Immunophenotypic features and t(14;18) (q32;q21) translocation of Chinese follicular lymphomas helps to distinguish subgroups.

BACKGROUND: The revised 2008 World Health Organization classification maintains a histological grading system (grades 1-3) for follicular lymphoma (FL). The value of grading FL has been debated. This study will yield deeper insights into the morphologic, immunophenotypic characterization and t(14;18) translocation in FL and explore their significance of diagnosis of Chinese FL subgroups. METHODS: We retrospectively reviewed the FL diagnoses according to the 2008 WHO classification in all diagnostic specimens from a multicentric cohort of 122 Chinese patients. Upon review, 115 cases proved to be truly FL. CD10, BCL6, MUM1, BCL2 and t(14;18) (q32;q21) translocation were detected by Envision immunostaining technique and fluorescence in situ hybridization. RESULTS: FL1 has larger proportion of follicular pattern (93.0%) than that of FL2 (73.7%, P = 0.036), FL3B (63.6%, P = 0.003) and FL3A (77.4%, P = 0.053), although the last P value was more than 0.05 (Pearson's chi-squared test). Areas of DLBCL were present in 25.8% (8/31) of FL3A and more frequent in FL3B (59.1%, 13/22; P = 0.015). The positivity of CD10 and BCL2 in FL1-2 were significantly higher than those in FL3 (P < 0.001, P = 0.043, respectively). The positivity of MUM1 in FL1-2 was significantly lower than that in FL3 (10.2% vs. 51.0%; P < 0.001). Furthermore the positivity of MUM1 in FL3A was significantly lower than that in FL3B (37.9% vs. 68.2%; P = 0.032). The positivity of t(14;18) was higher in FL1-2 than in FL3 (73.5% vs. 35.6%, P < 0.001), and was higher in FL3A than in FL3B (51.9% vs. 11.1%, P = 0.005). t(14;18) was significantly correlated with CD10+ (R = 0.453, P < 0.001) and MUM1+ (R = -0.482, P < 0.001). CONCLUSIONS: FL1 and FL2 were immunophenotypically and genomically similar, while FL3A and FL3B were partly immunophenotypically similar but morphologically, genomically distinct. FL3A was genomically closer to FL1-2, whereas FL3A was genomically closer DLBCL. Thus we hypothesize that FL may in fact be a heterogeneous indolent lymphoma encompassing entities with distinct molecular pathogenesis and genetic characteristics. Immunohistochemical and genetic characterization helps to distinguish subgroups of FLs. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1334018129864616.

[To emphasize the importance of specimen fixation and improve quality of diagnosis of gastrointestinal stromal tumor].

The most common problem in the diagnosis of gastrointestinal stromal tumor (GIST) is inadequate specimen fixation. The paper focused on specimen fixation and standardized protocol in immunohistochemistry staining and gene mutation detection. We have adjusted some procedure used in immunohistochemistry staining and c-kit gene detection to improve the quality of inadequately fixed specimen. It maybe useful for clinicians, pathologists and technicians working in immunohistochemistry labs and gene detection labs.

Primary cutaneous blastic plasmacytoid dendritic cell neoplasm without extracutaneous manifestation: case report and review of the literature.

Blastic plasmacytoid dendritic cell (BPDC) neoplasm is a rare, highly aggressive hematopoietic malignancy with involvement of bone marrow and peripheral blood. We present 2 cases of primary cutaneous BPDC neoplasm without extracutaneous manifestation during the course of disease. A 36-year-old and a 51-year-old male presented with erythematous patches, purple plaques, and nodules on the head, trunk, and extremities. Skin biopsies revealed that the lesions of both cases were composed of diffusely medium-sized monomorphic blastoid cells infiltrating into the dermis and subcutis. The neoplastic cells were strongly positive for CD4, CD56, CD123, and TdT, whereas other T-cell markers and EBV markers were not expressed. The patients underwent polychemotherapy with hyper-CVAD regimen and obtained a remarkable clinical response with regression of skin lesions. No sign of recurrence and extracutaneous manifestation was found during the period of follow-up. We presume that the favorable prognosis of our cases might result from the presentation only with a skin lesion, diffuse TdT expression in tumor cells, and aggressive chemotherapy with hyper-CVAD regimens. Laboratory examination for blood and bone marrow should be performed every 3-6 months during the first period of follow-up to monitor the progression of disease even if the patients had complete remission at initial chemotherapy.

[Clinicopathologic study of 963 cases of mature T-cell and natural killer/T-cell lymphoma with respect to 2008 WHO classification of lymphoid neoplasms].

