免疫组织化学在淋巴组织疾病诊断中的应用与结果评价

目的提高对合理应用免疫组化标记及正确评价免疫组化结果重要性的认识。方法回顾性分析3例淋巴组织疾病的临床资料,进行组织形态学观察并增加相关抗体标记予以鉴别诊断。结果例l为弥漫大B细胞性淋巴瘤,梭形细胞变异型,误诊为未分化肉瘤与形态学观察遗漏中心和中心母细胞特征的淋巴样细胞,没考虑到淋巴瘤的梭形细胞形态变异以及免疫标记的选择失误有关。例2为急性淋巴结T区增生误诊为T细胞淋巴瘤与忽略患者临床信息及对免疫组化阳性细胞量变及强弱评价有误相关。例3为经典型霍奇金淋巴瘤误诊为间变性大细胞淋巴瘤与对霍奇金淋巴瘤组织形态认识不足及淋巴瘤标记抗体的配伍不当有关。结论淋巴组织疾病的诊断建立在形态学、免疫组化、临床资料和遗传学基础上,免疫组化标记的选择与结果的正确分析对淋巴瘤确诊至关重要。     

经典型霍奇金淋巴瘤CD30、CD15表达差异性的研究

探讨经典型霍奇金淋巴瘤(cHL)及其亚型的R-S细胞CD30、CD15蛋白表达的差异性.将32例cHL全部按WHO新分类标准进行病理分型,采用免疫组织化学SABC法进行CD30和CD15染色,观察每一病例及不同亚型R-S细胞阳性表达率.32例cHL中R-S细胞CD30阳性表达率为93.8%(30/32),CD15为78.1%(25/32),各亚型间均无显著性差异(P>0.05).CD30、CD15两者的阳性表达率无显著性差异性(P>0.05),但CD30的表达程度高于CD15(P<0.001).CD15和CD30可作为诊断cHL的参考依据,在R-S细胞中的表达差异可能与肿瘤细胞的异质性有关.     

滤泡树突细胞肉瘤二例报道并文献复习

目的探讨滤泡树突细胞肉瘤（FDCS）的临床表现、病理形态、免疫组化、生物学行为和预后,提高对该病的认识。方法对2例滤泡树突细胞肉瘤进行报道,并结合临床资料、肿瘤大体及镜下特征、免疫组化标记结果及预后和国内外文献进行分析。结果2例FDCS,1例发生于颈部淋巴结,另1例发生于肠系膜,镜检肿瘤由梭形及卵圆形细胞组成,呈束状、漩涡状、编织状排列;免疫组化显示肿瘤细胞CD21、CD23和CD35阳性;患者预后不良。结论滤泡树突细胞肉瘤是一种罕见且容易误诊的低度恶性肿瘤,组织化学和免疫组化染色有助于该肿瘤的诊断及鉴别诊断,减少误诊。     

肝脏假性淋巴瘤的临床病理学研究及分析

目的:探讨肝脏假性淋巴瘤的临床病理特征、免疫表型及鉴别诊断。方法:在光学显微镜下对肝脏假性淋巴瘤进行组织学形态观察,并借助免疫组化进一步对其形态进行分析。由于原发于肝脏的假性淋巴瘤极为罕见,故本文将报道一例发生于肝脏的假性淋巴瘤,结合文献探讨其临床病理特点,以提高诊断及鉴别诊断水平。结果:大体上为切面可见灰白结节,结节切面灰白色,质地中等,与周围分界清,周围肝组织灰红质软。显微镜下组织学表现为肝周边淋巴结淋巴组织增生,细胞大小较一致,核圆形,细胞无异型,未见明显核分裂,其中见嗜酸性白细胞浸润及少数异型细胞。免疫组化显示肿瘤细胞表达CD3、CD20及CD30。结论:肝脏假性淋巴瘤为罕见的、良性淋巴组织增生性病变,形态学特征、免疫组化染色在肝假性淋巴瘤诊断中具有重要价值。在临床病理实践中,必须首先与常见的发生于该部位恶性肿瘤如霍奇金淋巴瘤肿瘤等鉴别。     

