Luteolin and its analog luteolin-7-methylether from Leonurus japonicus Houtt suppress aromatase-mediated estrogen biosynthesis to alleviate polycystic ovary syndrome by the inhibition of tumor progression locus 2.

ETHNOPHARMACOLOGICAL RELEVANCE: Leonurus japonicus Houtt (L. japonicus, Chinese motherwort), known as Yi Mu Cao which means "good for women", has long been widely used in China and other Asian countries to alleviate gynecological disorders, often characterized by estrogen dysregulation. It has been used for the treatment of polycystic ovary syndrome (PCOS), a common endocrine disorder in women but the underlying mechanism remains unknown. AIM OF THE STUDY: The present study was designed to investigate the effect and mechanism of flavonoid luteolin and its analog luteolin-7-methylether contained in L. japonicus on aromatase, a rate-limiting enzyme that catalyzes the conversion of androgens to estrogens and a drug target to induce ovulation in PCOS patients. MATERIALS AND METHODS: Estrogen biosynthesis in human ovarian granulosa cells was examined using ELISA. Western blots were used to explore the signaling pathways in the regulation of aromatase expression. Transcriptomic analysis was conducted to elucidate the potential mechanisms of action of compounds. Finally, animal models were used to assess the therapeutic potential of these compounds in PCOS. RESULTS: Luteolin potently inhibited estrogen biosynthesis in human ovarian granulosa cells stimulated by follicle-stimulating hormone. This effect was achieved by decreasing cAMP response element-binding protein (CREB)-mediated expression of aromatase. Mechanistically, luteolin and luteolin-7-methylether targeted tumor progression locus 2 (TPL2) to suppress mitogen-activated protein kinase 3/6 (MKK3/6)-p38 MAPK-CREB pathway signaling. Transcriptional analysis showed that these compounds regulated the expression of different genes, with the MAPK signaling pathway being the most significantly affected. Furthermore, luteolin and luteolin-7-methylether effectively alleviated the symptoms of PCOS in mice. CONCLUSIONS: This study demonstrates a previously unrecognized role of TPL2 in estrogen biosynthesis and suggests that luteolin and luteolin-7-methylether have potential as novel therapeutic agents for the treatment of PCOS. The results provide a foundation for further development of these compounds as effective and safe therapies for women with PCOS.

Discovery of Novel ERα and Aromatase Dual-Targeting PROTAC Degraders to Overcome Endocrine-Resistant Breast Cancer.

Estrogen receptor α (ERα) plays a pivotal role in the proliferation, differentiation, and migration of breast cancer (BC) cells, and aromatase (ARO) is a crucial enzyme in estrogen synthesis. Hence, it is necessary to inhibit estrogen production or the activity of ERα for the treatment of estrogen receptor-positive (ER+) BC. Herein, we present a new category of dual-targeting PROTAC degraders designed to specifically target ERα and ARO. Among them, compound 18c bifunctionally degrades and inhibits ERα/ARO, thus effectively suppressing the proliferation of MCF-7 cells while showing negligible cytotoxicity to normal cells. In vivo, 18c promotes the degradation of ERα and ARO and inhibits the growth of MCF-7 xenograft tumors. Finally, compound 18c demonstrates promising antiproliferative and ERα degradation activity against the ERαMUT cells. These findings suggest that 18c, being the inaugural dual-targeting degrader for ERα and ARO, warrants further advancement for the management of BC and the surmounting of endocrine resistance.

Developmental Delay and Male-Biased Sex Ratio in esr2b Knockout Zebrafish.

