Detection of oligoclonal T lymphocytes in lymph nodes draining from advanced non-small-cell lung cancer

Despite the combined use of surgery and chemoradiotherapy, the poor prognosis of advanced non-smallcell lung cancer (NSCLC) requires the definition of new therapeutic approaches. The presence of T lymphocytes, with peculiar phenotypic, functional and molecular characteristics within the tumour, suggested the possible use of these cells, expanded in vitro, in protocols of adoptive immunotherapy. We have described how a population of oligoclonal T lymphocytes, derived from advanced NSCLC, can be expanded in vitro and has the capability of lysing autologous cancer cells. What is more important, we observed that patients with advanced NSCLC, treated with TIL expanded in vitro and recombinant interleukin-2, seemed to have a disease-free period longer than that of patients treated with conventional chemoradiotherapy. in an attempt to find new sources of specific lymphocytes for immunotherapy, we describe the analysis of the phenotypic, functional and molecular characteristics of T lymphocytes, derived from lymph nodes draining advanced NSCLC. In this paper we show that these cells, have restriction patterns of T cell receptor chain similar to those detectable in the population of infiltrating T lymphocytes. This finding suggests that T cells derived from draining lymph nodes of advanced NSCLC have peculiar characteristics and can be a suitable source of effector cells for protocols of adoptive immunotherapy in lung cancer treatment.     

大鼠胸部照射后支气管肺泡灌洗液成分和肺组织纤溶活力的变化

大鼠胸部照射γ射线２０Ｇｙ,照射后观察支气管肺泡灌洗液（ＢＡＬＦ）中蛋白质含量、细胞总数和分类及巨噬细胞存活率,肺指数及肺组织纤溶活力,并做了大体解剖和组织学检查。结果为：照射后２周开始ＢＡＬＦ中蛋白质含量、细胞总数和中性粒细胞即明显增多,照射后１——２个月持续处于高水平,高峰在１．５─２个月,照射后３─４个月有所下降,肺指数有相应变化;肺组织纤溶活力则相反,从照射后２周开始持续下降,至照射后２个月已降至最低,接近于零。形态学观察也见到有早期肺水肿和晚期肺萎缩等变化。由上结果可见,大鼠胸部照射后的早期主要为渗出性病变和纤溶活力的降低。文中讨论了照射后血管内皮细胞的损伤在放射性肺损伤发病中的作用。     

Growth characteristics,morphology, and phospholipid composition of human type II pulmonary alveolar cells grown in a collagen-free microenvironment

Summary Human lung epithelial cells have been isolated and maintained in pure culture and characterized during their time in culture. Any residual fibroblasts were removed by selective trypsinization within the first 48 h in culture and the residual epithelial cells from the primary culture grew to confluent density. The epithelial cells at Passage 2 or greater were serially subpassaged when cultures reached ca. 80% confluency. This procedure permitted us to conduct biochemical and structural studies of starting materials and subsequent population doublings. Electron microscope evaluation of both initial monolayers and cell suspensions showed cultures to be composed of a single cell type. These cells had microvilli on their free or apical surface. Subsequent population doubling level 1 up to 5 exhibited the same structures. They contained lamellar inclusions, which are typical of Type II alveolar epithelial cells. Fetal lung (age 18 to 20 wk) cell suspensions processed for electron microscopy before culturing showed cells to be undifferentiated, epithelial-like with small microvilli along cell borders, and with desmosomes at cell junctions. Lamellar inclusions were not observed in these cells. Ultrastructural studies of the cultured epithelial cells demonstrated that the lamellar inclusions had a slightly positive reaction when tested for acid phosphatase. Phospholipid analysis of these lung epithelial cells showed a phospholipid composition consistent with that found in surfactant-containing Type II cells. Cultured epithelial cells stained with phosphine 3-R demonstrated a green fluorescent cytoplasm and nucleus with brightly fluorescent yellow-orange perinuclear particles. The preceding characterization of these cells leads us to conclude that they exhibit structural and biochemical features commensurate with Type II epithelial cells from human lung. Moreover, these selection techniques applied to the isolation of human lung Type II cells from the tissue permit us to study the differentiative function of these cells routinely under conditions of growth in vitro. This work was supported in part by grants from EPA, R 806638-01 and 131-640-1599A1     

Effect of lipids on activity and conformation of lysolecithin:lysolecithin acyltransferase from rabbit lung

Summary Lysolecithin:lysolecithin acyltranferase is an enzyme which in several previous studies has shown a dual behavior catalyzing two types of reaction, transacylation or hydrolysis, with the same substrate. Both activities have shown to be dependent on several environmental conditions and among them, the presence of lipids.The addition of several classes of lipids activated in all the cases the enzyme, decreasing the hydrolysis/transacylation molar ratio. This effect was higher for PC/PE/Chol mixture than for other lipids assayed. Circular dichroism spectra of the enzyme did not show any change with the addition of lipids, concluding that the effect of lipids was not due to any structural change in the protein. The hypothesis has been made of an influence of lipids on the physical state of the substrate as well as, possibly, on the enzyme-substrate interaction.The significance of these effects on the physiological role of lysolecithin:lysolecithin acyltransferase from soluble fraction of rabbit lung is discussed.Abbreviations Chol cholesterol - CMC critical micellar concentration - DPPC dipalmitoylphosphatidylcholine - FA fatty acid - H/T hydrolysis/transacylation molar ratio - LPC lysophosphatidylcholine - PC phosphatidylcholine - PE phosphatidylethanolamine - TG triglyceride - UV ultraviolet     

Endogenous proteinases modulate the function of the β-adrenergic receptor-adenylate cyclase system

