Evidence of cellular senescence during the development of estrogen-induced pituitary tumors

Abstract

Although pituitary adenomas represent 25% of intracranial tumors, they are usually benign, with the mechanisms by which these tumors usually avoid an invasive profile and metastatic growth development still remaining unclear. In this context, cellular senescence might constitute a plausible explanation for the benign nature of pituitary adenomas. In this study, we investigated the emergence of cellular senescence as a growth control mechanism during the progression of estrogen-induced pituitary tumors. The quantification of Ki67-immunopositive cells in the pituitaries of estrogenized male rats after 10, 20, 40, and 60 days revealed that the mitogenic potential rate was not sustained for the whole period analyzed and successively decreased after 10 days of estrogen exposure. In addition, the expression of cellular senescence features, such as the progressive rise in the enzymatic senescenceassociated b-galactosidase (SA-b-gal) activity, IL6, IL1b, and TGFb expression, was observed throughout pituitary tumor development. Furthermore, tumoral pituitary cells also displayed nuclear pATM expression, indicating activated DNA damage signaling, with a significant increase in p21 expression also being detected. The associations among DNA damage signaling activation, SA-b-gal expression, and p21 may provide a reliable combination of senescence-associated markers for in vivo pituitary senescence detection. These results suggest a role for this cellular process in the regulation of pituitary cell growth. Thus, cellular senescence should be conceived as a contributing component to the benign nature of pituitary adenomas, thereby influencing the capability of the pituitary gland to avoid unregulated cell proliferation.

Oxidative stress and mitochondrial adaptive shift during pituitary tumoral growth

“…We found an increased protein level of DRP-1 phosphorilated serine 637 ( Figure 1I), indicating the inhibition of mitochondrial fission [53]. These results, supported also by our previous findings regarding MFN-1 and DRP-1 proteins suggest an imbalance of the protein involved in mitochondrial dynamics, thereby favouring fusion processes [17].…”

“…Three-month-old Wistar strain male rats were maintained at controlled temperature (21±3 °C) and lighting conditions (14 h light: 10 h darkness cycle), with free access to commercial laboratory chow and tap water. Taking into account that exogenous estrogen excess induces pituitary tumours [17,32], intact animals were treated with estradiol benzoate (Sigma Aldrich, St. Louis, MO, USA) for 10, 20, 40, and 60 days (E10, E20, E40, and E60). Estrogen was implanted subcutaneously in slow releasing silastic brand capsules (Dow Corning, Medical grade, Midland, MI, USA) filled with 10 mg of estrogen crystals and sealed with silastic cement.…”

“…Paraffin-embedded pituitary glands were processed by immunohistochemistry for 8-OHG as described previously [17]. Briefly, an antigen retrieval method using a hot antigen unmasking solution was performed (0.01 M citrate buffer; pH 6), and in order to inhibit endogenous peroxidase, sections were treated with 3 % v/v hydrogen peroxide in methanol for 15 min.…”

“…Few attempts to elucidate mitochondrial health in pituitary tumours have been made [18][19][20][21][22], with currently little being known about mitochondrial behaviour during endocrine tumour development. The pituitary gland mostly develops benign proliferative injuries with the coexistence of mitotic events together with cellular senescence arrested cells and practically null apoptotic rates [17][18][19][20][21][22][23][24][25][26]. As a dynamic and plastic tissue the pituitary gland constantly adapts hormonal and metabolic responses to the changing environments [27].…”

Inhibitory role of ERβ on anterior pituitary cell proliferation by controlling the expression of proteins related to cell cycle progression

“…The control rats showed serum E2 values of 829.67 ± 252.7 pg/ml, whereas in the development of the hyperplastic/adenomatous process the values of the serum E2 obtained were always higher than 2000 pg/ml. The effect of estrogen as an inducer of pituitary tumor development was documented in semithin sections from epoxy resin-embedded glands by high-resolution light microscopy, as described previously (Sabatino et al, 2015). Briefly, pituitaries were fixed in a mixture of 4% v/v formaldehyde and 2% v/v glutaraldehyde in 0.1 M cacodylate buffer, and then treated with 1% OsO 4 before being stained in block with 1% v/v uranyl acetate.…”