The use of 7-amino actinomycin D in identifying apoptosis: simplicity of use and broad spectrum of application compared with other techniques

The detection and quantitation of apoptotic cells is becoming increasingly important in the investigation of the role of apoptosis in cellular proliferation and differentiation. The pathogenesis of hematologic disorders such as aplastic anemia and the development of neoplasia are believed to involve dysregulation of apoptosis. To quantitate accurately the proportion of apoptosis cells within different cell types of a heterogeneous cell population such as blood or bone marrow, a method is required that combines the analysis of large numbers of cells with concurrent immunophenotyping of cell surface antigens. In this study, we have evaluated such a method using the fluorescent DNA binding agent, 7-amino actinomycin D (7AAD), to stain three diverse human cell lines, induced to undergo apoptosis by three different stimuli. Flow cytometric analysis defines three populations on the basis of 7AAD fluorescence and forward light scatter. We have shown by cell sorting and subsequent morphological assessment and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling that the populations defined by 7AAD represent live, apoptotic, and late-apoptotic/dead cells. This method is quick, simple, reproducible, and cheap and will be a valuable tool in the investigation of the role of apoptosis in normal physiology and in disease states.     

A change in pituitary magnetic resonance imaging protocol detects ACTH‐secreting tumours in patients with previously negative results

Objective While detection of pituitary tumours with magnetic resonance imaging (MRI) may reduce diagnostic costs and improve surgical outcomes for patients with Cushing’s disease, the optimal T1‐weighted spin‐echo (SE) MRI protocol remains unknown. We hypothesized that specific MR scanning parameters influence detection of corticotropinomas. Design and patients Between December 1997 and November 2004, 21 of 84 consecutive patients with Cushing’s disease had a falsely negative initial pituitary MRI study and a lesion identified subsequently at the National Institutes of Health Clinical Center. This study retrospectively reviewed and compared technical parameters used for the two pituitary T1‐weighted SE MRIs in 18 patients with available scans. Measurements Repetition time (TR)/echo times (TE), field of view (FOV), matrix size, magnetic field strength, slice thickness, use of gadolinium contrast and the time interval between studies were recorded. Results The MRI interscan interval was 5·4 ± 1·1 months. All scans used gadolinium, matrix sizes were similar and nearly all had 3‐mm thick slices. Parameters that differed between the NIH‐ and externally performed scans were: TR (400 ms vs. 492 ± 19 ms, P = 0·0002); TE (10·3 ± 0·5 vs. 17·2 ± 1·2 ms, P = 0·0003); FOV (12 × 12 cm vs.17 ± 0·6 × 18 ± 0·7 cm, P < 0·0001). Immunohistochemistry of tumours resected at transsphenoidal surgery confirmed all to be corticotropinomas. Conclusions Not all ‘T1‐weighted SE’ scans are equally accurate. MRI technique, particularly FOV and TR/TE value, influences results. We recommend that endocrinologists consider pituitary MRI parameters when interpreting the results.     

Mixing during Intravertebral Arterial Infusions in an in vitro Model

Regional delivery of drugs can offer a pharmacokinetic advantage in the treatment of localized tumors. One method of regional delivery is by intra-arterial infusion into the basilar/vertebral artery network that provides local access to infratentorial tumors, which are frequent locations of childhood brain cancers. Proper delivery of drug by infused solutions requires adequate mixing of the infusate at the site of infusion within the artery lumen. Our mixing studies with an in vitro model of the vertebral artery network indicate that streaming of drug solution is likely to occur at low, steady infusion rates of 2ml/min. Streaming leads to maldistribution of drug to distal perfused brain regions and may result in toxic levels in some regions while concurrently yielding subtherapeutic levels in adjacent regions. According to our model findings, distribution to both brain hemispheres is not likely following infusion into a single vertebral artery even if the infusate is well-mixed at the infusion site. This outcome results from the unique fluid flow properties of two converging channels, which are represented by the left and right vertebral branches converging into the basilar. Fluid in the model remains stratified on the side of the basilar artery served by the infused vertebral artery. Careful thought and planning of the methods of intravertebral drug infusions for treating posterior fossa tumors are required to assure proper distribution of the drug to the desired tissue regions. Improper delivery may be responsible for some noted toxicities or for failure of the treatments.     

