肺腺癌细胞微环境及转移与黏附分子表达的关系

目的研究肺腺癌细胞生长环境及转移性与黏附分子CD44v6和CD29的表达关系。方法将起源相同、转移性不同的两个肺腺癌细胞系AGZY和Anip分别用简便肿瘤多细胞球体(MTS)培养法培养，并设常规单层贴壁细胞培养对照。通过倒置显微镜、扫描及透射电镜观察MTS形成情况，并用免疫组化法分别对MTS及贴壁细胞上CD44v6和CD29表达进行检测。结果MTS培养成功，贴壁细胞与MTS在细胞结构及细胞连接结构上相似，两种MTS在形态及结构上差异无显著性。免疫组化结果显示，CD29在高转移性的Anip细胞及其MTS上呈阳性表达；在低转移性的AGZY细胞及其MTS上阴性表达。CD44v6在Anip和AGZY细胞及MTS上均呈阳性表达，差异无显著性。贴壁细胞与MTS上两种黏附分子表达均无差异。结论成功建立了一种简易制备MTS的方法。细胞生长方式(单层贴壁与MTS)可能不影响CD44v6和CD29的表达。CD29表达可能与肺腺痛转移性相关；CD44v6表达可能与肺腺癌转移无关。     

A girl with 1p36 deletion syndrome and congenital fiber type disproportion myopathy

Chromosome 1p36 deletion syndrome is characterized by hypotonia, moderate to severe developmental and growth retardation, and characteristic craniofacial dysmorphism. Muscle hypotonia and delayed motor development are almost constant features of the syndrome. We report a 4-year-old Japanese girl with 1p36 deletion syndrome whose muscle pathology showed congenital fiber type disproportion (CFTD) myopathy. This is the first case report of 1p36 deletion associated with CFTD. This association may indicate that one of the CFTD loci is located at 1p36. Ski proto-oncogene −/− mice have phenotypes that resemble some of the features observed in patients with 1p36 deletion syndrome. Because fluorescent in situ hybridization analysis revealed that the human SKI gene is deleted in our patient, some genes in 1p36, including SKI proto-oncogene, may be involved in muscle hypotonia and delayed motor development in this syndrome. Received: March 4, 2002 / Accepted: July 7, 2002     

TGF-β1 Null Mutation Leads to CD154 Upregulated Expression in Affected Tissues

The TGF-1(–/–) mouse is a murine model for systemic autoimmune disease. The aim of this study is to elucidate the immunological mechanism that leads to multifocal tissue inflammation and autoantibody production in TGF-1(–/–) mice. Heart, lung, liver, and salivary gland from TGF-1(–/–) were assessed for CD154 expression by RT-PCR and immunohistochemistry. Compared to wild-type littermates, CD154 expression was elevated in all tissues studied. Furthermore, IL-12 mRNA was expressed in the salivary gland and heart of TGF-1(–/–) mice and not in wild-type littermates. This suggests that the CD154 pathway is activated in these tissues. This shows that TGF-1 regulates CD154 expression leading to spontaneous IL-12 production and autoimmunity.     

Enhancement of cALL immunogenicity by co-culture with a CD154 expressing 293 cell line

Pre-B cell acute lymphoblastic leukaemia (cALL) commonly occurs in young patients and although successful conventional therapies are available (such as cytotoxic drugs and bone marrow transplantation) for a proportion of patients (approximately 30%) these are ultimately unsuccessful. Recurrence of disease is a result of the failure of the immune system to recognize these abnormal cells and down-regulation of crucial molecules required for cognate CD4(+) T cell recognition has been postulated as a means of immune escape. In this study we show that an embryonic kidney cell line (293 cells) transfected with CD154 (40 L.1) are capable of not only maintaining the viability of primary ALL cells in culture but can also up-regulate the expression of a number of crucial molecules involved in antigen recognition. We show that 40 L.1 cell stimulation of primary ALL cell cultures can not only enhance the allogeneic and autologous MLR response to such cells but will also induce CTL effectors which are capable of lysing wild-type autologous ALL cells. It is therefore conceivable that such an approach could be used to generate an active anti-tumour response in patients, following conventional therapy, reducing the incidence of recurrence.     

