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ORIGINAL ARTICLE Table of Contents   
Year : 2010  |  Volume : 53  |  Issue : 4  |  Page : 723-728
Study of the morphological patterns and association of Epstein-Barr virus and human herpes virus 8 in acquired immunodeficiency deficiency syndrome-related reactive lymphadenopathy


1 Department of Pathology, Tata Memorial Hospital, Mumbai, India
2 Department of Hematopathology Laboratory, Tata Memorial Hospital, Mumbai, India

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Date of Web Publication27-Oct-2010
 

   Abstract 

Aims: Study of the morphological patterns of acquired immunodeficiency syndrome (AIDS)-related lymphadenopathy. Settings and Design: We retrospectively selected cases of AIDS-related benign lymphadenopathy. Cases with lymphomas, frank granulomas and necrosis were excluded. We analyzed different morphological patterns and correlated these with immunophenotypic markers along with viral markers human herpesvirus 8-latency-associated nuclear antigen (HHV8-LANA), and Epstein-Barr virus-encoded ribonucleic acid (EBER) studies via in situ hybridization (EBER-ISH). Materials and Methods: We present the morphological patterns of 13 cases of human immunodeficiency virus (HIV)-reactive lymph nodes and their clinical, hematological, biochemical and radiological parameters with special emphasis on the presence or absence of viral markers, including HHV8 and EBV. Results: Common patterns included follicular hyperplasia only (five cases), mixed pattern of follicular hyperplasia with burnt-out germinal centres (four cases), completely atretic follicle (two cases), folliculolysis (11 cases), dumbbell-shaped follicles (three each), progressive transformation of germinal centers (four cases), T-zone expansion (two cases), Reed Sternberg (RS) cells like immunoblasts (two cases), Castleman's-like features with lollipop-like follicles (three cases) and a spindle cell prominence (one case). CD8+ T-cells were predominant in 12 cases. CD8+ T-cells were prominent in germinal centers (eight cases). Plasmablasts were seen in four cases within the perigerminal center area. Immunohistochemistry for HHV8, i.e. HHV8-LANA were negative in all cases while EBER was detected in 11 cases in the centrocyte-like B cells. Two cases of multicentric Castleman's disease expressed EBER; however, they did not express HHV8. Conclusion: The wide spectrum of histological changes in HIV-associated lymphadenopathy requires recognition. The histological changes can mimic those of other infective lymphadenitis, follicular lymphoma, Castleman's disease, progressive transformation of germinal center, Hodgkin's disease and spindle cell neoplasms. Presence of EBV is common while HHV8 was not seen.

Keywords: AIDS, EBV, lymphadenopathy

How to cite this article:
Gujral S, Gandhi J S, Valsangkar S, Shet T M, Epari S, Subramanian P G. Study of the morphological patterns and association of Epstein-Barr virus and human herpes virus 8 in acquired immunodeficiency deficiency syndrome-related reactive lymphadenopathy. Indian J Pathol Microbiol 2010;53:723-8

How to cite this URL:
Gujral S, Gandhi J S, Valsangkar S, Shet T M, Epari S, Subramanian P G. Study of the morphological patterns and association of Epstein-Barr virus and human herpes virus 8 in acquired immunodeficiency deficiency syndrome-related reactive lymphadenopathy. Indian J Pathol Microbiol [serial online] 2010 [cited 2019 Dec 10];53:723-8. Available from: http://www.ijpmonline.org/text.asp?2010/53/4/723/72055



   Introduction Top


Acquired immunodeficiency deficiency syndrome (AIDS)-related lymphadenopathy is well known, tuberculosis being one of the most common causes. [1] Lymphadenopathy may be localized or generalized. Persistent generalized lymphadenopathy (PGL) is defined as lymphadenopathy of two or more noncontiguous sites of at least 3 months duration in the absence of intercurrent illness or drug use associated with lymphadenopathy. [2] There is a paucity of data from the Indian literature on this aspect. We present the histomorphological spectrum of human immunodeficiency virus (HIV)-reactive lymphadenopathy and its association with Epstein-Barr virus (EBV) and human herpes virus 8 (HHV8).