EB病毒诱发人B细胞淋巴瘤的分子病理特性

EB病毒 (EBV ,Epstein Barrvirus)与人类多种肿瘤有关 ,尤其是与鼻咽癌和淋巴瘤的关系密切。为此研究了EB病毒在huPBL SCID嵌合体小鼠体内诱发人B细胞淋巴瘤的分子特性及肿瘤发生机制。从健康成人外周血分离出淋巴细胞 ,将之移植到SCID小鼠腹腔内 ,实验感染EBV ,观察肿瘤的形成 ;采用单向免疫扩散法连续监测小鼠血清中人IgG的含量。分别用PCR方法检测肿瘤组织中是否存在人Alu序列 ,原位杂交检测肿瘤组织中EB病毒小RNA分子EBER 1;免疫组织化学方法检测人白细胞分化抗原 (CD4 5、CD2 0、CD4 5RO、CD3) ,病毒基因(LMP1、EBNA2、BZLF1)的表达 ,以及细胞瘤基因蛋白 (p5 3、C myc、Bcl 2、Bax)在诱发肿瘤中的表达情况。结果发现 ,实验中 34只感染EBV的huPBL SCID小鼠有 2 4只诱发出肿瘤 ,根据病理形态学特征、Alu PCR和免疫标志均证实诱发瘤是人源性B淋巴细胞肿瘤。原位分子杂交显示肿瘤细胞核内存在EBER 1,少数瘤细胞表达EB病毒BZLF1蛋白阳性 ,部分瘤细胞表达LMP1和EBNA2蛋白阳性。连续监测 12只huPBL SCID小鼠血清中人IgG含量 ,发现IgG水平随诱瘤时间延长和肿瘤生长有逐渐增高趋势。免疫组化显示诱发的 2 4例淋巴瘤组织p5 3、C myc、Bcl 2和Bax蛋白表达阳性率分别为 83.33%、10 0 %、95 .83%、91.6 7%。结果     

咽部淋巴结外滤泡树突细胞肉瘤诊治分析

目的:探讨咽部淋巴结外滤泡树突细胞肉瘤(Follicular dendritic cell sarcoma,FDCS)的临床病理特点、生物学行为及诊断与治疗要点。方法:对1例咽部结外FDCS进行临床病理观察、免疫组织化学及原位杂交检测,并结合文献进行分析。结果:肿瘤组织位于咽部粘膜下,呈巢团状、片状排列,瘤组织间见多少不一的淋巴细胞浸润;免疫组化显示瘤细胞CD21、CD35、CXCL-13弥漫阳性,CD23、Podoplanin(D2-40)灶性阳性;EBER原位杂交阴性。结合文献分析发现咽部结外FDCS多发生于中青年,无明显性别差异。大多数肿瘤病变比较局限,体积相对较小,可能属于一类低度恶性肿瘤。由于对其认识不足,常常疏于考虑而误诊。据现有病例资料推荐采用根治性手术切除,目前尚没有证据支持术后的辅助放化疗可以改善预后。结论:咽部结外FDCS是一种罕见且容易误诊的低度恶性肿瘤,加深临床及病理医师对该肿瘤的认识有助于减少误诊。     

流式细胞术检测外周血淋巴细胞诊断淋巴瘤的应用研究

目的:探讨流式细胞术(FCM)检测外周血淋巴细胞在淋巴瘤诊断中的应用价值。方法:通过筛选2011年8月至2017年8月期间初诊的皮肤淋巴瘤病例25例,淋巴节良性病变6例,采用FCM检测外周血淋巴细胞表面抗原分子,通过与病理切片HE染色和免疫组化法(金标准)比较,分析两种检测方法之间的差异。结果:在31例检测病例中,FCM检测结果与金标准检测结果一致性较高(Kappa=0.61):26例检查结果相同,5例检查结果不一致;检测19例T淋巴细胞淋巴瘤,FCM检测结果与金标准检测结果一致性也较高(Kappa=0.57):检测14例初诊为T细胞淋巴瘤病例,FCM检测T淋巴瘤细胞的表面抗原标志CD3分子为阳性,与组织学结果相符,另有5例T细胞淋巴瘤病例HE染色和免疫组化诊断明确,而FCM未能检出。检测6例B细胞淋巴瘤病例,6例淋巴瘤病例FCM检测结果都为阳性,FCM检测B淋巴瘤细胞的表面抗原标志CD19分子为阳性,与金标准检测结果符合率为100%。6例淋巴节良性病变病例FCM检测结果与金标准检测结果一致。结论:通过FCM检测外周血可以检测出部分皮肤淋巴瘤,FCM在皮肤淋巴瘤诊断和分型中有一定的临床价值,是检测皮肤淋巴瘤的有效的辅助方法。     