Activation and internalization of p56lck upon CD45 triggering of Jurkat cells

Relationships between CD45 and p56^Ick have been suggested by co-immunoprecipitation of both proteins and by dephosphorylation of the p56^lck regulatory site, Tyr 505, by CD45 in vitro. We investigated whether the kinase activity of p56^lck is modulated in T cells triggered via CD45. We showed that incubation of Jurkat cells with a combination of two anti-CD45 monoclonal antibodies (mAb) (MC5/2 + D3/9) induced an increase in p56^lck kinase activity, while a single mAb did not. Under these conditions, p56^lck underwent two consecutive waves of activation. This was accompanied by internalization of the kinase and by a time-dependent increased accessibility of CD45 phosphatase at the plasma membrane. Similarly, activation and internalization of p56^lck were observed using a combination of anti-CD45 (MC5/2) and anti-CD2(T11₂) mAb, suggesting that a functional complex consisting of CD45, CD2 and p56^lck was formed upon cell triggering. Taken together, these results suggests that: (i) CD45 participates in the regulation of p56^lck kinase activity in vivo and that (ii) CD45 could play a mediator role in the stimulation and endocytosis of p56^lck through the CD2 pathway.     

Adjustment of metabolism,catecholamines and β-adrenoceptors to 90 min of cycle ergometry

Adrenaline infusion of 0.1 g · kg^–1 · min^–1 in healthy volunteers results in an increase of hepatic glucose production, an increase of the absolute number of occupied -adrenoceptors and specific changes in metabolism. To compare these effects with the changes induced by an endogenous catecholamine release, we investigated healthy volunteers during cycle ergometry. After fasting at least 14 h seven healthy subjects exercised for 90 min at an intensity of 20% below their individual anaerobic threshold. The rate of glucose production as well as the turnover rates of alanine and leucine were calculated using stable isotope tracers. High and low affinity -adrenergic binding sites on lymphocytes were determined by an equilibrium binding assay with (–)¹²⁵ Iodocyanopindolol. After 90 min of cycling the rate of appearance of glucose increased significantly from means of 2.0 (SD 0.2) to 2.65 (SD 0.50) mg · kg^–1 · min^–1 with unchanged blood concentrations of glucose and lactate. The flux of the amino acids alanine and leucine decreased significantly from means of 0.91 (SD 0.21) to 0.62 (SD 0.14) mg · kg^–1 · min^–1 and from 0.40 (SD 0.05) to 0.32(SD 0.04) mg · kg^–1 · min^–1, respectively. The mean free fatty acid concentration increased significantly from 0.65 (SD 0.33) to 1.27 (SD 0.45) mmol · l^–1 during the endurance trial. The increase of glucose turnover and the decrease of amino acid flux point to a metabolic shift towards enhanced utilization of free fatty acids. Adrenaline and noradrenaline concentrations showed a moderate but significant increase from means of 0.61 (SD 0.20) to 0.99 (SD 0.36) nmol · l^–1 and from 2.27 (SD 0.75) to 3.46 (SD 0.38) nmol · 1^–1, respectively. The number of high affinity -adrenergic binding sites per cell (-adrenoceptors) nearly doubled from 770 (SD 130) to 1490 (SD 150) during 90 min of cycling. The observed endogenous plasma catecholamine concentrations were not sufficient to change significantly the relative receptor occupancy. This would seem to indicate that the aerobic exercise induced effects depended more on the absolute number of occupied -adrenoceptors than on their relative receptor occupancy. When compared to the results of the adrenaline infusion experiment the increases of the hepatic glucose production and the increase of -adrenoceptors were very similar in both groups despite ten times higher adrenaline plasma concentrations in the infusion group. This would seem to indicate that -adrenoceptors mediated effects do not correlate with catecholamine plasma concentrations.     

Variability of house-dust-mite allergen levels within carpets

Sensitization and exposure to house-dust-mite allergens is an important cause of asthma. Standardized, reliable, and reproducible methods for measuring exposure are essential for the assessment of the relationship between exposure, sensitization, and asthma. This study investigated the variability of the house-dust-mite allergen Der p 1 concentration in reservoir dust collected within whole carpets in living rooms and bedrooms. The carpets of nine bedrooms and 11 living rooms were sampled. Each room was divided into 1 m² areas measured from wall to wall where the carpet was accessible. Reservoir dust samples were collected by vacuuming each 1 m² area for 2 min. Der p 1 was assayed by a two-site monoclonal-antibody-based immunometric ELISA. Der p 1 was detectable in the carpets of nine bedrooms and six of the 11 living rooms. Within these 15 rooms, there was a wide range of Der p 1 levels. The smallest range of allergen within single room was 0.9 μg Der p 1/g dust (0.2 and 1.1 ng/g; 5.5-fold difference), and the largest was 149.2 μg Der p 1/g dust (0.8 and 150 μg/g; 192-fold difference). The mean range of allergen levels in the living rooms was 11.5 jig Der p 1/g of dust, and the mean coefficient of variation of these rooms was 80.2%. illustrating the huge variation of mite allergen levels within each room. The variation within bedrooms was also large, with a mean coefficient of variation value of 88.7%. The coefficient of variation was significantly lower around soft furnishings or beds (57%) than in the rest the room (89.3%), with the mean difference being 32% (95% CI 27ndash;63%; P=0.04). In conclusion, this study has shown that there is a great variation of Der p 1 levels between areas within a room. No consistent pattern of distribution of mite allergen within a room was found. Der p 1 levels in areas around soft furnishings and beds varied less than the levels in the rest of room.     

