The latent membrane protein-1 in Epstein-Barr virus-transformed lymphoblastoid cells is found with ubiquitin-protein conjugates and heat-shock protein 70 in lysosomes oriented around the microtubule organizing centre.

Immunofluorescence studies on Epstein-Barr virus (EBV)-transformed lymphoblastoid cells have previously shown that the latent membrane transforming protein (LMP-1) is found in patch-like inclusions which also immunostain for vimentin. We now show that EBV transformation causes a major reorganization of intermediate filaments, microtubules, mitochondria, and lysosomal elements, which generally become oriented around the microtubule organizing centre. Immunogold electron microscopy shows that LMP-1 is primarily concentrated in secondary lysosomes together with ubiquitin-protein conjugates and heat-shock protein 70. Intermediate filament inclusion formation with the above characteristics may be a general response triggered by other membrane glycoproteins; as seen, for example, in major human neurodegenerative diseases such as diffuse Lewy body disease.     

Immunogenic and antigenic epitopes of immunoglobulins--VII. The topographical distribution of Fc gamma epitopes and the relationship of an iso-allotypic specificity to the presence of histidine 435

Epitopes recognised by a panel of 23 anti-Fc gamma monoclonal antibodies (McAbs) have been subdivided into three groups each having a distinct topographical distribution. One group of mutually inhibitory McAbs are reactive with epitopes expressed on the fy "surface" of the C gamma 2 domain. A second group recognises epitopes in the region of arginine 355 of the C gamma 3 domain whilst the third group recognises epitopes expressed in the inter C gamma 2/C gamma 3 domain region--as evidenced by inhibition of Staphylococcus aureus protein A binding. An antibody of the latter group reactive with IgG1, 2, 4 and IgG3m(15,16) proteins but not IgG3m(5) or IgG3m(21) proteins allows histidine 435 to be identified as a critical residue for expression of the epitope recognised by this antibody.     

Apoptosis of malignant cells in Hodgkin's disease is related to expression of the cdk inhibitor p27KIP1

Previous results from B-cell chronic lymphocytic leukaemia suggest that expression of p27KIP1 might be important in protection from apoptosis. Given the relevance of apoptosis to the pathogenesis of Hodgkin's disease (HD), it was decided to examine the expression of p27KIP1 in relation to apoptosis in these lesions. Paraffin-wax sections from a total of 65 histologically confirmed HD tumours were used to derive apoptotic index (AI) and DNA fragmentation index (DFI) scores, which were compared with the expression of various cell-cycle-regulating proteins, including p27KIP1 (p27), p21WAF1/CIP1 (p21) and cyclin D1, and with Epstein-Barr virus (EBV) status. The DFI was measured by TdT-mediated dUTP-FITC nick end-labelling (TUNEL), and the AI by conventional morphology. Cells showing the typical morphology of apoptosis, together with those resembling so-called 'mummified' Hodgkin/Reed-Sternberg (HRS) cells, were included in AI measurements. Increasing numbers of p27-positive HRS cells were associated with lower levels of apoptosis in these cells, as indicated by significantly lower AI and DFI scores. There was a trend towards poorer survival in those patients with the highest numbers of p27-positive HRS cells and with lower AI and DFI scores, but these differences were not statistically significant. p21-positive HRS cells were significantly more frequent in those cases with lower AI scores. A similar trend was observed for p21 and DFI, although this relationship was not statistically significant. There was also a trend towards higher levels of cyclin D1 protein in HD cases with high AI and DFI values. A tendency for increasing numbers of p27-positive and p21-positive HRS cells in EBV-positive cases was noted, but this relationship was not statistically significant. EBV status did not correlate with either AI or DFI scores. The results of this study suggest that p27, and possibly also p21, may be involved in protection from apoptosis in HD.     

Separation distress and attachment in surrogate-reared squirrel monkeys.

Surrogate-reared infant squirrel monkeys were exposed to various conditions of separation from their surrogate. Infants showed significant increases in plasma levels of cortisol when they were placed in an unfamiliar environment during the separation period. Changes in behavior, but not cortisol, were observed under conditions in which the surrogate was removed and the infant left in the home cage. These results differ from those previously obtained with mother-reared infants. It is concluded that surrogate-reared infant squirrel monkeys do not show the same separation response or attachment to their rearing figure as do mother-reared infants.     

