Siew Meng Chong

Tuberculous and nontuberculous cervical lymphadenitis: a clinical review.

OBJECTIVES : The aim of the present study was to identify differences in clinical characteristics between patients with tuberculous cervical lymphadenitis and those with nontuberculous cervical lymphadenitis and to determine the diagnostic accuracy of fine needle aspiration (FNA) cytology. STUDY DESIGN AND SETTING : Seventy-two patients with inflammatory cervical lymphadenitis were studied retrospectively. They were divided into 2 groups: group 1 consisted of those with tuberculous lymphadenitis and group 2 consisted of those with non-tuberculous lymphadenitis. The demographic characteristics, clinical parameters, and hematological and cytological results of the 2 groups were compared. RESULTS : Other than there being a significantly higher proportion of foreign-born patients in group 1, there were no differences in clinical characteristics between the 2 groups. The sensitivity and specificity of FNA cytology in the diagnosis of tuberculous lymphadenitis were 88% and 96%, respectively. CONCLUSION : It is difficult to clinically differentiate tuberculous from nontuberculous lymphadenitis. FNA cytology is useful in the diagnosis of tuberculous lymphadenitis. SIGNIFICANCE : In regions where tuberculosis is endemic, treatment can be instituted without the need for excisional biopsy if the FNA results show characteristic caseating granuloma.

Expression of CD137 on Hodgkin and Reed–Sternberg Cells Inhibits T-cell Activation by Eliminating CD137 Ligand Expression

Hodgkin lymphoma is caused by a minority population of malignant Hodgkin and Reed-Sternberg (HRS) cells that recruit an abundance of inflammatory cells. The long-term survival of HRS cells among the vast majority of immune cells indicates that they have developed potent immune escape mechanisms. We report that the TNF receptor family member CD137 (TNFRSF9) is expressed on HRS cells, while normal B cells, from which HRS cells are most often derived, do not express CD137. In 48 of 53 cases of classical Hodgkin lymphoma, CD137 was detected on HRS cells. Ectopically expressed CD137 transferred by trogocytosis from HRS cells to neighboring HRS and antigen-presenting cells, which constitutively express the CD137 ligand (CD137L and TNFSF9), became associated with CD137L and the CD137-CD137L complex was internalized. Disappearance of CD137L from the surface of HRS and antigen-presenting cells led to reduced costimulation of T cells through CD137, reducing IFN-γ release and proliferation. Our results reveal a new regulatory mechanism for CD137L expression that mediates immune escape by HRS cells, and they identify CD137 as a candidate target for immunotherapy of Hodgkin lymphoma.

Increased production of interferon-γ by tumour infiltrating T lymphocytes in nasopharyngeal carcinoma: indicative of an activated status

Undifferentiated nasopharyngeal carcinomas (UNPC) are characterised by an association with Epstein-Barr virus and an abundant lymphoid stroma. We studied the functional status of the infiltrating T cells in ten UNPC biopsies using an immunohistochemical approach. Twelve non-NPC biopsies were included as controls. Tumour cells of UNPC were positive for HLA class I (10/10) and II (8/10), LMP1 (3/10), and CD86 (6/10). Tumour infiltrating T cells (TILs) were detected with antibodies directed at CD3, CD4, and CD8, and shown to be comparable to that in the control biopsies. Although expression of CD28 was shown to be decreased in TILs, expression of CD25 and IFN-gamma at a relatively high percentage could be consistently detected in the UNPC biopsies. These data suggest that TILs in UNPC are in an activated status, and this T cell response is possibly directed at the tumour cells.

A Diffuse Large B-Cell Lymphoma Classification System That Associates Normal B-Cell Subset Phenotypes with Prognosis

It is known that cells within the inflammatory background in classical Hodgkin lymphoma (cHL) provide signals essential for the continual survival of the neoplastic Hodgkin and Reed-Sternberg (HRS) cells. However, the mechanisms underlying the recruitment of this inflammatory infiltrate into the involved lymph nodes are less well understood. In this study, we show in vitro that HRS cells secrete lymphotoxin-α (LTα) which acts on endothelial cells to upregulate the expression of adhesion molecules that are important for T cell recruitment. LTα also enhances the expression of hyaluronan which preferentially contributes to the recruitment of CD4(+) CD45RA(+) naïve T cells under in vitro defined flow conditions. Enhanced expression of LTα in HRS cells and tissue stroma; and hyaluronan on endothelial cells are readily detected in involved lymph nodes from cHL patients. Our study also shows that although NF-κB and AP-1 are involved, the cyclooxygenase (COX) pathway is the dominant regulator of LTα production in HRS cells. Using pharmacological inhibitors, our data suggest that activity of COX1, but not of COX2, directly regulates the expression of nuclear c-Fos in HRS cells. Our findings suggest that HRS cell-derived LTα is an important mediator that contributes to T cell recruitment into lesional lymph nodes in cHL.

