Aliesha González-Arenas

Changes in Progesterone Receptor Isoforms Content in the Rat Brain During the Oestrous Cycle and After Oestradiol and Progesterone Treatments

We studied the effects of oestradiol and progesterone on progesterone receptor (PR) isoform content in the brain of ovariectomized rats and in intact rats during the oestrous cycle by Western blot analysis. In the hypothalamus and the preoptic area of ovariectomized rats, PR‐A and PR‐B content was increased by oestradiol, whereas progesterone significantly diminished the content of both PR isoforms after 3 h of treatment in the hypothalamus, but not in the preoptic area. In the hippocampus, only PR‐A content was significantly increased by oestradiol while progesterone significantly diminished it after 12 h of treatment. In the frontal cortex, no treatment significantly modified PR isoform content. During the oestrous cycle, the lowest content of PR isoforms in the hypothalamus was observed on diestrus day and, by contrast, in the preoptic area, the highest content of both PR isoforms was observed on diestrus day. We observed no changes in PR isoform content in the hippocampus during the oestrous cycle. These results indicate that the expression of PR isoforms is differentially regulated by sex steroid hormones in a regionally specific manner.

Macrophage--Mycobacterium tuberculosis interactions: role of complement receptor 3.

Tuberculosis is the leading infectious disease in the world. Mycobacterium tuberculosis, the causal agent of this disease, invades macrophages and can replicate inside them. Because invasion of macrophages is a critical step for establishing a mycobacterial infection, there is much interest in understanding the mechanisms for M. tuberculosis entry into macrophages. Complement receptor 3 (CR3) is a heterodimeric surface receptor with multiple binding sites, which can mediate complement-opsonized as well as nonopsonic entrance of M. tuberculosis into macrophages. Here, we describe and discuss the role of CR3 in macrophage[bond]M. tuberculosis interactions. The actual information suggests that CR3 mediates a substantial amount of M. tuberculosis binding to macrophages, but CR3 is not related to the mechanisms that allow mycobacteria to survive and replicate intracellularly. Understanding the mechanisms of macrophage[bond]M. tuberculosis interaction will help developing more effective methods to prevent and treat tuberculosis in the future.

Participation of the 26S proteasome in the regulation of progesterone receptor concentrations in the rat brain.

The aim of this study was to investigate the participation of the 26S proteasome in the regulation of progesterone receptor (PR) concentrations in the rat brain in vivo. Ovariectomized adult female rats were treated with estradiol (10 µg/100 g s.c.), estradiol + progesterone (400 µg/100 g), and vehicle (corn oil/10% ethanol) in the presence or absence of the proteasome inhibitor Z-Ile-Glu (OBu1)-Ala-Leu-H (PSI, 300 µg/100 g). Proteins were extracted from the preoptic area, the hippocampus, and the frontal cortex, and processed for Western blot. Estradiol-induced PR expression in the preoptic area and the hippocampus, whereas progesterone did not modify the effect of estradiol. Neither estradiol nor progesterone modified PR content in the frontal cortex. PSI treatment increased PR content in the preoptic area and the hippocampus. This increase was significant in both regions after 24 h of the treatment with progesterone + PSI in the animals primed with estradiol. In this case, the content of both PR isoforms (PR-A and PR-B) was increased in a similar manner by PSI in the preoptic area (90 and 97%) and in the hippocampus (49 and 50%). PSI did not affect PR content in the frontal cortex. Our results suggest that the 26S proteasome could participate in the turnover of PR in the preoptic area and the hippocampus of the rat in vivo.

