Differential expression of p53 gene family members p63 and p73 in head and neck squamous tumorigenesis

p73 and p63 are recently cloned genes that share considerable structural and functional homologies with the p53 tumor suppressor gene. These genes, unlike p53, express multiple mRNA isoforms with variable biologic functions, and their suppressor nature has yet to be confirmed. To determine the interrelationship between these genes in the tumorigenesis of head and neck squamous carcinoma (HNSC), we performed immunohistochemical analyses of their protein products and compared the data with clinicopathologic parameters in 38 patients. In histologically normal epithelium, p53 and p73 showed similar basal and/or parabasal expression, but that of p53 was weaker and discontinuous. p63 staining was noted in more suprabasal cellular layers and was stronger. In dysplasias, all three markers manifested variable but gradual increase in extent and intensity of cellular expression with histologic progression. In carcinomas, p63 was the most frequently expressed (94.7%), followed by p73 (68.4%) and p53 (52.6%). Significant statistical correlation was noted only between p63 and p73 expressions (P =.04). Although no statistical correlation was found between p53 and p63 or p73, p53-negative tumors overexpressed either p63 or p73. p73 expression was associated with distant metastasis and perineural/vascular invasion. Our study indicates that (1) p63 and p73 expression may represent an early event in HNSC tumorigenesis, (2) the lack of correlation between p73 or p63 and p53 expression suggests an independent and/or compensatory functional role, (3) p73 expression may play a part in HNSC progression, and (4) p73 and p63 may function as oncogenes in the development of these tumors.     

HLA antigens in ulcerative colitis: a study in the Sicilian population

HLA antigens were investigated in 41 Sicilian patients with ulcerative colitis and in 151 healthy controls. Frequencies of HLA-B5 and DR2 were increased in the group of patients with ulcerative colitis whereas the DR3 antigen frequency was decreased. However the corrected p values were not significant. Thus, present results indicate that in ulcerative colitis HLA linked genetic factors play a marginal role, if any.     

Primary osteoblasts response to shock wave therapy using different parameters

Over the past decade extracorporeal shock-wave therapy (ESWT) has been increasingly applied to orthopaedic and musculoskeletal pathologies, the aim of this study was to assess how the energy density of the shock waves and the number of impulses affect viability, differentiation and synthetic activity of osteoblasts. Primary sheep osteoblasts cultures were treated with ESWT with an electro-hydraulic shock wave generator by selecting three different energy levels (14-21-28 kV corresponding at 0.15-0.31-0.40 mJ/mm2) and two different total numbers of impulses (500, 1000) for each level. At the end of treatment, cell counts and viability were recorded. Cells were then cultivated for 48 hours starting from a concentration of 1 x 10(4) cells/ml. The biological activity and viability were evaluated at 24 and 48 hours after treatment. No cytodestructive effects were observed in Group A, while a cytodestructive effect of ESWT was seen in cultures receiving the highest energy treatments. The different shock wave treatment induced differences in MTT assays after 24 and 48 hours, in particular the highest level showed a detrimental effect on cell respiration at both experimental times as compared to the Control Group and the protein metabolism was generally depressed by ESWT with impulses at the highest energy level. After 24 hours such effect further increased with the growing number of impulses. The lowest energy level appeared to significantly improve the metabolic parameter in primary cell cultures as compared to controls when 500 impulses were selected. The current study has demonstrated that one of the most important aspects to be considered is not the total number of impulses used but the energy level of the shock waves, thus confirming that ESWT has a dose-dependent effect on cells.     

