Myostatin: A Skeletal Muscle Chalone.

Myostatin (GDF-8) was discovered 25 years ago as a new transforming growth factor-ß family member that acts as a master regulator of skeletal muscle mass. Myostatin is made by skeletal myofibers, circulates in the blood, and acts back on myofibers to limit growth. Myostatin appears to have all of the salient properties of a chalone, which is a term proposed over a half century ago to describe hypothetical circulating, tissue-specific growth inhibitors that control tissue size. The elucidation of the molecular, cellular, and physiological mechanisms underlying myostatin activity suggests that myostatin functions as a negative feedback regulator of muscle mass and raises the question as to whether this type of chalone mechanism is unique to skeletal muscle or whether it also operates in other tissues.

Novel donepezil-chalcone-rivastigmine hybrids as potential multifunctional anti-Alzheimer's agents: Design, synthesis, in vitro biological evaluation, in vivo and in silico studies.

Alzheimer's disease (AD) is a chronic, progressive brain neurodegenerative disorder. Up to now, there is no effective drug to halt or reverse the progress of AD. Given the complex pathogenesis of AD, the multi-target-directed ligands (MTDLs) strategy is considered as the promising therapy. Herein, a series of novel donepezil-chalone-rivastigmine hybrids was rationally designed and synthesized by fusing donepezil, chalone and rivastigmine. The in vitro bioactivity results displayed that compound 10c was a reversible huAChE (IC50 = 0.87 µM) and huBuChE (IC50 = 3.3 µM) inhibitor. It also presented significant anti-inflammation effects by suppressing the level of IL-6 and TNF-α production, and significantly inhibited self-mediated Aß1-42 aggregation (60.6%) and huAChE-mediated induced Aß1-40 aggregation (46.2%). In addition, 10c showed significant neuroprotective effect on Aß1-42-induced PC12 cell injury and activated UPS pathway in HT22 cells to degrade tau and amyloid precursor protein (APP). Furthermore, compound 10c presented good stabilty in artificial gastrointestinal fluids and liver microsomes in vitro. The pharmacokinetic study showed that compound 10c was rapidly absorbed in rats and distributed in rat brain after intragastric administration. The PET-CT imaging demonstrated that [11C]10c could quickly enter the brain and washed out gradually in vivo. Further, compound 10c at a dose of 5 mg/kg improved scopolamine-induced memory impairment, deserving further investigations.

Discovery of new chalone adamantyl arotinoids having RXRα-modulating and anticancer activities.

In the present study, a new series of chalcone adamantly arotinoids (chalcone AdArs) derived from RAR antagonist MX781, are synthesized, characterized, and evaluated for the biological activities in vitro. The studies of antiproliferative activity and RXRα-binding affinity of target compounds result in the discovery of a lead candidate (WA15), which is a good RXRα binder (Kd = 2.89 × 10-6 M) with potent antiproliferative activity against human cancer cell lines (IC50 ≈ 10 µM) and low toxic to normal LO2 and MRC-5 cells (IC50 > 50 µM). Different from MX781, WA15 eliminates RARα antagonist activity but inhibits 9-cis-RA-induced RXRα transactivation activity in a dose-dependent manner. Compound WA15 is found to be a good apoptosis inducer in various cancer cells and promotes cell apoptosis in an RXRα-independent manner. Besides, WA15 shows the induction of proteasome-dependent RXRα degradation which might enhance the WA15-induced apoptosis. Finally, the immunoblotting indicates that WA15 can inhibit the TNFα-induced IKK activation and IκBα degradation, suggesting that the anticancer activity of WA15 might be related to the inhibition of IKK/NF-κB signal pathway.

Phenols displaying tyrosinase inhibition from Humulus lupulus.

