The all-trans-retinoic acid (atRA) isomer, 9-cis-retinoic acid (9cRA), activates retinoic acid receptors (RARs) and retinoid X receptors (RXRs) in vitro. RARs control multiple genes, whereas RXRs serve as partners for RARs and other nuclear receptors that regulate metabolism. Physiological function has not been determined for 9cRA, because it has not been detected in serum or multiple tissues with analytically validated assays. Here, we identify 9cRA in mouse pancreas by liquid chromatography/tandem mass spectrometry (LC/MS/MS), and show that 9cRA decreases with feeding and after glucose dosing and varies inversely with serum insulin. 9cRA reduces glucose-stimulated insulin secretion (GSIS) in mouse islets and in the rat β-cell line 832/13 within 15 min by reducing glucose transporter type 2 (Glut2) and glucokinase (GK) activities. 9cRA also reduces Pdx-1 and HNF4α mRNA expression, ∼8-and 80-fold, respectively: defects in Pdx-1 or HNF4α cause maturity onset diabetes of the young (MODY4 and 1, respectively), as does a defective GK gene (MODY2). Pancreas β-cells generate 9cRA, and mouse models of reduced β-cell number, heterozygous Akita mice, and streptozotocin-treated mice have reduced 9cRA. 9cRA is abnormally high in glucose-intolerant mice, which have β-cell hypertropy, including mice with diet-induced obesity (DIO) and ob/ob and db/db mice. These data establish 9cRA as a pancreas-specific autacoid with multiple mechanisms of action and provide unique insight into GSIS.retinol | vitamin A | rexinoids I mpaired glucose-stimulated insulin secretion (GSIS) develops through multiple mechanisms, including actions of metabolic hormones and inflammatory cytokines, products of metabolic overload, and endoplasmic reticulum stress; however, mechanisms of GSIS and impaired glucose tolerance remain incompletely understood (1-4). Also uncertain is the contribution of impaired glucose tolerance to diminished pancreatic β-cell function and mass associated with type 2 diabetes (5). GSIS relies on the pancreas, and pancreas development, islet formation, and function require normal vitamin A nutriture (6-8). Vitamin A restriction during development impairs islet development and promotes glucose intolerance in adult rodents. On the other hand, restricting vitamin A in mature diabetes-prone rats reduces diabetes and insulitis, possibly through enhancing glucose sensing and metabolism. Alltrans-retinoic acid (atRA), an activated metabolite of vitamin A, regulates pancreas development, and atRA does not enhance the incidence of diabetes in diabetes-prone rats fed a vitamin Adeficient diet (7, 9, 10). Although the contribution of vitamin A to pancreas development through atRA seems clear, mechanisms whereby vitamin A affects mature pancreas function have not been determined in depth, nor have the specific vitamin A metabolites been identified that contribute to GSIS control.atRA induces differentiation and regulates cell processes by activating the nuclear receptors RAR α, -β, and -γ, which regulate transcription and translation (11...

The c-Myc (Myc) oncoprotein is a high-value therapeutic target given that it is deregulated in multiple types of cancer. However, potent small molecule inhibitors of Myc have been difficult to identify, particularly those whose mechanism relies on blocking the association between Myc and its obligate heterodimerization partner, Max. We have recently reported a structure-activity relationship study of one such small molecule, 10074-G5, and generated an analog, JY-3-094, with significantly improved ability to prevent or disrupt the association between recombinant Myc and Max proteins. However, JY-3094 penetrates cells poorly. Here, we show that esterification of a critical para-carboxylic acid function of JY-3-094 by various blocking groups significantly improves cellular uptake although it impairs the ability to disrupt Myc-Max association in vitro. These pro-drugs are highly concentrated within cells where JY-3-094 is then generated by the action of esterases. However, the pro-drugs are also variably susceptible to extracellular esterases, which can deplete extracellular reservoirs. Furthermore, while JY-3-094 is retained by cells for long periods of time, much of it is compartmentalized within the cytoplasm in a form that appears to be less available to interact with Myc. Our results suggest that persistently high extracellular levels of pro-drug, without excessive susceptibility to extracellular esterases, are critical to establishing and maintaining intracellular levels of JY-3-094 that are sufficient to provide for long-term inhibition of Myc-Max association. Analogs of JY-3-094 appear to represent promising small molecule Myc inhibitors that warrant further optimization.