Photoaffinity labeling techniques have recently demonstrated that mammalian β₁- and β₂-adrenergic receptors reside on peptides of M_r 62 000–64 000. These receptor peptides are susceptible to endogenous metalloproteinases which produce peptides of M_r 30 000–55 000. Several proteinase inhibitors markedly attenuate this process, specifically EDTA and EGTA. In this study we investigated the functional significance of this proteolysis (and its inhibition) in the β₂-adrenergic receptor-adenylate cyclase system derived from rat lung membranes. Membrane preparations containing proteolytically derived fragments of the receptor of M_r 40000–55 000 are fully functional with respect to their ability to bind β-adrenergic antagonist radioligands such as [³H]dihydroalprenolol and β-adrenergic antagonist photoaffinity reagents such as p-azido-m-[^{125I]iodobenzylcarazolol}. They retain the ability to form a high-affinity, agonist-promoted, guanine nucleotide-sensitive complex thought to represent a ternary complex of agonist, receptor and guanine nucleotide regulatory protein. Nonetheless, after proteolysis, GTP is less able to revert this high-affinity receptor complex to one of lower affinity, and all aspects of adenylate cyclase stimulation are reduced. In addition, the functional integrity of the N protein in membranes prepared without proteinase inhibitors is reduced as assessed by reconstitution studies with the cyc[su− variant of S49 lymphoma cell membranes. These results suggest that endogenous proteolysis does not directly impair the ability of β-adrenergic receptors to either bind ligands or interact with the guanine nucleotide regulatory protein. However, they imply that endogenous proteolysis likely impairs the functionality of other components of the adenylate cyclase system, such as the nucleotide regulatory protein.     

Influence of 0.5 ppm nitrogen dioxide exposure of mice on macrophage congregation in the lungs

Summary The lungs of 12 mice, half of which were exposed to continuous 0.5 ppm nitrogen dioxide for 3 weeks, were explanted in culture, and the instances of macrophage congregation were quantitated according to numbers of target cells involved, categories of congregation from three to 11 or more, numbers of macrophages participating in each category for the total cultures, and the influence of delaying explantation for 24 and 96 hr. A total of 9042 macrophages and 2140 epithelial and spindle target cells were counted in the outgrowths from 306 explants. The incidence of macrophage congregation (or numbers of target cells) was greater for the cultures from the NO₂-exposed animals, both with respect to total incidences between groups (p→0), and the 0-hr (p<0.001) and 24-hr (p<0.01) culture subgroups. In addition, the values for T₃ to T₆ macrophage congregation were individually and consistently greater for the exposed animal group. Postmortem interval stress at 96 hr appeared to result in large colonies, but they were reduced greatly in number. Also the incidence of macrophage congregation fell by 28% as compared to 0-hr and 24-hr subgroups. Supported by Grants NHLI No. HL 17412 and EPA No. R. 800881.     

A continuous-flow method of organ culture

Summary A method of perfusion organ culture is described in which explants cultured at the airmedium interface are bathed by a continuous flow of nutrient medium. Morphological studies on the fetal rat lung indicate that explant development in this system is comparable to that obtained using standard organ-culture dishes. Medium supply is easily manipulated and continuous sampling of the effluent stream is possible without disturbing the immediate explant environment. The basic design facilitates secretory-response studies on cultured organ explants as demonstrated by a study of glucose-stimulated insulin release by the neonatal rat pancreas. This work was supported by U. S. Public Health Service Training Grant No. GM 00114.     

Fetal mouse lung in circumfusion system cultures

Summary Fetal mouse lungs were cultivated, using the dual-rotary circumfusion system for tissue culture, and their histotypic development was surveyed for 75 days by phase-contrast and electron microscopy. Alveoli, terminal bronchioles and alveolar macrophages were photographed periodically with still and time-lapse phase-contrast microscopy. Their histotypic appearance was confirmed by electron micrographs of the 1- and 2 1/2-month-old specimens. These revealed typical alveoli surrounded by a basal lamina and composed of types I and II pneumocytes containing various lamellar-body forms within the type II cells, the alveolar lumen, and the alveolar macrophages. There was a shift from almost all type II cells in the 1-month-old alveoli to the presence of frequent type I cells as constituents of the alveoli in the 2 1/2-month-old cultures. The terminal bronchioles were tubules consisting of ciliated cells with Clara cells interspersed between them. The ciliated cells contained as many as 30 cilia or basal bodies per section and numerous microvilli. They were attached to each other and to the Clara cells by junctional complexes and accessory desmosomes which were generally in the apical ends of the cells. The Clara cells typically had glycogen granules interspersed between lamellae of the endoplasmic reticulum, contained numerous well dispersed mitochondria, occasional lysosome-like granules and crystalloid bodies which appeared to be tubular. Some Clara cells presented a moderately dense secretory granule in the center of the whorl of the endoplasmic reticulum. This work supported by Grant HL19684 from the National Heart and Lung Institute, National Institutes of Health. Pregnant Strong A mice were kindly supplied by Dr. Henry Browning of the Department of Anatomy.     

转移及非转移肿瘤移植后615小鼠血液流变学变化的研究

血道高转移瘤株FC、淋巴合并血道高转移瘤株U14、淋巴道高转移瘤株H22、非转移瘤株P615分别接种于336只纯系近交615小鼠.不同时间取血并处死动物,进行组织学及血液流变学检查.将转移瘤发展过程分为潜伏期、侵袭期、转移早、中、晚期,非转移瘤发展过程分为潜优期、增殖期、囊腔形成期及中心坏死期.本实验结果显示,不同转移能力及途径肿瘤发展的不同时期血液流变学变化规律不同,因而表明肿瘤侵袭、转移与血液流变学变化之间存在互为因果的紧密关系.其临床诊断及治疗意义被讨论.