Storage pool deficiency in cattle with the Chediak-Higashi syndrome results from an absence of dense granule precursors in their megakaryocytes

Platelets from cattle with the Chediak-Higashi syndrome (CHS) have a storage pool deficiency and virtual absence of platelet dense granules. Megakaryocytes (MKs) from five control (n = 135) and five CHS (n = 133) cattle were evaluated using standard transmission electron microscopy. Osmiophilic dense granules were not observed in control or CHS MKs. In MKs from normal cattle, clear vesicles of 200- to 650-nm diameter bounded by a sharp membrane were observed. They were easily differentiated from the demarcation membrane system, endoplasmic reticulum, and alpha granules. The clear vesicles were virtually absent in MKs from CHS cattle at all stages of maturation. MKs in bone marrow samples from two control (n = 91) and two CHS (n = 61) cattle that had been processed for the uranaffin reaction were also evaluated. The clear vesicles were replaced by uranaffin-positive granules in MKs from control cattle, but positive uranaffin granules were not observed in CHS MKs. These findings indicate that the platelet dense granule storage pool deficiency in CHS cattle results from an anatomic absence of dense granule precursors in maturing and mature CHS MKs.     

Diadenosine 5',5'-p1,p4-tetraphosphate deficiency in blood platelets of the Chediak-Higashi syndrome

Diadenosine tetraphosphate (AP4A) is an unusual nucleotide found in a variety of cells, including platelets. It has been suggested that platelet AP4A is stored in the dense granules and is metabolically inactive. We have studied the AP4A content of blood platelets in two patients and three cattle with Chediak-Higashi syndrome (CHS), a hereditary platelet defect with dense granule deficiency. Acid-soluble extractions of whole blood and platelets were neutralized. The adenosine triphosphate (ATP) level was measured by luminescence technique. To measure the AP4A content, the neutralized extract was treated with phosphomonoesterase for removal of ATP. The AP4A content was then measured by coupling the phosphodiesterase and luciferase reaction. The AP4A content was 0.43 nmol/mg protein for normal human platelets and 0.004 nmol/mg protein for CHS platelets. The ATP/AP4A ratio was 67 for normal and 3,023 for CHS platelets. The whole blood AP4A was reduced by 89% in CHS patients who had only a slight decrease in ATP level (26% reduction). Similarly, bovine platelets with CHS showed a marked decrease of AP4A content and a moderate reduction of the ATP level. The platelet ATP/AP4A ratio was 351 and 3,133 for normal and CHS cattle, respectively. Results demonstrate a marked reduction of AP4A in CHS platelets and suggest that AP4A may be a useful marker for the measurement of dense granule content in platelets.     

Induction of mutations in Ki-ras and INK4a in liver tumors of mice exposed in utero to 3-methylcholanthrene

An understanding of the basic mechanisms responsible for the pathogenesis of liver neoplasms is needed in order to develop better therapeutic strategies. The present study utilized a pharmacogenetic mouse model to assess the role of cytochrome P4501A1 (Cyp1a1) in modulating genetic damage to oncogenic and tumor suppressor loci following in utero exposure to the polycyclic aromatic hydrocarbon, 3- methylcholanthrene (MC). Analysis of the Ha-ras, Ki-ras, INK4a and p53 genes was carried out with lysates from paraffin-embedded liver tissue from transplacentally-treated mice. The lysates were subjected to DNA amplification by the PCR technique followed by allele-specific oligonucleotide hybridization screening and SSCP analysis. All of the 26 neoplasms screened (23 hepatocellular carcinomas, two hepatocellular adenomas and one sarcoma) exhibited a GGC-->CGC (GLY13-->ARG13) transversion at the Ki-ras gene locus. None of the tumors had Ki-ras mutations at codon 12 of exon 1. Approximately 12% (3/26) of the liver tumors exhibited point mutations in exon 1 of the INK4a gene, with each of the three tumors exhibiting two point mutations. Analysis of exon 2 of the INK4a gene showed the presence of a CCG-->CTG (PRO73-->LEU73) transition in two of the 26 neoplasms. No mutations were found in exons 1 or 2 of the Ha-ras gene, or in exons 5-8 of the p53 gene. Analysis of tumor RNAs showed overexpression of Ha-ras, cip1 and c-jun in approximately 38% of the liver tumor samples. The results of this study suggest that mutagenic damage to oncogenes and tumor suppressor genes may be critical factors in mediating transplacentally-induced liver tumorigenesis. The fact that Ki-ras mutations were found in all of the tumors suggests that mutation at this gene locus may be an early event in liver tumor pathogenesis, while mutation in tumor suppressor genes may occur later during tumor progression. These combined results are consistent with the pathogenesis of cancer in humans.      