Ligand activation of peroxisome proliferator-activated receptor (PPAR)␥ and retinoid X receptor (RXR) induces antitumor effects in cancer. We evaluated the ability of combined treatment with nanomolar levels of the PPAR␥ ligand rosiglitazone (BRL) and the RXR ligand 9-cis-retinoic acid (9RA) to promote antiproliferative effects in breast cancer cells. BRL and 9RA in combination strongly inhibit of cell viability in MCF-7, MCF-7TR1, SKBR-3, and T-47D breast cancer cells, whereas MCF-10 normal breast epithelial cells are unaffected. In MCF-7 cells, combined treatment with BRL and 9RA up-regulated mRNA and protein levels of both the tumor suppressor p53 and its effector p21 WAF1/Cip1 . Functional experiments indicate that the nuclear factor-B site in the p53 promoter is required for the transcriptional response to BRL plus 9RA. We observed that the intrinsic apoptotic pathway in MCF-7 cells displays an ordinated sequence of events, including disruption of mitochondrial membrane potential, release of cytochrome c, strong caspase 9 activation, and, finally, DNA fragmentation. Breast cancer is the leading cause of death among women in the world. The principal effective endocrine therapy for advanced treatment on this type of cancer is anti-estrogens, but therapeutic choices are limited for estrogen receptor (ER)␣-negative tumors, which are often aggressive. The development of cancer cells that are resistant to chemotherapeutic agents is a major clinical obstacle to the successful treatment of breast cancer, providing a strong stimulus for exploring new approaches in vitro. Using ligands of nuclear hormone receptors to inhibit tumor growth and progression is a novel strategy for cancer therapy. An example of this is the treatment of acute promyelocytic leukemia using all-trans retinoic acid, the specific ligand for retinoic acid receptors.1-3 A further paradigm for the use of retinoids in cancer therapy is for early lesions of head and neck cancer 4 and squamous cell carcinoma of the cervix. 5The retinoic acid receptor, retinoid X receptor (RXR), and peroxisome proliferator receptor (PPAR)␥, ligandactivated transcription factors belonging to the nuclear hormone receptor superfamily, are able to modulate gene networks involved in controlling growth and cellular differentiation.6 Particularly, heterodimerization of PPAR␥ with RXR by their own ligands greatly enhances DNA binding to the direct-repeated consensus sequence AGGTCA, which leads to transcriptional activation.7 Previous data show that PPAR␥, poorly expressed in normal breast epithelial cells, 8 is present at higher levels in Supported by AIRC, MURST, and Ex 60%.Portions of this work were presented as an Abstract at Società Italiana di Patologia XXIX

Cisplatin and other platinum-based drugs are well-known valid anticancer drugs. However, during chemotherapy, the presence of numerous side effects and the onset of frequent phenomena of resistance has pushed many research groups to devise new metal-based compounds holding improved anticancer properties and fewer undesired effects. Amongst the variety of synthesized compounds, significant antiproliferative effects have been obtained by employing organometallic compounds, particularly those based on silver and gold. With this in mind, we synthesized four compounds, two silver complexes and two gold complexes, with good inhibitory effects on the in vitro proliferation of breast and ovarian cancer-cell models. The antitumor activity of the most active compound, that is, AuL4, was found to be ninefold higher than that of cisplatin, and this compound induced dramatic morphological changes in HeLa cells. AuL4 induced PARP-1 cleavage, caspases 3/7 and 9 activation, mitochondria disruption, cytochrome c release in cancer-cell cytoplasm, and the intracellular production of reactive oxygen species. Thus, AuL4 treatment caused cancer-cell death by the intrinsic apoptotic pathway, whereas no cytotoxic effects were recorded upon treating non-tumor cell lines. The reported outcomes may be an important contribution to the expanding knowledge of medicinal bio-organometallic chemistry and enlarge the available anticancer toolbox, offering improved features, such as higher activity and/or selectivity, and opening the way to new discoveries and applications.

Background: The Ketogenic Diet (KD) promotes metabolic changes and optimizes energy metabolism. It is unknown if microRNAs (miRs) are influenced by KD in obese subjects. The screening of circulating miRs was performed with the FDA approved platform n-counter flex and blood biochemical parameters were dosed by ADVIA 1800. </P><P> Objectives: The aim of this study was to evaluate mir profile under 6 weeks of biphasic KD in obese subjects. We enrolled 36 obese subjects (18 females and 18 males) in stage 1 of Edmonton Obesity Staging System (EOSS) parameter. </P><P> Result: Any correlation was found between biochemical parameter and three miRs, hsa-let-7b-5p, hsa-miR-143-3p and hsa-miR-504-5p influenced in an equal manner in both sexes. The KD resulted safe and ameliorate both biochemical and anthropometric factors in obese subjects re-collocating them into stage 0 of EOSS parameters. Conclusion: The miRs herein identified under KD might be a useful tool to monitor low carbohydrate nutritional regimens which reflect indirectly the regulatory biochemical mechanisms and cell signaling that orchestrate metabolic and signaling pathways.