Regulation of progesterone receptor isoforms content in human astrocytoma cell lines

Progesterone regulates several functions through the interaction with its intracellular receptor (PR) which expresses two isoforms with different functions and regulation: PR-A and PR-B. Both PR isoforms have been detected in human astrocytomas, the most common and aggressive primary brain tumours, but their regulation and function are unknown. We studied the effects of estradiol, progesterone and their receptor antagonists (ICI 182,780 and RU 486) on PR isoforms content in U373 and D54 human astrocytoma cell lines, respectively derived from grades III and IV astrocytomas, by Western blot analysis. In U373 cells we also evaluated the effects of PR-A overexpression on cell growth. We observed that in U373 cells estradiol increased the content of both PR isoforms whereas in D54 cells it had no effects. Estradiol effects were blocked by ICI 182,780. In both cell lines, PR isoforms content was down-regulated by progesterone after estradiol treatment. This effect was blocked by RU 486. We observed that overexpression of PR-A significantly diminished the increase in U373 cells number produced after progesterone treatment. Our results suggest a differential PR isoforms regulation depending on the evolution grade of human astrocytoma cells, and an inhibitory role of PR-A on progesterone effects on astrocytomas cell growth.

Regulation of the Phosphoinositide-3 Kinase and Mitogen-Activated Protein Kinase Signaling Pathways by Progesterone and Its Reduced Metabolites in the Rat Brain

Several growth factors, such as vascular endothelial growth factor, brain‐derived neurotrophic factor, and insulin‐like growth factor‐I are involved in the actions of progesterone in the central nervous system. Previous studies in neuronal and glial cultures have shown that progesterone may regulate growth factor signaling, increasing the phosphorylation of extracellular‐signal regulated kinase (ERK) and the phosphorylation of Akt, components of the mitogen‐activated protein kinase (MAPK) and the phosphoinositide‐3 kinase (PI3K) signaling pathways, respectively. In this study, we have evaluated whether progesterone and its reduced metabolites, dihydroprogesterone and tetrahydroprogesterone, regulate PI3K and MAPK signaling in the brain of ovariectomized rats in vivo. Significant increases in the phosphorylation of ERK, in the expression of the catalytic (p110) and the regulatory (p85) subunits of PI3K and in the phosphorylation of Akt were observed in the hypothalamus, the hippocampus, and the cerebellum 24 hr after progesterone administration. Progesterone metabolites partially mimicked the effect of progesterone and had a stronger effect on MAPK and PI3K signaling in the hypothalamus than in the other brain regions. These findings suggest that progesterone regulates MAPK and PI3K signaling pathways in the central nervous system in vivo by direct hormonal actions and by mechanisms involving progesterone metabolites.

Changes in the content of steroid receptor coactivator-1 and silencing mediator for retinoid and thyroid hormone receptors in the rat brain during the estrous cycle ☆

In this work, we determined the variations in the content of the steroid receptor coactivator (SRC-1) and the silencing mediator for retinoic acid and thyroid hormone receptors corepressor (SMRT) in the hypothalamus, the preoptic area, and the hippocampus of adult intact rats during the estrous cycle by Western blot. SRC-1 content changed only in the hypothalamus where its lowest content was found on diestrus day with a significant increase at proestrus. This increase was maintained on estrus day. In contrast, SMRT content changed only in the preoptic area where it diminished at metestrus in comparison with the other days of the cycle. SRC-1 content was higher than that of SMRT in the hypothalamus throughout the estrous cycle, whereas SMRT content was higher in the preoptic area. In the hippocampus, there were no significant differences in the content of any cofactor. These results demonstrate that SRC-1 and SMRT content change in a tissue-specific manner in the rat brain during the estrous cycle, and suggest that the transcriptional activity of steroid hormone receptors in the rat brain in physiological conditions is regulated by changes in SRC-1 and SMRT content.

Estradiol increases cell growth in human astrocytoma cell lines through ERα activation and its interaction with SRC-1 and SRC-3 coactivators ☆

Estradiol (E2) regulates several cellular functions through the interaction with estrogen receptor subtypes, ERα and ERβ, which present different functional and regulation properties. ER subtypes have been identified in human astrocytomas, the most common and aggressive primary brain tumors. We studied the role of ER subtypes in cell growth of two human astrocytoma cell lines derived from tumors of different evolution grades: U373 and D54 (grades III and IV, respectively). E2 significantly increased the number of cells in both lines and the co-administration with an ER antagonist (ICI 182, 780) significantly blocked E2 effects. ERα was the predominant subtype in both cell lines. E2 and ICI 182, 780 down-regulated ERα expression. The number of U373 and D54 cells significantly increased after PPT (ERα agonist) treatment but not after DPN (ERβ agonist) one. To determine the role of SRC-1 and SRC-3 coactivators in ERα induced cell growth, we silenced them with RNA interference. Coactivator silencing blocked the increase in cell number induced by PPT. The content of proteins involved in proliferation and metastasis was also determined after PPT treatment. Western blot analysis showed that in U373 cells the content of PR isoforms (PR-A and PR-B), EGFR, VEGF and cyclin D1 increased after PPT treatment while in D54 cells only the content of EGFR was increased. Our results demonstrate that E2 induces cell growth of human astrocytoma cell lines through ERα and its interaction with SRC-1 and SRC-3 and also suggest differential roles of ERα on cell growth depending on astrocytoma grade.

