Comparison of the IHC Markers CD138 and CD43 in Oral Leukoplakia: An Original Research

ABSTRACT

Background:

In the clinical practice, one of the most common suspicious lesions that may be potentially malignant is oral leukoplakia. Globally, the rate at which it turns malignant varies. This study examines the levels of markers CD138 and 43 in oral leukoplakia.

Materials and Methods:

Twenty archival blocks of confirmed epithelial dysplasia were taken from the Department of Oral Pathology. These were processed for the identification of markers CD138 and 43 through Immuno Histo Chemistry (IHC). The blocks were divided equally for both the markers.

Results:

There was a noticeable difference in staining intensity between dysplastic tissue and nondysplastic epithelium. However, CD138 expression was low or weak in dysplastic epithelium. CD43 expression was negative in all nonhematopoietic tissues.

Conclusion:

Genes that are cancer associated have been found to have incredibly different impacts in numerous tissues during the multistep process of oral carcinogenesis. In tissues undergoing dysplastic changes, CD138 expression was shown to be decreased, which could point out the malignant changes initiated in the epithelium of the oral tissues.

INTRODUCTION

The colored lesions of the oral cavity may usually be docile; however, they have to be always watched for any malignant transformation (MT). The spectrum of oral potentially malignant disorders (OPMDs) includes conditions such as oral submucous fibrosis, oral leukoplakia (OL), erythroleukoplakia, erythroplakia, oral lichen planus, oral lichenoid reactions, palatal lesions in reverse smokers, graft versus host disease, and oral lupus erythematosus.[1] OL was described by the World Health Organization (WHO) as the clinical manifestations that raise the possibility of MT in the oral cavity.[2] OPMDs were replaced by potentially premalignant oral epithelial lesions (PPOELs) in 2018, since these diseases have the potential to develop into malignancies.[1] With a global frequency of 4.11%, OL is the second most dominant OPMD after oral submucous fibrosis.

OL manifests as white plaques with a dubious possibility of MT.[3,4] The transition from normal to dysplastic to malignant mucosa involves a complicated interplay between the immune system, host genetics, and environmental variables such as exposure to carcinogens like tobacco, alcohol, and betel nuts. The risk factors of OL are age/gender, clinical appearance, size exceeding >2 cm, location, type of nonhomogeneous lesion, and most significantly, the degree of epithelial dysplasia. The range of transformation into malignancy varies from 0.13% to 34.0% globally. It is unknown what proportion of oral squamous cell carcinoma (OSCC) originates from OPMDs. The chances of transformation of OPMD to a cancer cannot be ignored.[5-10] The malignant potential of PPOELs cannot be predicted solely on the clinical traits/histological features of the lesion or patients who are more likely to progress to MT. Establishing prognostic indicators which can recognize lesions that may transform to malignancy is, therefore, crucial.[3,9] Hence, this study examines the levels of markers CD138 and 43 in OL.

MATERIALS AND METHODS

The study was performed after taking approval from the ethics committee as human tissue was involved. However, only archival blocks of the tissue were procured for the study. Twenty blocks were taken from the Department of Oral Pathology. The relevant cases were identified from their ID number of the blocks. All the lesions were collected from the patients who had white lesions that were suspicious and had been biopsied. These blocks were reported to exhibit dysplasia. These were fixed in formalin, and the routine lab procedure was followed for the fixing and embedding in wax. The markers that were tested were CD138 and 43. Ten blocks were assigned for each of the markers. Following deparaffinization and rehydration, epitope retrieval was carried out in hot citrate buffer for 30 min as advised. Following the manufacturer’s instructions, the slides were stained with the primary antibodies CD138 and 43 from Dako Corporation (USA). These were counterstained with Mayer’s hematoxyl in and mounted on a coverslip with DPX. To determine where exactly immunostaining was located inside the epithelial tissue, the immune-stained slices were inspected under light microscopy that was examined at 10×. Positive controls included the use of tonsil and Squamous Cell Carcinoma (SCC), and negative controls involved not using the main antibody at all. The percentage of cells that were positive (grade I: 0%–<25%, grade II: 25%–50%, grade III: 50%–75%, and grade IV: 75%–100%), with staining localization cytoplasmic, membranous, or both and staining intensity mild, moderate, or strong were documented.

RESULTS

The immune system cell served as an internal control for CD43 because there was no evidence of CD43 expression in any of the cases where it was investigated. It was clearly evident that CD43 was negatively expressed in epithelial dysplasia for all the 10 blocks. This can be compared to the adjacent CD43 expression in the immune cells, where this is abundant [Figure 1].

Figure 1.

(a–d) CD43 (red arrows- immune system cells; black arrows- CD43 negative expression)

When the 10 blocks were compared for the expression of the CD138, it was seen that in the dysplastic epithelium, the marker was weakly expressed. Normal epithelium exhibited primarily cytoplasmic and membrane CD138 expression, although dysplastic changes resulted in its absence or weak expression [Figure 2].

