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Year : 2020  |  Volume : 47  |  Issue : 3  |  Page : 153-157

Human herpesvirus-8 latent nuclear antigen-1 expression in kaposi's sarcoma cases as seen in Nnewi, Anambra State: (A 10-year retrospective study)

1 Department of Anatomic Pathology and Forensic Medicine, Nnamdi Azikiwe University, Nnewi Campus, Awka, Anambra, Nigeria
2 Department of Morbid Anatomy and Forensic Medicine, Obafemi Awolowo University Teaching Hospital Complexes, Ile-Ife, Osun, Nigeria

Date of Submission27-Dec-2019
Date of Acceptance12-Sep-2020
Date of Web Publication21-Jan-2021

Correspondence Address:
Dr. Felix Emeka Menkiti
Department of Anatomic Pathology and Forensic Medicine, Nnamdi Azikiwe University, Nnewi Campus, PMB 5025, Awka, Anambra
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jss.JSS_46_19

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Background: Human herpesvirus-8 (HHV-8) is a recognized major causative agent of Kaposi sarcoma (KS). Latent nuclear antigen-1 (LNA-1) is constitutively expressed in all HHV-8-infected cells. The index study emphasizes the role of immunohistochemistry in the diagnosis of KS in Nnewi, Anambra State, Nigeria. After thorough searches in Google Scholar and PubMed, we discovered that our study is the first to employ HHV-8 LNA-1 immunohistochemistry in the diagnosis of KS in Nigeria and other parts of Africa. Materials and Methods: It is a 10-year retrospective analysis of all histopathologically diagnosed KS cases from the archives of both Nnamdi Azikiwe University Teaching Hospital and Pathocon Specialist Clinic and Research Institute Nnewi. Pathology report forms of all KS cases and their blocks were retrieved and recut, and the recut slides were reviewed to confirm the cases. Of the 69 cases of KS retrieved, only 58 that met the inclusion criteria were analyzed using HHV-8 LNA-1 immunohistochemical stains. Results: Of these 58 cases, 20.7' were KS mimickers misdiagnosed as KS while 79.3' were true KS. Confirmed or HHV-8 LNA-1-positive KS cases accounted for 1.3' of all solid malignancies and 15.7' of all sarcomas, with majority presenting as cutaneous lesions mainly involving the lower limb (52.2') and predominantly (69.6') in nodular stage. Conclusion: HHV-8 LNA-1 is an important immunohistochemical marker in confirming diagnosis of KS.

Keywords: Formalin-fixed paraffin-embedded tissue blocks, human herpesvirus-8 latent nuclear antigen-1, immunohistochemistry, Kaposi's sarcoma

How to cite this article:
Menkiti FE, Ukah CO, Adelusola KA, Ezejiofor IF. Human herpesvirus-8 latent nuclear antigen-1 expression in kaposi's sarcoma cases as seen in Nnewi, Anambra State: (A 10-year retrospective study). J Sci Soc 2020;47:153-7

How to cite this URL:
Menkiti FE, Ukah CO, Adelusola KA, Ezejiofor IF. Human herpesvirus-8 latent nuclear antigen-1 expression in kaposi's sarcoma cases as seen in Nnewi, Anambra State: (A 10-year retrospective study). J Sci Soc [serial online] 2020 [cited 2021 May 7];47:153-7. Available from: https://www.jscisociety.com/text.asp?2020/47/3/153/307599

  Introduction Top

Kaposi sarcoma (KS) is a locally aggressive often multi-centric tumor of unknown histogenesis, previously thought to be vasoformative.[1] It was first described as “idiopathic multiple pigmented sarcoma of the skin” by Moriz Kaposi in 1872, and it was known as a relatively rare tumor of the elderly until the advent of Acquired Immune Deficiency Syndrome (AIDS) and organ transplantation.[2] It presents often as a multi-centric cutaneous plaque, patch, or nodular lesion.[1],[3] Recent data have shown uniform association with KS-associated herpes virus or human herpesvirus-8 (HHV-8) infection,[3],[4],[5] thus representing an example of virus-induced vascular proliferation.[3]