The estrogen receptor signaling pathway plays an important role in vertebrate embryonic development and sexual differentiation. There are four major estrogen receptors in zebrafish: esr1, esr2a, esr2b and gper. However, the specific role of different estrogen receptors in zebrafish is not clear. To investigate the role of esr2b in zebrafish development and reproduction, this study utilized TALENs technology to generate an esr2b knockout homozygous zebrafish line. The number of eggs laid by esr2b knockout female zebrafish did not differ significantly from that of wild zebrafish. The embryonic development process of wild-type and esr2b knockout zebrafish was observed, revealing a significant developmental delay in the esr2b knockout zebrafish. Additionally, mortality rates were significantly higher in esr2b knockout zebrafish than in their wild-type counterparts at 24 hpf. The reciprocal cross experiment between esr2b knockout zebrafish and wild-type zebrafish revealed that the absence of esr2b resulted in a decline in the quality of zebrafish oocytes, while having no impact on sperm cells. The knockout of esr2b also led to an abnormal sex ratio in the adult zebrafish population, with a female-to-male ratio of approximately 1:7. The quantitative PCR (qPCR) and in situ hybridization results demonstrated a significant downregulation of cyp19ab1b expression in esr2b knockout embryos compared to wild-type embryos throughout development (at 2 dpf, 3 dpf and 4 dpf). Additionally, the estrogen-mediated induction expression of cyp19ab1b was attenuated, while the estradiol-induced upregulated expression of vtg1 was disrupted. These results suggest that esr2b is involved in regulating zebrafish oocyte development and sex differentiation.

Small angle scattering reveals the orientation of cytochrome P450 19A1 in lipoprotein nanodiscs.

Human aromatase (CYP19A1), the cytochrome P450 enzyme responsible for conversion of androgens to estrogens, was incorporated into lipoprotein nanodiscs (NDs) and interrogated by small angle X-ray and neutron scattering (SAXS/SANS). CYP19A1 was associated with the surface and centered at the edge of the long axis of the ND membrane. In the absence of the N-terminal anchor, the amphipathic A'- and G'-helices were predominately buried in the lipid head groups, with the possibly that their hydrophobic side chains protrude into the hydrophobic, aliphatic tails. The prediction is like that for CYP3A4 based on SAXS employing a similar modeling approach. The orientation of CYP19A1 in a ND is consistent with our previous predictions based on molecular dynamics simulations and lends additional credibility to the notion that CYP19A1 captures substrates from the membrane.

CYP19A1 Expression Is Controlled by mRNA Stability of the Upstream Transcription Factor AP-2γ in Placental JEG3 Cells.

CYP19A1 encodes aromatase, which converts testosterone to estrogen, and is induced during placental maturation. To elucidate the molecular mechanism underlying this function, histone methylation was analyzed using the placental cytotrophoblast cell line, JEG3. Treatment of JEG3 cells with 3-deazaneplanocin A, an inhibitor of several methyltransferases, resulted in increased CYP19A1 expression, accompanied by removal of the repressive mark H3K27me3 from the CYP19A1 promoter. However, this increase was not observed in cells treated with GSK126, another specific inhibitor for H3K27me3 methylation. Expression of TFAP2C, which encodes AP-2γ, a transcription factor that regulates CYP19A1, was also elevated on 3-deazaneplanocin A treatment. Interestingly, TFAP2C messenger RNA (mRNA) was readily degraded in JEG3 cells but protected from degradation in the presence of 3-deazaneplanocin A. TFAP2C mRNA contained N6-methyladenosines, which were reduced on drug treatment. These observations indicate that the TFAP2C mRNA undergoes adenosine methylation and rapid degradation, whereas 3-deazaneplanocin A suppresses methylation, resulting in an increase in AP-2γ levels. We conclude that the increase in AP-2γ expression via stabilization of the TFAP2C mRNA is likely to underlie the increased CYP19A1 expression.

Synthesis, in vitro and in silico anticancer evaluation of novel pyridin-2-yl estra-1,3,5(10)-triene derivatives.

Aim: The aim of this study was the synthesis of steroid compounds with heterocyclic rings and good anticancer properties. Materials & methods: The synthesis, in silico and in vitro anticancer testing of novel pyridin-2-yl estra-1,3,5(10)-triene derivatives was performed. Results: All synthesized compounds have shown promising results for, antiproliferative activity, relative binding affinities for the ligand binding domains of estrogen receptors α, ß and androgen receptor, aromatase binding potential, and inhibition of AKR1C3 enzyme. Conclusion: 3-Benzyloxy (17E)-pycolinilidene derivative 9 showed the best antitumor potential against MDA-MB-231 cell line, an activity that can be explained by its moderate inhibition of AKR1C3. Molecular docking simulation indicates that it binds to AKR1C3 in a very similar orientation and geometry as steroidal inhibitor EM1404.

