Upload
cristiane-martins
View
215
Download
1
Embed Size (px)
Citation preview
Argyrophilic nucleolar organizer regions(AgNORs) in odontogenic myxoma (OM)and ameloblastic fibroma (AF)
Cristiane Martins1
Yasmin Rodarte Carvalho2
Maria Auxiliadora Vieira do Carmo1
1Department of Oral Pathology, School ofDentistry, Federal University of Minas Gerais(UFMG) and2Department of Oral Pathology, Sao PauloState University (UNESP), Brazil
Correspondence to:Maria Auxiliadora Vieira do CarmoDepartment of Oral Pathology, School of DentistryFederal University of Minas Gerais (UFMG), AvenidaAntonio Carlos, 6627 – Pampulha, CEP: 31270-901 – Belo Horizonte – Minas Gerais, Brazile-mail: dorinhav/freemail.com.br
Accepted for publication January 11, 2001
Copyright C Munksgaard 2001J Oral Pathol Med . ISSN 0904-2512
Printed in Denmark . All rights reserved
489
AbstractTen cases of odontogenic myxoma (OM) and six cases ofameloblastic fibroma (AF) were subjected to comparative analy-sis by the AgNOR technique, in order to determine a possibledifference in cell proliferation index between these lesions. Themean AgNOR number of the mesenchymal component of AFwas compared with its epithelial component and the differencewas not found to be statistically significant. The mean AgNORindex of the AF group was significantly higher than that of theOM group. Moreover, the mesenchymal component of AF dem-onstrated increased AgNOR numbers compared with that of OM(P∞0.05). These results suggest that the epithelial and mes-enchymal components of AF may have similar cell proliferativeactivity. However, the cell proliferative index of this lesionseems to be higher than that of OM.
Key words: ameloblastic fibroma; argyrophilic nucleolar organ-izer regions (AgNORs); odontogenic myxoma
J Oral Pathol Med 2001: 30: 489–93
Odontogenic myxoma (OM) is an uncommon, locally invasive, be-
nign neoplasm, consisting of rounded and angular cells lying in an
abundant mucoid stroma (1). Its odontogenic origin has been justi-
fied by its association with missing or unerupted teeth, the sparse
presence of islands of odontogenic epithelium within the neoplastic,
myxomatous tissue, and its localization in the maxilla and mandible
(1, 2). Microscopically, the tumour is composed of a myxomatous
tissue made up of an acidic mucopolysaccharide ground substance,
containing a sparse population of stellate or spindle cells with long
anastomosing processes. The tumour cells commonly demonstrate
moderate pleomorphism, but rarely exhibit mitoses (1).
Ameloblastic fibroma (AF) is a rare benign odontogenic tumour
with a characteristic histologic pattern, including ameloblastic epi-
thelium embedded in fibromyxoid connective tissue without forma-
tion of enamel or dentin (3). This lesion is not aggressive and does
Martins et al.
not infiltrate (4, 5). Nonetheless, AF is a rare tumour that does recur
if not completely removed (4).
Nucleolar organizer regions (NORs) are loops of DNA with genes
of ribosomal RNA (rRNA). These genes are highly active during
metaphase and during the reorganization of the nucleolus at the
end of telophase (6). AgNORs are NOR-associated nonhistone argyr-
ophilic acidic proteins. Thus, some authors have proposed that inter-
phase AgNOR quantity is related to the cell duplication time and
rate of proliferation (7).
Some authors have utilized the AgNOR technique in odontogenic
lesions, and they have encountered controversial results (8, 9). The
aim of our study was to investigate probable differences in the cell
proliferation index between OM and AF by means of a comparative
AgNOR quantification.
Material and methods
Ten cases of OM and six cases of AF, from the files of the Oral
Pathology Laboratory, School Dentistry, Federal University of
Minas Gerais (UFMG) and from the Department of Oral Pathology
of Sao Paulo State University (UNESP), were analysed.
Two adjacent sections of 4 mm and 3 mm from each paraffin
block were obtained. The first one was stained routinely with hema-
toxylin and eosin (H-E), and the other by the AgNOR method (10).