Sites of involvement other than the skin include mucosal sites, lymph nodes, and visceral organs.[3] KS occurs in four (4) epidemio-clinical forms: classic (European), iatrogenic (immunosuppression related), endemic (African), and HIV/AIDS-associated KS.[1] Irrespective of the clinical form of KS, the morphologic features are identical, progressing through three stages - patches, plaques, and nodules. The most typical histologic feature of KS is the presence of spindle cells, with mild-to-moderate mitotic activity, forming slits containing red blood cells admixed with lymphocytes, hemosiderin-laden macrophages, and other inflammatory cells; and variously sized periodic acid–Schiff positive hyaline bodies, often seen in the cytoplasm of the proliferating cells and sometimes extracellularly.[6] The early lesions are uncharacteristic and appear like granulation tissue with subtle vascular proliferation which may be limited to the upper reticular dermis.[7],[8] Several lesions, both neoplastic and nonneoplastic, such as pyogenic granuloma, bacillary angiomatosis, spindle cell hemangioma, and pilar leiomyoma can present histologic features similar to the different stages of KS.[3],[6],[7] The distinction between KS and these other lesions based only on morphology cannot be made with certainty, and detection of HHV-8 antigen immunohistochemically is recommended for accurate diagnosis of KS.[5],[9]

  Materials and Methods Top

Our study is a retrospective one involving 69 diagnosed cases of KS from both Pathocon Specialist Clinic and Research Institute, Nnewi and Histopathology Department of Nnamdi Azikiwe University Teaching Hospital (NAUTH), Nnewi in Anambra State, Nigeria, between January 2007 and December 2016. The pathology report forms were retrieved, and information about the age, sex, anatomical sites, and clinical and histological diagnoses were extracted. The formalin-fixed paraffin-embedded (FFPE) tissue blocks of all available cases of KS that met inclusion criteria diagnosed in the study period were retrieved. Fresh sections were prepared and stained with hematoxylin and eosin (H and E) stains and examined using multiheaded compound light microscope (®CARL ZEISS).

Immunohistochemical studies were done by the indirect immunoperoxidase method on the FFPE tissue blocks. Four micrometer (4 μm) thick sections of fresh tissue sections were made from the selected FFPE tissue blocks. The tissue sections were deparaffinized by passing it through xylene and then rehydrated in decreasing alcohol concentrations and mounted on positively charged glass slides. Antigens were retrieved from the tissues by heat-induced epitope retrieval method using a pressure cooker. Endogenous peroxidase activity was blocked using 3' hydrogen peroxide. The tissues were then incubated with the primary antibody, rinsed and then followed by the use of secondary detection system using diaminobenzene as chromogen. Immunohistochemical staining was performed using HHV-8 monoclonal antibody to latent nuclear antigen-1 (LNA-1) in 1:200 dilution and incubated for 3o min with a positive tissue control in parallel. A known HHV-8-positive tissue that stained positive for HHV-8 LNA-1 was used as control. All the aforementioned steps were carried out at room temperature. HHV-8 LNA-1 stains only HHV-8-associated lesions with a reported specificity of near 100'.[10]

  Interpretation of result Top

HHV-8 LNA-1 is normally expressed in the nuclei of positive cells. In this index study, KS tumor cells were considered “positive” when the stain was exclusively observed in the tumor cell nuclei including neoplastic spindle and endothelial cells as a granular or diffuse appearance without concomitant cytoplasmic staining.

The data analysis was done using the Statistical Package for Social Sciences software version 20.0 (IBM, Chicago, IL, USA), and the result presented with tables and charts.

  Results Top

A total of 15,530 surgical specimens were received in both facilities (7772 [50.1'] from NAUTH and 7758 [49.9'] from Pathocon) during the 10-year period under review. Of these number of surgical specimens, 3505 (22.6') were solid malignancies comprising 3201 (91.3') carcinomas and 304 (8.7') sarcomas. Of the 304 cases of sarcomas, 69 cases were diagnosed as KS, but only 58 cases (25 cases from Pathocon and 33 cases from NAUTH) fulfilled the inclusion criteria and were used in this index study.

