
The toxins ricin and abrin are potent inhibitors of protein synthesis. Apoptosis has been shown to be induced in some cells by cycloheximide and actinomycin D whereas the process is prevented in other cells by the same agents, both inhibitors of protein synthesis. We were interested to find whether ricin and abrin caused any apoptotic changes in rapidly dividing tissues where we believed that these toxins concentrate. Rats were injected intramuscularly with toxin and killed at time intervals, tissues being removed and examined by light and electron microscopy. Apoptotic-like bodies were abundant in para-aortic lymph nodes, Peyer's patches and ideal crypts of ricin or abrin intoxicated rats. Abrin was found to cause markedly more pronounced changes in these tissues, when compared with a similar dose of ricin. Prior to this, these toxins have been reported as causing necrosis in animal tissues.
Immunocytochemical detection of ricin. II. Further studies using the immunoperoxidase method.
Radio-iodinated ricin was injected into rat muscle in vivo to establish the distribution of the toxin at various time intervals after injection. Injection site muscle and para-aortic lymph nodes were selected for localization of ricin by the immunoperoxidase technique. Sections of snap-frozen tissues were fixed using a variety of methods to establish the best protocol for the immunodetection method. This was found to be with an ether-ethanol mixture. Ricin was detected in tissue at the site of injection taken from rats sacrificed 1, 4, 8 and 24 h after injection and in tissue from animals dying from ricin intoxication after about 30 h. This method, however, failed to demonstrate unequivocally the presence of ricin in lymphoid tissue which had been indicated by the radiotracer study. The significance of these findings and their relevance to forensic diagnosis are discussed.
Rabbit peroxidase-antiperoxidase complex (PAP) as a model for the uptake of immunoglobulin G by the human placenta.
Rabbit peroxidase-antiperoxidase complex (PAP) has been shown to bind to IgG receptors on the human placental syncytiotrophoblast microvillar membrane. Its binding characteristics suggest that it is suitable as a probe for studies on the uptake of IgG by the human placenta. A novel assay system was developed to measure the dissociation constants (Kd) of the binding of PAP and of unlabelled human IgG to purified placental microvillar membranes. The Kd for PAP was found to be 54 nM, while that for unlabelled IgG was found to be 17.5 nM. The uptake of PAP by placental tissue slices was observed using peroxidase histochemistry and electron microscopy. In initial experiments, reaction product was confined to the peripheral regions of the syncytiotrophoblast. Assaying a placental homogenate for catalase activity showed that it contained 250 units of activity per g wet weight of tissue (compared with 680 units/g for rat liver). Treatment of fixed tissue with the catalase inhibitor 3-amino-1, 2, 4-triazole allowed the localization of peroxidase reaction product in deeper regions of the syncytiotrophoblast. Based on observations of the localization of reaction product, we propose that PAP is taken up in coated pits, transferred into large apical multivesicular bodies, segregated into small vesicles which then transport it to the Golgi. From here the PAP is directed to the basal membrane by a mechanism as yet unknown.
Comparison of three methods used for the measurement of the avidity of antibody to DNA.
Three methods for determining the avidity of antibodies to single-stranded DNA (ssDNA) have been compared. None of the 3 methods was totally satisfactory but for theoretical and practical reasons the measurement of avidity by determination of dissociation rate is considered the most appropriate.