The series of pyridine-containing estra-1,3,5(10)-triene derivatives was synthesized. One novel derivative stood out by its excellent activity against the MDA-MB-231 cell line. This activity can be explained by its moderate inhibition of the AKR1C3 enzyme.

A Novel Investigation for Early Sex Determination in Alive Adult European Seabass (Dicentrarchus labrax) Using cyp19a1a, dmrt1a, and dmrt1b Genes Expression in Tail Fin tissues.

This study is the first investigation for using sex-related gene expression in tail fin tissues of seabass as early sex determination without killing the fish. The European seabass (Dicentrarchus labrax) is gonochoristic and lacks distinguishable sex chromosomes, so, sex determination is referred to molecular actions for some sex-related genes on autosomal chromosomes which are well known such as cyp19a1a, dmrt1a, and dmrt1b genes which play crucial role in gonads development and sex differentiation. cyp19a1a is expressed highly in females for ovarian development and dmrt1a and dmrt1b are for testis development in males. In this study, we evaluated the difference in the gene expression levels of studied genes by qPCR in tail fins and gonads. We then performed discriminant analysis (DA) using morphometric traits and studied gene expression parameters as predictor tools for fish sex. The results revealed that cyp19a1a gene expression was significantly higher in future females' gonads and tail fins (p ≥ 0.05). Statistically, cyp19a1a gene expression was the best parameter to discriminate sex even the hit rate of any other variable by itself could not correctly classify 100% of the fish sex except when it was used in combination with cyp19a1a. In contrast, Dmrt1a gene expression was higher in males than females but there were difficulties in analyzing dmrt1a and dmrt1b expressions in the tail because levels were low. So, it could be used in future research to differentiate and determine the sex of adult fish using the cyp19a1a gene expression marker without killing or sacrificing fish.

New enantioenriched ß-indolyl ketones as aromatase inhibitors: Unraveling heme-ligand interactions by MD simulation and MMPBSA analysis.

A series of enantioenriched ß-indolyl ketones as aromatase inhibitors (AI) is synthesized through the Michael-type Friedel-Crafts alkylation of indole. A highly efficient bifunctionalized amino catalyst is developed to access structurally diverse ß-indolyl ketones in high yields (up to 91%) and excellent enantioselectivity (enantiomeric ratio up to 98:2). All the synthesized compounds demonstrated promising aromatase inhibitory potential, where ortho-substituted analogs (3c and 3e) were found most active with IC50 values of 0.68 and 0.90 µM, respectively. Both of these compounds exhibited significant cytotoxicity (IC50 = 0.34 and 0.37 µM) against the MCF-7 breast cancer cell line in the (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay. Molecular docking studies of the synthesized compounds demonstrate favorable binding interactions with the estrogens controlling CYP19A1 (3EQM) and metabolizing CYP3A4 (5VCC) enzymes. Molecular dynamic (MD) simulation analysis revealed the essentiality of heme-ligand interactions to build a stable protein-ligand complex. An average root mean square deviation of 0.35 nm observed during a 100-ns MD simulation and binding free energy in the range of -190 to -227 kJ/mol calculated by g_mmpbsa analysis authenticated the stability of the 3c-3EQM complex. ADMET and drug-likeness parameters supported the suitability of these indole derivatives as the drug lead to develop potent inhibitors for estrogen-dependent breast cancer.

The expression profiles of cyp19a1, sf-1, esrs and gths in the brain-pituitary during gonadal sex differentiation in juvenile Japanese eels.

Eels are gonochoristic species whose gonadal differentiation initiates at the yellow eel stage and is influenced by environmental factors. We revealed some sex-related genes were sex dimorphically expressed in gonads during gonadal sex differentiation of Japanese eel (Anguilla japonica); however, the expression of sex-related genes in the brain-pituitary during gonadal sex differentiation in eels is still unclear. This study aimed to investigate the sex-related gene expressions in the brain-pituitary and tried to clarify their roles in the brain and gonads during gonadal sex differentiation. Based on our previous histological study, the control eels developed as males, and estradiol-17ß (E2) was used for feminization. Our results showed that during testicular differentiation, the brain cyp19a1 transcripts and aromatase proteins were increased significantly; moreover, the cyp19a1, sf-1, foxl2s, and esrs (except gperb) transcripts in the midbrain/pituitary also were increased significantly. Forebrain gnrh1 transcripts increased slightly during gonadal differentiation of both sexes, but the gnrhr1b and gnrhr2 transcripts in the midbrain/pituitary were stable during gonadal differentiation. The expression levels of gths and gh in the midbrain/pituitary were significantly increased during testicular differentiation and were much higher in males than in E2-feminized females. These results implied that endogenous estrogens might play essential roles in the brain/pituitary during testicular differentiation, sf-1, foxl2s, and esrs may have roles in cyp19a1 regulation in the midbrain/pituitary of Japanese eels. For the GnRH-GTH axis, gths, especially fshb, may be regulated by esrs and involved in regulating testicular differentiation and development in Japanese eels.

