Material and Method: Ninety-three biopsies obtained from 71 patients were included in the study group. Hematoxylin-eosin stained slides were evaluated and graded according to architectural and cytological features. A total score was obtained for both architectural and cytological atypia in each case. Masson Trichrome stained sections were used to assess dermal fibroplasia and evaluated semiquantitatively.
Results: Twenty-eight (30.1%) cases had mild, 48 (51.6%) cases moderate, and 17 (18.3%) cases had severe atypia regarding architectural features. Nine (9.7%) cases were scored having mild, 28 (30.1%) cases moderate, and 56 (60.2%) cases having severe atypia regarding cytologic features. There was a significant correlation between the degree of architectural and cytologic atypia. Six (21.4%) cases with mild, 31 (64.6%) cases with moderate,, and 12 cases (70.6%) with severe architectural atypia had dermal fibroplasia. One case (11.1%) with mild, 10 cases (% 35.7) with moderate, and 10 cases (67.9%) with severe cytological atypia had dermal fibroplasia. A significant relationship was found between the presence of fibroplasia and the degree of cytological and architectural atypia.
Conclusion: A statistically significant relationship was revealed between the increase of architectural and cytological atypia and dermal fibroplasia. Detection and evaluation of stromal changes by histochemical methods, and scoring of atypia in dysplastic nevi may be helpful to distinguish dysplastic from other banal nevi.
Many recent epidemiological[1,7,8], morphological[1,7-10], immunohistochemical[11-14] and genetic[15-20] studies support the notion that DN is a major risk factor for melanoma even if not a direct precursor.
Many studies support atypia grading for DN but there is marked variability both for intra-observer and interobserver values[21-24]. Neoplastic stromal changes progress together with the characteristic parenchymal changes in melanocytic tumor progression[6]. A band-like lymphocytic infiltrate together with diffuse fibroplasia is seen more commonly in MM[25]. There are many opinions on the development mechanism for the dermal fibroplasia during the MM tumorigenesis[6,25] but there is no study on the relationship between architectural and cytological atypia grading in DN's and dermal fibroplasia. The aim of this study was to evaluate atypia according to architectural and cytological features of DN and to demonstrate the potential relationship of this atypia with dermal fibroplasia.
Table I: Architectural and cytological atypia grading criteria
The architectural features, circumscription, symmetry, cohesion, suprabasal melanocytes, confluence, and single cell proliferation were scored and evaluated. The total of obtained scores gave the degree of architectural atypia. 0-1= Mild architectural atypia (MiAA), 2-3 = Moderate architectural atypia (MoAA), 4-6 = Severe architectural atypia (SAA). The cytological features were evaluated by scoring the nuclear shape and staining, nuclear size, nucleolar prominence and melanocyte cell size.
The total of obtained scores gave the degree of cytological atypia as 0-1: Mild cytological atypia (MiCA), 2: Moderate cytological atypia (MoCA) 3-4: Severe cytological atypia (SCA). Masson's Trichrome histochemical technique was used to show dermal fibroplasia in DN[27]. The sections stained with Masson's Trichrome stain were evaluated under light microscopy by 2 pathologists and were scored semiquantitatively as fibroplasia present (1) or absent (0).
The architectural and cytological grades and histochemical staining characteristics of the cases were evaluated with statistical analysis. The association between the degree of atypia in the DN and the Masson Trichrome staining scores was investigated. Data were analyzed using Scientific Package for Social Sciences Software (SPSS) and Epi Info 2002 Statcalc programs. Pearson Chi square, Chi square test for trend and Spearman correlation tests were used. A p value less than 0.05 was accepted as significant.
Table II shows the distribution of the cases by anatomic localization.
Table II: Distribution of the cases by anatomical localization
The most common architectural disorders were confluence in the junctional melanocytes (Figure 1), the loss of termination with a nest in one or both ends and the presence of single cell melanocytic proliferation.
Figure 1: Confluence of the junctional melanocytes in dysplastic nevus (Hematoxylin eosin, x200).
The architectural atypia features of the cases have been summarized in Table III. We found MiAA in 28 cases (30.1%), MoAA in 48 cases (51.6%) and SAA in 17 cases (18.3%).
Table III: Architectural atypia features and scores of the cases
The most common cytological features were abnormal nuclear shape and staining pattern (Figure 2), large cell diameter, nucleolar prominence and marked nuclear enlargement. Table IV summarizes the cytological atypia features and scores. We found MiCA in 9 cases (9.7%), MoCA in 28 cases (30.1%) and SCA in 56 cases (60.2%).
Table IV: Cytological atypia features and scores of the cases
A total of 6 MiAA cases (21.4%), 31 MoAA cases (64.6%), and 12 SSA cases (70.6%) showed marked dermal fibroplasia with Masson Trichrome histochemistry (Figure 3) (Table V).
Table V: Association between the degree of architectural atypia and dermal fibroplasia
Marked dermal fibroplasia on Masson Trichrome histochemistry was also found in 1 MiCA case (11.1%), 10 MoCA cases (35.7%) and 10 SCA cases (67.9%) (Figure 4) (Table VI).
Table VI: Association between the cytological atypia grades and dermal fibroplasia
The presence of dermal fibroplasia changed with the degree of architectural atypia (p<0.001, r= 0.354). The increased incidence of fibrosis with increasing architectural atypia degree (χ2= 15.88, p<0.0001) was found to be statistically significant.
The presence of dermal fibroplasia changes with the degree of cytological atypia (p<0.001, r= 0.371). We found the increasing incidence of fibrosis with increasing degree of cytological atypia to be statistically significant (χ2= 14.64, p<0.001).
