Kaempferol as a flavonoid induces osteoblastic differentiation via estrogen receptor signaling

<p>Abstract</p> <p>Background</p> <p>Flavonoids, a group of compounds mainly derived from vegetables and herbal medicines, chemically resemble estrogen and some have been used as estrogen substitutes. Kaempferol, a flavonol derived from the rhizome of <it>Kaempferia galanga </it>L., is a well-known phytoestrogen possessing osteogenic effects that is also found in a large number of plant foods.</p> <p>The herb <it>K. galanga </it>is a popular traditional aromatic medicinal plant that is widely used as food spice and in medicinal industries. In the present study, both the estrogenic and osteogenic properties of kaempferol are evaluated.</p> <p>Methods</p> <p>Kaempferol was first evaluated for its estrogenic properties, including its effects on estrogen receptors. The osteogenic properties of kaempferol were further determined its induction effects on specific osteogenic enzymes and genes as well as the mineralization process in cultured rat osteoblasts.</p> <p>Results</p> <p>Kaempferol activated the transcriptional activity of pERE-Luc (3.98 ± 0.31 folds at 50 μM) and induced estrogen receptor α (ERα) phosphorylation in cultured rat osteoblasts, and this ER activation was correlated with induction and associated with osteoblast differentiation biomarkers, including alkaline phosphatase activity and transcription of osteoblastic genes, <it>e.g</it>., type I collagen, osteonectin, osteocalcin, Runx2 and osterix. Kaempferol also promoted the mineralization process of osteoblasts (4.02 ± 0.41 folds at 50 μM). ER mediation of the kaempferol-induced effects was confirmed by pretreatment of the osteoblasts with an ER antagonist, ICI 182,780, which fully blocked the induction effect.</p> <p>Conclusion</p> <p>Our results showed that kaempferol stimulates osteogenic differentiation of cultured osteoblasts by acting through the estrogen receptor signaling.</p

A study of the membrane interactions of K-ras

Ras proteins are monomeric GTPases which operate in cellular signal transduction pathways to regulate cell growth, differentiation and apoptosis. Mutations at amino acid residue 12, 13 or 61 render the protein constitutively active and oncogenic. These mutations are found in 30% of human cancers. There are three isoforms of the Ras protein: H-ras, N-ras and K-ras. All three isoforms localize to the inner leaflet of the plasma membrane, and this localization is important for their biological function. H-ras and N-ras traffic to the plasma membrane via the classical exocytic pathway, while the K-ras trafficking pathway is non-vesicular and is currently poorly characterized. In order to identify the proteins that may be involved in the K-ras trafficking pathway, an affinity chromatography approach was developed. The method utilized the C-terminal membrane-targeting regions of K-ras (CTK) and H-ras (CTH) as baits to purify cellular proteins from mouse brain cytosol. Proteins which selectively interacted with CTK but not CTH were identified by mass spectrometry. Four of these proteins, B23, Ncl, LANCL1 and laminin receptor-1, were further characterized and evaluated as putative K-ras transport factors. B23 localizes to P100 and nuclear fractions, and selectively interacts with K-ras in vivo. However, overexpression or siRNA-mediated knockdown of B23 does not affect plasma membrane localization of K-ras. Ncl localizes to the P100, S100 and nuclear fractions. Nuclear Ncl, but not extranuclear Ncl, selectively interacts with K-ras in vivo. Overexpression of Ncl does not affect the intracellular localization of either H-ras or K-ras. LANCL1 is associated with the P100, S100 and nuclear fractions, and preliminary results show that LANCL1 may selectively interact with K-ras in vivo. Selective interaction between laminin receptor-1 and K-ras could not be confirmed, either in vitro or in vivo. Also, overexpression of laminin receptor-1 has no effect on the P100/S100 distribution of K-ras, or its downstream MEK/ERK signalling. In conclusion, this thesis represents the first known proteomic approach dedicated to the discovery of K-ras transport factors. Three novel selective interacting partners of the membrane targeting domain of K-ras are identified, although their precise function in K-ras trafficking remains to be elucidated