免疫磁珠法分离鼻咽癌基质的T淋巴细胞

肿瘤内环境与肿瘤的发生密切相关.肿瘤细胞周围的组织在癌变发生时不会是一个沉默的旁观者,可能在肿瘤的发生和发展中扮演十分重要的角色.本研究分别采用不同的磁珠分选技术分离T淋巴细胞.采用CK LMP1,CD105和成纤维细胞表面蛋白,结合全血总T细胞试剂盒,间接法分离鼻咽癌基质的T淋巴细胞;采用CD3直接磁分选法分离鼻咽癌基质的T淋巴细胞,然后用免疫组化法鉴定分选的效果和细胞的质量.结果表明,免疫组化显示间接磁分选法分离出来的T淋巴细胞不能完全去除肿瘤细胞,RNA的质量不佳;而直接磁分选分离出来的T淋巴细胞为纯净的T淋巴细胞,而且RNA的质量良好.提示直接磁分选技术是分离鼻咽癌基质T淋巴细胞的首选方法.     

抗CD3/抗CD20双特异性单链抗体介导的B淋巴瘤细胞体外裂解作用

在构建并成功表达抗CD3/抗CD20双特异性单链抗体(bscCD3×CD20)的基础上,对其在体外介导T淋巴细胞杀伤Ramous B淋巴瘤细胞的生物活性进行了分析。Annexin V/PI(AV/PI)染色和形态学观察及扫描电镜分析表明bscCD3×CD20介导的B淋巴瘤细胞体外裂解作用是通过先诱导靶细胞凋亡而继发坏死、裂解的方式实现的。非放射性细胞毒性分析表明bscCD3×CD20介导的T淋巴细胞杀伤活性随抗体浓度、反应时间和效靶比的升高而增加。在抗体浓度为5μg/mL、作用时间为24h、效靶比为10∶1时,杀伤活性最高可达87·3%。采用美国SuperArray人细胞凋亡芯片检测细胞杀伤起始阶段细胞凋亡相关基因的表达水平变化,许多凋亡相关基因的表达均发生了不同程度的上调或下调,其中ATM基因表达升高了187倍,p53基因升高了15倍,提示ATM-p53途径可能是bscCD3×CD20介导T细胞诱导B淋巴瘤细胞凋亡的主要途径。     

甲状腺浆细胞瘤1 例及文献复习

目的:探讨甲状腺浆细胞瘤的病理学特点及临床表现。方法:对1例甲状腺浆细胞瘤进行组织学表现、免疫组化染色观察及文献复习。结果:组织学特点:肿瘤细胞成分单一,大多数为分化较为成熟的肿瘤性浆细胞,胞核圆形或卵圆形,大小较一致,常偏位,染色质呈车辐状或钟面状。免疫组化特点:肿瘤细胞表达CD20(++)、CD79a(+)、CD38(++)、CD138(+)、κ链(++)。结论:甲状腺浆细胞瘤是甲状腺一种少见的肿瘤,应同炎症反应的浆细胞增生及低分化癌、淋巴瘤、髓样癌等相鉴别,可以通过免疫组化染色及形态学观察进行鉴别诊断。     

Anaplastic large cell lymphoma of the mediastinum diagnosed by transbronchial scratch cytology. A case report

BACKGROUND: Anaplastic large cell lymphoma (ALCL) is a subtype of non-Hodgkin's lymphoma characterized by CD30 antigen-positive, large neoplastic cells. We describe a case of ALCL suggested by cytologic examination of the tumor cells obtained from bronchial scratch preparations. CASE: A 26-year-old woman had had a dry cough since November 1996. Chest radiography in May 1997 revealed an abnormal shadow in the mediastinum extending to the pulmonary hilar region. The patient was hospitalized in June 1997. Computed tomography revealed a neoplastic lesion in the anterior mediastinum invading the right lung. Transbronchial scratch cytology revealed large, atypical lymphoid cells expressing CD30 and CD3 on immunocytochemical examination. A transcutaneous mediastinal biopsy was performed and a diagnosis of ALCL made. CONCLUSION: Differentiation from Hodgkin's disease was the most difficult point in this case. Detailed cytologic observation and CD3-positive immunocytology led to the correct diagnosis. The cell transfer technique of Sherman et al was very useful for immunocytologic staining. Thus, transbronchial scratch cytology was an especially valuable and effective procedure in this case.     