The degree of architectural atypia changed with the degree of cytological atypia (p=0.05).
The presence of single cell proliferation that is the architectural atypia grading parameter changed with the presence of hyperchromatic or irregular shape nuclei that are features of cytologic atypia (p=0.013, r=0.233). Similarly, the termination of the junctional component with a nest at both ends, that is a parameter of architectural atypia, changed significantly with the presence of a hyperchromatic or irregular shaped nuclei, that are cytological atypia parameters (p=0.005, r=0.154).
Nucleolar prominence changed with discohesion of nests (p=0.013, r=0.202) and single cell proliferation (p=0.000, r=-0.449).
A cell diameter over twice the basal keratinocyte nucleus diameter changed significantly with more than 50% bridge formation between junctional melanocytic nests or the presence of continuous single melanocytic cellular proliferation (p=0.011, r=0.230).
We also found a significant relationship between nucleolar prominence and the lack of termination of the junctional component with a nest at both ends (p=0.008, r=0.153).
Pozo et al.[24] have graded DN cases as low, moderate or high degree of dysplasia. We found severe dysplasia in 99.5% of DN cases using this criteria. However, we were unable to define significant architectural features that differentiated between a low and moderate degree of dysplasia. All the cases in the series had both architectural and cytological atypia. This study did not find a significant relationship between the nuclear pleomorphism and chromatin pattern and the severe atypia degree, in contrast to our findings. We separately scored the architectural and cytological features in DN cases and graded the atypia. We also evaluated the dermal fibroplasia. We found a significant relationship between both architectural and cytological atypia and dermal fibroplasia by comparing the architectural and cytological atypia degree and dermal fibroplasia. As the severity of architectural and cytological atypia increased, we observed that the fibroplasia rate also increased. We suggest that as different architectural disorders and cellular changes may be seen in the same case, the architectural and cytological atypia grading should be performed separately and these atypia grades should be interpreted together.
The scoring and grading of the degree of atypia of our cases was performed according to the criteria defined by Shea et al.[26]. We found a significant association and correlation between the architectural and cytological degrees of atypia in this study. Although there are many studies on grading in the literature, there is no consensus on the descriptive criteria that constitute the basis of the grading. Some of the studies degree melanocytic dysplasia by only cytological characteristics[29] while others grade by both architectural and cytological characteristics[23,26]. Our findings show a significant relationship between architectural and cytological atypia similar to many studies in the literature. A larger number of studies with a larger number of cases are required for the standardization of grading and establishment of reliable criteria. Although we found a significant relationship between the degrees of architectural and cytological atypia in our study, we also had cases with different architectural and cytoogical atypia degrees. Although investigators report an increase in the incidence of a history of melanoma with increasing degree of dysplasia, it is possible that high grade melanocytic dysplasia cases without fibroplasia are present[23]. Our results support the idea that an increasing grade of melanocytic dysplasia leads to a significant increase in the presence of dermal fibroplasia.
When grading this cytological atypia, we used the term “severe cytological atypia” in the presence of three or more nuclear criteria. We observed that dermal fibroplasia changed with the degree of cytological atypia. This finding may indicate that dermal fibroplasia may provide an additional criteria when interpreting the degree of cytological atypia in DN cases.
The dermal fibroplasia in DN can be concentric or lamellar fashion[22,25,33,34]. Concentric eosinophilic fibroplasia is generally associated with melanocytic atypia or aytpical lentiginous epidermal or melanocytic hyperplasia and is most commonly seen in precursor nevi. Lamellar fibroplasia is the most prominent stromal pattern. It can also be seen with invasive melanomas[6]. It strongly indicates parenchymal extracellular matrix and tumor interaction. Lamellar fibroplasia indicates the end point of the progression of melanocytic precursor lesions. These changes are seen in a marked manner in lesions that are between DN and early melanoma[7]. Light microscopy studies have shown that the nests and melanoma nests consisting of nevus cells are surrounded by basal membrane material consisting of type-IV collagen and laminin. The basal membrane material is of varying width and has an amorphous appearance. Type-IV collagen, laminin and MMP-2 are synthesized both by melanoma cells and the neighboring fibroblasts. It is not known whether basal membrane production is mostly from melanocytic cells or fibroblasts. The interaction between basal membrane melanocytes and ECM possibly develops as a result of the backwards migration of neural crest-derived melanocytes[35]. This view partially explains the dermal fibroplasia found in DN cases.
The fibroplasia is thought to develop from the interaction of some growth factors secreted by proliferated melanocytes, adjacent keratinocytes and fibroblasts with autocrine or paracrine effect[36]. Various stromal patterns have been defined in melanocytic dysplasia development[6,37]. We had no case of fibroplasia without cytological atypia in our study. The significantly increased rate of dermal fibroplasia with increasing cytological and architectural atypia supports these views. We found dermal fibroplasia together with nuclear atypia in 49 cases (52.7%). However, we did not find dermal fibroplasia despite the presence of nuclear atypia in 44 cases (47.3%). These findings indicate that the degree of stromal changes in DN can help in differentiating DN cases from other basal nevi.
Dermal fibroplasia is a valuable marker that reflects the neoplastic cell-stroma relationship in DN cases. Evaluation of dermal fibroplasia and determining its relationship with the degree of atypia in DN cases will contribute to new studies directed towards understanding the progression risk of DN to malignant melanoma.
ACKNOWLEDGEMENT
We would like to thank Prof. Uğur Pabuççuoğlu, M.D., for
his valuable help in writing the article.
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