   Materials and Methods Top


We received 8,840 cases of suspected lymphoma over a 6-year period (2003-2008), which included 7,836 cases of hematolymphoid neoplasms and another 1,006 cases of a benign nature. There were 130 cases of AIDS-related lymphadenopathy, which included 38 cases of benign lymphadenopathy and 92 cases of hematolymphoid neoplasms. Twelve of these 38 cases of benign adenopathy underwent fine needle aspiration cytology for a diagnosis, while 26 cases had a lymph node biopsy. These 26 cases included 13 cases of necrotizing lymphadenitis (seven cases in addition revealed epithelioid cell granulomas). The remaining 13 cases of reactive lymphadenopathy were included for this study. Thus, we excluded all cases with necrosis, frank epithelioid cell granulomas and hematolymphoid neoplasms. We analyzed different morphological patterns based on hematoxylin and eosin (H and E)-stained lymph node biopsy sections. Special stains for fungus (Gomori methenamine silver stain, periodic acid Schiff stain [PAS]) and mycobacterium (Ziehl-Neelsen stains) were performed. Immunohistochemistry (IHC) panel included leukocyte common antigen (LCA), CD3, CD20, CD23, CD4, CD8, CD138, CD10, bcl2, Mib1, CD15 and CD30. CD23 was employed to study the follicular dendritic cell network in reactive HIV lymph nodes. CD138 was used to highlight the plasma cells and the plasmablasts within the cords and perigerminal centers. CD4 and CD8 were used to highlight the presence of CD4+ and CD8+ T-cells. HHV8-latency-associated nuclear antigen (HHV8-LANA) was performed by IHC, which is a nuclear stain, highlighting infected HHV8 cells. For demonstration of the EBV-infected cells, Epstein Barr virus encoded RNA (EBER) was used via in situ hybridization (EBER-ISH). DAKO (Mumbai, India) and Biogenix (Hyderabad, India) reagents were used with proper standardization procedures as per the kit instructions along with positive and negative controls. HIV (1 and 2) testing was carried out by immunoassays and was positive in all cases included in the study. Other tests like routine hematology, biochemistry, bone marrow aspirate and biopsy, X-ray chest and computerized tomography (CT) scan findings were correlated wherever available.


   Results Top


There were 38 cases of benign lymphadenopathy of a total of 130 cases of AIDS-associated lymphadenopathy. Only 13 cases of AIDS-related benign lymphadenopathy could be included. There were 10 males and three females, age range from 5 to 61 years (median, 31 years). All of these were symptomatic with common complaints of fever (4/13), weight loss (4/13) and chronic cough (3/13). PGL was seen in nine cases and localized lymphadenopathy was seen in four cases. Size of the lymph nodes ranged from 2.0 to 6.5 cm. Cervical lymphadenopathy (7/13) was the most common site followed by axillary (4/13) and inguinal nodes (2/13). One patient had enlarged tonsil and bilateral parotids. One patient had lymphoma of the fourth lumbar vertebra and, in addition, had a benign cervical lymphadenopathy. Serum globulin levels were available in nine cases, and were normal in six, revealing polyclonal hypergammaglobulinemia in three cases. The b2-microglobulin in available cases was elevated in seven of eight cases, with a range of 3.9-5.8 mg/dl. All our cases were HIV-1 subtype. Serological tests for hepatitis C virus and hepatitis B virus were negative in all cases. Chest X-ray was available in six cases and showed multiple hilar lymph nodes (two cases), bilateral lung consolidation (two cases) and normal chest X-ray (two cases). CT scan findings were available in only eight cases, which revealed hepatosplenomegaly and paraaortic lymph nodes in four cases and the four had normal radiological findings. Bone marrow biopsy was available in 11 of 13 cases and revealed a normocellular marrow in all cases. Lymphocytosis was seen in six cases (more than 15% lymphocytes in the bone marrow, range 10-42%). One case revealed an ill-defined granuloma and two cases showed the presence of micromegakaryocytes.

Different morphological patterns were seen on histological evaluation of these lymph nodes [Table 1]. The prominent pattern was folliculolysis, which was seen in 11 cases [Figure 1]. Five cases showed follicular hyperplasia and revealed hyperplastic germinal centers with widespread apoptosis, prominence of tingible body macrophages and brisk mitotic activity. Three of these cases showed dumbbell-shaped follicles [Figure 2]. The mantle zone was diminished or absent in these cases. Four cases revealed a mixture of follicles showing hyperplasia and the others showing burnt-out germinal centers [Figure 3]. Two cases had completely atretic follicles (burnt-out germinal centers) [Figure 4]. Castleman's-like features with lollipop-like follicles were seen in these two cases [Figure 5]. Both these cases had generalized persistent lymphadenopathy and were labelled as multicenteric Castleman's disease (MCCD). Another case had completely atretic follicles and showed a prominence of spindle cells in the T-zone. Multinucleated giant cells (polykaryocytes) were seen in two cases. Progressive transformation of germinal centers (large-sized nodules rich in CD20-positive B-cells) was seen in four cases [Figure 6]. No Reed Sternberg (RS)-like or popcorn-like cells were seen in these cases. RS cells like immunoblasts were seen in a further two cases [Figure 7] (both with folliculolysis), which were highlighted with LCA, CD20 and CD30. Paracortical T-zone expansion was seen in two cases.
Table 1 :Morphological features in cases of AIDS-related reactive lymphadenopathy