Intramedullary spinal cord metastasis from renal cell carcinoma: detection by positron emission tomography

Although positron emission tomography (PET) is an established diagnostic method in brain and lung cancer, its use is often confined to research. The authors report a case of a minimally symptomatic intramedullary spinal cord metastasis, an uncommon and often diagnostically challenging lesion, that was confirmed by PET. A 37-year-old man with a history of metastatic renal cell carcinoma treated with systemic agents, an autologous stem cell transplant, and local palliative radiotherapy with a 2-month history of vague right foot numbness and right leg dysesthesias was found to have an intramedullary lesion at the level of T12. Although the findings of magnetic resonance imaging suggested central necrosis, a PET scan revealed a metabolically active lesion and confirmed the diagnosis of intramedullary metastasis. PET can be used to detect and confirm intramedullary spinal cord metastatic carcinoma. PET imaging may have a vital role in clinical diagnosis by helping to distinguish diagnostically troublesome lesions based on metabolic activity.     

Evaluation of attempted prevention of unexpected infant death in very high-risk infants by planned health care

Three hundred and ninety-six babies born in Sheffield between 1982 and 1990 identified as being at "very high risk" of unexpected infant death by means of a scoring system, received an intensive programme of health care including a case discussion between a paediatrician, the GP and the health visitor held in the family doctor's surgery, weekly visits from the health visitor and informal hospital admission. Significantly fewer sudden unexpected infant deaths occurred in this group than were expected by logistic regression anlysis or occurred in the best available control group with comparable scores ( p = 0.024). Problems in evaluation include identification of an adequate control population, ethical difficulties in introducing a controlled study when the programme is already perceived as effective, and the calculation of "expected death rates". The results of this study indicate that very energetic programmes of intervention may prevent some deaths in vulnerable infants.     

Outcome of congenital lung abnormalities detected antenatally

To determine the outcome of congenital lung abnormalities, data were collected retrospectively between January 1991 and December 1996 on any foetus found to have a lung lesion on antenatal ultrasound. A total of 23 foetuses had lung lesions on antenatal ultrasound. In two foetuses the antenatal ultrasound showed bilateral enlarged "bright" echogenic lungs with evidence of hydrops. Both pregnancies were terminated and tracheal atresia was confirmed. In 15 foetuses the antenatal ultrasound appearance was of a unilateral "bright" echogenic lung. There was one case of bronchial atresia and two cases of congenital lobar emphysema, which all had surgery. In nine cases there was a reduction in the size of the lesion on serial antenatal ultrasounds and no lesion was detected after birth. In three cases a small lesion was present after birth on chest radiography. In six foetuses the antenatal ultrasound appearance was of unilateral cystic or mixed cystic and echogenic lung lesions. Two pregnancies were terminated; both had congenital cystic adenomatoid malformation. Four pregnancies were continued and three infants had surgery soon after birth and were confirmed to have had congenital cystic adenomatoid malformation. One infant has been managed conservatively. In conclusion, a definitive diagnosis cannot usually be made antenatally. A large lesion on initial scan does not necessarily predict a poor outcome. The natural history of small asymptomatic postnatal lesions is unknown and a long-term prospective study is needed to determine the outcome of these lesions.     

Hyperemic and ischemic problems of surgical treatment of arteriovenous malformations.

Three patients with arteriovenous malformations are described who showed signs of massive hyperemia in the vascular territory of the normal brain proximal to arterial ligation. One additional patient had evidence of ischemia of the brain in the territory distal to ligation (steal), and in another both mechanisms were considered as operative hazards.     

Microvascular embolization of cerebral arteriovenous malformations: A technical variation.

A useful variation of an established technique is described for embolization of cerebral arteriovenous malformations. Silastic sponge emboli that fit into No. 16, 17, and 18 stub adapters are passed through standard-sized transfemoral catheters. Of 28 treated patients, obliteration was regarded as very successful in 16. Partial success was achieved in four. Eight were regarded as failures because the reticulum was too large for these microemboli.