Olfactory neuroblastoma. Cytodiagnostic features in a case with ultrastructural and immunohistochemical correlation

In a case of olfactory neuroblastoma, originally misdiagnosed as an undifferentiated carcinoma, cytologic examination of material scraped from the superior nasal vault revealed tumor cells suggestive of neuroblastoma. The most significant cytodiagnostic feature was the presence of a fibrillary cytoplasm with ill-defined borders. Also noteworthy were the smudged hyperchromatic nuclei and structures resembling rosettes or pseudorosettes. The diagnosis was confirmed by electron microscopy, which revealed the presence of dense-core neurosecretory granules, clear vesicles, neurotubules and neurofilaments, and by immunohistochemistry, which showed positive staining for neuron-specific enolase but negative staining for keratin and glial fibrillary acidic protein. Since olfactory neuroblastoma has a relatively good prognosis and aggressive surgical resection may be curative, it is important that this tumor be distinguished from other small cell malignancies arising in the nasal cavity. The present case shows that the diagnosis can be made by the cytologic examination of scrapings from the tumor.     

Study of Regulatory T-Cells in Patients with Gastric Malt Lymphoma: Influence on Treatment Response and Outcome

Purpose

FOXP3+ regulatory T cells (Treg) play an essential role in modulating host responses to tumors and infections. The role of these cells in the pathogenesis of MALT lymphomas remains unknown. The aims of the study were to quantify the number of infiltrating FOXP3+ and CD3+ cells in patients with gastric MALT lymphoma at diagnosis and to study kinetics of these cells and CD20+ tumor cells after treatment and during long-term follow-up.

Methods

FOXP3+, CD3+ and CD20+ cells were analyzed by immunohistochemistry and the number of cells was quantified using a micrometric ocular. Samples of 35 patients with gastric MALT lymphoma at diagnosis and after treatment were included. Diagnostic samples were compared to 19 cases of chronic gastritis and diffuse large B-cell lymphoma (DLBCL) of the stomach.

Results

The median number of FOXP3+ infiltrating cells was higher (27 cells/cm²) in gastric MALT patients than in DLBCL (10 cells; p = 0.162) but similar to chronic gastritis (20 cells; p = 0.605). No characteristic or specific distribution pattern of infiltrating FOXP3+ cells was found. Gastric MALT lymphoma patients responding to bacterial eradication therapy had higher number of FOXP3+ cells at study entry. Kinetics of both infiltrating FOXP3+ cells and tumor CD20+ cells were strongly dependent on the treatment administered.

Discussion

Gastric MALT lymphomas have a number of Treg cells more similar to chronic gastritis than to DLBCL. Patients with higher number of tumor infiltrating FOXP3+ cells at study entry seem to have better response to antibiotics. Kinetics of Treg and tumor cells are influenced by type of treatment.     

Clinical Significance of sIL-2R Levels in B-Cell Lymphomas

Elevated soluble interleukin-2 receptor (sIL-2R) in sera is observed in patients with malignant lymphoma (ML). Therefore, sIL-2R is commonly used as a diagnostic and prognostic marker for ML, but the mechanisms responsible for the increase in sIL-2R levels in patients with B-cell lymphomas have not yet been elucidated. We first hypothesized that lymphoma cells expressing IL-2R and some proteinases such as matrix metalloproteinases (MMPs) in the tumor microenvironment can give rise to increased sIL-2R in sera. However, flow cytometric studies revealed that few lymphoma cells expressed IL-2R α chain (CD25) in diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL), and most CD25-expressing cells in the tumor were T-cells. Distinct correlations between CD25 expression on B-lymphoma cells and sIL-2R levels were not observed. We then confirmed that MMP-9 plays an important role in producing sIL-2R in functional studies. Immunohistochemical (IHC) analysis also revealed that MMP-9 is mainly derived from tumor-associated macrophages (TAMs). We therefore evaluated the number of CD68 and CD163 positive macrophages in the tumor microenvironment using IHC analysis. A positive correlation between the levels of sIL-2R in sera and the numbers of CD68 positive macrophages in the tumor microenvironment was confirmed in FL and extranodal DLBCL. These results may be useful in understanding the pathophysiology of B-cell lymphomas.     