Of these 58 cases, 46 (79.3') were positive to HHV-8 LNA-1 immunohistochemical stain, hence confirmed KS cases while 12 (20.7') were mimickers [Table 1]. Among the mimickers seen in this study, 1 is a sarcomas while 11 were benign lesions [Table 2]. HHV-8 LNA-1-positive KS cases accounted for of 1.3' all solid malignancies and 15.7' of all sarcomas. The age range of the confirmed KS cases was 7–70 years with mean age, median age, and modal age group of 37.61 (±15.96) years, 36.00 years and third decade, respectively [Figure 1]. KS was also found to be more common in males, accounting for 24 (52.2') cases, than in females, accounting for 22 (47.8') cases, with a male: female ratio of 1.1:1.
Table 1: Final diagnosis following human herpesvirus-8 latent nuclear antigen-1 immunohistochemistry

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Table 2: The reviewed morphologic Kaposi's sarcoma mimics

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Figure 1: Age range distribution of Kaposi's sarcoma

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The KS mimics identified in this study following morphologic review of the HHV-8 LNA-1 negative cases included acroangiodermatitis, spindle cell hemangioma, bacillary angiomatosis, spindled angiosarcoma, gastrointestinal stromal tumor (GIST), pyogenic granuloma, and Kaposiform hemangioendothelioma [Table 2].

Kaposi sarcoma site distribution

Cutaneous involvement was found in 38 (82.6') of the cases, 24 (52.2') of which occurred in the lower limb accounting for most of the KS cases [Table 3]. There was generalized and visceral involvement in 2 (4.3') and 8 (8.7') of the confirmed KS cases, respectively.
Table 3: Anatomical site distribution of Kaposi's sarcoma

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Kaposi sarcoma morphologic stage at diagnosis

Morphologically, 32 (69.6') of the cases were in the nodular stage of the disease, 12 (26.1') cases were plaques while 2 (4.3') cases were in the patch stage [Figure 2].
Figure 2: Histologic stage distribution of Kaposi's sarcoma

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Determination of the presence of human herpesvirus-8 latent nuclear antigen-1 in the hematoxylin and eosin, diagnosed Kaposi's sarcoma cases

Following HHV8 LNA-1immunohistochemical staining of the H and E, diagnosed cases of KS [Figure 3], we observed that the true KS cases expressed strong nuclear immunoreactivity of the neoplastic spindle and endothelial cells [Figure 4] while the KS mimics showed no staining [Figure 5].
Figure 3: H and E-stained high magnification of Kaposi's sarcoma lesion that shows intersecting fascicles of mildly atypical spindle cells with numerous slit-like spaces (arrows) containing red cells (×400)

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Figure 4: A true Kaposi's sarcoma case that shows positive nuclear staining for human herpesvirus-8 latent nuclear antigen-1 immunohistochemical staining (×400)

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Figure 5: Representative section of Kaposi's sarcoma mimics (spindle cell hemangioma) that shows negative human herpesvirus-8 latent nuclear antigen-1 staining (×100)

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  Discussion Top

KS is of public health concern, described in many literatures as a common neoplasm among black populations especially in equatorial region of sub-Saharan Africa.[11],[12] Parkin et al. in Cancer in Indigenous Africans, estimated about 66,200 KS cases worldwide, with about 58,800 (88.8') occurring in Africa.[13] Of these, about 39,500 cases in males and 17,100 cases in females occurred in sub-Saharan Africa compared to 102 males and 17 females in northern Africa.[13] With the increasing burden of this malignant neoplasm among African populace, especially in sub-Saharan countries to which Nigeria belongs. As a result, there is need for precise diagnoses of KS cases with the aid of immunohistochemistry.

Confirmed or HHV-8 LNA-1-positive KS cases accounted for 1.3' of all solid malignancies and 15.7' of all sarcomas in this research. This frequency of 1.3' is lower than the reported 3.0', 2.5', and 1.4' in Lagos, Zaria, and Jos studies, respectively.[14],[15],[16] Similarly, a 10-year study done in Tanzania also showed a frequency of 2.4' of all malignancies;[17] while reports from Harare, Zimbabwe between 1993 and 1995 showed KS to account for 31.1' of all cancers.[18] The difference between the figures from these other studies and that observed in this current study may be explained by the fact that diagnosis were either clinical and/or morphologic using H and E stains only. None of these studies employed HHV-8 LNA-1 immunohistochemistry for KS confirmation. It is possible therefore that some KS mimics were also erroneously classified as KS.

Our study revealed that a significant percentage (20.7') of cases diagnosed as KS based only on H and E, stains were KS mimics following HHV-8 LNA-1 immunohistochemical staining. This finding underscores the importance of routinely using HHV-8 LNA-1 immunohistochemistry in confirming suspected morphologic KS cases as either true KS or its mimics. There is well-documented evidence that there are several KS mimics, including benign and malignant lesions, which can only be distinguished from KS by subjecting them to HHV-8 LNA-1 immunohistochemistry.[5],[10],[19],[20] Therefore, studying these neoplasms with accurate immunoconfirmation of the lesions is of paramount importance not only for diagnosis but also for prompt and appropriate treatment. Our study is the first in Nigeria that employed the use of HHV-8 LNA-1 immunohistochemical stain in confirming the diagnosis of KS and eliminating its mimics.

