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Aims and Background: Altered oncogenic activity is a feature associated with many malignant and premalignant conditions. Among the many oncogenes, ras and myc are commonly altered in many tumors. This study aims to evaluate the expression of ras and c-myc oncoproteins in a total of 204 cervical tissue samples, including premalignant and malignant lesions as well as apparently normal cervical tissue., Methods and Study Design: Mouse monoclonal antibodies against the three mammalian ras gene products (c-H-ras, c-K-ras, c-N-ras) and the c-myc protein were used to evaluate oncoprotein expression by immunocytochemistry., Results: None of the samples analyzed displayed immunoreactivity for H-ras and K-ras. Normal cervical epithelium showed minimal immunoreactivity for N-ras with about 33% of the samples expressing the protein. More conspicuous expression in normal tissue was displayed by c-myc, with about 90% of the samples expressing the protein (mean value of cells positive=34%). The immunoreactivity for N-ras increased with increasing histological abnormality from low-grade squamous intraepithelial lesions (SIL) to invasive carcinoma. Increased immunoreactivity for N-ras was evident in the basaloid cells of malignant lesions, with the maximum value of 66% found in poorly differentiated squamous cell carcinoma (PDSCC). The percentage of nuclei positive for c-myc also showed a gradual increase from low-grade SIL onwards, the highest positivity being found in PDSCC, where the mean value was 85%. Statistical analysis revealed a good correlation between the expression of N-ras (r=0.8922, P=0.001) and c-myc (r=0.8856, P=0.001) and various histological stages of tumor progression in the cervical epithelium., Conclusions: These results therefore suggest that c-myc and N-ras oncoproteins are important during tumor progression in the uterine cervix.

The differential expression of cytokeratins in epithelial or squamous cells has been demonstrated to be altered during the process of carcinogenesis. This altered expression of cytokeratins (CKs) may be closely related with epithelial differentiation and may remain stable in malignant tumors. In the present study an analysis using two monoclonal antibodies, CK 8.12 antibody specific for CK type 13 and 16 and CK 8.60 antibody specific for CK type 1, 10 and 11 was done in different grades of lesions in the uterine cervix. Changes from the normal expression pattern were seen in high grade Squamous intraepithelial lesions (SIL) (CIN-2/3) and invasive squamous cell carcinoma (SCC). No conspicuous difference in the staining expression between normal/benign cervical tissue and low grade SIL (CIN-I) was evident. Statistical analysis also revealed a significant correlation between the expression of these CK types to the differentiation status of the cervical lesions analyzed. Alterations in the expression of these CKs can be correlated to the differentiation pathway which may be deregulated during cervical carcinogenesis. The findings of the present study suggest that the expression of CK types 13 and 16 and 1, 10 and 11 using CK 8.12 and CK 8.60 antibodies respectively may serve as markers of differentiation in cervical squamous neoplasms.

Immunocytochemical localization of the basement membrane (BM) proteins laminin, type-IV collagen and fibronectin were analyzed in normal cervical epithelium, low grade squamous intraepithelial lesions (SILs), high grade SILs and invasive squamous cell carcinoma (SCC) of the uterine cervix. A regular, thick and continuous BM was present in normal cervical epithelium and low grade SIL. Interruptions and discontinuity of the BM were more evident in high grade SILs. There was a good correlation between increasing severity of the lesion and increasing number of breaks. In SCC, the distribution of laminin, collagen IV and fibronectin was related to the degree of cellular differentiation, with decreased immunoreactivity being evident in moderately and poorly differentiated tumors. As the invasive potential of the tumor increased, the fragmentation and loss of BM was more evident. Fibronectin showed only moderate to mild immunoreactivity in normal cervical epithelium and low grade SILs. However, the intensity of expression increased in high grade SILs especially in the peritumoral stroma. It may therefore be concluded from these results that snythesis and reabsorption of BM proteins may be related to shifts in cellular metabolism during tumorigenesis.

The expression of cytokeratins (CKs) in normal cervical epithelium, low grade squamous intraepithelial lesions (SIL), high grade SILs and squamous cell carcinoma (SCC) were analyzed using four different monoclonal antikeratin antibodies. In normal cervical epithelium, CK 18 showed strong immunoreactivity in basal and parabasal layers. CK 19 and 14 were expressed only in the basal layer while CK 13 was found selectively n the spinal cells. As the lesions progressed from low grade SIL to high grade SIL, immunoreactivity of CK 18, 19 and 14 in the basal cell compartment increased while the expression of CK 13 decreased. In SCC, as well-differentiated tumors showed decreased immunoreactivity for CK 18, 19 and 14 with CK 13 showing a strong and focal (localized) immunoreactivity. Undifferentiated carcinomas totally lacked CK 13 reactivity. Our findings therefore suggest that expression of CK 18, 19 and 14 may be directly related to tumor grade and CK 13 may be a marker of differentiation in cervical lesions.

The expression of involucrin, a cytoplasmic protein synthesized during squamous maturation, was assessed by immunocytochemistry in different grades of cervical lesions. In normal/benign cervical epithelium and low-grade squamous intraepithelial lesions [SILS or cervical intraepithelial neoplasia (CIN)-1] involucrin showed intense and homogenous cytoplasmic expression in the spinal layers of 75 and 57% of samples, respectively. The basal cell layers showed no expression of involucrin. In high-grade SILs (CIN-2/3) 40% of the samples showed diffuse and focal cytoplasmic expression of involucrin in the differentiated basaloid cells. In the squamous cell carcinomas (SCCs) analyzed, well-differentiated tumors showed intense focal expression in 61% of the cases, moderately differentiated SCCs showed intense expression in 33% of the cases, while poorly differentiated SCCs (PDSCC) showed only a mild focal expression in 7% of cases. With increasing severity of the lesions, patchy expression of involucrin with a mixture of reactive and nonreactive cells predominated. Patterns of immunocytochemical staining for involucrin in cervical lesions of different grades, from low-grade to high-grade SILs, and invasive carcinoma may be of critical importance, if loss of involucrin expression is used as a criterion for neoplastic transformation in cervical epithelium. Our findings suggest that involucrin may be a sensitive marker in identifying the differentiation status of the lesion while the absence of involucrin in PDSCC may be helpful in differential diagnosis.