J-chain-producing immunoblastic lymphoma in a case of Richter's syndrome

Summary A 66-year old male with Richter's syndrome died 52 month after diagnosis of chronic lymphocytic leukaemia (CLL). The clinical course was characterized by a marked IgM hypoglobulinaemia which paralleled a chronically relapsing Herpes simplex infection. Autopsy showed a large retroperitoneal and intraabdominal tumour mass and well defined supradiaphragmatic lymphomas. Histological examination revealed a composite tumour consisting of CLL B-cell type (B-CLL) and immunoblastic malignant lymphoma of B-cell type (B-IbL). The lymphocytes bear -chains on their surface and to a lesser extend within their cytoplasm, the obviously defective immunoblasts produce J chains exclusively. Flow cytophotometric data seem to indicate an identical diploid stem line of the two tumours. The majority of the cells are in G_0/1 phase. The CLL rarely produces mitoses, however, the IbL has a mitotic rate of 7% and a considerable proportion (33%) of cells in the phase of DNA-synthesis. This is the fourth malignant lymphoma and the second immunoblastic lymphoma to be reported that produces J chain in the absence of immunoglobulin.     

Immunophenotypic and molecular analysis of a case of lymphocytic interstitial pneumonia

A case of lymphocytic interstitial pneumonia was studied immunophenotypically and with molecular methods in order to clarify its lymphocytic clonality. The patient, a 43 year old Japanese female, underwent lobectomy for a suspected malignant lymphoma as no clear diagnosis could be made from the biopsy specimen. An ill-demarcated, yellowish and elastic firm lesion measuring 60 times 35 times 20 mm in size was located in the peripheral part of the middle lobe of the right lung. Histologically, the alveolar, peribronchial-vascular and subpleural interstitia within the lesion were thickened markedly with severe cellular infiltration largely composed of small lymphocytes with germinal centers. lmmunostaining revealed immunoglobulin (lg) kappa and lg lambda-bearing cells to be evenly distributed, suggestive of polyspecificity. Immunoglobulin gene analysis further demonstrated the unrearranged germ-line DNA but no rearranged band. These results strongly indicated a reactive process rather than a neoplastic nature for the lesion.     

Southern blot analysis in a case of Richter's syndrome. Evidence for a postrearrangement heavy chain gene deletion associated with the altered phenotype.

Richter's syndrome (RS) can be defined as the emergence of an aggressive lymphoma in patients suffering from chronic lymphocytic leukemia (CLL). The authors performed immunophenotypic and Southern blot analysis of the peripheral blood and tissue specimen of a patient with RS. Using immunoperoxidase and immunogold-silver staining techniques and a panel of monoclonal antibodies, the authors found that the large cells characteristic of RS showed an altered immunophenotype as compared with the CLL cells and did not express mu heavy chain. Southern blot analysis revealed identical kappa light chain rearrangements in both tumoral cell populations consistent with a common clonal origin. Using the JH probe and several restriction enzymes, the authors also found evidence for a postrearrangement deletion of the heavy chain mu gene. These findings suggest that in this case of RS, a deletion of the heavy chain mu gene resulted in loss of mu expression by the larger cells that were characteristic of RS and was associated with their altered phenotype.     

经典型Richter综合征转化前后IgVH基因克隆重排及突变分析

目的 检测经典型Richter综合征的IgVH基因克隆重排及突变状态,分析转化前后IgVH基因的分子特征及其与预后的关系,并对可能涉及的分子机制进行初步探讨.方法 用基因扫描和测序分析IgVH基因.另外用免疫组织化学LAB-SA法检测两种肿瘤中zeta链结合蛋白激酶70(ZAP70)、p53、干扰紊调节因子(IRF)-4等可能潜在危险因子的表达,并结合随访资料进行生存率分析,筛选危险因子.结果 (1)B-CLL/DLBCL克隆相关(18/23,78.3%),克隆不相关(5/23,21.7%);(2)在16例克隆相关中,12例转化前B-CLL及转化后DLBCL携带未突变IgVH基因;(3)转化前后IgVH基因使用是非随机的,在克隆相关的病例中,B-CLL/DLBCL最常使用VH3-23、VH3-74、VH1-2,各占11.1%;(4)转化后DLBCL仅部分表达CD5(32.1%)和CD23(14.3%)及ZAP70(23.8%),绝大多数表达p53(80.6%)和IRF-4(82.6%);(5)17例转化后DLBCL患者中位生存时间为7个月;(6)统计学分析生存时间与B-CLl/DLBCL转化前后克隆相关与否、IgVH基因的突变状态、ZAP70、p53、IRF-4的表达不相关.结论 (1)转化后DLBCL中克隆相关与克隆不相关的比例为2:1;(2)克隆相关的DLBCL多由携带未突变型IgVH基因的B-CLL患者转化而来;(3)发生转化的B-CLL中IgVH基因使用的偏向性提示了抗原在转化中的可能作用;(4)转化后DLBCL与普通DLBCL在IgVH基因的使用、突变状态,免疫表型及预后的不同,提示其可能是一种新的DLBCL亚类.     