Most often, the histologic features of KS show overlapping features with other benign or malignant vascular tumors as well as other nonvascular spindle cell soft-tissue neoplasms, thus leading to diagnostic dilemmas. With the discovery of HHV-8 in all forms of KS, it became possible to consider virus detection as a potential diagnostic test.[21] In Nigeria, where facilities for molecular study are not available, immunohistochemical detection of HHV-8 LNA-1 antigen in suspected cases of KS is a cost-effective and reliable alternative. The morphologic mimickers of KS - acroangiodermatitis, spindle cell hemangioma, bacillary angiomatosis, spindled angiosarcoma, GIST, pyogenic granuloma, and kaposiform hemangio-endothelioma - seen in this index research were easily distinguished from true KS due to negative reactions demonstrated by these mimickers to HHV-8 LNA-1 immunohistochemical stain [Figure 5].

Works done by Yves-Marie et al., on formalin fixed tissues in the University Institute of Pathology, Lausanne, Switzerland, and work done by Speicher et al. in resource-constrained settings (University of Nairobi, Grif?th University and Department of Anatomical Pathology Kenyatta National Hospital) all showed anti–HHV-8 antibody as a reliable marker for all variants of KS and proved its usefulness in eliminating KS mimickers.[5],[10]

This index study further revealed slight male preponderance (male: female ratio of 1.1:1) as against the finding of female preponderance reported in earlier studies done in other parts of Nigeria: Benin, Lagos, and Zaria. The observed difference could be due to none application of HHV-8 LNA-1 immunohistochemical stain in those earlier studies.[15],[22],[23] We also found that most KS occur as cutaneous lesions and in individuals between the ages of 21 and 30 years. These observations corroborate the findings of earlier national (in Ilorin, Lagos, and Zaria) and international studies on KS.[8],[15],[17],[23]

  Conclusion Top

HHV-8 LNA-1 immunohistochemistry is essential for the correct diagnosis of KS, especially in a poor resource setting like ours where molecular diagnosis is out-of-reach and accurate diagnosis and appropriate treatment are needed to avoid wasting already limited resources. Therefore, we recommend the routine use of HHV-8 LNA-1 in conjunction with clinical and morphologic features in KS diagnosis in poor resource countries. The index study also reaffirms the suitability of long archived FFPE tissue blocks for immunohistochemical staining.

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Conflicts of interest

There are no conflicts of interest.

  References Top

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Mentzel T, Knuutila S, Lamovec S. Kaposi sarcoma. In: Fletcher DM, Julia AB, Hogendoorn PC, Mertens F, editors. WHO Classification of Tumours of Soft Tissue and Bone. 4th ed.. Lyon: IARC Press; 2012. p. 151-3.  Back to cited text no. 3
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Sasco AJ, Jaquet A, Boidin E, Ekouevi DK, Thouillot F, Lemabec T, et al. The challenge of AIDS-related malignancies in sub-Saharan Africa. PLoS One 2010;5:e8621.  Back to cited text no. 11
Krown SE. Treatment strategies for Kaposi sarcoma in sub-Saharan Africa: Challenges and opportunities. Curr Opin Oncol 2011;23:463-8.  Back to cited text no. 12
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Akinde O, Obadofin O, Adeyemo T, Omoseebi O, Ikeri N, Okonkwo I, et al. Kaposi sarcoma among HIV infected patients in Lagos University Teaching Hospital, Nigeria: A 14-year retrospective clinicopathological study. J Skin Cancer 2016;2016:1-6.  Back to cited text no. 14
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Chalya PL, Mbunda F, Rambau PF, Jaka H, Masalu N, Mirambo M, et al. Kaposi's sarcoma: A 10-year experience with 248 patients at a single tertiary care hospital in Tanzania. BMC Res Notes 2015;8:440.  Back to cited text no. 17
Chokunonga E, Levy LM, Bassett MT, Mauchaza BG, Thomas DB, Parkin DM. Cancer incidence in the African population of Harare, Zimbabwe: Second results from the cancer registry 1993-1995. Int J Cancer 2000;85:54-9.  Back to cited text no. 18
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  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]

  [Table 1], [Table 2], [Table 3]


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