Molecular characterization of Waldenstrom's macroglobulinemia reveals frequent occurrence of two B-cell clones having distinct IgH VDJ sequences.

PURPOSE: Malignant B lineage cells in Waldenstrom's macroglobulinemia (WM) express a unique clonotypic IgM VDJ. The occurrence of biclonal B cells and their clonal relationships were characterized. EXPERIMENTAL DESIGN: Bone marrow and blood from 20 WM patients were analyzed for clonotypic VDJ sequences, clonal B-cell frequencies, and the complementary determining region 3 profile. RESULTS: Two different clonotypic VDJ sequences were identified in 4 of 20 WM. In two cases, partner clones had different VDJ rearrangements, with one clonotypic signature in bone marrow and a second in blood. For both cases, the bone marrow clone was hypermutated, whereas the blood clone was germ line or minimally mutated. In two other cases, partner clones shared a common VDJ rearrangement but had different patterns of somatic mutations. They lacked intraclonal diversity and were more abundant in bone marrow than in blood. VDJ mutation profiles suggested they arose from a common IgM progenitor. Single-cell analysis in one case indicated the partner clones were reciprocally expressed, following rules of allelic exclusion. CONCLUSIONS: The existence of two B-cell clones having distinct VDJ sequences is common in WM, suggesting that frequent transformation events may occur. In two cases, the partner clones had distinct tissue distributions in either blood or bone marrow, were of different immunoglobulin isotypes, and in one case exhibited differential response to therapy. The contributions of each clone are unknown. Their presence suggests that WM may involve a background of molecular and cellular events leading to emergence of one or more malignant clones.     

Cell generation within human thymic subsets defined by selective expression of CD45 (T200) isoforms

Although the thymus is the source of all mature peripheral T lymphocytes, the majority of thymocytes die intrathymically. Until recently, there has been no phenotypic marker to allow definition of the generative thymocyte lineage, thereby distinguishing those thymocytes committed to death from those which will evenually give rise to thymic emigrants. We believe that expression of the high-molecular-mass isoforms (p190, p205, and/or p220) of the leukocyte common antigen (CD45) distinguishes the thymic generative lineage from the vast majority of thymocytes expressing the low-molecular-mass isoform (p180) of CD45 and committed to die within the thymus. The thymocytes defined by their lack of CD45 p180, the low-molecular-mass isoform, comprise all thymocytes with clonogenic potential and include all major subsets defined by CD4 and CD8. We have proposed that a CD45 p180⁻ lineage exists in the human thymus and that this lineage results in the production of mature thymocytes and thymic emigrants. The objective of the present study was to determine by DNA analysis whether the degree of cell cycling in subsets of human thymus, defined by selective expression of high-molecular-mass isoforms of CD45, was sufficient to account for the generation of thymic emigrants. Multicolor immunofluorescence analysis of surface markers and 7-amino actinomycin D as well as propidium iodide staing was used to measure the DNA content of thymic subsets. Negative depletion methods were used to isolate and characterize human thymocyte subsets defined by CD45 isoform, CD3, CD4, and CD8, and subsequently to determine the cell cycle status of the isolated subsets by flow-cytometric analysis of cellular DNA content. CD3^−/lo thymocytes had a high number and CD1^−/lo thymocytes a low number of cycling cells, consistent with murine data. CD45 p 180⁻ cells, as well as the CD4⁻8⁻ and CD3⁻4⁻8⁻ subsets which express high molecular-weight CD45 isoforms, exhibited a significant number of cycling cells. Since CD45 p180- thymocytes exhibited a significant number of cycling cells, based on numerical arguments we conclude that this cycling thymocyte fraction is capable of generating the daily requirements of mature thymocytes and thymic emigrants.     

Inhibition of experimental autoimmune encephalomyelitis in Lewis rats by nasal administration of encephalitogenic MBP peptides: synergistic effects of MBP 68-86 and 87-99