Tyrosinase is the rate-limiting enzyme for the production of melanin and other pigments via the oxidation of l-tyrosine. The methanol extract from Humulus lupulus showed potent inhibition against mushroom tyrosinase. The bioactivity-guided fractionation of this methanol extract resulted in the isolation of seven flavonoids (1-7), identified as xanthohumol (1), 4'-O-methylxanthohumol (2), xanthohumol C (3), flavokawain C (4), xanthoumol B (5), 6-prenylnaringenin (6) and isoxanthohumol (7). All isolated flavonoids (1-7) effectively inhibited the monophenolase (IC50s = 15.4-58.4 µM) and diphenolase (IC50s = 27.1-117.4 µM) activities of tyrosinase. Kinetic studies using Lineweaver-Burk and Dixon-plots revealed that chalcones (1-5) were competitive inhibitors, whereas flavanones (6 and 7) exhibited both mixed and non-competitive inhibitory characteristics. In conclusion, this study is the first to demonstrate that the phenolic phytochemicals of H. lupulus display potent inhibitory activities against tyrosinase.

A retinoblastoma orthologue is required for the sensing of a chalone in Dictyostelium discoideum.

Retinoblastoma-like proteins regulate cell differentiation and inhibit cell proliferation. The Dictyostelium discoideum retinoblastoma orthologue RblA affects the differentiation of cells during multicellular development, but it is unclear whether RblA has a significant effect on Dictyostelium cell proliferation, which is inhibited by the secreted proteins AprA and CfaD. We found that rblA⁻ cells in shaking culture proliferate to a higher density, die faster after reaching stationary density, and, after starvation, have a lower spore viability than wild-type cells, possibly because in shaking culture, rblA⁻ cells have both increased cytokinesis and lower extracellular accumulation of CfaD. However, rblA⁻ cells have abnormally slow proliferation on bacterial lawns. Recombinant AprA inhibits the proliferation of wild-type cells but not that of rblA⁻ cells, whereas CfaD inhibits the proliferation of both wild-type cells and rblA⁻ cells. Similar to aprA⁻ cells, rblA⁻ cells have a normal mass and protein accumulation rate on a per-nucleus basis, indicating that RblA affects cell proliferation but not cell growth. AprA also functions as a chemorepellent, and RblA is required for proper AprA chemorepellent activity despite the fact that RblA does not affect cell speed. Together, our data indicate that an autocrine proliferation-inhibiting factor acts through RblA to regulate cell density in Dictyostelium, suggesting that such factors may signal through retinoblastoma-like proteins to control the sizes of structures such as developing organs or tumors.

Cell lineages and the logic of proliferative control.

It is widely accepted that the growth and regeneration of tissues and organs is tightly controlled. Although experimental studies are beginning to reveal molecular mechanisms underlying such control, there is still very little known about the control strategies themselves. Here, we consider how secreted negative feedback factors ("chalones") may be used to control the output of multistage cell lineages, as exemplified by the actions of GDF11 and activin in a self-renewing neural tissue, the mammalian olfactory epithelium (OE). We begin by specifying performance objectives-what, precisely, is being controlled, and to what degree-and go on to calculate how well different types of feedback configurations, feedback sensitivities, and tissue architectures achieve control. Ultimately, we show that many features of the OE-the number of feedback loops, the cellular processes targeted by feedback, even the location of progenitor cells within the tissue-fit with expectations for the best possible control. In so doing, we also show that certain distinctions that are commonly drawn among cells and molecules-such as whether a cell is a stem cell or transit-amplifying cell, or whether a molecule is a growth inhibitor or stimulator-may be the consequences of control, and not a reflection of intrinsic differences in cellular or molecular character.

eIF2α kinases control chalone production in Dictyostelium discoideum.