一例性反转综合征SRY基因分析

目的 分析1例46,XY女性性反转综合征患者的性别决定基因SRY(sex-determining region of Y chromosome)的突变类型,探讨该患者发生性反转的分子机制.方法 收集患者临床资料,抽取静脉血.体外培养外周血淋巴细胞.采用染色体G显带分析患者核型.提取患者外周血DNA,采用PCR扩增SRY基因,并将回收产物直接测序.结果 患者核型为46,XY.PCR检测患者存在SRY基因.DNA序列分析显示患者SRY基因编码区第6位碱基缺失,发生移码突变,导致编码SRY蛋白第9位亮氨酸(L)的第2位碱基T、第3位碱基A与编码第10位丝氨酸(S)的第1位碱基A组合成为终止密码TAA,翻译过程提前终止,导致SRY蛋白缺乏.结论 SRY基因编码区第6位碱基缺失导致翻泽过程提前终止,患者不能合成完整的SRY蛋白,使其在胚胎期发生性逆转.     

2 cases of Richter's syndrome]

Two cases of Richter's syndrome are reported (in a 62 and 64 years old man) consistent with the appearance of B cell lymphoma of high malignancy in the course of CLL (low malignancy B cell lymphoma). In one patient, after 8-, and in the other one--after 53 months since the diagnosis of CLL, there was rapid clinical deterioration with lymphadenopathy, hepato- splenomegaly, fever and progressive cachexia, anemia and thrombocytopenia and leukopenia, unrelated to treatment. Both patients died, 4 and 3 months respectively, since the appearance of these symptoms. In the first cases Richter's syndrome was diagnosed histopathologically from the autopsy material. In the liver, spleen, adrenals and bone marrow, in addition to the characteristic infiltrates of CLL (small lymphocytes) there were areas of large cell proliferation consistent with high malignancy lymphoma. In the other case, the infiltrates of large cell lymphoma were found in the gall bladder removed because of acute cholecystitis, and in the lymph node from the hepatic hilar area. Immunocytochemical studies performed on the biopsy material indicated that the neoplastic cells had markers of B lymphocytes and cytoplasmic IgM kappa, as lymphocytes of CLL. In patients with CLL, who display rapid clinical deterioration and general symptoms with cachexia, the possibility of Richter's syndrome should be considered, and appropriate morphological studies performed.     

骨髓增生异常综合征患者骨髓涂片的免疫表型分析

目的 检测骨髓增生异常综合征（MDS）患者的免疫表型特征,探讨对其早期诊断及分型诊断的价值.方法 采用免疫细胞化学法中的生物素-亲和素桥联碱性磷酸酶酶标法（ABC-AP）对60例初诊的MDS患者、14例再生障碍性贫血（AA）患者及21例正常对照的骨髓单个核细胞（MNC）进行免疫表型检测.结果 CD41阳性的淋巴样小巨核细胞在MDS组中较特异出现,且以难治性细胞减少伴多系异常（RCMD）组出现率最高;与正常对照组比较,MDS组中的难治性贫血伴原始细胞增多（RAEB）组CD13、CD33和CD34抗原表达升高,CD3抗原表达下降;RCMD组CD13和CD33抗原表达升高,CD34抗原表达升高不明显,CD3抗原表达下降;难治性贫血/环形铁粒幼细胞性难治性贫血（RA/RAS）组CD13、CD33、CD34、CD3抗原表达均无明显特征性变化.结论 酶标淋巴样小巨核及免疫表型CD3、CD13、CD33、CD34检测有助于MDS的早期诊断及分型诊断.     

