Downregulation of calcineurin activity in cervical carcinoma

Abstract

BACKGROUND: Calcineurin (CaN) is an important serine-threonine phosphatase (PP2B), which plays a crucial role in calcium-calmodulin mediated signal transduction events. Calcineurin has been implicated in pathogenesis of various diseases cardiac hypertrophy, diabetic neuropathy and Alzheimer's, however its role in neoplasia remains unclear. RESULTS: In view of this we evaluated the calcineurin activity in serum and biopsy samples collected from women diagnosed with invasive squamous cell carcinoma of cervix. A significant reduction was observed in the calcineurin activity in cancer cervix patients compared to the control group. However the calcineurin activity remained unaltered in the cervical scrapes obtained from patients diagnosed with low-grade squamous intra epithelial lesions (LSIL). Interestingly the downregulation of calcineurin activity in squamous cell carcinomas was not accompanied by any significant change in DNA-binding affinity of the transcriptional factor NFAT (Nuclear Factor of Activated T-cells). All the squamous cell carcinoma samples used in the present study were positive for high-risk human papillomavirus (HPV) types. CONCLUSION: The present study demonstrates the downregulation of calcineurin activity in squamous cell carcinoma of cervix with high risk HPV infection. We conclude that perturbations in calcineurin-mediated pathway may be involved in development of cervical neoplasia.

Figure 1

Calcineurin activity in serum (a) and biopsy samples (b) of control and cervical cancer patients diagnosed with squamous cell carcinoma (SCC). The calcineurin activity is significantly reduced in serum samples [1a, p = 0.0001] and in tissue lysates [1b, p = 0.0037] obtained from cancer patients compared to the normal group. The calcineurin activity was calculated as nmoles of inorganic phosphate released per mg protein of tissues or per deciliter of serum.

Figure 2

Total calcineurin content in normal and cervical cancer tissue lysates [SCC]. The calcineurin content was assayed by using antibody specific to calcineurin [Sigma monoclonal anti-CaNα, 1:7500 dilution using a sandwich ELISA method.

Figure 3

Total calmodulin content in normal and cervical cancer tissue lysates (SCC) were determined by using antibody specific to calmodulin. Note a significant increase [p = 0.0263] in calmodulin content in cancer patients.

Figure 4

Calcineurin activity as determined in the cervical scrapes obtained from control (N = 25) and those diagnosed with low-grade intraepithelial squamous lesion (LSIL) or CIN1. Note, no significant change in calcineurin activity in control and LSIL group.

Figure 5

NFAT-DNA-binding activity in normal and cervical cancer nuclear extracts. Briefly 100 μg of nuclear extracts was incubated with ³²P-labelled NFATc oligonucleotide and Electrophoretic mobility shift assay was performed as described in Material and methods. Lane one contains free probe without nuclear extract and lane 2 contains 100-fold excess of unlabelleled NFAT oligonucleotide as a specific competitor. Specific NFAT- DNA complexes are indicated by arrow.