Detection of ALK Gene Rearrangements in Formalin-Fixed, Paraffin-Embedded Tissue Using a Fluorescence In Situ Hybridization (FISH) Probe

AbstractBackground: It is widely known that the efficiency of fluorescence in situ hybridization (FISH) probes applied to formalin-fixed, paraffin-embedded tissues is affected by the conditions under which the tissues are fixed and embedded. However, relatively few studies address exactly how tissue archiving conditions affect the performance of FISH probes. We report our experience based on use of an ALK FISH probe, during the validation of its diagnostic utility. Methods: We applied the probe to 77 formalin-fixed, paraffin-embedded tissue blocks archived from 1991 through to 2000, and studied the interrelationship between the archival age (which ranged up to 10 years), type and condition of tissue, duration required for optimum hydrolysis, and obtainability of hybridization signals. Results: We found that as archival age and tissue collagen content increased, not only did hydrolysis times have to be prolonged in order to yield interpretable hybridization signals, but also the likelihood of blocks becoming non-signaling increased. The most striking positive correlations were seen between the archival age of signaling lymphoid blocks and their requisite hydrolysis times. Conclusions: The difficulty in applying FISH on archival tissue increases with its archival age and collagen content, and may necessitate changes in laboratory protocol accordingly.

Rhinosporidiosis: Differential Diagnosis of a Large Nasal Mass:

the organism Rhinosporidium seeberi. This organism is classified as a fungus.1 It is endemic in South India and Sri Lanka and commonly affects the nasal and conjunctival mucosa.2 The condition should be included as a differential diagnosis in patients from the Indian subcontinent who present with a nasal mass. We present a case of rhinosporidiosis that was diagnosed and managed at the Department of Otolaryngology in Singapore.

Intralaryngeal thyroglossal duct cyst : Implications for the migratory pathway of the thyroglossal duct

An atypically located thyroglossal duct cyst in a 42-year-old man is described. A purely intralaryngeal thyroglossal duct cyst is extremely rare and can mimic other laryngeal lesions. This case demonstrates that thyroglossal duct cyst is a possible cause of intralaryngeal swellings and would have significant implications for the manner in which they are managed.

Multi-Lineage Interrogation of the Performance Characteristics of a Split-Signal Fluorescence In Situ Hybridization Probe for Anaplastic Lymphoma Kinase Gene Rearrangements

AbstractBackground: Fluorescence in situ hybridization (FISH) can identify chromosomal translocations on fixed archival tissue, but studies cross-validating the utility of FISH on lesions of different cell lineages that harbor similar translocations (e.g. those involving anaplastic lymphoma kinase [ALK]) have not been published. Aim: Our objective was to define the diagnostic utility, performance characteristics, and limitations of a commercially available, split-signal, FISH probe for ALK gene rearrangements on fixed, archived tissue from lesions of diverse cell lineage. Study design: The sensitivity, specificity, and positive and negative predictive values of the Vysis® ALK FISH probe were compared with those of the ALK-1 antibody (Dako®) in a series of 101 cases, comprising 43 hematolymphoid neoplasms, 4 reactive lymphoid controls, 50 non-hematolymphoid (including neuroectodermal, epithelial, myofibroblastic, and germ cell) lesions, and 4 early-trimester aborted fetuses that served as neuroblastic controls. Methods: The study involved a predominantly (72%) Singaporean Chinese population aged between 9 months and 88 years (excluding the aborted fetal controls). All cases were reviewed both histologically and immunohistochemically with a wide panel of antibodies using the standard protocols in order to diagnose them according to the latest WHO classification systems. A positive cut-off value was determined, both by comparison with diagnostic categories with and without ALK translocations, as well as with negative controls. Results: The ALK FISH probe suffered a 33% non-informative rate, but in informative cases it showed 94% concordance with the ALK-1 immunostain. A minimum cut-off value of 5 in 200 informative cells was adopted to make a positive call in each case. Of the ALK-1 immunoreactive lesions, nine lymphomas were concordantly ALK translocation-positive but one vesical inflammatory myofibroblastic tumor was discordantly FISH-negative. Among the ALK-1-immunonegative lesions, one case each of anaplastic lymphoma and pulmonary mycobacterial spindle cell pseudotumor were discordantly ALK FISH-positive, while a case each of intestinal myeloblastic tumor and ganglioglioma showed initial — but not reproducible — positive FISH readings. The remaining cases were concordantly negative. Discussion: The discrepancies between ALK FISH results and well-established immunomorphological parameters indicate that interpretation is not always straightforward. Notably, the derivation of threshold cut-off values for positive calls on FISH assays has seldom been addressed in the literature, and has raised issues in interpreting cases with borderline positivity in this study. The factors that may influence such cut-off values are extensively reviewed. Conclusions: We propose the term ‘conditional threshold positivity’ to encourage the adoption of different cut-off values for making positive calls in lesions of different origin.