Lipopolysaccharide-binding protein in follicular fluid is associated with the follicular inflammatory status and granulosa cell steroidogenesis in dairy cows.

Metabolic stress and subsequent hepatic dysfunction in high-producing dairy cows are associated with inflammatory diseases and declining fertility. Lipopolysaccharide (LPS)-binding protein (LBP) is produced by hepatocytes and controls the immune response, suggesting that it is involved in the pathophysiology of inflammation-related attenuation of reproductive functions during metabolic stress. This study investigated the effect of LBP on the inflammatory status, oocyte quality, and steroidogenesis in the follicular microenvironment of dairy cows. Using bovine ovaries obtained from a slaughterhouse, follicular fluid and granulosa cells were collected from large follicles to evaluate the follicular status of metabolism, inflammation, and steroidogenesis. Cumulus-oocyte complexes were aspirated from small follicles and subjected to in vitro embryo production. The results showed that follicular fluid LBP concentrations were significantly higher in cows with fatty livers and hepatitis than in those with healthy livers. Follicular fluid LBP and LPS concentrations were negatively correlated, whereas LPS concentration showed a positive correlation with the concentrations of non-esterified fatty acids (NEFA) and ß-hydroxybutyric acid in follicular fluid. The blastulation rate of oocytes after in vitro fertilization was impaired in cows in which coexisting large follicles had high NEFA levels. Follicular fluid NEFA concentration was negatively correlated with granulosa cell expression of the estradiol (E2) synthesis-related gene (CYP19A1). Follicular fluid LBP concentration was positively correlated with follicular fluid E2 concentration and granulosa cell CYP19A1 expression. In conclusion, follicular fluid LBP may be associated with favorable conditions in the follicular microenvironment, including low LPS levels and high E2 production by granulosa cells.

Role of neuroestrogens in the regulation of social behaviors - From social recognition to mating.

In this mini-review, we summarize the brain distribution of aromatase, the enzyme catalyzing the synthesis of estrogens from androgens, and the mechanisms responsible for regulating estrogen production within the brain. Understanding this local synthesis of estrogens by neurons is pivotal as it profoundly influences various facets of social behavior. Neuroestrogen action spans from the initial processing of socially pertinent sensory cues to integrating this information with an individual's internal state, ultimately resulting in the manifestation of either pro-affiliative or - aggressive behaviors. We focus here in particular on aggressive and sexual behavior as the result of correct individual recognition of intruders and potential mates. The data summarized in this review clearly point out the crucial role of locally synthesized estrogens in facilitating rapid adaptation to the social environment in rodents and birds of both sexes. These observations not only shed light on the evolutionary significance but also indicate the potential implications of these findings in the realm of human health, suggesting a compelling avenue for further investigation.

Neuropeptide W protects against ovarian oxidative injury and reinforces ovarian steroidogenic activity via the upregulation of ERα expression.

OBJECTIVES: The aim of this study was to investigate the protective effect of neuropeptide W (NPW) on ovarian ischemia-reperfusion-induced oxidative injury and ovarian steroid metabolism. METHODS: Rats were randomly divided into control and ischemia groups that received either saline or NPW (0.1 or 5 µg/kg/day). Bilateral ovarian ischemia was performed for 3 h followed by a 72-h reperfusion. Blood, ovary, and uterus samples were collected for biochemical and histological assessments. KEY FINDINGS: Treatment with either dose of NPW alleviated oxidative injury of the ovaries with a significant suppression in free radical formation and decreased histopathological injury in both the ovarian and uterine tissues, along with reduced lipid peroxidation and neutrophil accumulation in the uterus. Moreover, NPW treatment reversed the decrease in aromatase expression with a concomitant reduction in the expression of the inactivity enzyme estrogen sulfotransferase. Also, downregulation of estrogen receptor-α (ERα) expression in the injured ovarian tissue was abolished by NPW treatment, which implicates that the protective effect of NPW on the female reproductive system may involve the upregulation of the ERα-mediated signaling pathway. CONCLUSIONS: Our study demonstrated for the first time that NPW protects against ovarian oxidative injury and reinforces ovarian steroidogenic activity, which is accompanied by the upregulation of ERα expression in the ovaries.