B-1 cells are pivotal for in vivo inflammatory giant cell formation

The mechanisms that govern giant cell (GC) formation in inflammatory, neoplastic and physiologic conditions are far from being understood. Here, we demonstrate that B-1 cells are essential for foreign-body GC formation in the mouse. GCs were analysed on the surface of glass cover slips implanted into the subcutaneous tissue of the animals. It was demonstrated that GCs are almost absent on cover slips implanted into the subcutaneous tissue of BALB/c or CBA/N X-linked immunodeficient mice. As these animals do not have B-1 cells in the peritoneal cavity, they were reconstituted with B-1 cells obtained from cultures of adherent mouse peritoneal cells. Results showed that in B-1-reconstituted animals, the number of GCs on the implant surface surpassed the values obtained with preparations from wild animals. In animals selectively irradiated (pleural and peritoneal cavities) to deplete these cavities of B-1 cells, GCs were also not formed. Enriched suspensions of B-1 cells grown in culture were labelled with [(3)H]-tymidine and injected into the peritoneal cavity of naive mice before implantation of glass cover slips. After 4 days, about 17% of mononuclear cells had their nuclei labelled, and almost 70% of GCs had one or more of their nuclei labelled when analysed by histoautoradiographic technique. A few GCs expressed an immunoglobulin M when analysed by immunostaining and confocal microscopy. Overall, these data demonstrate that B-1 cells are pivotal in the mechanisms of foreign-body GC formation in the mouse.     

Microfilament system in the microvascular endothelium of the palmar fascia affected by mechanical stress applied from outside

The effect of externally applied mechanical stress was investigated by thin section electron microscopy of the microvessels in the unaffected palmar fascia in the carpal tunnel syndrome and in patients with Dupuytren's contracture before and after application of a continuous elongation device. In the unaffected palmar fascia the microfilaments of the endothelial cells were connected to a few adherens junctions and focal contacts; stress fibres were absent. In the cord of Dupuytren's disease the microfilaments were increased in quantity. The length ratios of the connections with the lateral and basal cell membrane were significantly higher than in the control group and increased to an even greater extent in the continuously extended fascia. Stress fibres appeared in the endothelial cells of postcapillary venules in the nonextended cord and in the endothelium of both arterioles and venules after extension elongation. The numerous intermediate filaments and the rare microtubules remained unchanged in the endothelial cells of all palmar fasciae analysed. In the endothelial cells of the microvessels the mechanical stress applied from outside mainly affected the contractile component of the cytoskeleton.     

Cell proliferation in breast cancer is a major determinant of clinical outcome in node-positive but not in node-negative patients.

The growth rate of a tumor cell population depends on two major factors: the percentage of proliferating cells (cell growth fraction) and the rapidity of their duplication (cell proliferation rate). The authors evaluated the prognostic and predictive value of both kinetics parameters in a large series of breast cancer patients (n=504). The cell growth fraction was determined by MIB-1 immunostaining, the cell proliferation rate by AgNOR analysis. Ki-67 LI (labeling index) and AgNOR area were significantly associated with histotype, histologic grade, tumor size, estrogen/progesterone receptor status, patient age, and lymph node involvement (P<0.005). In the entire series of patients, both kinetics variables were significantly and independently associated with the clinical outcome, but their prognostic relevance was quite different when node-negative and node-positive patients were considered separately. Although in node-positive patients Ki-67 LI and AgNOR area were the unique independent predictors of disease-free and overall survival, they were excluded by the multivariate Cox model in node-negative patients, where only tumor size and estrogen receptor status retained a significant P-value. These results show that in breast carcinoma the cell growth fraction and the cell proliferation rate have a different prognostic impact with respect to the lymph node status and are major determinants of clinical outcome in node-positive patients only. Within this subgroup, the rapidity of cell proliferation as assessed by AgNOR analysis also served as a sensitive predictor of the response to adjuvant treatments.     

The housekeeping gene xanthine oxidoreductase is necessary for milk fat droplet enveloping and secretion: gene sharing in the lactating mammary gland

Xanthine oxidoreductase (XOR) is the rate-limiting enzyme in purine catabolism occurring in most cell types. However, this housekeeping gene is expressed at very high levels in a number of mammalian tissues including the lactating mammary epithelium, suggesting additional roles for XOR in these tissues. Mice with targeted disruption of XOR were generated to assess these potential additional roles. XOR-/- mice are runted and do not live beyond 6 wk of age. Strikingly, however, XOR+/- females, although of healthy appearance and normal fertility, are unable to maintain lactation and their pups die of starvation 2 wk postpartum. Histological and whole-mount analyses showed that in XOR+/- females the mammary epithelium collapses, resulting in premature involution of the mammary gland. Electron microscopy showed that XOR is specifically required for enveloping milk fat droplets with the apical plasma membrane prior to secretion from the lactating mammary gland. We present evidence that XOR may have primarily a structural role, as a membrane-associated protein, in milk fat droplet secretion and thus XOR provides another example of "gene sharing". About 5% of women experience primary lactation insufficiency. The above observations suggest that human females suffering from xanthinuria, a deficiency in XOR, are potential candidates for lactation problems.     