Multi-Lineage Interrogation of the Performance Characteristics of a Split-Signal Fluorescence In Situ Hybridization Probe for Anaplastic Lymphoma Kinase Gene Rearrangements A Study of 101 Cases Characterized by Immunohistomorphology on Fixed Archival Tissue

archival tissue, but studies cross-validating the utility of FISH on lesions of different cell lineages that harbor similar translocations (e.g. those involving anaplastic lymphoma kinase [ALK]) have not been published. Aim: Our objective was to define the diagnostic utility, performance characteristics, and limitations of a commercially available, split-signal, FISH probe for ALK gene rearrangements on fixed, archived tissue from lesions of diverse cell lineage. Study design: The sensitivity, specificity, and positive and negative predictive values of the Vysis® ALK FISH probe were compared with those of the ALK-1 antibody (Dako®) in a series of 101 cases, comprising 43 hematolymphoid neoplasms, 4 reactive lymphoid controls, 50 non-hematolymphoid (including neuroectodermal, epithelial, myofibroblastic, and germ cell) lesions, and 4 early-trimester aborted fetuses that served as neuroblastic controls. Methods: The study involved a predominantly (72%) Singaporean Chinese population aged between 9 months and 88 years (excluding the aborted fetal controls). All cases were reviewed both histologically and immunohistochemically with a wide panel of antibodies using the standard protocols in order to diagnose them according to the latest WHO classification systems. A positive cut-off value was determined, both by comparison with diagnostic categories with and without ALK translocations, as well as with negative controls. Results: The ALK FISH probe suffered a 33% non-informative rate, but in informative cases it showed 94% concordance with the ALK-1 immunostain. A minimum cut-off value of 5 in 200 informative cells was adopted to make a positive call in each case. Of the ALK-1 immunoreactive lesions, nine lymphomas were concordantly ALK translocation-positive but one vesical inflammatory myofibroblastic tumor was discordantly FISHnegative. Among the ALK-1-immunonegative lesions, one case each of anaplastic lymphoma and pulmonary mycobacterial spindle cell pseudotumor were discordantly ALK FISH-positive, while a case each of intestinal myeloblastic tumor and ganglioglioma showed initial – but not reproducible – positive FISH readings. The remaining cases were concordantly negative. Discussion: The discrepancies between ALK FISH results and well-established immunomorphological parameters indicate that interpretation is not always straightforward. Notably, the derivation of threshold cut-off values for positive calls on FISH assays has seldom been addressed in the literature, and has raised issues in interpreting

Abstract A4: Hodgkin and Reed-Sternberg cells secrete soluble factors to modulate endothelial cell-T cell interactions in classical Hodgkin lymphoma

Introduction: Classical Hodgkin lymphoma (CHL) is characterized by the presence of a minority of malignant Hodgkin and Reed-Sternberg cells (H-RS cells) surrounded by an abundant mixed inflammatory infiltrate that includes CD4+ T helper 2 cells, regulatory T cells and CD8+ cytotoxic T cells. The mechanisms exploited by HRS cells to influence T cell recruitment into the involved lymph nodes are still unknown. Objective: The aim of this study is to elucidate whether H-RS cells can induce endothelial cell activation to modulate T cell recruitment in CHL. Procedures: Fresh cell culture supernatant (C/S) from the H-RS cell line, KM-H2, was used to stimulate endothelial cells (EC) in-vitro. Controls were EC incubated in medium alone; or with 10ng/ml TNF-α Up-regulation of adhesion molecule expression was assessed by ELISA and FACS. Using the transwell migration system and the parallel plate flow chamber apparatus respectivel, the ability of these stimulated EC to support T cell interactions in-vitro was assessed under static and defined flow conditions. Signaling pathways involved in C/S induced EC activation were examined by western blotting. Results: KM-H2 C/S-stimulated EC up-regulate ICAM-1, VCAM-1 and E-selectin to levels comparable to those of TNF-α-stimulated EC. Both C/S-stimulated EC and TNF-α stimulated-EC support naive and memory T-cell interactions under defined flow conditions. Furthermore, both CD4+ naive and memory T cell demonstrated enhanced transmigration across C/S-stimulated EC monolayers in response to SDF-1α, as compared to TNF-α-stimulated or unstimulated EC. The activation of EC by KM-H2 C/S is dependent on the NFkB pathway. Blocking of p65 nuclear translocation by the specific IκBα phosphorylation inhibitor (BAY11-8075) abrogated the stimulatory effect of C/S on EC. Phosphorylation of ERK, p38 and JNK were also detected in the KM-H2 C/S stimulated EC. Intracytoplasmic expression of TNF-α was detected in KM-H2 cells. The bioactivity assay showed that the TNF-α sensitive cell line, L929 is sensitive to KM-H2 C/S. However, treatment of cells with TNF-α neutralizing antibody did not effectively prevent L929 cell death, nor inhibit the stimulatory effect of KM-H2 C/S on EC. Conclusion: Our data suggest that the malignant Hodgkin and Reed-Sternberg cells secrete soluble mediators that modulate endothelial cell function and influence recruitment of T cells into the cHL lesion. While the C/S induced EC activation is dependent on the NFkB pathway, our data suggest that the dominant mediator involved is not TNF-α. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A4.