Using cell culture systems from the Persian Gulf Arabian yellowfin sea bream, Acanthopagrus arabicus, to assess the effects of dexamethasone on gonad and brain aromatase activity and steroid production.

Dexamethasone (DEX) is a synthetic glucocorticoid widely used as pharmaceutical and usually exists in effluents with varying degrees of concentrations. In this study, cultivated Brain, ovary and testis cells from Arabian Sea bream, Acanthopagrus arabicus, were treated by DEX at concentrations of 0, 0.3, 3.0, 30.0 and 300.0 µg/ml for 48 h. The aromatase activity and steroid (17-ß-estradiol (E2), progesterone (P) and testosterone (T)) production by cells were measured at 12, 24 and 48 h of the experiment. The results showed that the sensitivity of cultivated ovarian, testicular and brain cells to DEX increased dose dependently. DEX was potent inhibitor of aromatase activity at specially 30.0 and 300.0 µg/ml in the cultivated ovarian and testicular cells at different sampling time. On the other hand, DEX was found to stimulate the aromatase activity of fish brain. DEX also decreased E2, P and T production by cultivated ovarian and testicular cells during the experiment. While, DEX caused an increase in the production of E2 and P by brain cells, which seems logical considering the stimulating effect of this drug on brain aromatase activity. In conclusion, results highlight that DEX is able to change the activity of aromatase, and disrupt the biosynthesis of estrogens and thus affect reproduction in fish.

Sex-specific vulnerabilities in human astrocytes underpin the differential impact of palmitic acid.

Obesity and neurometabolic diseases have been linked to neurodegenerative diseases. Our hypothesis is that the endogenous estrogenic component of human astrocytes plays a critical role in cell response during lipotoxic damage, given that obesity can disrupt hormonal homeostasis and cause brain inflammation. Our findings showed that high concentrations of palmitic acid (PA) significantly reduced cell viability more in male astrocytes, indicating sex-specific vulnerabilities. PA induced a greater increase in cytosolic reactive oxygen species (ROS) production in males, while female astrocytes exhibited higher superoxide ion levels in mitochondria. In addition, female astrocytes treated with PA showed increased expression of antioxidant proteins, including catalase, Gpx-1 and Nrf2 suggesting a stronger cellular defence mechanism. Interestingly, there was a difference in the expression of estrogenic components, such as estrogen, androgens, and progesterone receptors, as well as aromatase and 5α-reductase enzymes, between males and females. PA induced their expression mainly in females, indicating a potential protective mechanism mediated by endogenous hormones. In summary, our findings highlight the impact of sex on the response of human astrocytes to lipotoxicity. Male astrocytes appear to be more susceptible to cellular damage when exposed to high concentrations of fatty acids.

The spatio-temporal distribution of aromatase cytochrome in ovary throughout the canine oestrous cycle.

CONTEXT: New animal welfare legislation and ethical guidelines encourage alternative approaches for canine contraception, instead of surgical gonadectomy which is considered invasive and unjustified in healthy dogs. AIMS: Reversible contraception might be achieved by inhibition of aromatase (CYP19), an enzyme catalysing the conversion of androgens to oestrogens. This study provides insights into the spatio-temporal expression and distribution of aromatase in canine ovarian tissue. METHODS: Ovarian tissue was collected from 39 healthy and sexually mature bitches during different stages of the oestrous cycle: pro-oestrus (n =8), oestrus (n =12), dioestrus (n =9) (luteal phase) and anoestrus (n =10). Localisation of cytochrome P450 aromatase was determined by immunohistochemistry. KEY RESULTS: Aromatase activity in the dog is high during pro-oestrus, ovulation and early dioestrus. Comparing types of follicles and corpora lutea, the highest aromatase abundance was found in antral follicles and luteinising follicles, whereas corpora lutea and early antral follicles showed an intermediate presence of the enzyme. Interesting was the high abundance of aromatase in luteinising theca interna cells, prevailing over granulosa cells. CONCLUSIONS AND IMPLICATIONS: Understanding of cells involved in oestradiol production is important for targeted inhibition of oestradiol synthesis, possibly offering an approach for contraception and suppression of oestrus.