Angiocentric T-cell lymphoma

SUMMARY: The CD20+ variant of angiocentric T-cell lymphoma is an unusual type of T-cell lymphomas that present cystic changes in organs because of ischaemic necroses. The purpose of this study was to describe a case of CD20+angiocentric T-cell lymphoma, discussing its clinical, histopathological and immunohistochemical features, to analyze its proliferation kinetics and to consider its possible relationship to the Epstein-Barr virus (EBV) to understand better the pathobiological nature of the disease. METHODS: The clinical, histopathological, immunohistochemical and single-cell DNA cytophotometric features of the case were analyzed. In addition in situ hybridization was performed to detect EBV. RESULTS: The 24 years old woman was admitted to our Institute because of pain in the abdominal region and weight loss. There were enlarged lymph nodes on the neck, and biopsy was done. Histological diagnosis: angiocentric T-cell lymphoma, CD20+ variant. CD3, CD43, CD45RA and CD45R0 antigens were positive in the atypic lymphoid cells of the tumour and in cells infiltrating the vascular wall. DNA index was 0.8589 (hypodiploid). Tumour cells in G1 phase: 47%, S phase: 45.4%, G2 phase: 7.6%. Combined chemotherapy was administered because of clinical stadium IV/B of malignant lymphoma (5 CHOP-Bleo, CEPP, CEP, CMVE treatment). The disease showed gradual progression and the patient died 14 months after the first symptoms had appeared. CONCLUSIONS: In the last 13 years there were 5 cases of angiocentric T-cell lymphoma at our Institute. The CD20+ variant is rare, its clinical symptoms are special, the prognosis is unfavourable. The cause why we demonstrate this case is to call attention to a new treatment for these patients by immunotherapy using monoclonal antibodies against CD20 antigen.     

Intraabdominal follicular dendritic cell sarcoma: report of a case with fine needle aspiration findings.

BACKGROUND: Follicular dendritic cell sarcoma is a rare tumor derived from dendritic cells of lymphoid follicles. Although it shows characteristic cytologic, histologic and immunohistochemical features, it may be misdiagnosed because of the lack of experience with this entity. There are few reports in the cytologic literature describing this entity. CASE: A left hypochondrial mass was found in an 80-year-old man who had a past history of colonic carcinoma. A computed tomography-guided fine needle aspiration biopsy was performed, and the findings were initially thought to be compatible with metastatic carcinoma. The mass was excised, and a diagnosis of follicular dendritic cell sarcoma was made. CONCLUSION: Follicular dendritic cell sarcoma is characterized by loosely cohesive or syncytial groups of dendritic cells that show oval, vesicular nuclei and ill-defined cytoplasm intimately admixed with small, mature lymphocytes. The diagnosis can be confirmed by positive immunostaining for CD21 and CD35 antigens and negative staining for cytokeratin. Unusual features in our case were the presence of prominent plasma cells and intracytoplasmic, periodic acid-Schiff stain, diastase-positive deposits, with the last not described before.     

Expression of glucose transporter-1 in follicular lymphoma affected tumor-infiltrating immunocytes and was related to progression of disease within 24 months