Chromosome neighborhood composition determines translocation outcomes after exposure to high-dose radiation in primary cells

Radiation exposure is an occupational hazard for military personnel, some health care professionals, airport security screeners, and medical patients, with some individuals at risk for acute, high-dose exposures. Therefore, the biological effects of radiation, especially the potential for chromosome damage, are major occupational and health concerns. However, the biophysical mechanisms of chromosome instability subsequent to radiation-induced DNA damage are poorly understood. It is clear that interphase chromosomes occupy discrete structural and functional subnuclear domains, termed chromosome territories (CT), which may be organized into ‘neighborhoods’ comprising groups of specific CTs. We directly evaluated the relationship between chromosome positioning, neighborhood composition, and translocation partner choice in primary lymphocytes, using a cell-based system in which we could induce multiple, concentrated DNA breaks via high-dose irradiation. We critically evaluated mis-rejoining profiles and tested whether breaks occurring nearby were more likely to fuse than breaks occurring at a distance. We show that CT neighborhoods comprise heterologous chromosomes, within which inter-CT distances directly relate to translocation partner choice. These findings demonstrate that interphase chromosome arrangement is a principal factor in genomic instability outcomes in primary lymphocytes, providing a structural context for understanding the biological effects of radiation exposure, and the molecular etiology of tumor-specific translocation patterns.     

COX-2+ myofibroblasts may play a key role in marked stromal fibrosis in strictured colorectal carcinomas

AIMS: To elucidate the mechanism of marked stromal fibrosis in strictured colorectal carcinomas (SC) that cause complete ileus. METHODS AND RESULTS: Sixteen cases of SC and 29 cases of non-strictured colorectal carcinoma (NSC) were studied. These carcinomas showed similar clinicopathological features except for bowel stricture. The stricture index (SI) showing the degree of bowel stricture was 59.8 +/- 12.1% in SC versus 20.8 +/- 24.6% in NSC (P < 0.001). The fibrosis index (FI), defined to reflect the extent of stromal fibrosis, was 56.3 +/- 8.8% in SC versus 21.9 +/- 10.6% in NSC (P < 0.001). COX-2+ myofibroblasts were detected in 13 cases (81.3%) in SC versus eight cases (27.6%) in NSC (P < 0.01). The COX-2+ myofibroblast density was 276.7 +/- 181.1 cells/mm(2) in SC versus 26.6 +/- 52.7 cells/mm(2) in NSC (P < 0.001). When all cases were divided into two groups with and without COX-2+ myofibroblasts, the SI was 48.8 +/- 19.1% in those with COX-2+ myofibroblasts versus 24.8 +/- 29.3% in those with COX-2- myofibroblasts (P < 0.001). CONCLUSION: COX-2+ myofibroblasts may play an important role in extensive bowel stricture in colorectal carcinomas.     

Strain-Dependent Migration of CD4 and CD8 Lymphocyte Subsets to Lymph Nodes in NOD (Nonobese Diabetic) and Control Mice

Subpopulations of lymphoid cells were compared with respect to their ability to migrate into peripheral lymphoid organs of nonobese diabetic (NOD) mice and various strains of control mice. In short-term, in vivo homing studies, no major differences in the pattern of homing of B and T cells were observed among all mouse strains studied. On the other hand, CD4 cells localized consistently more efficiently than CD8 cells in both PP and LN of adult NOD and BALB/c mice, whereas both populations migrated roughly equivalently in LN of adult DBA/2, CBA, and C57BL/6 mice. No age-dependent differences in the homing of CD4 and CD8 cells were observed in BALB/c mice. On the contrary, in 2-week-old NOD mice, CD4 and CD8 cells migrated equally well. The preferential entry of CD4 cells in adult NOD and BALB/c did not result from increased blood transit time of CD8 cells. On the other hand, the preferential migration of CD8 cells was observed in the liver, whereas the two T-cell subsets migrated equally well in the lungs. The differences in the homing characteristics of CD4 and CD8 cells among NOD, BALB/c, and C57BL/6 mice were not related to modifications in the level of expression of adhesion molecules such as MEL-14, LFA-1, and Pgp-1.