Regulation of progesterone receptor isoforms expression by sex steroids in the rat lung

In this work, we determined the expression pattern and the hormonal regulation of progesterone receptor (PR) isoforms in the rat lung of ovariectomized female rats after estradiol (E2) and progesterone (P4) treatments. We also evaluated the content of estrogen receptor beta (ER-beta) which is the ER isoform expressed in the lung. RNA and proteins were extracted and processed for reverse transcription (RT) coupled to polymerase chain reaction (PCR) and Western blot, respectively. The expression of both PR isoforms in the lung at mRNA and at protein levels was up-regulated by E2 while P4 down-regulated it at mRNA level. P4 did not modify PR isoforms protein content unlike its effect in the uterus where both PR isoforms were down-regulated by their ligand at mRNA and protein levels. PR-A was the predominant isoform, both in the lung and in the uterus. In the lung, ER-beta was down-regulated by E2 while P4 did not significantly modify the effect of E2. These results suggest that both PR isoforms should be expressed in the rat lung, and that their expression should be differentially regulated at mRNA and at protein levels by P4. We also suggest that the up-regulation of PR isoforms by E2 in the lung is mediated by ER-beta.

Progesterone Induces the Growth and Infiltration of Human Astrocytoma Cells Implanted in the Cerebral Cortex of the Rat

Progesterone (P4) promotes cell proliferation in several types of cancer, including brain tumors such as astrocytomas, the most common and aggressive primary intracerebral neoplasm in humans. In this work, we studied the effects of P4 and its intracellular receptor antagonist, RU486, on growth and infiltration of U373 cells derived from a human astrocytoma grade III, implanted in the motor cortex of adult male rats, using two treatment schemes. In the first one, fifteen days after cells implantation, rats were daily subcutaneously treated with vehicle (propylene glycol, 160 μL), P4 (1 mg), RU486 (5 mg), or P4 + RU486 (1 mg and 5 mg, resp.) for 21 days. In the second one, treatments started 8 weeks after cells implantation and lasted for 14 days. In both schemes we found that P4 significantly increased the tumor area as compared with the rest of the treatments, whereas RU486 blocked P4 effects. All rats treated with P4 showed tumor infiltration, while 28.6% and 42.9% of the animals treated with RU486 and P4 + RU486, respectively, presented it. Our data suggest that P4 promotes growth and migration of human astrocytoma cells implanted in the motor cortex of the rat through the interaction with its intracellular receptor.

Role of progesterone receptors during postpartum estrus in rats.

We studied the role of progesterone receptor (PR) in the display of female sexual behavior during postpartum estrus in rats. Adult female rats were treated with the PR antagonist, RU486 (1.25 and 5 mg), 3 h after parturition and sexual behavior was evaluated throughout the first postpartum day. Estradiol and progesterone serum levels changed during the first 24 h postpartum. The highest estradiol and progesterone levels were found at 9 and 12 h postpartum, respectively. The predominant PR isoform in the hypothalamus and the preoptic area was PR-A during postpartum day. The content of PR-A increased at 6 h postpartum in the hypothalamus and the preoptic area, and decreased in both regions at 9 h. PR-B content only increased in the preoptic area at 12 h postpartum. The highest display of lordotic and proceptive behaviors were found at 12 h postpartum. The treatment with 1.25 and 5 mg of RU486 respectively reduced lordosis by 61% and 92% at 12 h postpartum. These results suggest that PR is essential in the display of postpartum estrus in rats.