Immunophenotypic and antigen receptor gene rearrangement analysis in T cell neoplasia.

The author reviews the immunophenotypic profiles displayed by the major clinicopathologic categories of T cell neoplasia, the immunophenotypic criteria useful in the immunodiagnosis of T cell neoplasia, and the contributions made by antigen receptor gene rearrangement analysis to the understanding of T cell neoplasia. Neoplasms belonging to distinct clinicopathologic categories of T cell neoplasia often exhibit characteristic immunophenotypic profiles. Approximately 80% of lymphoblastic lymphomas and 20% of acute lymphoblastic leukemias express phenotypes consistent with prethymic and intrathymic stages of T cell differentiation, including intranuclear terminal deoxynucleotidyl transferase. Cutaneous T cell lymphomas of mycosis fungoides type usually express pan-T cell antigens CD2, CD5, and CD3, often lack the pan-T cell antigen CD7, and usually express the mature, peripheral helper subset phenotype, CD4+ CD8-. Cutaneous T cell lymphomas of nonmycosis fungoides type and peripheral T cell lymphomas often lack one or more pan-T cell antigens and, in addition, occasionally express the anomalous CD4+ CD8+ or CD4- CD8- phenotypes. T gamma-lymphoproliferative disease is divisable into two broad categories: those cases that are CD3 antigen positive and exhibit clonal T cell receptor beta chain (TCR-beta) gene rearrangements and those cases that are CD3 antigen negative and exhibit the TCR-beta gene germline configuration. Human T cell lymphotropic virus-I (HTLV-I) associated Japanese, Carribean, and sporadic adult T cell leukemia/lymphomas usually express pan-T cell antigens, the CD4+ CD8- phenotype, and various T cell-associated activation antigens, including the interleukin-2 receptor (CD25). Immunophenotypic criteria useful in the immunodiagnosis of T cell neoplasia include, in increasing order of utility, T cell predominance, T cell subset antigen restriction, anomalous T cell subset antigen expression, and deletion of one or more pan-T cell antigens. Only in exceptional circumstances do normal, non-neoplastic T cell populations express the CD4- CD8- or the CD4+ CD8+ phenotype and/or lack one or more pan-T cell antigens. T cell receptor beta chain gene rearrangement analysis represents an accurate, objective, and sensitive molecular genetic marker of T cell lineage and clonality that allows discrimination among non-T cell, polyclonal T cell and monoclonal T cell populations. Non-T cells exhibit the TCR-beta gene germline configuration.(ABSTRACT TRUNCATED AT 400 WORDS)     

Immunophenotypic analysis of sinonasal non-Hodgkin's lymphomas

A panel of paraffin effective antibodies recognizing B cells and T cells (LN-2, MB1, L26, MT1, UCHL1, kappa, lambda) was used to characterize the immunophenotypes of 26 sinonasal non-Hodgkin's lymphomas. Seventeen tumors were stage I, five were stage II, one was stage III, and three were stage IV. Nine lymphomas were classified morphologically as large cell, six were large cell immunoblastic, six were small cleaved cell, two were mixed small and large cell, two were small noncleaved cell, and one was lymphoblastic. None were follicular. Twenty-two lymphomas had a B cell immunophenotype, three were T cell neoplasms, and one was immunoreactive only for MT1. This predominance of sinonasal lymphomas with a B cell immunophenotype in patients residing in the United States contrasts with the almost exclusive occurrence of T cell sinonasal lymphomas in Chinese patients living in Hong Kong and Japanese patients residing in regions of Japan that are nonendemic for human T cell leukemia virus-1.     

Investigation of gene dosage imbalances in patients with Noonan syndrome using multiplex ligation-dependent probe amplification analysis