OBJECTIVE: To study the clinicopathologic features of various types of mature T-cell and natural killer (NK)/T-cell lymphoma in Guangdong, China, with respect to the 2008 WHO classification of lymphoid neoplasms. METHODS: Eleven hundred and thirty-seven (1137) cases of mature T-cell or NK/T-cell lymphoma diagnosed during the period from 2002 to 2006 in Guangzhou area were retrieved. The clinical data, histologic features and immunohistochemical findings were reviewed by a panel of experienced hematopathologists. Additional immunostaining was performed if indicated. The cases were re-classified according to the 2008 WHO classification of lymphoid neoplasms. RESULTS: Nine hundred and sixty-three (963) cases fulfilled the diagnostic criteria of mature T-cell or NK/T-cell lymphoma and accounted for 20.1% of all cases of lymphoma encountered during the same period (963/4801). A predominance of extranodal involvement was noted in 644 cases (66.9%), while 319 cases (33.1%) showed mainly nodal disease. The prevalence of various lymphoma subtypes was as follows: peripheral T-cell lymphoma, unspecified (PTCL, NOS) 293 cases (30.4%), extranodal NK/T-cell lymphoma, nasal type 281 cases (29.2%), anaplastic large cell lymphoma (ALCL) 198 cases (20.6%), and angioimmunoblastic T-cell lymphoma (AILT) 46 cases (4.8%). The male-to-female ratio was 1.99. The median age of the patients was 44 years, with the peak age of PTCL, NOS, extranodal NK/T-cell lymphoma, nasal type and AILT being 55 to 64 years, 25 to 54 years and 65 to 74 years, respectively. ALK-positive ALCL occurred more frequently in young age, while the ALK-negative ALCL cases occurred mainly in the elderly. CONCLUSIONS: Extranodal lesions predominate in mature T-cell and NK/T-cell lymphomas occurring in Guangzhou area. There is a male predominance and the overall incidence shows no increasing trend with age of the patient. The peak age of various subtypes however varies. The most common subtype was PTCL, NOS, followed by extranodal NK/T-cell lymphoma, nasal type, ALCL and AILT. The relatively frequent occurrence of extranodal NK/T-cell lymphoma, nasal type in Guangdong area is likely associated with the high incidence of Epstein-Barr virus infection there.

[Clinical features and prognosis of mucosa-associated lymphoid tissue lymphoma: a report of 90 cases].

BACKGROUND AND OBJECTIVE: Mucosa-associated lymphoid tissue lymphoma is a histological type of marginal zone non-Hodgkin's lymphoma (NHL). Its clinical features and prognosis have seldom been reported because of its indolent clinical course. This study was to explore the clinical features and prognosis of this disease. METHODS: Clinical data of 90 pathologically confirmed mucosa-associated lymphoid tissue lymphoma patients, treated from December 1997 to February 2007, were analyzed. RESULTS: Of the 90 patients, 23 (25.6%) had gastric lymphoma and 67 (74.4%) had non-gastric lymphoma, with a median age of 52 (range, 13-77); 75 (83.3%) had stage I-II disease and 15 (16.7%) had stage III-IV disease; 31 (34.4%) had multiple organ involvement and 40 (44.4%) had nodal involvement. The percentage of nodal involvement was significantly higher in non-gastric group than in gastric group (P=0.040). The complete remission (CR) rate after treatment was 72.1%. The patients were followed up for a median of 31.4 months. The 5-year overall survival rates of patients with and without nodal involvement were 58.7% and 88.4%, respectively (P=0.012). The median time to progression was significantly longer in patients with IPI score of 0-2 than in those with IPI score of > 2 (61.9 months vs. 5.2 months, P=0.005), and was significantly longer in patients who got CR after initial treatment than in those without CR (not reached vs. 15.0 months, P=0.030). In non-gastric lymphoma group, IPI score was an independent prognostic variable of overall survival (P=0.023). CONCLUSIONS: Mucosa-associated lymphoid tissue lymphoma should be considered as a kind of disseminated indolent lymphoma. The patients with non-gastric lymphoma are likely to have nodal involvement. Patients with poor prognostic factors should be treated more aggressively.

Effects of aspirin on the development of Helicobacter pylori-induced gastric inflammation and heterotopic proliferative glands in Mongolian gerbils.