Figure 2.

(a–d) CD138 (blue arrow- cytoplasmic and membrane expression in nondysplastic epithelium; black arrow- weak staining)

The staining intensity between nondysplastic and dysplastic epithelia was significantly different.

DISCUSSION

Many scientists believe that OL is the most common OPMD and it is well known. It is the primary cause of the majority of oral malignancies. Recent studies on genomic analysis have revealed that carcinogenesis is a multistep event and most of the cancer driver genes are altered in a tissue-dependent fashion. These are activated only in a few, but not all specific conditions; the reason is yet to be found.[6,11-13] The basal layer is the home for epithelial cells’ regeneration. It is the layer where the cells replicate, and hence hoards the mechanical keys that help in cellular replication. Any derailment in the cellular replicatory genes may lead to the development of dysplasia.[6] The microscopic assessment of the degree of dysplasia—the gold standard to estimate the likelihood of malignant changes—has generated debate among histopathologists. It is commonly accepted that the likelihood that a lesion would proceed to cancer increases with the severity of epithelial alterations. However, the degree of dysplasia is not always a factor in how quickly OL develops into malignancy. According to studies, MT can be seen in up to 30% of slightly dysplastic lesions D1 and 3% of hyperplastic lesions D0. A OSCC’s MT rate for OL varies from 0.13% to 34.0%. The yearly MT rate of OL ranges from 1.36% to 2.9%; however, a latest long-term research revealed a yearly MT rate of ~5%. The rates of MT vary between studies due to variations in the underlying pathophysiology and usage of suspected carcinogens such alcohol and tobacco, but the impact of these on the MT of OL is still debatable.[14-22] There is clear data on what stage or grade of dysplasia may progress to MT. It is observed that any stage of dysplasia may show MT. Hence, biomarkers are urgently required to know the progression.

OSCC is one of the solid tumors that express cancer stem cells (CSCs). In order to conclude on the subsets of OPMD that are most likely to advance into malignancy, a number of proteins have been examined as molecular biomarkers. According to the CSC hypothesis, each marker only identifies a certain group of cancer cells that resemble stem cells, leaving other separate subpopulation features undiscovered. Multiple biomarkers may be used to distinguish between OPMDs with low and high risk. Until date, predicting which OL lesion will advance to malignancy has been the most difficult problem for a clinician.[23-28] Leukosialin, lialophorin, or CD43 is a sialoglycoprotein that is routinely expressed on the exterior of white blood cells (WBCs). Movement, apoptosis regulation, differentiation, and immunological homeostasis are all impacted by CD43. Expression of CD43 may promote oncogenic cell survival in both hematopoietic and nonhematopoietic cancers. In epithelial neoplasms, it is seen to promote Wnt/APC/catenin sign aling pathway. It was determined that regardless of the tissue of origin, adenocarcinomas are frequently CD43 positive than OSCC, although its expression is more particularly always seen in hematological neoplasms.[18,20]

Syndecan-1, often referred to as CD138 or SDC1, is a family of integral membrane proteoglycans that participate in binding the growth factors and interacting with the cell matrix. They are connected to angiogenesis, apoptosis, cell proliferation, and tumor invasion and are altered during carcinogenesis. Their function in MT has been demonstrated by in vitro studies that link them to the preservation of epithelial shape, anchorage-dependent growth, and invasiveness inhibition. In a recent study, it was discovered that tonsil, lip, and oral cavity normal squamous epithelial cells all exhibit high levels of cell membrane immunoreaction. Numerous human malignancies have been thoroughly studied using CD138. CD138 expression is assumed to be prelude to MT. According to a recent study, CD138 can be a useful biomarker for spotting oral verrucous carcinoma microinvasion, since its expression is lowered in the epithelium that is going through neoplastic alterations.[28-31] Strong CD138 expression was seen in normal epithelium, and the expression varied or disappeared with oral epithelial dysplasia and was virtually nonexistent in cancer.[31] In cervical cancers, OSCC, and cutaneous SCC, the expression was dramatically reduced, indicating that CD138 can be used for the identification.[32,33] Reduced expression is considered a precursor to MT or a sign that malignancy is more likely to develop. Few studies have examined the immunoreactivity of CD138 and CD43 in OL. Also, CD43 and 138 might be useful in determining if OL has a low or high risk for developing cancer. Cancer cells are highly heterogeneous. As a result, screening of suspicious lesions for a combination of CSC markers may be essential.[23-33] It is common knowledge that not all dysplastic lesions will develop into cancer. Only OL tissues were evaluated in this retrospective investigation, which has a small sample size and could be a significant flaw. More studies with larger samples are required, particularly those on other potentially cancerous oral conditions including Oral Sub Mucous Fibrosis (OSMF) and lichen planus.

CONCLUSION

While CD138 expression is diminished or absent in hematopoietic cells and tumors derived from them, CD43 is a more specific and sensitive marker that helps us identify the variation between low-risk OL and a high-risk lesion that may advance to malig nancy.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.