Neonatal Aromatase Inhibition Blocked Defeminization of AVPV Kiss1 Neurons and LH Surge-Generating System in Male Rats.

The neuroendocrine system that controls the preovulatory surge of gonadotropin-releasing hormone (GnRH)/luteinizing hormone (LH), which triggers ovulation in female mammals, is sexually differentiated in rodents. A transient increase in circulating testosterone levels in male rats within a few hours of birth is primarily responsible for the defeminization of anteroventral periventricular nucleus (AVPV) kisspeptin neurons, which are critical regulators of the GnRH/LH surge. The present study aimed to determine whether neonatal estradiol-17ß (E2) converted from testosterone by aromatase primarily causes the defeminization of AVPV kisspeptin neurons and the surge of GnRH/LH in male rodents. The results of the present study showed that the neonatal administration of letrozole (LET), a nonsteroidal aromatase inhibitor, within 2 hours of birth rescued AVPV Kiss1 expression and the LH surge in adult male rats, while the neonatal administration of testosterone propionate (TP) irreversibly attenuated AVPV Kiss1 expression and the LH surge in adult female rats. Furthermore, the neonatal LET-treated Kiss1-Cre-activated tdTomato reporter males exhibited a comparable number of AVPV Kiss1-Cre-activated tdTomato-expressing cells to that of vehicle-treated female rats, while neonatal TP-treated females showed fewer AVPV Kiss1-Cre-activated tdTomato-expressing cells than vehicle-treated females. Moreover, neonatal TP administration significantly decreased the number of arcuate Kiss1-expressing and Kiss1-Cre-activated tdTomato-positive cells and suppressed LH pulses in adult gonadectomized female rats; however, neonatal LET administration failed to affect them. These results suggest that E2 converted from neonatal testosterone is primarily responsible for the defeminization of AVPV kisspeptin neurons and the subsequent GnRH/LH surge generation in male rats.

Charged Amino Acids in the Transmembrane Helix Strongly Affect the Enzyme Activity of Aromatase.

Estrogens play critical roles in embryonic development, gonadal sex differentiation, behavior, and reproduction in vertebrates and in several human cancers. Estrogens are synthesized from testosterone and androstenedione by the endoplasmic reticulum membrane-bound P450 aromatase/cytochrome P450 oxidoreductase complex (CYP19/CPR). Here, we report the characterization of novel mammalian CYP19 isoforms encoded by CYP19 gene copies. These CYP19 isoforms are all defined by a combination of mutations in the N-terminal transmembrane helix (E42K, D43N) and in helix C of the catalytic domain (P146T, F147Y). The mutant CYP19 isoforms show increased androgen conversion due to the KN transmembrane helix. In addition, the TY substitutions in helix C result in a substrate preference for androstenedione. Our structural models suggest that CYP19 mutants may interact differently with the membrane (affecting substrate uptake) and with CPR (affecting electron transfer), providing structural clues for the catalytic differences.

Exploring the Potential of Essential Oil from Plectranthus amboinicus Leaves against Breast Cancer: In vitro and In silico Analysis.