BACKGROUND: Helicobacter pylori infection is a major cause of gastritis and gastric carcinoma. Aspirin has anti-inflammatory and antineoplastic activity. The aim of the present study was to determine the effects of aspirin on H. pylori-induced gastritis and the development of heterotopic proliferative glands. METHODS: H. pylori strain SS1 was inoculated into the stomachs of Mongolian gerbils. Two weeks after inoculation, the animals were fed with the powder diets containing 0 p.p.m. (n = 10), 150 p.p.m. (n = 10), or 500 p.p.m. (n = 10) aspirin. Mongolian gerbils were killed after 36 weeks of infection. Uninfected Mongolian gerbils (n = 10) were used as controls. Histologic changes, epithelial cell proliferation and apoptosis, and prostaglandin E(2) (PGE(2)) levels of gastric tissue were determined. RESULTS: H. pylori infection induced gastric inflammation. Administration of aspirin did not change H. pylori-induced gastritis, but alleviated H. pylori-induced hyperplasia and the development of heterotopic proliferative glands. Administration of aspirin accelerated H. pylori-associated apoptosis but decreased H. pylori-associated cell proliferation. In addition, the increased gastric PGE(2) levels due to H. pylori infection were suppressed by treatment with aspirin, especially at the dose of 500 p.p.m. CONCLUSIONS: Aspirin alleviates H. pylori-induced hyperplasia and the development of heterotopic proliferative glands. Moreover, aspirin increases H. pylori-induced apoptosis. We demonstrated the antineoplastic activities of aspirin in H. pylori-related gastric carcinogenesis.

Expression of peroxisome proliferator-activated receptor gamma, E-cadherin and matrix metalloproteinases-2 in gastric carcinoma and lymph node metastases.

BACKGROUND: Peroxisome proliferator activated receptor gamma (PPARgamma) is a ligand-activated transcription factor. Activation of PPARgamma has recently been demonstrated to inhibit various tumor cells growth, progression and metastasis. E-cadherin-mediated cell adhesion system is now considered to be an "invasion suppressor system" in cancer tissues. Matrix metalloproteinases-2 (MMP-2) is a prerequisite for metastasizing tumor cells. However their correlation is still unknown in gastric carcinoma. The aim of this study was to assess the expression of PPARgamma, E-cadherin, MMP-2 and their correlation in gastric carcinoma and metastases. METHODS: Gastric carcinoma tissues and their corresponding lymph nodes with metastases and the adjacent non-tumor tissues were obtained from 54 patients with gastric cancer who underwent gastrectomy. Expression of PPARgamma, E-cadherin and MMP-2 was assessed by immunohistochemical staining. RESULTS: The nuclear expression level of PPARgamma in neoplastic cells was significantly lower than that in the normal controls (P < 0.001), with the expression of PPARgamma being weaker in primary tumors compared with that in metastases. In all neoplastic cells, E-cadherin was expressed with abnormal patterns (cytoplasm pattern, cytoplasm and membrane pattern or absent), compared with normal cells where E-cadherin was expressed with a normal pattern (membrane pattern). Compared with the normal tissues, the expression level of E-cadherin decreased in primary tumors and further decreased in metastases (P < 0.001). Membrane staining of MMP-2 was detected in the foveolar epithelia of normal gastric mucosa, whereas predominant cytoplasm staining of MMP-2 was found in malignant tissues. The expression of MMP-2 was stronger in metastatic tissues than in primary tumors. In neoplastic foci the expression of PPARgamma was negatively correlated with MMP-2 expression (P < 0.05). However, there was no correlation between E-cadherin and PPARgamma or MMP-2 expression. CONCLUSIONS: Down-regulation of PPARgamma and E-cadherin and up-regulation of MMP-2 in neoplastic foci might be helpful to gastric carcinogenesis and metastases. An inverse relationship between PPARgamma and MMP-2 in human gastric carcinoma suggests that PPARgamma might modulate MMP-2 expression and affect gastric cancer metastases.

[Expression of syndecan-1 at different stages in the course of gastric carcinoma and its significance].

OBJECTIVE: To investigate the expression of syndecan-1 protein at different stages in the course of gastric carcinoma and its significance in carcinogenesis and metastasis. METHODS: There were 56 cases of chronic gastritis, 50 cases of chronic atrophic gastritis, 59 cases of intestinal metaplasia, 61 cases of displasia, and 112 cases of gastric carcinoma. Among the carcinoma cases, 55 were without and 57 with lymph node metastases. All paraffin-embedded tissue samples were assessed by immunohistochemistry. RESULTS: The syndecan-1 positive rate was 96.43% (54/56) in gastritis, 98.00% (49/50) in chronic atrophic gastritis, 100.00% (59/59) in intestinal metaplasia, 91.80% (56/61) in displasia, 45.45% (25/55) in gastric carcinoma without, and 24.56% (14/57) in gastric carcinoma with lymph node metastases. There was no significant difference among chronic gastritis, chronic atrophic gastritis and intestinal metaplasia (P > 0.05). There was a significant difference between displasia group and gastric carcinoma group (P <0.05), as well as between gastric carcinoma with and without lymph node metastases. There was a significant difference among well, moderately and poorly differentiated carcinoma groups. CONCLUSION: A decreasing expression of syedecan-1 in the development of gastric carcinoma is related with gastric carcinogenesis, and it may further promote metastasis of gastric carcinoma.