In brief, hydrated sections were incubated in a freshly prepared
solution of one part made by dissolving 2 g of gelatin in 1% aque-
ous formic acid to two parts of 50% aqueous silver nitrate. Sections
were left in this solution for 30 min at 45 æC. The sections were then
washed with deionized water and mounted in Canada balsam.
Black-brown dots within nuclei were counted in 100 mesenchy-
mal cells of OM. Islands of odontogenic epithelium were not ob-
served in the OM lesions studied. In the cases of AF, the AgNORs
were evaluated in 100 cells of the epithelial component and in 100
cells of the ectomesenchymal component. The counting was done
twice by two different observers in characteristic areas of the
lesions. Cells where dots were not visualized were not considered.
Table 1. Mean AgNOR counts in odontogenic myxoma and ameloblasticfibroma
Lesion Means
Ameloblastic fibromaEpithelium 1.79a
Mesenchyme 1.60b
Odontogenic myxoma 1.37c
a,b P±0.05 n.s. b,c P∞0.05.
490 J Oral Pathol Med 30: 489–93
Aggregated dots without a ‘‘halo’’ of nucleoplasm around them were
considered as one (9).
Routine light microscopy under ¿1250 magnification and oil im-
mersion using an eyepiece graticule of 0.025 mm2 was performed.
After using Spearman’s correlation coefficient to establish both
inter- and intra-observer reliability (r50.91), the counts were per-
formed and the AgNOR means established. The non-parametric
Wilcoxon test was used for comparative statistical analysis of the
epithelial and the mesenchymal components of AF. The Wilcoxon-
Mann-Whitney test for independent samples was used between the
ectomesenchymal components of both lesions.
Results
AgNORs were visualized as black or brown, defined intranuclear
homogenous dots (Figs. 1 & 2). Table 1 demonstrates no statistically
Fig. 1. Visualization of AgNOR dots in epithelial (E) and mesenchymal (M)cells of ameloblastic fibroma (silver staining, ¿1250).
AgNORs in odontogenic tumours
Fig. 2. Visualization of AgNOR dots in cells of odontogenic myxoma (silverstaining, ¿1250).
significant difference in AgNOR means between epithelial and ecto-
mesenchymal components of AF, but a significant difference in
AgNOR means was observed between the ectomesenchymal cells
of both lesions.
Discussion
The AgNOR technique has been useful in differentiating benign
tumours from malignant tumours (6), although it has been of little
value in the diagnosis of some tumours. It is more useful in research
than in routine diagnosis. Many studies have associated the AgNOR
technique with other proliferative cellular indices, in lesions from
respiratory tract, skin tumours, prostate and bladder lesions and
491J Oral Pathol Med 30: 489–93
acute leukemias (11–13). Moreover, the AgNOR technique has been
useful both in diagnosis and in prognosis of salivary gland tumours
(14).
There are some controversial results in the literature about
studies using the AgNOR technique in oral mucosal carcinomas and
in odontogenic tumours. Some authors compared the mean AgNOR
index among odontogenic cysts and ameloblastomas and demon-
strated that AgNOR counts have neither a diagnostic nor a prognos-
tic value with respect to these lesions (8). However, other authors
compared odontogenic cysts and unicystic ameloblastoma, and ob-
served that there were statistically significant differences in AgNOR
counts among these lesions; according to these authors, this sug-
gests that these differences may or may not be an indication of
variations in metabolic, proliferative and transcriptional activities
(9).
The use of the AgNOR technique in comparative studies of
ameloblastomas and adenomatoid odontogenic tumours demon-
strated that both lesions have similar cell proliferation activity, im-
plying that the difference between their clinical behaviour is prob-
ably not correlated with their cell proliferation indices (15).
The use of the AgNOR technique with proliferating cell nuclear
antigen (PCNA) expression in ossifying fibroma (OF) and peripheral
ossifying fibroma (POF) has suggested an increased proliferative
activity in OF when compared with POF. This result is in accord-
ance with the more aggressive clinical behaviour of OF (16).
Some authors analysed ameloblastoma and basal cell carcinoma
(BBC) by AgNOR and observed that, although these lesions are
neoplasms with similar clinical behaviour, they have cell popula-
tions with statistically significant differences in AgNOR counts (17).