Numerical and structural chromosomal abnormalities detected in human sperm with a combination of multicolor FISH assays

A pair of multicolor FISH assays (X-Y-21 and A-M-16) was developed for human sperm to simultaneously measure sex ratios; aneuploidies involving chromosomes 1, 16, 21, X, and Y; meiotic diploidies; and structural aberrations involving chromosome 1p. Sex ratios in sperm were not significantly different from unity among healthy men. Baseline frequencies of disomic sperm for chromosomes 1, 8, and 21 were similar (6.7 per 10(4) sperm, 95% CI of 5.6-8.1), suggesting that among these three chromosomes, chromosome 21 was not especially prone to nondisjunction. Frequencies of disomy 16 sperm were significantly lower, however (3.5 per 10(4) sperm, 95% CI of 2.0-6.2; P < 0.02). The baseline frequencies of sperm disomy by FISH for chromosomes 16 and 21 were validated against aneuploidy data obtained by the hamster-egg technique for human sperm cytogenetics. The frequencies of X-X, Y-Y, X-Y ("Klinefelter") sperm and sex-null ("Turner") sperm were 5.5, 5.1, 5.5, and 7.8 per 10(4) sperm, respectively. For chromosomes 16 and 21, the frequencies of nullisomic and disomic sperm were similar, suggesting that gain and loss events occurred symmetrically. However, more gain than loss was reported for chromosomes 1, X, and Y. The frequency of MI and MII diploid sperm (with flagella) was approximately 12 per 10(4) (range 8.3-16.7 per 10(4) sperm). Based on flagella data, the frequency of somatic cells in the semen was estimated to be approximately 1.8 per 10(4) sperm. Loss or gain of a portion of chromosome-arm 1p occurred in 5.5 per 10(4) sperm, and the percentage of sperm carrying structural aberrations within the haploid genome as calculated from FISH (1.4%), was similar to that obtained with the hamster-egg technique. These complementary sperm FISH assays have promising applications in studies of chromosomally abnormal sperm after exposure to occupational, medical, and environmental toxicants.     

Relation between high energy phosphate and lethal injury in myocardial ischemia in the dog.

The relationship between progressive depletion of high energy phosphate and the onset of lethal cell injury in ischemic myocardium following coronary occlusion has been evaluated. Myocardial ischemia was induced by proximal occlusion of the circumflex coronary artery for 15, 30, 40, or 60 minutes. Cell injury in the severely ischemic posterior papillary muscle (PP) was evaluated by electron microscopy and by measuring the capacity of slices of the injured PP to maintain electrolytes, resynthesize high energy phosphate, and exclude inulin during in vitro incubation. ATP content in the ischemic myocardium decreased to 35%, 9%, 7%, and 5% of control values after 15, 30, 40, and 60 minutes of ischemia, respectively, and was associated with a corresponding depletion of total adenine nucleotides. The loss of 65% of the ATP after 15 minutes of ischemia (reversible injury) was associated with only minimal ultrastructural changes and no significant defects of electrolytes in incubated slices. However, the depletion of over 90% of the ATP after 40 minutes of ischemia (irreversible injury) was associated with significant fine structural changes and markedly altered cell volume regulation. The results suggest a close relationship between the marked depletion of high energy phsophates and the development of lethal injury in acutely ischemic myocardium.     

Functional characterization of peripheral blood lymphocytes in chronic HBsAg carriers

Increased proportions of suppressor/cytotoxic T cells have been identified in the peripheral blood of chronic HBsAg carriers and to investigate a possible relationship to T cell cytotoxicity against autologous hepatocytes, suppressor cell activity, viral replication and the histological type of disease, 42 consecutive HBsAg carriers undergoing a liver biopsy have been investigated. The proportion of suppressor/cytotoxic lymphocytes directly correlated with T cell cytotoxicity to autologous hepatocytes and both were higher in those with HBeAg in serum than in those with anti-HBe or those on corticosteroid therapy. There was no relationship to underlying histological classification. In contrast, suppressor cell regulation of IgG producing cells was unrelated to the proportion of suppressor/cytotoxic lymphocytes in peripheral blood or HBeAg status, but impaired function was associated with chronic hepatitis, particularly chronic active hepatitis. These data suggest that the increased proportion of suppressor/cytotoxic lymphocytes in the peripheral blood of HBsAg carriers represents an increase in the cytotoxic and not the suppressor cell subset and that this is a consequence of active viral replication and not of the severity of hepatic inflammation. Defective suppressor cell function may be one factor in the development of chronic active hepatitis, but is not reflected by alterations in the T4:T8 ratio.