[Subgrouping and outcome prediction of diffuse large B-cell lymphoma by immunohistochemistry].

OBJECTIVE: To categorize diffuse large B-cell lymphoma (DLBCL) into germinal center B cell-like (GCB) and non-germinal center B cell-like (non-GCB) subgroups by immunohistochemistry; and to investigate the underlying prognostic significance. METHODS: Immunohistochemical study for CD10, bcl-6 and MUM1 was performed on 133 cases of DLBCL. The cases were then categorized into GCB and non-GCB subgroups. The 5-year overall survival and 5-year progression-free survival rates were compared between the GCB and non-GCB groups, and among the cases with different immunohistochemical expression or with different IPI. RESULTS: Amongst the 133 case studied, CD10 was expressed in 33.1%, while bcl-6 was positive in 34.6% and MUM1 in 45.1%. CD10 expression had a favorable impact on 5-year overall survival (P=0.041) and 5-year progression-free survival (P=0.031). On the other hand, bcl-6 expression had a favor able impact on 5-year progression-free survival (P=0.044). Expression of MUM1 carried an adverse effect on 5-year overall survival (P=0.031) and 5-year progression-free survival (P=0.028). GCB immunophenotype was demonstrated in 40.6% of the cases, while 59.4% showed a non-GCB profile. GCB DLBCL had a significantly longer 5-year overall survival (P=0.004) and 5-year progression-free survival (P=0.003), as compared with the non-GCB group. When dividing the cases into two groups according to their IPI score (IPI=0 to 1 and IPI=2 to 5), it turned out that the 5-year overall and progression-free survival rates of the GCB group were significantly higher than those of the non-GCB group (P=0.019 and 0.014 respectively in cases with IPI of 0 to 1 and P=0.006 and 0.009 respectively in cases with IPI of 2 to 5). The non-GCB cases with a IPI of 2 to 5 had the poorest prognosis. CONCLUSION: DLBCL subgrouping by immunohistochemistry and analysis of the subgrouping with IPI is feasible and useful in predicting clinical outcome.

[Effect of shRNA-mediated survivin gene silencing on apoptosis and proliferation of leukemia cell line].

OBJECTIVE: To transfect a short hairpin RNA (shRNA) against survivin gene into human T lymphoblastic leukemia cell line Jurkat, and to explore the effects on apoptosis and proliferation of transfected cells. METHODS: The survivin-shRNA expression vector were constructed and transfected into Jurkat cells. Expression of survivin mRNA and protein were assessed by RT-PCR and Western blot analysis respectively. Apoptosis index of transfected Jurkat cells was quantified by flow cytometry. The potential of cell proliferation was described by cell growth curves. RESULTS: In survivin-shRNA transfected Jurkat cells, survivin mRNA levels were significantly reduced by 66.67% ( transient transfection) and 60.69% ( stable transfection) respectively, compared with that in control-shRNA treated group and PBS treated group (P < 0.05); and the levels of survivin protein were significantly reduced by 63.41% (transient transfection) and 60.18% (stable transfection), compared with that in the two control groups (P < 0.05). Apoptosis index was significantly increased during both transient and stable transfection, respectively [(22. 41 +/- 2.83)% and (20.73 +/- 2.56)% (P < 0.05)]. Survivin-shRNA also inhibited the proliferation of Jurkat cells. CONCLUSIONS: Vector-based survivin-shRNA can effectively reduce the expression of survivin gene, induce apoptosis

Knockdown of survivin gene by vector-based short hairpin RNA technique induces apoptosis and growth inhibition in Burkitt's lymphoma Raji cell line.

