We studied the circadian rhythm of plasma melatonin, growth hormone (GH), prolactin (PRL), adrenocorticotropic hormone (ACTH), and cortisol in 52 mentally healthy old subjects, 35 old demented patients, and 22 clinically healthy young controls. When compared to young controls, the circadian profile of plasma melatonin of old subjects, both demented or not, was clearly flattened, particularly during the night. The selective impairment of nocturnal melatonin secretion was significantly related to both the age and the severity of mental impairment (Mini Mental State Examination [MMSE] score). The PRL and GH circadian profiles were similar in the three groups during the day, but a significant lowering of the values recorded during the night occurred with aging. The impairment of the nocturnal secretion was related to the subjects' age and, for the GH secretory pattern only, also to the MMSE score. The ACTH circadian profile was similar in the three groups studied, even when old subjects exhibited higher ACTH levels throughout the 24 h cycle, compared to young controls. Significantly higher cortisol values at evening- and nighttime occurred in elderly subjects and particularly in the demented group. Both the mean levels and the nadir values of plasma cortisol were positively related to age and negatively to MMSE score. In order to verify the sensitivity of the hypothalamo-pituitary-adrenal (HPA) axis to the steroid feedback, the circadian profile of plasma cortisol was evaluated also after dexamethasone (DXM) administration (1 mg at 23:00 h); the sensitivity of the HPA axis was significantly impaired in old subjects and particularly in the demented ones. These findings suggest that the neuroendocrine alterations already present in physiological aging, due to both anatomical damages and unbalanced central neurotransmitters, are enhanced in senile dementia.

The circadian organization of adrenal secretion was studied in 23 healthy elderly subjects, 23 elderly demented patients and 10 healthy young subjects, in order to investigate the relationships between the hypothalamic-pituitary-adrenal axis and some cerebral morphometric parameters. The cerebral morphometric analysis was performed in some subjects of the three groups by MRI. A significant increase in cortisol levels during evening and nighttime was found in both groups of the aged subjects. In elderly subjects, particularly if demented, the mean serum dehydroepiandrosterone sulfate (DHEAs) levels throughout the 24-hour cycle were significantly lower than in young controls. A significant reduction of the hippocampal and temporal volume and an enlargement of the lateral ventricles were found in aged subjects, these changes being significantly related to subjects’ age. Moreover, the hippocampal volume was positively correlated with the circadian mesor of DHEAs (i.e., the circadian rhythm adjusted mean) and with the cortisol nocturnal increase. Our data may suggest the existence of a link between the selective impairment of cortisol secretion and DHEAs levels, and the progression of hippocampal degeneration.

Parathyroid tumors deregulate microRNAs belonging to the two clusters on the chromosome 19, the C19MC and miR-371-373 clusters. Here, we report that the embryonic miR-372 is aberrantly expressed in half of parathyroid adenomas (PAds) in most of atypical adenomas and carcinomas ( = 15). Through hybridization, we identified that miR-372-positive parathyroid tumor cells were scattered throughout the tumor parenchyma. In PAd-derived cells, ectopic miR-372 inhibited the expression of its targets/p21 and LATS2 at both mRNA and protein levels. Although the viability of parathyroid cells was not affected by miR-372 overexpression, the miRNA blunted camptothecin-induced apoptosis in primary PAd-derived cultures. miR-372 overexpression in parathyroid tumor cells increased parathormone () mRNA levels, and it positively correlated with circulating PTH levels. Conversely, the parathyroid-specific genes and were not affected by miR-372 mimic transfection. Finally, miR-372 dampened the Wnt pathway in parathyroid tumor cells through DKK1 upregulation. In conclusion, miR-372 is a novel mechanism exploited by a subset of parathyroid tumor cells to partially decrease sensitivity to apoptosis, to increase PTH synthesis and to deregulate Wnt signaling.

(250 words) 53Parathyroid tumors display reduced sensitivity to extracellular calcium ([Ca 2+ ] o ). [Ca 2+ ] o 54 activates calcium-sensing receptor (CASR), which interacts with the scaffold protein filamin A 55 (FLNA). The study aimed to investigate: 1) the FLNA expression in human parathyroid tumors, 2) 56 its effects on the CASR mRNA and protein expression, and 3) on ERK signaling activation, 4) the 57 effect of the carboxy-terminal CASR variants and 5) of the treatment with the CASR agonist R568 58 on FLNA-mediated ERK phosphorylation in HEK293 cells. 59Full-length FLNA immunostaining was variably reduced in parathyroid tumors. 60Immunofluorescence showed that FLNA localized in membrane and cytoplasm and co-localized 61 with CASR in parathyroid adenomas (PAds)-derived cells. Cleaved C-terminus FLNA fragment 62 could also be detected in PAds nuclear protein fractions. In HEK293 cells transfected with 990R-63 CASR or 990G-CASR variants, silencing of endogenous FLNA reduced CASR mRNA levels and 64 total and membrane-associated CASR proteins. In agreement, FLNA mRNA levels positively 65 correlated with CASR expression in a series of 74 PAds; however, any significant correlation with 66 primary hyperparathyroidism severity could be detected and FLNA transcript levels did not differ 67 between PAds harboring 990R or 990G CASR variants. R568 treatment was efficient in restoring 68 990R-CASR and 990G-CASR sensitivity to [Ca 2+ ] o in absence of FLNA. 69In conclusion, FLNA is down-regulated in parathyroid tumors and parallels the CASR 70 expression levels. Loss of FLNA reduces CASR mRNA and protein expression levels and the 71 CASR-induced ERK phosphorylation. FLNA is involved in receptor expression, membrane 72 localization and ERK signaling activation of both 990R and 990G CASR variants.

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