Protein kinase A regulatory subunit distribution in medulloblastoma

Abstract

Background: Previous studies showed a differential distribution of the four regulatory subunits of cAMP-dependent protein kinases inside the brain, that changed in rodent gliomas: therefore, the distribution of these proteins inside the brain can give information on the functional state of the cells. Our goal was to examine human brain tumors to provide evidence for a differential distribution of protein kinase A in different tumors.

Methods: The distribution of detergent insoluble regulatory (R1 and R2) and catalytic subunits of cAMP dependent kinases was examined in pediatric brain tumors by immunohistochemistry and fluorescent cAMP analogues binding.

Results: R2 is organized in large single dots in medulloblastomas, while it has a different appearance in other tumors. Fluorescent cAMP labelling was observed only in medulloblastoma.

Conclusions: A different distribution of cAMP dependent protein kinases has been observed in medulloblastoma.

Figure 1

Medulloblastoma cases 1 (A to C) and 2 (D to F). A: SAF-cAMP labelling. B: R2 immunolabelling. C: merge of A and B: SAF-cAMP labelling does not overlap to R2. D: SAF-cAMP labelling. E: R2 immunolabelling. F: merge of D and E: the two signals appear separated. Bar = 10 μm, 100× objective.

Figure 2

Medulloblastoma, case 3. A: SAF-cAMP labelling. B: R2 immunolabelling. C: merge of A and B, showing that SAF-cAMP does not overlap to R2. D: SAF-cAMP labelling. E: immunohistochemistry to reveal PKA catalytic subunit. F: merge of D and E: most of SAF-cAMP labelled structures are labelled also by antibody that reveals PKA catalytic subunit. Bar = 10 μm, 100× objective.

Figure 3

Immunohistochemistry on different samples. A: MB relapse, R2 immunolabelling, similar to primitive MB specimens, 40× objective. B: ETANTR, R2 immunolabelling (red). Arrows indicate three structures not seen in MB specimens. 100× objective. C to E: Ewing's sarcoma relapse. C and D: same field, 40× objective C: R1 immunolabelling in the form of fine dots; this kind of labelling was not present in MB samples. D: small autofluorescence granuli, albeit not numerous, were present in this specimen; autofluorescence was not present in primitive MB that did not undergo therapy. E: R2 immunolabelling, 20× objective. F: primary culture obtained from Ewing's sarcoma relapse; anti-R1 immunohistochemistry, 40× objective. Bar = 10 μm (B); bar = 25 μm (A, C, D, F); bar = 50 μm (E).