Survivin is the smallest member of mammalian IAP (inhibitor of apoptosis) family. It is ubiquitous during embryonic development but is not expressed in normal post-natal tissues, except the thymus, colonic epithelial cells and CD34+ hematopoietic stem cells. However, its expression is upregulated during neoplastic transformation in both solid organ and hematological malignancies, including leukemia and lymphoma. In this study, we used RNA interference with short hairpin RNA (shRNA) technique to inhibit survivin expression in a Burkitt's lymphoma cell line Raji and validated its effects on apoptosis and cell proliferation. A survivin-shRNA expression vector were constructed and introduced into Raji cells. Expression of survivin mRNA and protein was assessed by RT-PCR and western blot analysis. Apoptosis index of transfected cells was quantified by flow cytometry and cell proliferation was enumerated by trypan blue exclusion. In Raji cells treated with survivin-shRNA expression vector, survivin mRNA levels were significantly reduced by 67.14% (transient transfection) and 64.28% (stable transfection) respectively, compared with control-shRNA treated group and PBS treated group (p<0.05). The levels of survivin protein were significantly reduced by 62.50% (transient transfection) and 60.93% (stable transfection), compared with the two control groups (p<0.05). Apoptosis index was significantly increased during transient transfection and stable transfection, respectively 31.20+/-2.45% and 29.40+/-1.72% (p<0.05). Survivin-shRNA inhibited the proliferation of Raji cells of stable transfection. In conclusion, the vector-based survivin-shRNA can effectively reduce the expression of survivin gene and induce apoptosis and growth inhibition of transfected Raji cells. We suggest that survivin can be regarded as an ideal target for new anticancer intervention of NHL.

Effects of cyclooxygenase-1 and -2 gene disruption on Helicobacter pylori-induced gastric inflammation.

BACKGROUND: Cyclooxygenases (COXs) play important roles in inflammation and carcinogenesis. The present study aimed to determine the effects of COX-1 and COX-2 gene disruption on Helicobacter pylori-induced gastric inflammation. METHODS: Wild-type (WT), COX-1 and COX-2 heterozygous (COX-1+/- and COX-2+/-), and homozygous COX-deficient (COX-1-/- and COX-2-/-) mice were inoculated with H. pylori strain TN2 and killed after 24 weeks of infection. Uninfected WT and COX-deficient mice were used as controls. Levels of gastric mucosal inflammation, epithelial cell proliferation and apoptosis, and cytokine expression were determined. RESULTS: COX deficiency facilitated H. pylori-induced gastritis. In the presence of H. pylori infection, apoptosis was increased in both WT and COX-deficient mice, whereas cell proliferation was increased in WT and COX-1-deficient, but not in COX-2-deficient, mice. Tumor necrosis factor (TNF)-alpha and interleukin-10 mRNA expression was elevated in H. pylori-infected mice, but only TNF-alpha mRNA expression was further increased by COX deficiency. Prostaglandin E2 levels were increased in infected WT and COX-2-deficient mice but were at very low levels in infected COX-1-deficient mice. Leukotriene (LT) B4 and LTC4 levels were increased to a similar extent in infected WT and COX-deficient mice. CONCLUSIONS: COX deficiency enhances H. pylori-induced gastritis, probably via TNF-alpha expression. COX-2, but not COX-1, deficiency suppresses H. pylori-induced cell proliferation.

[Differential diagnosis between nodular lymphocyte-predominant Hodgkin lymphoma and T-cell/histiocyte-rich B-cell lymphoma].

OBJECTIVE: To study the differential diagnosis between nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) and T-cell/histiocyte-rich B-cell lymphoma (TCRBCL). METHODS: 15 cases of NLPHL and 16 cases of TCRBCL were studied on both morphology and immunophenotype according to the WHO classification of lymphoid neoplasms. SP-immunohistochemical staining were performed on paraffin sections. In situ hybridization for EBER1/2 and gene rearrangement of immunoglobulin heavy chain (IgH) were carried out in 3 cases of NLPHL and 4 cases of TCRBCL, respectively. RESULTS: Histologically, a few atypical large cells scattered in a background of small lymphocytes with or without histiocytes were a common finding in both NLPHL and TCRBCL. Of NLPHL, nodular pathern predominated in 11 cases, diffuse patterns without nodules in 3 cases and one case showed nodular and diffuse pattern intermixed with a increased number of large cells. 14 cases of TCRBCL showed diffuse pattern. One case with micronodular pattern involving the splenic white pulp. One case showed a combination of nodules of NLPHL, diffuse areas of TCRBCL and a sheet of large cells of diffuse large B-cell lymphoma (DLBCL) within the same lymph node biopsy specimen. Immunophenotypically, the large cells showed and CD20, CD79a, bcl-6 and EMA positive, and CD15, CD30, CD3, CD45RO and LMP-1 negative. In NLPHL, small B cells and CD57 positive cells were common, whereas in TCRBCL, TIA-1 positive cytotoxic cells and histiocytes dominated, small B cells were scarce or absent. EBER1/2 were negative and gene rearrangement of IgH was found in all tested 3 cases of NLPHL and 4 cases of TCRBCL, respectively. CONCLUSION: There are some morphologic and immunophenotypic resemblance between NLPHL and TCRBCL. A combination of the morphological characteristics and the reactivity of the background cells for CD57 and TIA-1 seem to reliably discriminate between the entities and should therefore help to increase the interobserver reproducibility of diagnosis in the gray zone around Hodgkin lymphomas.