The detection by enzyme-linked immunosorbent assays of non-complement- fixing HLA antibodies in transfusion medicine

BACKGROUND: Noncomplement-fixing white cell antibodies have been demonstrated by the use of immunofluorescence flow cytometry against intact lymphocytes. However, such antibodies may be either HLA-specific or directed against other white cell antigens. Commercial enzyme-linked immunosorbent assay (ELISA) kits, using solubilized HLA molecules as targets, enable such HLA-specific antibodies to be detected in patients who are refractory to platelet transfusion, patients experiencing febrile transfusion reactions, and patients whose sera give nonspecific hemagglutination in indirect antiglobulin tests. STUDY DESIGN AND METHODS: Sera from all three groups of patients, previously screened for cytotoxic antibodies by using complement-dependent lymphocytotoxicity, were re-investigated with commercial ELISA kits for HLA antibody screening and identification using the manufacturers' recommended test methods. RESULTS: Non-complement fixing HLA antibodies were detected by ELISA in many sera that were lymphocytotoxicity test- negative; that is, 14 (17.5%) of 80 from refractory patients, 8 (23.5%) of 34 from those with febrile reactions, and 11 (22.4%) of 49 from those with nonspecific hemagglutination in the direct antiglobulin test. However, not all cytotoxic white cell antibodies were detectable by ELISA: only 19 (82.6%) of 23, 19 (67.8%) of 28, and 11 (73.6%) of 49, respectively in the three groups. Similarly, only 143 (79.4%) of 181 cytotoxic sera with clear-cut HLA-A or -B locus specificities were detectable by ELISA. CONCLUSION: ELISAs detect some but not all clinically significant HLA antibodies, irrespective of their ability to fix complement in vitro.     

Active MR visualization of a vascular guidewire in vivo

The purpose of this study was development of an actively visualized .035-inch vascular guidewire for use in MR-guided interventions. The guidewire was actively visualized by inclusion of a 6-cm-long radiofrequency coil in its tip. A high contrast outline of the distal tip of the guidewire was obtained by acquiring an image with the radiofrequency coil as the receiving antenna. The position of the guidewire relative to the surrounding anatomy was determined by overlaying the guidewire image on a previously acquired road map. The guidewire was evaluated in vivo in the abdominal vessels of a rabbit and swine at 1.5 T. The built-in radiofrequency coil delivered a high contrast signal over its full length, enabling visualization of the position and curvature of the tip of the guidewire. The ability to see the curvature of the guidewire over several centimeters significantly eased manipulation into targeted vessels and represents an important advance toward MR-guided vascular interventions.     

MRI of absent left pulmonary artery.

Unilateral absence of a pulmonary artery, more accurately referred to as unilateral proximal interruption of a pulmonary artery, is a rare congenital anomaly that may occur as an isolated lesion or in association with other congenital cardiovascular abnormalities. Diagnosis of associated lesions is imperative as early detection and intervention may significantly improve the patient's prognosis. We present the case of an adult patient who had come to our attention after suffering neurological decompression illness related to scuba diving. The patient's cardiopulmonary anatomy was evaluated using MRI gated spin echo, cine, and breath-held fast spoiled recalled echo sequences.     

Normal myocardial perfwsion assessed with multishot echo-planar imaging

A new magnetic resonance imaging strategy is presented for accessing myocardial perfusion. Most previous work has relied on using T₁-weighted fast gradient-echo imaging to monitor dynamically the signal changes during the passage of a contrast media bolus. However, the gradient-echo approach is limited by an inability to image the entire heart with adequate temporal resolution. This paper focuses on a electrocardiogram-gated multishot echo-planar imaging sequence, using the simple strategy of using the intrinsic T₁ weighting produced by a repetition time equal to the heart period. To quantitate the sequence's performance with respect to normal myocardial perfusion, seven volunteers were imaged, each with three different doses of the contrast medium gadolinium diethylenetriamine penta-acetic acid (Gd-DTPA). The first-pass dynamics of the contrast were quantified in 13 regions per heart for each examination. In all volunteers, the complete heart could be covered, with five to seven slices, every two heartbeats. Enhancement was homogeneous throughout the left ventricular myocardium, with an enhancement of approximately 50% for the optimum contrast dose of 0.05 mmol/kg Gd-DTPA.     

On postural stability and variability

The stability of posture is often assessed directly by the variability of certain centre of pressure parameters, such as length or area of the centre of pressure profiles. Increased variability of these parameters is usually taken as an index of decreased postural stability. In this paper we present data from normal and tardive dyskinetic adult subjects that support the notion that centre of pressure variability is not a sufficient measure of postural stability. It is shown that variability in a given centre of pressure parameter does not necessarily distinguish the different attractor dynamics that support postural control. Dimension estimates of the centre of pressure time series showed that: (1) there is more structure in the centre of pressure pattern of normal subjects than is traditionally interpreted; (2) the dimension of the centre of pressure in tardive dyskinetic individuals is systematically lower than in normals. The implication of these findings for assessing the relation between the variability and stability of posture is discussed. It is proposed that postural stability can only be assessed by considering the attractor dynamics of the postural control system.