Growing Dictyostelium cells secrete CfaD and AprA, two proteins that have been characterized as chalones. They exist within a high-molecular-weight complex that reversibly inhibits cell proliferation, but not growth, via cell surface receptors and a signaling pathway that includes G proteins. How the production of these two proteins is regulated is unknown. Dictyostelium cells possess three GCN2-type eukaryotic initiation factor 2 α subunit (eIF2α) kinases, proteins that phosphorylate the translational initiation factor eIF2α and possess a tRNA binding domain involved in their regulation. The Dictyostelium kinases have been shown to function during development in regulating several processes. We show here that expression of an unregulated, activated kinase domain greatly inhibits cell proliferation. The inhibitory effect on proliferation is not due to a general inhibition of translation. Instead, it is due to enhanced production of a secreted factor(s). Indeed, extracellular CfaD and AprA proteins, but not their mRNAs, are overproduced in cells expressing the activated kinase domain. The inhibition of proliferation is not seen when the activated kinase domain is expressed in cells lacking CfaD or AprA or in cells that contain a nonphosphorylatable eIF2α. We conclude that production of the chalones CfaD and AprA is translationally regulated by eIF2α phosphorylation. Both proteins are upregulated at the culmination of development, and this enhanced production is lacking in a strain that possesses a nonphosphorylatable eIF2α.

A Dictyostelium chalone uses G proteins to regulate proliferation.

BACKGROUND: Several studies have shown that organ size, and the proliferation of tumor metastases, may be regulated by negative feedback loops in which autocrine secreted factors called chalones inhibit proliferation. However, very little is known about chalones, and how cells sense them. We previously identified two secreted proteins, AprA and CfaD, which act as chalones in Dictyostelium. Cells lacking AprA or CfaD proliferate faster than wild-type cells, and adding recombinant AprA or CfaD to cells slows their proliferation. RESULTS: We show here that cells lacking the G protein components Galpha8, Galpha9, and Gbeta proliferate faster than wild-type cells despite secreting normal or high levels of AprA and CfaD. Compared with wild-type cells, the proliferation of galpha8-, galpha9- and gbeta- cells are only weakly inhibited by recombinant AprA (rAprA). Like AprA and CfaD, Galpha8 and Gbeta inhibit cell proliferation but not cell growth (the rate of increase in mass and protein per nucleus), whereas Galpha9 inhibits both proliferation and growth. galpha8- cells show normal cell-surface binding of rAprA, whereas galpha9- and gbeta- cells have fewer cell-surface rAprA binding sites, suggesting that Galpha9 and Gbeta regulate the synthesis or processing of the AprA receptor. Like other ligands that activate G proteins, rAprA induces the binding of [3H]GTP to membranes, and GTPgammaS inhibits the binding of rAprA to membranes. Both AprA-induced [3H]GTP binding and the GTPgammaS inhibition of rAprA binding require Galpha8 and Gbeta but not Galpha9. Like aprA- cells, galpha8- cells have reduced spore viability. CONCLUSION: This study shows that Galpha8 and Gbeta are part of the signal transduction pathway used by AprA to inhibit proliferation but not growth in Dictyostelium, whereas Galpha9 is part of a differealnt pathway that regulates both proliferation and growth, and that a chalone signal transduction pathway uses G proteins.

Anti-African trypanocidal and antimalarial activity of natural flavonoids, dibenzoylmethanes and synthetic analogues.

OBJECTIVES: The known anti-protozoal activity of flavonoids has stimulated the testing of other derivatives from natural and synthetic sources. METHODS: As part of our efforts to find potential lead compounds, a number of flavonoids isolated from Neoraputia paraensis, N. magnifica, Murraya paniculata, (Rutaceae), Lonchocarpus montanus, L. latifolius, L. subglaucescens, L. atropurpureus, L. campestris, Deguelia hatschbachii (Leguminosae), dibenzoylmethanes from L. subglaucescens and synthetic analogues were tested for in-vitro activity against chloroquine-sensitive Plasmodium falciparum and Trypanosoma brucei rhodesiense bloodstream form trypomastigotes. An assay with KB cells has been developed in order to compare in-vitro cytotoxicity of flavonoids with a selective action on the parasites. KEY FINDINGS: Thirteen of the compounds tested had IC50 values ranging from 4.6 to 9.9 microm against T. brucei rhodesiense. In contrast, a small number of compounds showed significant activity against P. falciparum; seven of those tested had IC50 values ranging from 2.7 to 9.5 microm. Among the flavones only one had IC50 < 10 microm (7.6 microm), whereas against T. brucei rhodesiense seven had IC50 < 10 microm. Synthetic dibenzoylmethanes were the most active in terms of number (five) of compounds and the IC50 values (2.7-9.5 microm) against P. falciparum. CONCLUSIONS: Dibenzoylmethanes represent a novel class of compounds tested for the first time as antimalarial and trypanocidal agents.