Correlation of AIB1 overexpression with advanced clinical stage of human colorectal carcinoma.

AIB1, a member of the steroid receptor coactivator 1 family, has been cloned on 20q12 and is a candidate oncogene in human breast cancer. It is commonly amplified and overexpressed in several types of human cancers. In this study, we examined the expression of AIB1, as related to clinicopathologic features, in 85 human colorectal cancers (CRCs). The status of the number of AIB1 copies, p53 expression, and DNA ploidy was also analyzed. The overexpression of AIB1 was detected in 35% of CRCs. Amplification of AIB1 was observed in 10% of CRCs. In addition, the overexpression of AIB1 was observed more frequently in CRCs in later clinical stages (T3 N1 M0/T3 N0 2M1), compared with that in T3 N0 M0 stage (P < .05). These results suggest that overexpression of AIB1 might provide a selective advantage for the developmental growth and/or progression of subsets of CRCs. In addition, a significant correlation (P < .05) of overexpression of AIB1 with p53 overexpression as well as with aneuploid DNA content was observed in these CRCs. The overexpression of p53 was also correlated significantly with CRC DNA ploidy (P < .05). Furthermore, there was a substantial population of CRCs showing overexpression of both AIB1 and p53 protein and all had aneuploid DNA content; most of these were in the later clinical stage. These findings suggest a possible convergence of AIB1 with a pathway involving p53, which might induce chromosomal instability and affect the clinical phenotype of a subset of CRCs.

Oncogenic role of clusterin overexpression in multistage colorectal tumorigenesis and progression.

AIM: To investigate the expression pattern of clusterin in colorectal adenoma-carcinoma-metastasis series, and to explore the potential role of clusterin in multistage colorectal tumorigenesis and progression. METHODS: A colorectal carcinoma (CRC)-tissue microarray (TMA), which contained 85 advanced CRCs including 43 cases of Dukes B, 21 of Dukes C and 21 of Dukes D tumors, were used for assessing the expression of clusterin (clone 41D) and tumor cell apoptotic index (AI) by immunohistochemistry and TUNEL assay, respectively. Moreover the potential correlation of clusterin expression with the patient's clinical-pathological features were also examined. RESULTS: The positive staining of clusterin in different colorectal tissues was primarily a cytoplasmic pattern. Cytoplasmic overexpression of clusterin was detected in none of the normal colorectal mucosa, 17% of the adenomas, 46% of the primary CRCs, and 57% of the CRC metastatic lesions. In addition, a significant positive correlation between overexpression of clusterin and advanced clinical (Dukes) stage was observed (P<0.01). Overexpression of cytoplasmic clusterin in CRCs was inversely correlated with tumor apoptotic index (P<0.01), indicating the anti-apoptotic function of cytoplasmic clusterin in CRCs. CONCLUSION: These data suggests that overexpression of cytoplasmic clusterin might be involved in the tumorigenesis and/or progression of CRCs. The anti-apoptotic function of cytoplasmic clusterin may be responsible, at least in part, for the development and biologically aggressive behavior of CRC.

Nucleotide sequences and functions of the Epstein-Barr virus latent membrane protein 1 genes isolated from salivary gland lymphoepithelial carcinomas.