The RAS-MAPK syndromes are a group of clinically and genetically related disorders caused by dysregulation of the RAS-MAPK pathway. A member of this group of disorders, Noonan syndrome (NS), is associated with several different genes within the RAS-MAPK pathway. To date, mutations in PTPN11, SOS1, KRAS, RAF1 and SHOC2 are known to cause NS and a small group of patients harbour mutations in BRAF, MEK1 or NRAS. The majority of the mutations are predicted to cause an up-regulation of the pathway; hence they are gain-of-function mutations. Despite recent advances in gene identification in NS, the genetic aetiology is still unknown in about ¼ of patients.To investigate the contribution of gene dosage imbalances of RAS-MAPK-related genes to the pathogenesis of NS, a multiplex ligation-dependent probe amplification (MLPA) assay was developed. Two probe sets were designed for seven RAS-MAPK-syndrome-related candidate genes: PTPN11, SOS1, RAF1, KRAS, BRAF, MEK1 and MEK2. The probe sets were validated in 15 healthy control individuals and in glioma tumour cell lines. Subsequently, 44 NS patients negative for mutations in known NS-associated genes were screened using the two probe sets. The MLPA results for the patients revealed no gene dosage imbalances. In conclusion, the present results exclude copy number variation of PTPN11, SOS1, RAF1, KRAS, BRAF, MEK1 and MEK2 as a common pathogenic mechanism of NS. The validated and optimised RAS-MAPK probe sets presented here enable rapid high throughput screening of further patients with RAS-MAPK syndromes.     

The clonal origin of two cell populations in Richter's syndrome

A case of Richter's syndrome was studied by morphology, immunohistochemistry, flow cytometry, and immunoglobulin gene rearrangement. Flow cytometric study clearly demonstrated two monoclonal populations. The use of double staining with CD 5/CD 19 antibodies accompanied by two-color flow cytometric analysis clearly defined the chronic lymphocytic leukemia population and separated it from the lymphoma population. Immunoglobulin heavy-chain gene analysis of blood and lymph node specimens revealed nonidentical as well as identical nongermline bands in these two populations. However, light-chain gene analysis demonstrated that both populations shared a common clonal origin. This result underscores the unreliability of using heavy chain genotype alone to identify clonal origin. Since post-rearrangement deletion, point mutation, and heavy chain switching occur in heavy chain genes, but are seldom seen in light chain genes, it is important to analyze both heavy and light chain genes to conclusively determine clonal origin.     

一例Nicolaides-Baraitser综合征的临床与遗传学分析

目的探讨1例表现为癫痫、语言发育迟缓、智力发育轻度迟缓的女性患儿的遗传学病因。方法采集患儿及其父母的外周血样并提取基因组DNA,应用高通量测序技术对患儿进行检测,对疑似致病变异进行Sanger测序验证及生物信息学分析。结果测序结果显示患儿SMARCA2基因存在c.3592 G>A(p.V1198M)杂合变异,生物信息学分析预测其为致病性;在患儿父母的外周血DNA中未发现相同的变异。结论该患儿被确诊为SMARCA2基因杂合变异所致的Nicolaides-Baraitser综合征。     

Immunophenotypic analysis of human spleen compartments.

Microanatomical compartments of the human spleen are yet under evaluation as most of the present information comes from experiments on animals with different anatomical structures. Immune staining of stromal and blood-born cells by cell surface antigens facilitates the differentiation of functional microanatomical compartmentalization of immune organs, including the spleen. Twenty-two specimens from healthy adult subjects with the average age of 35.6 +/- 13.8 (Range 17 to 58) years were included in this study. Monoclonal antibodies used in this study were supplied from the 5th, 6th and 7th International Workshops and Conferences on Human Leukocyte Differentiation Antigens. Tetraspan antigens presented a rather unique staining pattern in the human spleen, suggesting special roles for each (CD9, CD53, CD63, CD151 and CD231) in certain locations. Sinus lining cells presented a distinctive antigenic profile, sharing both endothelial cell (CD31, CD36, CD54, CD62P, CD102, CD105, CD106 and CD146) and macrophage lineage characteristics. The sheathed capillaries were not restricted to the perifollicular zone alone. Extracellular matrix receptors (CD49 a, CD49 b, CD49 c, CD49 e, CD49f, CD29 and CD44) stained the penicillary arterioles and vascular smooth muscle. These molecules were also found on the vascular endothelium. Leukocyte antigens (CD11a, CD11b, CD22, CD43, CD45, CD45RB, CD45RO and CD50) were mainly expressed in the white and red pulp of the spleen at different intensities, excluding the penicillary arterioles. Activation antigens (CD26, CD71 and CD98) presented a diffuse and broad staining pattern. In conclusion, microanatomical compartmentalization, microcirculation and function of the human spleen were evaluated using a wide panel of monoclonal antibodies.     