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Figure 1 :Lymph node section showing folliculolysis in the form of small lymphocytes penetrating the germinal centers causing their disruption (H and E, ×100)

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Figure 2 :Dumbell-shaped follicle as highlighted with CD23 immunostains (follicular dendritic cell network) (immunohistochemistry, ×200)

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Figure 3 :Lymph node showing both folliculolysis as well as atretic follicles (H and E, ×100)

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Figure 4 :Stains for CD23 highlight the follicular dendritic cell network of atretic follicles (immunohistochemistry, ×100)

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Figure 5 :Lymph node section showing Castleman's-like features with lollipop follicle (H and E, ×200)

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Figure 6 :Section from the lymph node showing large-sized nodules with progressive transformation of germinal centers with thick capsule (H and E, ×100)

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Figure 7 :Reed Sternberg-like cells in a case of reactive adenopathy (H and E, ×400)

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The CD4:CD8 ratio in the paracortical T-zone was 2:1 (one case), 1:1 (three cases), 1:2 (four cases), 1:4 (three cases) and 1:8 (two cases). In addition to the paracortical region, CD8+ T-cells were also increased within the germinal centers (eight cases) [Figure 8]. Six cases showed a mild to moderate increase in the plasma cell percentages, highlighted with CD138. Plasmablasts were highlighted in four cases only in the perigerminal center. HHV8-LANA immunostains were negative in all cases. EBV via EBER-ISH expression was seen in 11 cases, mainly in the centrocyte-like B-cells, both within the germinal center as well as in the paracortical region [Table 2], [Figure 9].
Table 2 :Results of IHC/ISH in studying viral markers in cases of AIDS-related reactive lymphadenopathy


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Figure 8 :Section shows prominence of CD8-positive cells in the paracortical zone as well as within the germinal centers (immunohistochemistry, ×200)

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Figure 9 :Section showing Epstein-Barr virus-encoded ribonucleic acid positivity in centrocyte like B in a diffuse pattern (in situ hybridization, ×200)

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   Discussion Top


HIV 1 and 2 , the etiologic agents for HIV infection, are members of the lentivirus, which is a subfamily of retroviruses. [3] HIV manifests as a strong tropism for the lymphoid tissue, particularly CD4+ T-cells, dendritic cells and monocytes. [4] As a result of continuous infection, the CD4+ T-cells are destroyed by cytopathic mechanisms. [1]

Patients may present with localized or generalized lymphadenopathy. The histopathology of HIV lymphadenitis has been considered nonspecific. [5],[6] However others have shown characteristic histopathological features associated with HIV-related reactive adenopathy. [1],[7] Ioachim et al.[1] have described HIV lymphadenopathy on histology as type A (follicular hyperplasia, folliculolysis), type C (atrophied burnt-out germinal centers, Castleman's-like features) and type B (in between A and C types). These subtypes generally correspond to the clinical stages of acute, chronic and burn-out phases. [1] The histologic patterns of HIV lymphadenitis (types A, B and C) has been shown to correlate with lymphocyte evaluations as expressed by the total number of CD4+ T-cells, CD4+/CD8+ cell ratios and even levels of antilymphocyte globulins, as previously shown. [8] However, no statistically significant correlation was seen in our study. Wannakrairot et al. have described six reaction patterns in AIDS-related lymphadenopathy, including classic necrotizing granulomas, extensive necrosis with minimal granulomatous response, sarcoid-like nonnecrotizing granulomas, foamy macrophage or pseudogaucher cell response, inflammatory pseudotumor-like proliferation and nonspecific lymphoid hyperplasia. [7] Common patterns seen in our series included follicular hyperplasia, mixed pattern of follicular hyperplasia with burnt-out germinal centers, completely atretic follicle, folliculolysis, dumbbell-shaped follicles, progressive transformation of germinal centers, T-zone expansion, RS cells like immunoblasts, Castleman's-like features with lollipop-like follicles. We did not find pseudoGaucher-like cells. Follicular hyperplasia and folliculolysis were the most common features. The paracortical zone revealed a relative increase in CD8+ T-cells. The CD4:CD8 ratio was 2:1 (one case), 1:1 (three cases), 1:2 (four cases), 1:4 (three cases) and 1:8 (two cases). There was a relative predominance of CD8+ T-cells in the germinal center.

Apart from Mycobacterium, Cryptococcus has been identified in such biopsies. [7] Organisms may be identified in the cases showing lymphoid hyperplasia, extensive necrosis and minimal granulomatous response. [7] However, we could not demonstrate any organisms in any of these 13 cases. HIV presence in the affected lymph nodes may be demonstrated with antibodies to the core protein p24, which is localized to the reactive germinal centers. [1] We did not study the presence of p24 in our cases.