A comparative study of tumor-suppression effects of enterotoxin B and Chalone 19-peptide on experimental gastric carcinoma.

BACKGROUND/AIMS: In this paper, the in vivo tumor suppression effects of Staphyococal enterotoxin B (SEB) and Chalone 19-peptide, a form of Tumstain with modified coding sequence, on poorly differentiated human gastric carcinoma cells were compared. METHODOLOGY: Animal model for studying human gastric carcinoma was established by injecting tumor cells (Human poorly differentiated gastric carcinoma cell line, SGC-7901) underneath the gastric serosa of BALB/c nude mice. RESULTS: Results demonstrated that SEB induced tumor cell death on a large scale and destroyed surrounding normal tissue at the same time, leading to tumor cluster breaking down and seeding. SEB had no effect on lymphovascular metastasis. The administration of 19-peptide on gastric carcinoma resulted in sheets-like tumor central necrosis, decreased angiogenesis and a moderate tumor infiltration into surrounding tissue without distant metastasis. Therefore, both SEB and 19-peptide could suppress the local growth, distant metastasis and invasion of poorly differentiated gastric carcinoma cells into surrounding tissues. CONCLUSIONS: Data suggested that this model could effectively simulate the microenvironment of human gastric carcinoma, hence providing a platform for study on this cancer.

The terms 'autacoid', 'hormone' and 'chalone' and how they have shifted with time.

The increase of knowledge in a particular field (endocrinology) can be understood if one follows how certain key concepts were constructed and transformed over time. To explore such construction and transformation (shifts in meaning), we studied the use of the concepts 'autacoid' and 'chalone' in a period of one century (1916-2016), since the introduction of these concepts by the British professor of physiology Sir Sharpey-Schäfer. We could identify that the use of 'autacoid' shifted from a very broad category encompassing both stimulating and inhibiting hormones, in the period 1916-1960, to a much more specific use of the term for locally produced bioactive molecules, from the 1960s onwards. Histamine was the first compound seen as an 'autacoid', followed by prostaglandins, ATP, ADP and bradykinin, and from 1993 onwards, compounds such as 'palmitoylethanolamide' were also classified as 'autacoids'. For 'chalone', a comparable shift was noticed around the 1960s, when the concept suddenly changed from the category of inhibiting hormones into a substance that is produced within a tissue, inhibiting mitosis of the cells of that tissue. For both concept shifts, we could not find any argument. Around 1980, authors started to relate autacoids to various promising indications in the field of inflammation and immune modulation. The Nobel laureate Rita Levi-Montalcini gave an extra dimension to the concept autacoid in 1993, and introduced a new class of compounds modulating mast cells, the ALIAmides (from Autacoid Local Inflammation Antagonist), of which palmitoylethanolamide was the prototype. Our exploration demonstrates that biomedical concepts can be constructed and defined differently as time goes by, while concept transformations seem to emerge without arguments.

The epidermal G1-chalone: an endogenous tissue-specific inhibitor of epidermal cell proliferation.

An apparently macromolecular factor is isolated from aqueous skin extracts which inhibits DNA synthesis in vivo and in vitro with high efficacy (ID50 in vivo 0.2 pmol/g, in vitro 0.2 pM) and in a highly specific manner showing a point of attack in the late G1-phase of the cell cycle (epidermal G1-chalone). Preliminary characterization indicates an unusual highly amphipathic structure consisting of amino acids and carbohydrate. Despite its apparent molecular weight of approximately 10 kD the chalone is stable against denaturing agents and most enzymes, including proteases. An inverse correlation between chalone responsiveness of mouse epidermis in vivo and the development of hyperplasia due to injury indicates an important role of the factor in the regulation of tissue homeostasis. According to its physicochemical and biological properties the epidermal G1-chalone appears not to be related to other endogenous inhibitors of epidermal cell proliferation such as the pentapeptide pyroGlu-Glu-Asp-Ser-GlyOH and transforming growth factor beta (TGF beta).