There was no statistically significant difference in AgNOR
counts between the epithelial and the mesenchymal components of
AF. This observation suggests that these components have similar
cell proliferation indices that could be related to the mixed nature
of AF where both the epithelial and the mesenchymal components
are considered to be neoplastic (18). In spite of its benign nature,
the rare cases of recurrences with possible malignant transform-
ation would be an important factor in the neoplastic characteristic
of AF (19).
According to the literature, the neoplastic characteristic of the
ectomesenchyme of AF was due to interactions of epithelium and
mesenchyme, which are related to tumour growth factor b2 (TGF-
b2) (19). It is present in AF, but absent in OM. It was suggested
that absence of TGF-b2 in OM could result in loss of inductive
capacity of the epithelium and its involution or disappearance (20).
It has also been suggested that a possible explanation for that dif-
ference in epithelial and mesenchymal interactions is due to tenascin
and cellular fibronectin in AF, regulating the growth and fetal pheno-
Martins et al.
type of neoplastic cells. These proteins are absent in OM. It was
further suggested that an early odontogenic induction may result in
a transformation of the fibroblasts into cells with enzymatic activity
similar to mature odontoblasts. These fibroblasts might lose their
ability to produce and secrete collagen, resulting in either a fibroblast
with low differentiation or a metabolically immature fibroblast (20,
21). The synthesis and degradation rate of the collagen is the basis of
the equilibrium of the connective tissue. In OM, the presence of cavi-
tation areas suggests that degradation is higher than synthesis, while
in AF the organization of the connective tissue rich in fibroblasts sug-
gests that synthesis is higher than degradation (22).
According to some authors, the mean AgNOR counts may be
correlated more to synthesis than to the rate of cellular proliferation
(6). Thus, our results may reflect either the higher proliferation
index or a greater capacity for synthesis of AF than of OM.
In the present study the mean AgNOR counts in AF was higher
than in OM. Thus, our results suggest that, although AF is com-
monly delimited by a fibrous capsule, the rate of growth of AF is
probably higher than that of OM. Furthermore, the proliferative
index of AF is probably not related to its inability to infiltrate ad-
jacent tissues. Therefore, these factors seem to indicate that the
infiltrative and recurrent features of OM are probably not related to
its cell proliferation index.
Some authors consider the infiltrative and recurrent features of
OM to be related to its gelatinous consistency and lack of an envel-
oping capsule. Likewise, mitosis is rarely seen, and the growth ratio
of the OM depends on the accumulation of myxoid substance (1, 2).
As a consequence, its low metabolic activity and the absence of
mitosis may result in a lower AgNOR index. Although it is not
compatible with its infiltrative and recurrent growth, it is compat-
ible with its slow growth. Moreover, the neoplastic cells of OM have
a high alkaline phosphatase and ATPase activity and a low acidic
phosphatase activity. These observations suggest a low metabolic
activity, which is compatible with its slow growth (19).
In conclusion, our results suggest that the cellular proliferation
ratio of AF is higher than that of OM. Thus, the different clinical
behaviours between these lesions with regard to their infiltrative
ability is probably not related to the proliferative index of their
tumour cells.
References
1. Piattelli A, Scarano A, Antinori A, Trisi P. Odontogenic myxoma of themandible: report of a case and review of the literature. Acta StomatolBelg 1994; 91: 101–10.
492 J Oral Pathol Med 30: 489–93
2. Schneck DL, Gross PD, Tabor MW. Odontogenic myxoma: report of twocases with reconstruction considerations. J Oral Maxillofac Surg 1993;51: 935–40.
3. Dallera P, Bertoni F, Marchetti C, Bachini P, Campobassi A. Ameloblasticfibroma: a follow-up of six cases. Int J Oral Maxillofac Surg 1996; 25:199–202.
4. Mosby EL, Russell D, Noren S, Barker BF. Ameloblastic fibroma in a 7week-old infant: a case report and review of the literature. J Oral Maxil-lofac Surg 1998; 56: 368–72.
