The pathogenesis of palmar fibromatosis remains uncertain. The disease contains two fibrotic structures, the nodule and the cord. As the disease progresses, the nodule, which contains proliferative fibroblasts, with a high proportion being myofibroblasts, develops into the cord, a collagen-rich and acellular structure [15
]. The appearance and disappearance of myofibroblasts in the lesions are similar to those observed in other tissues in which fibroblasts are present, such as granulation tissue. Interleukin-1(IL-1), platelet-derived growth factor-BB (PDGF-BB), and transforming growth factor-β (TGF-β) stimulate the growth of fibroblasts [17
]. The effect of TGF-β on the proliferation of myofibroblasts has been studied in vitro [16
]. The results indicate that TGF-β could induce fibroblast proliferation and increases the expression of α-SMA.
Desmoid fibromatosis (aggressive fibromatosis) is clonal fibroblastic proliferation. Despite the lack of the ability to metastasize, local recurrence is frequent and some desmoids prove fatal due to the madly local recurrence, especially in the significant viscera. In contrast, the rate of local recurrence of palmar fibromatosis is much lower, and the process of involution, from a cellular proliferation of nodule to the acellular scar-like tissue, is also different from desmoid fibromatosis.
The present study of fibromatosis shows that clonal chromosome changes are another feature of the disease process. Wever [19
] found that the frequency of cytogenetic abnormalities varied with the type of fibromatosis. The results demonstrated that more than a half of samples from desmoid fibromatosis had clonal chromosome aberrations, and only 3 out of 28 specimens of superficial fibromatosis had aberrations. The frequent finding of clonal chromosome changes in desmoid fibromatosis confirms the neoplastic nature of demoid fibromatosis, which is in line with molecular data showing that demoid fibromatosis is monoclonal [6
]. Our results suggested that palmar fibromatosis is a polyclonal reactive proliferation. It explains the infrequent chromosome aberrations in superficial fibromatosis, and the different clinic outcome and prognosis between palmar fibromatosis and desmoid fibromatosis. The both lesions belong to subtypes of fibromatosis, however, they are hyperplasia with completely different characters, namely one is reactive proliferation while the other is monoclonal proliferation.
Although the technique of clonality analysis based on the pattern of X-chromosome inactivation is powerful, there are still certain limitations. First, the purity of the cells we investigated could be reduced by the contamination of the normal cells, such as endothelial cells of vasculatures and fatty cells. Hence, it is unclear whether the appearance of polyclonality is genuine or due to contamination. In order to minimize any possibility of contamination, we used laser cut microdissection to gain the spindled cells, which allowed for rapid and accurate acquisition of cells that we were interested in [20
], and tried to reduce the contamination of endothelial cells and fatty cells. Only a single band was observed in the sample of rectum adenocarcinoma, which was used as positive control. The result suggested that tumor cells are accurately harvested, none normal tissue components is existed, and the amount of methylation-sensitive restriction endonuclease we used is suitable, by which DNA template was completely digested. The test samples DNA were digested in identical fashion as the positive control. Second, being based on the pattern of X-chromosome inactivation, the technique of clonal analysis is only applied to female patients. The application of the lesions that are more common in men than women is limited, such as palmar fibromatosis. The third limitation is the occurrence of nonrandom X-chromosome inactivation (also known as skewing or unequal Lyonization) in healthy females. The skewed X-chromosome inactivation pattern mimics clonal derivation of cells, and makes clonality results non-informative [23
]. In our study, eight informative samples were successful in amplification after digestion with methylation-sensitive restriction endonuclease HpaII, and presented two allelic bands of approximately equal intensity. The results not only assess palmar fibromatosis is polyclonality, but also show that none of the eight patients is the skewed X-chromosome inactivation pattern.
DNA from both frozen tissues and formalin-fixed, paraffin-embedded tissues were suitable for clonality analysis [24
]. In the current study, only formalin-fixed, paraffin-embedded tissues due to the rare of palmar fibromatosis in women and the limited number of specimens suitable for analysis. This approach take the advantage of the large archival paraffin-embedded specimens that spanned many years and are easy to collect. In addition, the structure of tissues and the shapes of cells on paraffin-embedded specimens were clearer than frozen tissue, which made the diagnosis more credible and isolation of spindle cells of interest more precise. However, partial DNA template were degraded and damaged during process of formalin-fixed and paraffin-embedded, therefore, relative large amounts of DNA are needed from paraffin-embedded tissues comparing with frozen tissues. In our study, we cut fifteen 10 μm thick sections from each block, which ensured the amounts of DNA template, and we extracted DNA three times with Tris-buffer-saturated phenol-chloroform and once with chloroform, which purified crude DNA samples to ensure successful PCR amplification. Nevertheless, three of the twelve samples (cases 5, 10, and 11) fail to be amplified. We analyze the possible reasons for this failure. Both of lesions from patient 10 and patient 11 were in the residual stage, which was characterized by less cellular and increased amounts of dense collagen, therefore, there were so limited amounts of DNA template that PCR failed to amply. Surely, the damage of DNA template was not excluded. However, case 5 was in the proliferative phase, we speculated that there was scant optimal DNA template, which made a failure of PCR amplification. Because of the rupture of DNA, the PCR amplicon should be controlled within less than 500 base pairs. In our study, the DNA sequence we chosen to amplified was less than 300 base pairs, which ensured the success of PCR.