[Detection of squamous cell differentiation in experimental tumors of urothelium by using epidermal G2 chalone]. / Vyiavlenie ploskokletochnoi differentsirovki v éksperimental'nykh opukholiakh uroteliia s pomoshch'iu épidermal'nogo G2-keilona.

The presence of epidermal G2 chalone was investigated by different immunochemical methods in normal urothelium and in tumors of the urinary bladder induced in rats with N-butyl-N-butanol-(4)-nitrosamine. This chalone was not revealed in normal urothelium but found in 37 out of 56 tumors examined. In all these 37 tumors, squamous urothelial metaplasia was detected by electron or even light microscopy. The extent of morphological manifestations of squamous metaplasia showed a definite correlation with the concentration of chalone in tissue extracts. The method used may be applied as a subsidiary test for the determination of focal squamous metaplasia in biopsy material of human urothelial tumors which is of importance for prognosis of the disease.

[Several approaches to purifying rat liver G2-chalone]. / Nekotorye podkhody k ochistke G2-khalona pecheni krys.

An attempt was made to isolate the liver tissue chalone blocking the transition of hepatocytes from G2 period of the cell cycle into mitosis/g2-chalone/. The purification was carried out using gel filtration through Sephadex G-75 and hydroxyapatite chromatography as well as by means of ultrafiltration through Diaflo membranes XM-300, XM-100 and PM-10. The biologically active preparation, obtained by these two procedures, was purified 100-fold. Molecular mass of G2 chalone was apparently less then 10,000; it penetrated through PM-10 membrane. The chalone appears to form complexes with high molecular carriers.

Novel prenylated bichalcone and chalcone from Humulus lupulus and their quinone reductase induction activities.

A new prenylated chalcone xanthohumol M (1), a novel prenylated bichalcone humulusol (2) and six known chalcones (3-8) were found from Humulus lupulus. Their structures were determined by spectroscopic methods. All the chalcones' electrophilic abilities were assessed by GSH (glutathione) rapid screening, and their QR (quinone reductase) induction activities were evaluated using hepa 1c1c7 cells. The results of electrophilic assay and QR induction activity assay were quite well. New compounds 1 and 2, along with some known prenylated chalcones, displayed certain QR induction activity.

Synthesis and biological evaluation of novel pyranochalcone derivatives as a new class of microtubule stabilizing agents.

Twenty-five novel pyranochalcone derivatives were synthesized and evaluated for their in vitro and in vivo antiproliferative activities. Among them, compound 10i exhibited superior potent activity against 21 tumor cell lines including multidrug resistant phenotype with the IC50 values ranged from 0.09 to 1.30 µM. In addition, 10i significantly induced cell cycle arrest in G2/M phase, promoted tubulin polymerization into microtubules and caused microtubule stabilization. Further studies confirmed that 10i significantly suppressed the growth of tumor volume in HepG2 xenograft tumor model. Our study demonstrated that 10i could have beneficial antitumor activity as a novel microtubule stabilizing agent.

Design, synthesis, and structure-activity relationship studies of novel millepachine derivatives as potent antiproliferative agents.

In this paper, 38 millepachine derivatives have been designed, synthesized and evaluated for their in vitro and in vivo antiproliferative activity. Among these novel derivatives, 15 displayed more potent antiproliferative activity than millepachine against HepG2, K562, SK-OV-3, HCT116, HT29, and SW620 tumor cells (mean IC(50) = 0.64 vs. 2.86 µM, respectively). Furthermore, 15 could effectively inhibit tubulin polymerization in HepG2 cells, and induce the HepG2 cell cycle arrest at the G2/M phase in a concentration-dependant manner. Further studies confirmed that 15 significantly suppressed the growth of tumor volume and exerted more potent anticancer potency than millepachine and anticancer drug cisplatin in A549 lung xenograft tumor model.