Eleven (83%) cases expressed EBER-ISH, of which eight had generalized lymphadenopathy (PGL). A Ugandan study by Kalungi et al. has shown 52% of such cases expressing EBER. [9] However, an Italian study revealed no association of EBV in PGL of HIV lymph nodes. [10] EBER positivity was seen in small centrocyte-like B-cells in the paracortical T-zone and also in the centrocytes and centroblasts in the germinal center. EBV-positive cases have been associated with hyperplastic germinal centers and follicular fragmentation is characteristic of HIV infection. [9]

When EBV infection occurring in an immunocompromised situation may lead to uncontrolled lymphoproliferation and subsequent development of non-Hodgkin lymphomas (NHL). [11] Interestingly, in HIV infection, both deregulation of the normal B-cell biology and a reduction in immunity play a role in developing NHL. [11] At our center, there has been a recent spurt in plasmablastic lymphomas associated with seropositive HIV patients with a variable morphological pattern (plasmacytic, plasmablastic and Burkitt's like). [12] EBER-ISH expression was reported in five of 13 cases of plasmablastic lymphomas. [12] EBV association in HIV-positive patients with lymphomas from India is common in Hodgkin disease and Burkitt's lymphoma and is also seen in immunoblastic lymphoma. [13] The present study is a first of its kind in reactive lymphoid proliferations. A wide variety of histological features in our study were not specific for the presence or absence of EBV. Moreover, there was no correlation of presence or absence of EBV in A, B and C subtypes as defined by Ioachim et al. [1] The lacuna in the present study is that we did not have any follow-up of these patients. Moreover, a correlation of CD4 counts could not be performed. All these AIDS-related lymphadenopathy patients were referred to our center to exclude the possibility of a hematolymphoid neoplasm. Once the biopsy report was negative for malignancy, patients were lost to further follow-up.

HHV8-LANA positivity in primary reactive HIV nodes are generally seen in the plasmablasts that show lambda light-chain restriction, localize in the mantle zone of B-cell follicles and coalesce to form microscopic lymphomas in some cases. [14] HHV8 has been commonly associated in MCCD, [15] but in our study all cases, including two cases of MCCD, were negative for HHV8. It has been reported that the sensitivity for detection of HHV8 is more with polymerase chain reaction as compared with the immunostains (HHV8-LANA). [16] Although we have seen a few cases of Kaposi sarcoma with the expression of HHV8 in our routine practice (unpublished data), there are no studies from India reporting the presence of HHV8 in HIV-related lymphoid proliferations.


   Conclusions Top


The wide spectrum of histological changes in AIDS-related lymphadenopathy requires recognition, common features being follicular hyperplasia and folliculolysis. The histological changes mimic those of other infective lymphadenitis, follicular lymphoma, Castleman's disease, progressive transformation of germinal centers, Hodgkin's disease and spindle cell neoplasms. EBER-ISH expression is common while HHV8-LANA expression was not detected in any of our cases. There were, in addition, two cases of MCCD that expressed EBER-ISH. However, these did not express HHV8-LANA.


   Acknowledgments Top


The authors would like to acknowledge the help rendered by Dr. Anita Borges, Dr. Jai Mehta and Dr. Laxmi Narayan from Nicholas Piramal Wellspring Diagnostics, Parel, Mumbai, in performing HHV8 stains.

 
   References Top

1.Ioachim HL, Medeiros LJ. Ioachim's lymph node pathology. In: Ioachim HL, Medeiros LJ, editors. Human immunodeficiency virus lymphadenitis. 4th ed. Philadelphia: Lippincott William and Wilkins; 2009. p. 99-105.  Back to cited text no. 1
    
2.Gellar SA, Said JW, Joshi VV. Anderson's Pathology. In: Damjanov I, Linder J editors. Acquired Immunodeficiency Syndrome (AIDS). 10th ed. Missouri: Mosby; 1996. p. 632-3.  Back to cited text no. 2
    
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11.Pietersma F, Piriou E, van Baarle D. Immune surveillance of EBV-infected B cells and the development of non-Hodgkin lymphomas in immunocompromised patients. Leuk Lymphoma 2008;49:1028-41.  Back to cited text no. 11
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14.Dupin N, Diss TL, Kellam P, Tulliez M, Du MQ, Sicard D, et al. HHV-8 is associated with a plasmablastic variant of Castleman disease that is linked to HHV-8 positive plasmablastic lymphoma. Blood 2000;95:1406-12.  Back to cited text no. 14
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Correspondence Address:
S Gujral
Department of Pathology, Tata Memorial Hospital, Mumbai
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0377-4929.72055

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    Figures

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