Epstein-Barr virus (EBV) infection is associated with salivary gland lymphoepithelial carcinoma (SLEC) and nasopharyngeal carcinoma (NPC). EBV is a ubiquitous herpes virus world wide, but EBV-associated SLEC and NPC are prevalent in restricted regions such as south areas of China, Southeastern Asia and Greenland (Eskimos). To examine whether particular EBV variants play roles in the development of SLEC and NPC, we isolated the complete EBV LMP1 genes from 12 paraffin-embedded biopsy samples of SLECs isolated from China, Taiwan and Russia, and compared these LMP1 genes with those of NPC (CAO) and the prototype B95-8 EBV. Nucleotide sequence analysis showed that SLECs LMP1 is more similar to that of CAO than that of prototype B95-8. The analysis also identified several conserved (67-100%) variations in SLEC-LMP1 and CAO-LMP1 distinct from B95-8-LMP1. These included 10-amino acid deletion, 5-amino acid deletion and 12-single amino acid variations. A SLEC-LMP1 gene with the aforementioned conserved variations inhibited the growth of an embryonic kidney cell line (293T), highly activated the NF-kappaB pathway, and these activities were equivalent to those of B95-8 and CAO. These findings suggest that the biological functions of SLEC-LMP 1 are similar to those of B95-8-LMP1 and CAO-LMP1, and that these amino acid variations including the well-known 10-aa deletion did not affect these two prominent activities. While the present results could not uncover functional differences between SLEC-LMP1 and B95-8-LMP1, the nucleotide sequences and the molecular clone of LMP1 directly isolated from SLEC patients will be a useful tool to identify the high-pathogenic EBV strain(s), associated with SLEC and NPC.

[T-cell/histiocyte-rich B-cell lymphoma: histology, immunophenotype and differential diagnosis].

OBJECTIVE: To study the histology, immunophenotype and differential diagnosis of T-cell/histiocyte-rich B-cell lymphoma (TCRBCL). METHODS: A review of 245 cases of so-called Hodgkin lymphoma diagnosed during the period from 1980 to 2000 in 3 hospitals in Guangzhou, 8 cases were reclassified as TCRBCL, according to the 2001 World Health Organization classification of lymphoid neoplasms. An additional 8 cases of TCRBCL were retrieved from consultation files, as well as routine biopsy cases encountered between 2000 and 2004. Immunohistochemical studies were performed on paraffin-embedded tissue by SP technique in order to study the immunophenotype of the large neoplastic cells (CD20, CD79a, CD3, CD45RO, CD15, CD30, CD10, bcl-6 and EMA) and background non-neoplastic cells (CD3, CD8, CD20, CD45RO, CD79a, CD57, CD68, CD21, CD35, cyclin D1, TIA-1). In-situ hybridization for EBER 1/2 and immunoglobulin heavy chain gene rearrangement study were also performed in 4 and 4 cases respectively. RESULTS: Among the TCRBCL cases studied, there were 8 males and 8 females. The age of patients ranged from 10 to 68 years old (mean = 40.3 years old). All had lymphadenopathy and hepatosplenomegaly. On presentation, 3 cases belonged to stage II, 10 cases stage III and 3 cases stage IV. Histologically, scattered atypical large neoplastic cells were seen in a background of small lymphocytes and sometimes histiocytes. The large cells exhibited CD20+, CD79a+, EMA+, CD15- and CD30- phenotype. On the other hand, the background small lymphocytes were CD3 and CD45RO-positive. Most of these background T cells expressed CD8 and TIA-1, while they were mostly CD57-negative. The histiocytic cells were CD68-positive; and CD21 and CD35-positive follicular dendritic cell meshworks were absent. In-situ hybridization for EBER 1/2 showed negative nuclear signals. Immunoglobulin heavy chain gene rearrangement study revealed clonal pattern in all the 4 cases tested. CONCLUSIONS: TCRBCL is a rare subtype of lymphoma, with distinctive histology and immunophenotype. The above features are helpful in delineating this entity from Hodgkin lymphoma, reactive lymphoid hyperplasia and lymphomatoid granulomatosis.

[Effect of survivin antisense mRNA transfection on the growth and chemotherapy sensitivity of lymphoma cells].

OBJECTIVE: To study the effect of transfecting survivin antisense mRNA on growth and chemotherapy sensitivity of lymphoma cells. METHODS: Eukaryotic expression plasmid pcDNA3. 1-antisense (As) survivin was constructed and transfected into Jurkat T lymphoblastic lymphoma cell lines with high expression survivin mRNA by use of lipofectmine gene transfer technique. Expression of survivin mRNA and protein were detected by reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemical and Western blot. The effect of transfecting survivin antisense mRNA on the growth of Jurkat cell lines was monitored by population doubling time (PDT) and Apoptotic indexes (AI). The morphologic features were observed in transfected cells by light and electric microscopes. MTT assay was used to analyze the response of transfected cells to CTX and MTX. RESULTS: Compared with the control cells, the expression of survivin mRNA and protein were reduced after transfected pcDNA3. 1-Assurvivin 48 h, 5 w and 6 w, PDT (52 h) was prolonged. Apoptotic indexes were higher in transfected antisense survivin mRNA cells [20.2% (48 h)], 6.2% (5 w) and 6.8% (6 w) than control ones [2.1%, 1.3% (48 h)] and [1.3% (5 w) and 1.0% (6 w)]. The cells grow slowly and the dead cells increase and some swelling and apoptotic cells were observed in transfected pcDNA3. 1-Assurvivin groups by invert, light and electric microscopes. The Jurkat cell line of transfected pcDNA3. 1-Assurvivin had higher sensitivity to CTX and MTX. The rate of inhibition was higher in transfected group. There is a significant difference between the transfected group and untransfected one, P < 0.05. CONCLUSIONS: The result indicated that survivin gene was very important for growth of Jurkat cells. To inhibit the expression of survivin will be significant in therapy of T lymphoblastic lymphoma. Survivin gene might be a target of therapy.