Cellular retinol-binding protein type I (CrbpI), encoded by Rpb1, serves as a chaperone of retinol homeostasis, but its physiological effects remain incompletely understood. We show here that the Rbp1 ؊/؊ mouse has disrupted retinoid homeostasis in multiple tissues, with abnormally high 9-cis-retinoic acid (9cRA), a pancreas autacoid that attenuates glucose-stimulated insulin secretion. The Rbp1 ؊/؊ pancreas has increased retinol and intense ectopic expression of Rpb2 mRNA, which encodes CrbpII: both would contribute to increased ␤-cell 9cRA biosynthesis. 9cRA in Rbp1 ؊/؊ pancreas resists postprandial and glucose-induced decreases. Rbp1 (27,36). The relative amounts of apo-and holo-CrbpI facilitate cellular retinol uptake, modulate retinol storage as retinyl esters (RE), and affect retinoid homeostasis by regulating enzyme activity differentially (25, 34). Retinoic acid receptor (RAR) activation depends on CrbpI-mediated retinol uptake. CrbpI mutants with low retinol-binding affinity reduce RAR function in human mammary epithelial cells, leading to a loss of differentiation (10). Nevertheless, Rbp1 Ϫ/Ϫ mice are seemingly healthy and show no gross abnormalities characteristic of overt retinoid deficiency (13). All-trans-retinoic acid (atRA) in serum and several target tissues does not differ significantly between Rbp1 Ϫ/Ϫ and wild-type (WT) mice, likely accounting for lack of gross abnormalities (20, 29). Despite these insights, physiological effects of CrbpI remain to be elucidated fully.Retinol functions primarily through its metabolite atRA, which has diverse effects on energy metabolism. atRA induces pancreas development and differentiation into acini (18,24,28,32), but restricting dietary vitamin A in diabetes-prone rats reduces diabetes and insulitis, an effect not reversed by dosing atRA, suggesting contributions of additional retinoids (9). atRA arrests differentiation of preadipocytes into mature white adipocytes, early in the differentiation process (43, 48). Ablation of Rdh1, which encodes one of several short-chain dehydrogenases that catalyze the first and rate-limiting step in atRA biosynthesis from retinol, increases adiposity in mice fed a nonobesogenic diet (50). Dosing atRA ameliorates weight gain and represses insulin resistance in mice fed an obesogenic (high-fat) diet (3). atRA induces Ucp1, which encodes an uncoupling protein expressed in brown adipose tissue that supports thermogenesis (31).Although atRA mediates most vitamin A functions, 9-cis-RA (9cRA) occurs as an endogenous pancreas retinoid with unique actions independent of atRA (22). 9cRA, biosynthesized primarily by ␤-cells and detected only in pancreas, varies rapidly and inversely with serum glucose. 9cRA attenuates glucose-stimulated insulin secretion (GSIS): a decrease in 9cRA allows optimum glucose-stimulated insulin secretion, whereas dosing with 9cRA impairs insulin secretion and glucose tolerance. 9cRA acts, in part, through reduction of Glut2 and GK activities and through repressing expression of genes that function to pro...

Considering, this evidence the variation in immune-related miRNAs should be take into account and could be relevant for preterm and hospitalized infants who usually received pasteurized HM from donors.

Varroa destructor is the most important ectoparasitic mite of honey bees that has a negative impact on bee health and honey production. The control programs are mainly based on the use of synthetic acaricides that are often administered indiscriminately. All this has led to drug resistance that now represent a great concern for honey bee farming. The research for alternative products/methods for mites’ control is now mandatory. The aim of this study was to test whether Citrus spp. essential oils could diminish the growth of the V. destructor mite. In Calabria (southern Italy), plants of the Citrus genus are very common and grow both spontaneously and cultured. The essential oils used in this study were extracted from bergamot (Citrus bergamia), grapefruit (Citrus paradisi), lemon (Citrus limon), orange (Citrus sinensis), and mandarin (Citrus reticulata) by hydrodistillation. Every EO was in vitro tested against V. destructor. Each experimental replicate was performed using 35 viable adult female mites (5 for each EO) collected the same day from the same apiary and included negative controls (5 individuals exposed to acetone only) and positive controls (5 individuals exposed to Amitraz diluted in acetone). The essential oils (Eos) were diluted (0.5 mg/mL, 1 mg/mL, and 2 mg/mL) in HPLC grade acetone to obtain the working solution to be tested (50 µL/tube). Mite mortality was manually assessed after 1 h exposure under controlled conditions. The essential oils that showed the best effectiveness at 0.5 mg/mL were bergamot, which neutralized (dead + inactivated) 80% (p ≤ 0.001) of the parasites; grapefruit, which neutralized 70% (p ≤ 0.001); and lemon, which neutralized 69% of them. Interestingly, the positive control (Amitraz) at the same concentration neutralized 60% of the parasites. These results demonstrate that Calabrian bergamot, grapefruit, and lemon Eos consistently reduced V. destructor viability and open the possibility for their utilization to control this parasite in honey bee farming.