Prognostic factors for outcome of pituitary surgery in dogs with corticotroph adenomas

Abstract

Pituitary-dependent hypercortisolism (PDH) is a common endocrinopathy in dogs, caused by an ACTH secreting adenoma in the pituitary gland. Dogs with PDH can be treated with medication, radiation therapy or surgery. Although postoperative results are good, long-term recurrences do occur in around 25% of the dogs. The aims of this thesis were to analyze the long-term follow-up of dogs with PDH treated with transsphenoidal hypophysectomy over a 20-year period and to study the prognostic value of peri-operative measurement of plasma ACTH and cortisol concentrations in predicting recurrence of hypercortisolism. The aims also included the identification of possible prognostic pituitary tissue markers with immunohistochemistry and gene expression analysis, and the identification and characterization of stem cells in the normal canine pituitary gland.

We showed that dogs with a larger pituitary have shorter survival and disease free periods after surgery, and that the pituitary size of patients at time of surgery significantly increased over the 20-year period. The evaluation of the individual peri-operative hormone curves of ACTH and cortisol of hypophysectomy patients provided valuable information about the risk of recurrence. However, it was not possible to identify an exact cut-off point to identify all recurrences with a single hormone measurement. Therefore, analysis of other clinical parameters, like pituitary size, remains essential.

The second part of the thesis focused on the expression of possible prognostic markers in pituitary tissue. We found that the expression of Ki-67 and PCNA (proliferation markers) were not significantly different between enlarged and non-enlarged pituitaries. The cell-cycle inhibitor p27kip1 trended to be expressed less in enlarged pituitaries compared to the non-enlarged pituitaries. We found Pax7 [removed]specific marker for intermediate lobe cells) in adenomas originating in the anterior and intermediate lobe, but we found no correlation between Pax7 expression or activation of the Pax7 signaling pathway to clinical parameters and outcome. In the process, also pituitary tissue specific reference genes were established.

Isolation of stem cells from pituitary gland tissue is complicated by the lack of specific stem cell markers. We were not able to relate Sox2 [removed]possible pituitary stem cell marker) to clinical parameters in dogs with pituitary adenomas. Fluorescence activated cell sorting (FACS) was used to sort a side population (SP) of cells. FACS was previously used successfully in multiple tissue types to isolate a SP containing stem cells. In this thesis, we successfully isolated and characterized the SP-cells from six healthy canine pituitary glands.

In conclusion, transsphenoidal hypophysectomy is an effective treatment for dogs with PDH. It remains unclear why, after initial remission, some dogs develop recurrence of hypercortisolism, whereas others do not. Of the markers studied in this thesis, p27kip1 and Pax7 remain interesting for future studies, indicating a disruption of cell-cycle control as the basis for pituitary tumorigenesis. However, these markers cannot be used as absolute prognostic markers. Continued studies are needed to unravel the role of stem cells in the pathogenesis of canine pituitary adenomas. What is urgently needed is a specific biomarker to predict surgical outcome, i.e. remission or recurrence.