Inter-relationships between pituitary-adrenal hormones and catecholamines during a 6-day Nordic ski race

Summary The aim of the study was to investigate the inter-relationships between pituitary-adrenal hormones and catecholamines during a prolonged competition over 6 days. Plasma adrenocorticotropic hormone (ACTH), cortisol (C), -endorphin (EP), free and sulphated adrenaline (A) and noradrenaline (NA) were measured in 11 volunteer male subjects during a national Nordic-ski race (323 km). Blood samples were obtained before the competition in the evening as control (D₀), and before and after each day's racing (D₁–D₆). The mean daily heart rate (f _c) was calculated fromf _c values recorded every minute during the race. The results showed the following: changes in meanf _c [from 147 (SEM 3) to 156 (SEM 3) beats · min^–1 according to the day] were not significant during the race. Diurnal variations in ACTH, EP and C were no longer apparent after the race: evening levels were higher than their respective D₀ values during the race, except on D₃ when there was a lack of response to exercise in the three hormones. Unlike ACTH and EP, pre- and postexercise C values on D₁ and D₂ were higher than those on the subsequent days (P<0.001). In contrast, there was a progressive accumulation of A and NA in pre-and postrace concentrations which reached a plateau in about 4 days. Positive correlations between exercise responses in ACTH, C and EP were found especially on D₃ and D₆ (P<0.001) but there were no significant correlations between catecholamines and the other three hormones. Thus, prolonged competition over 6 days evoked different control mechanisms for hormones of the pituitary-adrenal axis and catecholamines. A sustained catecholamine release and sympathetic activation induced a progressive NA and A accumulation during the race. In contrast, the lack of a response to exercise in ACTH, EP and C on D₃ suggested a dissociated central command for pituitary axis hormones and sympathetic adrenal activation. On the following days, the response to a lack of exercise, in spite of ACTH stimulation, may have reflected an adaptation of adrenal glands to prolonged stress.     

Reactivation of Epstein-Barr virus in Sj?gren''s syndrome

Summary Sjögren's syndrome (SS) is a chronic autoimmune disease characterized by severe dryness of the eyes and mouth, resulting from lymphocytic infiltration of the lacrimal and salivary glands. SS may exist as a primary condition (primary SS, 1° SS) or as a secondary condition (2° SS) in association with rheumatoid arthritis, systemic lupus erythematosus, or progressive systemic sclerosis. In some 1° SS patients, there may be involvement of the extraglandular organs, including skin, kidney, liver, lung and nervous system. Furthermore, these patients may develop a lymphoproliferative syndrome that includes lymphadenopathy and increased risk of lymphoma. In the pathogenesis of SS, a role for Epstein-Barr virus (EBV) has been suggested because: (a) EBV is present in salivary gland epithelial cells of normal individuals and exaggerated immune responses against EBV could play a role in the destruction of salivary glands in SS; (b) SS salivary gland biopsies contain increased levels of EBV DNA in comparison to normal salivary glands, indicating viral reactivation and inability of lymphoid infiltrates to control EBV replication in SS patients; and (c) salivary gland epithelial cells in SS patients express high levels of HLA-DR antigens and may present EBV-associated antigens to immune T cells in SS patients. Therefore, SS may represent a situation in which genetically predisposed individuals (i. e., HLA-DR3-DQA4-DQB2) have a persistent but ineffectual T cell immune response against EBV at its site of latency. Among 14 non-Hodgkin's lymphomas that developed in SS patients, EBV DNA was detected in increased amounts in the tumor tissue of one patient. Characterization of this tumor DNA revealed: (a) polyclonal immunoglobulin gene rearrangements; (b) EBV DNA with an unusual restriction fragment length polymorphism pattern involving the Bam M fragment; and (c) EBV terminal repeat sequences suggestive of viral replication, similar to those reported in EBV lymphomas occurring in other immunocompromised individuals. Early recognition of this clinical problem may allow beneficial use of antiviral agents.