Induction of mucosal tolerance by inhalation of soluble peptides with defined T cell epitopes is receiving much attention as a means of specifically down-regulating pathogenic T cell reactivities in autoimmune and allergic disorders. Experimental autoimmune encephalomyelitis (EAE) induced in the Lewis rat by immunization with myelin basic protein (MBP) and Freund's adjuvant (CFA) is mediated by CD4+ T cells specific for the MBP amino acid sequences 68-86 and 87-99. To further define the principles of nasal tolerance induction, we generated three different MBP peptides (MBP 68-86, 87-99 and the non- encephalitogenic peptide 110-128), and evaluated whether their nasal administration on day -11, -10, -9, -8 and -7 prior to immunization with guinea pig MBP (gp-MBP) + CFA confers protection to Lewis rat EAE. Protection was achieved with the encephalitogenic peptides MBP 68-86 and 87-99, MBP 68-86 being more potent, but not with MBP 110-128. Neither MBP 68-86 nor 87-99 at doses used conferred complete protection to gp-MBP-induced EAE. In contrast, nasal administration of a mixture of MBP 68-86 and 87-99 completely blocked gp-MBP-induced EAE even at lower dosage compared to that being used for individual peptides. Rats tolerized with MBP 68-86 + 87-99 nasally showed decreased T cell responses to MBP reflected by lymphocyte proliferation and IFN-gamma ELISPOT assays. Rats tolerized with MBP 68-86 + 87-99 also had abrogated MBP-reactive IFN-gamma and tumor necrosis factor-alpha mRNA expression in lymph node cells compared to rats receiving MBP 110-128 nasally, while similar low levels of MBP-reactive transforming growth factor-beta and IL-4 mRNA expressing cells were observed in the two groups. Nasal administration of MBP 68-86 + 87-99 only slightly inhibited guinea pig spinal cord homogenate-induced EAE, and passive transfer of spleen mononuclear cells from MBP 68-86 + 87-99-tolerized rats did not protect naive rats from EAE. Finally, we show that nasal administration of MBP 68-86 + 87-99 can reverse ongoing EAE induced with gp-MBP, although higher doses are required compared to the dosage needed for prevention. In conclusion, nasal administration of encephalitogenic MBP peptides can induce antigen-specific T cell tolerance and confer incomplete protection to gp-MBP-induced EAE, and MBP 68-86 and 87-99 have synergistic effects. Non-regulatory mechanisms are proposed to be responsible for tolerance development after nasal peptide administration.      

Localization of endogenous galactoside-binding lectin during morphogenesis ofXenopus laevis

Summary A monoclonal antibody has been produced againstXenopus laevis galactoside-binding neural-creststage lectin. This antibody inhibits lectin-mediated hemagglutination. Using this antibody in conjunction with immunohistochemical techniques, lectin deposition has been studied in embryos and tadpoles at different stages of morphogenesis, from initial neural crest migration, up to the formation of a swimming tadpole. Lectin levels change during development in different regions of the embryo and tadpole, decreasing in migratory cells, and increasing in sites where cells become more adhesive to one another. The results suggest that galactoside-binding lectins may be an important class of cellular adhesion molecules during these stages of development.     

Apoptotic cell death in mouse models of GM2 gangliosidosis and observations on human Tay-Sachs and Sandhoff diseases

Tay-Sachs and Sandhoff diseases are autosomal recessive neurodegenerative diseases resulting from the inability to catabolize GM2 ganglioside by beta-hexosaminidase A (Hex A) due to mutations of the alpha subunit (Tay-Sachs disease) or beta subunit (Sandhoff disease) of Hex A. Hex B (beta beta homodimer) is also defective in Sandhoff disease. We previously developed mouse models of both diseases and showed that Hexa-/- (Tay-Sachs) mice remain asymptomatic to at least 1 year of age while Hexb-/- (Sandhoff) mice succumb to a profound neurodegenerative disease by 4-6 months of age. Here we find that neuron death in Hexb-/- mice is associated with apoptosis occurring throughout the CNS, while Hexa-/- mice were minimally involved at the same age. Studies of autopsy samples of brain and spinal cord from human Tay-Sachs and Sandhoff diseases revealed apoptosis in both instances, in keeping with the severe expression of both diseases. We suggest that neuron death is caused by unscheduled apoptosis, implicating accumulated GM2 ganglioside or a derivative in triggering of the apoptotic cascade.      

奥曲肽治疗肝硬变食管静脉曲张破裂出血

0　引言　我院 1996 - 0 6 / 1998- 0 7用奥曲肽治疗肝硬变食管静脉曲张破裂出血 19例 ,并与垂体后叶素作了对照 .1　对象和方法1.1　对象　经内镜证实食管静脉曲张破裂出血 37例 ;男 32例 ,女 5例 ;年龄 42～ 6 9岁 .肝炎后肝硬变 32例 ,乙醇性肝硬变 3例 ,血吸虫性肝硬变 2例 .按就诊顺序随机分为治疗组19例 ,对照 18例 .估计出血量 :治疗组 (146 3.6± 375 .8) m L;对照组 (144 9.5± 36 8.4) m L .出血至治疗间隔时间 :治疗组(11.6± 3.5 ) h;对照组 (11.2± 3.6 ) h.两组的年龄、性别、肝功 Child分级、食管静脉曲张程度、出血量、出…     

五灵胶囊治疗慢性肝炎的临床随访观察

0　引言　五灵丸治疗慢性病毒性肝炎取得了较好的疗效 [1 ] ,但五灵丸为大蜜丸 ,不易被患者所接受 ,且其消化道副作用较明显 .为此 ,我们改进剂型 ,研制成五灵胶囊 ,为明确其能否保持远期疗效 ,我们对用该药治疗的患者进行随访观察 ,同时与五灵丸的疗效进行比较 .1　对象和方法1.1　病例选择　按慢性肝炎的诊断标准选择住院患者 10 0例 ,随机分为两组 .治疗组为五灵胶囊组 ,对照组为五灵丸组 .每组各 5 0例 .治疗组轻度 19例 ,中度 2 0例 ,重度 11例 .对照组轻度 2 0例 ,中度 18例 ,重度 12例 .两组均选择 18～5 8岁 ,病程 1～ 2 0 a.平均 4…     

Smoking, alcohol, diet, dentition and sexual practices in the epidemiology of oral cancer in Poland.