ObjectiveFollicular lymphoma (FL) occurring progression within 24 months (POD24) after initial immunochemotherapy has poor prognosis. GLUT1 affects glycolysis within tumor microenvironment (TME) and promotes tumor progression. However, its specific mediated mechanism remains unclear in FL.MethodsBaseline GLUT1 expression, infiltrations of M2 macrophage, and CD8+ T-cells were assessed by immunohistochemistry in FL with POD24 and long-term remission respectively. The spatial features of TME were assessed by MIBI-TOF and proteomics. Predictive immunophenotypes for POD24 occurrence was analyzed by random forest algorithm. The lactate production and the induction of M2 macrophages were detected when GLUT1 was transfected or knocked down in DOHH2. The activation of PI3K/Akt/mTOR signaling in DOHH2 and WSU-FSCCL cells co-cultured with induced inhibitory immunocytes was tracked by western blotting.ResultsThe FL with POD24 exhibited higher baseline GLUT1 expression and increased infiltration of various inhibitory immunocytes. Spatial signatures of 69 immunophenotypes could predict POD24 occurrence. The activation of PI3K/ Akt /mTOR signaling pathway was not significant in both groups. The supernatant of DOHH2-GLUT1 cells which had more lactate content could induce more M2-type macrophages than that of DOHH2/siRNA GLUT1 cells. When co-cultured with exhausted CD8+ T cells, M2-type macrophages and Tregs, compared with WSU-FSCCL cells, DOHH2 cells with high GLUT1 expression induced more M2-type macrophages and was triggered activation of PI3K/ Akt /mTOR signaling pathway.ConclusionTumor cells overexpressing GLUT1 could domesticate immunocytes to form an immunosuppressive TME, which promotes occurrence of POD24 and gradually activates PI3K/ Akt /mTOR pathway of tumor cells in FL.SignificanceTumor cells overexpressing GLUT1 could domesticate immunocytes to form an immunosuppressive microenvironment, which in turn promoted the growth of tumor cells and was related to the progression of disease within 24 months in FL. Suppressive immunocytes gradually activated PI3K/ Akt /mTOR pathway of tumor cells in later stage. Distinguishing spatial features of immunocytes could well predict POD24 occurrence, hoping to benefit these patients from early anti-metabolism therapy based on GLUT1 in the future.     

Intravascular lymphomatosis of the brain. Report of a case using an intraoperative cytologic preparation

BACKGROUND: Intravascular lymphomatosis (IVL) of the brain is a rare non-Hodgkin's lymphoma characterized by proliferation of tumor cells in the lumina of blood vessels. Although an intraoperative cytologic examination of the brain is routinely done, the cytologic features of IVL are rarely encountered by pathologists. We report a case of IVL diagnosed by an intraoperative cytology preparation. CASE: A 62-year-old woman presented with a seizure and fever. Computed tomography and magnetic resonance imaging of the brain were insufficient to establish a diagnosis but suggested a cerebral infarction, so a stereotactic brain biopsy was performed. On an intraoperative cytologic examination, a few small and medium-sized vessels were filled with tumor cells showing atypical, pleomorphic, large nuclei. Immunohistochemical staining using paraffin-embedded tissue revealed intraluminal tumor cells expressing leukocyte common antigen and CD20 but not CD3. CONCLUSION: This disease must be considered one of the diagnostic possibilities in any patient with rapidly progressive dementia and cerebral infarction. Increased awareness of this disease and intraoperative early diagnosis are important for its successful management.     

Immunofluorescent labeling of CD20 tumor marker with quantum dots for rapid and quantitative detection of diffuse large B-cell non-Hodgkin's lymphoma

Fluorescent semiconductor quantum dots (QDs) are newfound nanocrystal probes which have been used in bioimaging filed in recent years. The purpose of this study is to evaluate the diagnostic value of specific QDs coupled to rituximab monoclonal antibody against CD20 tumor markers for patients with diffuse large B-cell lymphoma (DLBCL). In current study rituximab-conjugated quantum dots (QDs-rituximab) were prepared against CD20 tumor markers for detection of CD20-positive cells (human Raji cell line) using flowcytometry. A total of 27 tumor tissue samples were collected from patients with DLBCL and 27 subjects with negative pathological tests as healthy ones, which stained by QD-rituximab. The detection signals were obtained from QDs using fluorescence microscopy. The flowcytometry results demonstrated a remarkable difference in fluorescent intensity and FL2-H + (CD20-positive cells percentage) between two groups. Both factors were significantly higher in Raji in comparison with K562 cell line (P < 0.05). Lot of green fluorescence signals was observed due to the selectively binding of QD-rituximab to CD20 tumor markers which overexpressed in tumor tissues and a few signals observed on the defined healthy ones. Based on these observations the cut-off point was 46.8 dots and the sensitivity, specificity, positive predictive value, and negative predictive value were 100%, 89.5%, 91.3%, and 100%, respectively (LR+, 9.52; LR−, 0). The QD - rituximab could be beneficial as a bioimaging tool with high sensitivity to provide an accurate molecular imaging technique for identifying CD20 tumor markers for early diagnosis of the patients with DLBCL.