[Analysis of survivin expression in subtypes of lymphoma].

BACKGROUND & OBJECTIVE: It is difficult to diagnose and classify lymphoma in clinical pathology. This study was designed to examine the expression of survivin, an important anti-apoptosis gene, in subtypes of lymphoma,and to investigate its value in classification of lymphoma. METHODS: Biopsies from 83 cases of lymphoma and 5 cases of lymph node reactive proliferation were collected from the First Affiliated Hospital and Cancer Center of Sun Yat-sen University from January 2001 to June 2003. Reverse transcription- polymerase chain reaction (RT-PCR) and immunohistochemical staining were performed to determine the mRNA and protein expression of survivin. In addition, the survivin expression in four tumor cell lines (K562, HL60, Raji, and Jurkat cell line) were also determined by RT-PCR and immunohistochemical staining. Semi-quantity assay was used to evaluate the quantity of survivin protein and mRNA expression in subtypes of lymphoma. RESULTS: Protein expression of survivin was high and strong in diffuse large B-cell lymphoma (DLBL) (87.2%,34/39), Burkitt lymphoma (BL) (100%,2/2), and lymphoblastic lymphoma (LBL) (85.7%,6/7), while their expression were lower and weaker in follicular lymphoma (FL)(22.2%), extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) (33.3%), and marginal zone lymphoma (MZL)(40.0%). There exist a significant difference between the higher expression group (DLBL, BL, and LBL) and lower one (FL, MZL, and MALT) in expression of survivin. Chi-square test,Chi(2)=24.77,P< 0.01. Furthermore, survivin protein expression level in older patients (media age: 57-year old) with DLBL was higher than that in younger patients (media age: 41-year old). Almost all of Reed-Sternberg cells (R-S cells) in Hodgkin's lymphoma (HL) showed strongly positive expression of survivin. The protein expression of survivin was positively correlated with mRNA (r=0.627 0,P< 0.01). CONCLUSION: The expression level of survivin mRNA and protein shows significant difference in subtypes of lymphoma,and it might be act as a biomarker to classify the subtypes of lymphoma.

Heterogeneous expression and association of beta-catenin, p16 and c-myc in multistage colorectal tumorigenesis and progression detected by tissue microarray.

Most colorectal carcinomas (CRCs) arise from adenomas through an archetypal pathogenic pathway, the adenoma-carcinoma-metastasis sequence. Aberrant expression of beta-catenin, p16, E-cadherin and c-myc appears to have played important roles in the development and/or progression of CRC, but their precise distribution pattern and associations in different pathologic loci along CRC's pathogenic pathway have not been thoroughly examined. In this study, a tissue microarray (TMA) containing 85 advanced CRCs in different Dukes stages was constructed. In each of 85 cases, tissue specimens from normal mucosa and primary carcinomas in different layers of the bowel wall were included in the TMA. Tissue specimens from matched adenoma, lymph node metastases and distant metastases were obtained from 22, 21 and 21 cases, respectively. Expression patterns of beta-catenin, p16, E-cadherin and c-myc were evaluated by immunohistochemistry. The results revealed that nuclear expression of beta-catenin, p16 and c-myc was quantitatively increased from normal mucosa to premalignant adenoma, primary carcinoma and lymph node metastatic carcinoma; the frequency of nuclear overexpression of beta-catenin and p16 in lymph node metastases was significantly higher than that in distant metastases (p < 0.05). These results suggest an association between nuclear overexpression of beta-catenin and/or p16 and CRC lymph node metastasis but not distant metastasis. The results also showed that correlative high nuclear expression of beta-catenin and c-myc was observed in primary carcinomas involving the serosa and lymph node metastases (p < 0.05) but not in other pathologic regions of CRCs, suggesting that the tumor microenvironment in different pathologic loci of colorectal tumorigenesis and progression may influence c-myc responsiveness to beta-catenin/Tcf activation.