一例罕见的TCF4基因部分缺失型Pitt-Hopkins综合征的遗传学分析

目的探讨1例脑发育落后患儿的遗传学病因。方法采用常规G显带技术分析患儿及其双亲的染色体核型,应用单核苷酸多态性微阵列(single nucleotide polymorphism array,SNP-array)技术对患儿进行全基因组拷贝数变异(copy number variations,CNVs)筛查,并对其双亲进行验证。结果患儿及其双亲染色体G显带核型分析均未见异常。SNP-array检测双亲均未见异常,发现患儿染色体18q21.2区存在172 kb(52957042~53129237)的新发缺失,仅涉及OMIM基因TCF4,导致TCF4第6~8外显子的缺失。结论染色体18q21.2区的缺失与Pitt-Hopkins综合征密切相关。SNP-array检测为该患儿的确诊提供了依据。     

Hodgkin's disease variant of Richter's syndrome. Two cases and literature review

We report the clinical and immunohistological features of two cases of chronic lymphocytic leukaemia (CLL) with Hodgkin's transformation. These cases occurred in a 70-year-old man with a three-year history of CLL and in a 76-year-old man with a few months history of CLL. Microscopic examination showed the presence of large tumor cells with the morphological and immunophenotypic features of classical Hodgkin and Reed-Sternberg (R-S) cells, in a background of otherwise typical B-CLL. The transformation of CLL into large B cell lymphoma (Richter's syndrome) is a well-documented phenomenon. Only rarely does CLL transform into Hodgkin's lymphoma, but this diagnosis is often easy and offers few differential diagnoses. The major points of interest lie in the pathogenetic relationship between CLL and Hodgkin's disease, and in the potential clinical implications of this peculiar condition. Literature on the subject indicates that identical IgH gene rearrangements in micromanipulated R-S and CLL cells have been identified in 7/12 cases. In these patients, the R-S and CLL cells belong to the same clonal population, suggesting a progression from the underlying CLL cells. This group appears to have a poor prognosis, identical to classical Richter's syndrome. In other cases, the R-S cells were often Epstein-Barr virus (EBV) positive and did not share the clonal rearrangements identified in CLL cells, suggesting that Hodgkin's disease in these patients could represent a second malignancy, EBV-related and favored by immunosuppression, associated with a better prognosis.     

一例因基因突变致LIG4综合征病例的临床特点及分析

目的探讨DNA连接酶Ⅳ(LIG4)综合征的临床特点。方法分析一例LIG4综合征病例的临床资料,对患儿及其父母进行二代高通量基因测序。结合文献总结该病的临床特点。结果本例患儿的临床表现有小头畸形,特殊面容,发育迟缓,免疫功能缺陷,反复呼吸道感染,慢性腹泻。基因检测结果显示患儿LIG4基因存在纯合性改变:c.833G>T(p.R278L),故诊断LIG4综合征。结合文献复习包括本研究在内的共42例LIG4综合征病例的临床特点。结论LIG4综合征临床特点主要有小头畸形,免疫功能缺陷,特殊面容,血细胞下降和发育迟缓。临床上反复呼吸道感染、小头畸形伴免疫缺陷患儿应警惕此病,基因检测可协助诊断。     

一例MEGDEL综合征患儿的临床及遗传学分析

目的:探讨1例MEGDEL综合征患儿的临床及基因变异特点。方法:收集患儿的临床资料,采集患儿及其父母的外周血样,应用二代测序技术对患儿进行线粒体基因组及全外显子组分析,用Sanger测序以及荧光定量PCR验证候选变异及其来源。结果:患儿为男性,2岁6个月,主要表现为新生儿期低血糖、智力运动发育落后伴倒退。头颅磁共振成像...     

Immunoblastic lymphosarcoma in chronic lympholeukemia and prolymphocytic lymphosarcoma (Richter's syndrome)

Available are three cases of Richter's syndrome, i.e. immunoblastic lymphosarcoma with plasmocytic differentiation which developed in patients with chronic lymphocytic leukemia and prolymphocytic (lymphocytic) lymphosarcoma. Clinical, anatomical and autopsy findings are analysed. The immunoblastic lymphosarcoma is suggested to arise either because of transformation of the tumor prolymphocytes (lymphocytes) due to the disease progression or can be promoted by a second tumor.