The effect of smoking, drinking, diet, dental care and sexual habits on the risk of oral and pharyngeal cancer was investigated in a case-control study conducted in Warsaw, Poland. The study comprised 122 patients (including 44 females) aged 23-80 years with histologically confirmed cancer of oral cavity and pharynx. Controls were 124 subjects (including 52 females) admitted to the hospital for different non-neoplastic conditions unrelated to tobacco and alcohol consumption, with frequency matched to cases by age and sex. Smoking and drinking were strongly associated with an increased risk of oral cancer. Among consumers of both products, risks of oral cancer tended to combine in a multiplicative fashion and were increased more than 14-fold among those who consumed more than 15 cigarettes and seven or more drinks per day. Cessation of smoking was associated with reduced risk of this cancer. The risks varied by type of cigarettes smoked, being lower among those consuming filtered cigarettes only (OR = 1.6) than nonfilter (OR = 6.5) or mixed (OR = 4.2) cigarettes. High fruit intake was associated with significantly decreased risk (OR = 0.4) with the strongest significant inverse association found for fruit juices and citrus fruits ( < 0.01). After adjustment for tobacco smoking and alcohol drinking, poor dentition as evidenced by missing teeth, frequency of dental check-ups and frequency of teeth brushing emerged as a strong risk factor. Number of missing teeth and frequency of dental check-ups and frequency of tooth brushing showed increased ORs of 9.8, 11.9 and 3.2, respectively. Denture wearing did not affect oral cancer risk. No differences were detected in sexual practices (including oral sex and intercourse with prostitutes). In terms of attributable risk, smoking accounted for 57% of oral cancer cases in Poland, alcohol for 31% and low fruit intake for 12%. Attributable risks for low frequency of tooth brushing and dental check-ups were 56% and 47%, respectively. In conclusion, smoking and drinking cessation and increase of fresh fruit intake are likely to be effective preventive measures against oral cancer. These findings indicate also that poor oral hygiene may be an independent risk factor.     

Drug resistance in multiple myeloma: novel therapeutic targets within the malignant clone

Myeloma is incurable because the malignant stem cell is not eradicated by treatment. Thus, identification of the malignant hierarchy of B lineage cells in myeloma is required to identify potentially generative components and to evaluate their drug resistance properties. BM plasma cells are usually depleted by chemotherapy, but clonotypic B cells survive melphalan/prednisone as well as combination chemotherapy. In vitro, circulating and bone marrow-localized myeloma plasma cells show defective drug export, despite their phenotypic expression of P-glycoprotein, the mdr1 gene product. In contrast to plasma cells, circulating myeloma clonotypic B cells exhibit very efficient drug export. This suggests that circulating clonotypic MM B cells comprise a reservoir of drug resistant disease in myeloma although their stem cell potential remains to be confirmed. The malignant clone in each myeloma patient is defined by a unique IgH VDJ gene rearrangement. Using methods that exclude the possibility that a frequent but non-malignant clone has inadvertently been identified, and after confirming that the sequence identified is expressed by nearly all bone marrow plasma cells, we show that the drug resistant set of myeloma B cells is clonally related to the malignant plasma cells in myeloma. Clonotypic MM B cells survive chemotherapy, persist during clinically defined "minimal residual disease" and remain after autologous transplantation. Thus their malignant status is an important consideration. If malignant, they must be considered in the design of therapy. If non-malignant, they would be expected to have minimal impact on the disease process. A variety of evidence provides strong support for the view that clonotypic drug resistant B cells are malignant and may include the generative compartment of myeloma. The P-gp+ set of clonotypic B cells is extensively DNA aneuploid, an attribute of malignancy. All clonotypic B cells overexpress RHAMM, a novel oncogene involved in malignant spread. Finally, the population of clonotypic B cells lacks intraclonal heterogeneity. Since intraclonal heterogeneity is driven by the response to antigens, its absence in these cells indicates that they are no longer antigen-responsive. Since antigen-independent clonal expansion is characteristic of lymphoid malignancies, these observations provide further proof that clonotypic B cells in myeloma are malignant. Thus, the drug resistance of these cells is highly relevant to understanding why myeloma remains incurable despite the initial chemosensitivity of most bone marrow plasma cells.