5. Muller S, Parker DC, Kapadia SB, Budnick SD, Barnes EL. Ameloblasticfibrosarcoma of the jaws; a clinicopathologic and DNA analysis of fivecases and review of the literature with discussion of its relationship toameloblastic fibroma. Oral Surg Oral Med Oral Pathol Oral Radiol En-dod 1995; 79: 469–77.
6. Derenzini M, Ploton D. Interphase nucleolar organizer regions in cancercells. Int Rev Exp Pathol 1991; 32: 149–92.
7. Derenzini M, Trere D. AgNOR proteins as a paramater of the rapidityof cell proliferation. Zentralbl Pathol 1994; 140: 7–10.
8. Allison R T, Spencer S. Nucleolar organizer regions in odontogenic cystsand ameloblastomas. Br J Biomed Sci 1993; 50: 309–12.
9. Coleman HG, Altini M, Groeneveld H. Nucleolar organizer regions(AgNORs) in odontogenic cysts and ameloblastomas. J Oral Pathol Med1996; 25: 436–40.
10. Ploton D, Menanger M, Jeannensson P, Himber G, Pigeon Fadnet JJ. Im-provement in the staining and the visualization of the argyrophilic pro-teins of the nucleolar organizer region at the optical level. Histochem J1986; 18: 5–14.
11. Pich A, Chiusa L, Margaria E. Role of the visualization of the argyrophil-ic nucleolar organizer regions in tumour detection and prognosis. CancerDetect Prev 1995; 19: 282–91.
12. Ahiskali R, Alican Y, Ekicioglu G, Cevik I, Kullus S, Akdas A. Evalu-ation of three different AgNOR counting methods in advanced carci-noma of the prostate. Prostate 1995; 26: 105–10.
13. Loran D, Metze I, Carvalho MA, Metze K. Relationship between morpho-metric analysis of AgNOR and cell proliferation in acute leukemias.Cytometry 1998; 32: 51–6.
14. Freitas RA, Araujo VC, Araujo NS. Argyrophilia in nucleolar organizerregions (AgNOR) in adenomatoid carcinoma and polymorphous low-grade adenocarcinoma of the salivary glands. Eur Arch Otorhinolaringol1993; 250: 213–17.
15. do Carmo MAV, Silva EC. Argyrophilic nucleolar organizer regions(AgNORs) in ameloblastomas and adenomatoid odontogenic tumours(AOTs). J Oral Pathol Med 1998; 27: 153–6.
16. Mesquita RA, Souza SCOM, Araujo NS. Proliferative activity in periph-eral ossifying fibroma (POF) and ossifying fibroma (OF). J Oral PatholMed 1998; 27: 64–7.
17. Rosa LEB, Jaeger MMM, Jaeger RG. Morphometric study of nucleolarorganizer regions in ameloblastoma and basal cell carcinoma. Oral Oncol1997; 33: 209–14.
18. Sano K, Yoshida S, Ninomiya H, et al. Assessment of growth potentialby MIB-1 immunohistochemistry in ameloblastic fibroma and relatedlesions of the jaws compared with ameloblastic fibrosarcoma. J OralPathol Med 1998; 27: 59–63.
19. Heikinheimo K, Happonen RP, Miettinen PJ, Ritvos O. Transforminggrowth factor B2 in epithelial differentiation of developing teeth andodontogenic tumours. J Clin Invest 1993; 91: 1019–27.
20. Auriol M, Chomette G, Martino R, Bertrand JC, Gulbert F. Myxome odon-togene; etude histoenzimologique et ultrastructurale; a propos de 5 ob-servations. J Biol Buccale 1986; 14: 215–22.
AgNORs in odontogenic tumours
21. Schmidt-Westhausen A, Becker J, Schuppan D, Burkhardt A, ReichartPA. Odontogenic myxoma: characterisation of the extracellular matrix(ECM) of the tumour stroma. Eur J Cancer B Oral Oncol 1994; 30: 377–80.
493J Oral Pathol Med 30: 489–93
22. Gotjamanos, T. Intracellular collagen in recurrent ameloblastic fibroma.J Oral Pathol 1979; 8: 277–83.