Plectranthus amboinicus leaves were subjected to hydrodistillation to obtain essential oil (EO). Phytochemical analysis using gas chromatography-mass spectrometry revealed a diverse range of compounds in the EO, with p-cymen-4-ol (18.57%) emerging as the most predominant, followed by isocaryophyllene (12.18%). The in vitro antiproliferative activity of EO against breast cancer was assessed in MCF-7 and MDA-MB-231 cell lines. The MTT assay results revealed that EO showed IC50 values of 42.25 µg/mL and 13.44 µg/mL in MCF-7 cells and 63.67 µg/mL and 26.58 µg/mL in MDA-MB-231 cells after 24 and 48 h, respectively. The in silico physicochemical and pharmacokinetic profiles of the EO constituents were within acceptable limits. Molecular docking was conducted to investigate the interactions between the constituents of the EO and protein Aromatase (PDB ID:3S79). Among the EO constituents, 4-tert-butyl-2-(5-tert-butyl-2-hydroxyphenyl)phenol (4BHP) exhibited the highest dock score of -6.580 kcal/mol when compared to the reference drug, Letrozole (-5.694 kcal/mol), but was slightly lesser than Anastrozole (-7.08 kcal/mol). Molecular dynamics simulation studies (100 ns) of the 4BHP complex were performed to study its stability patterns. The RMSD and RMSF values of the 4BHP protein complex were found to be 2.03 Å and 4.46 Å, respectively. The binding free energy calculations revealed that 4BHP displayed the highest negative binding energy of -43 kcal/mol with aromatase protein, compared to Anastrozole (-40.59 kcal/mol) and Letrozole (-44.54 kcal/mol). However, further research is required to determine the safety, efficacy, and mechanism of action of the volatile oil. Taking into consideration the key findings of the present work, the development of a formulation of essential oil remains a challenging task and novel drug delivery systems may lead to site-specific and targeted delivery for the effective treatment of breast cancer.

PPARγ antagonists induce aromatase transcription in adipose tissue cultures.

Aromatase is the rate-limiting enzyme in the biosynthesis of estrogens and a key risk factor for hormone receptor-positive breast cancer. In postmenopausal women, estrogens synthesized in adipose tissue promotes the growth of estrogen receptor positive breast cancers. Activation of peroxisome proliferator-activated receptor gamma (PPARγ) in adipose stromal cells (ASCs) leads to decreased expression of aromatase and differentiation of ASCs into adipocytes. Environmental chemicals can act as antagonists of PPARγ and disrupt its function. This study aimed to test the hypothesis that PPARγ antagonists can promote breast cancer by stimulating aromatase expression in human adipose tissue. Primary cells and explants from human adipose tissue as well as A41hWAT, C3H10T1/2, and H295R cell lines were used to investigate PPARγ antagonist-stimulated effects on adipogenesis, aromatase expression, and estrogen biosynthesis. Selected antagonists inhibited adipocyte differentiation, preventing the adipogenesis-associated downregulation of aromatase. NMR spectroscopy confirmed direct interaction between the potent antagonist DEHPA and PPARγ, inhibiting agonist binding. Short-term exposure of ASCs to PPARγ antagonists upregulated aromatase only in differentiated cells, and a similar effect could be observed in human breast adipose tissue explants. Overexpression of PPARG with or without agonist treatment reduced aromatase expression in ASCs. The data suggest that environmental PPARγ antagonists regulate aromatase expression in adipose tissue through two mechanisms. The first is indirect and involves inhibition of adipogenesis, while the second occurs more acutely.

Transforming Growth Factor α Evokes Aromatase Expression in Gastric Parietal Cells during Rat Postnatal Development.

Estrogen, well known as a female hormone, is synthesized primarily by ovarian aromatase. However, extra-glandular tissues also express aromatase and produce estrogen. It is noteworthy that aromatase in gastric parietal cells begins expression around 20 days after birth and continues secreting considerable amounts of estrogen into the portal vein throughout life, supplying it to the liver. Estrogen, which is secreted from the stomach, is speculated to play a monitoring role in blood triglyceride, and its importance is expected to increase. Nevertheless, the regulatory mechanisms of the aromatase expression remain unclear. This study investigated the influence of transforming growth factor α (TGFα) on gastric aromatase expression during postnatal development. The administration of TGFα (50 µg/kg BW) to male Wistar rats in the weaning period resulted in enhanced aromatase expression and increased phosphorylated ERK1+2 in the gastric mucosa. By contrast, administration of AG1478 (5 mg/kg BW), a protein tyrosine kinase inhibitor with high selectivity for the epidermal growth factor receptor and acting as an antagonist of TGFα, led to the suppression of aromatase expression. In fact, TGFα expression in the gastric fundic gland isthmus began around 20 days after birth in normal rats as did that of aromatase, which indicates that TGFα might induce the expression of aromatase in the parietal cells concomitantly.