Giant axonal neuropathy diagnosed on skin biopsy

Evaluation of hereditary axonal neuropathy in childhood is complex. Often, the child has to be subjected to general anaesthesia for a nerve biopsy to guide further genetic testing, which may or may not be readily available. We describe a toddler with clinical features suggesting giant axonal neuropathy (GAN), whose diagnosis was confirmed by minimally invasive skin biopsy and corroborated by the finding of compound heterozygous mutations involving the GAN gene, including a novel interstitial microdeletion at 16q23.2 detected by microarray and a point mutation detected by direct sequencing

A Multi-Disciplinary Team Approach to Genomic Testing for Drug-Resistant Epilepsy Patients-The GENIE Study

Background. The genomic era has led to enormous progress in clinical care and a multi-disciplinary team (MDT) approach is imperative for integration of genomics into epilepsy patient care. Methods. The MDT approach involved patient selection, genomic testing choice, variant discussions and return of results. Genomics analysis included cytogenomic testing and whole exome sequencing (WES). Neurologist surveys were undertaken at baseline and after genomic testing to determine if genomic diagnoses would alter their management, and if there was a change in confidence in genomic testing and neurologist perceptions of the MDT approach. Results. The total diagnostic yield from all genomic testing was 17% (11/66), with four diagnoses from cytogenomic analyses. All chromosomal microarray (CMA) diagnoses were in patients seen by adult neurologists. Diagnostic yield for WES was 11% (7/62). The most common gene with pathogenic variants was DCX, reported in three patients, of which two were mosaic. The genomic diagnosis impacted management in 82% (9/11). There was increased confidence with integrating genomics into clinical care (Pearson chi square = 83, p = 0.004) and qualitative comments were highly supportive of the MDT approach. Conclusions. We demonstrated diagnostic yield from genomic testing, and the impact on management in a cohort with drug-resistant epilepsy. The MDT approach increased confidence in genomic testing and neurologists valued the input from this approach. The utility of CMA was demonstrated in epilepsy patients seen by adult neurologists as was the importance of considering mosaicism for previously undiagnosed patients.</p

Chemical and Biological Assessment of Ziziphus jujuba Fruits from China: Different Geographical Sources and Developmental Stages

Chinese date, the fruit of Ziziphus jujuba Mill., has thousands of years cultivation history, and about 700 cultivars of dates in China. Two types of dates are commonly found in the market: (i) fresh immature dates consumed as fruits, and (ii) dried mature dates used as Chinese medicines. Here, chemical and biological properties of these dates were revealed. Different sources of dates showed similar chemical profiles; however, the amounts of identified chemicals showed a great variation. The amount of nucleotides, flavonoids and polysaccharides in dates could be affected by its maturity and drying process. In parallel, the antioxidative functions of their extracts were compared. The date extracts protected PC12 cells against tBHP-induced cytotoxicity, and which also stimulated the transcriptional activity of antioxidant response element. The antioxidative effects were varied among different dates. The current results suggested the optimization of sources and specific usage of different maturity dates

Nucleophosmin and Nucleolin Regulate K-Ras Plasma Membrane Interactions and MAPK Signal Transduction*

The spatial organization of Ras proteins into nanoclusters on the inner leaflet of the plasma membrane is essential for high fidelity signaling through the MAPK pathway. Here we identify two selective regulators of K-Ras nanoclustering from a proteomic screen for K-Ras interacting proteins. Nucleophosmin (NPM) and nucleolin are predominantly localized to the nucleolus but also have extranuclear functions. We show that a subset of NPM and nucleolin localizes to the inner leaflet of plasma membrane and forms specific complexes with K-Ras but not other Ras isoforms. Active GTP-loaded and inactive GDP-loaded K-Ras both interact with NPM, although NPM-K-Ras binding is increased by growth factor receptor activation. NPM and nucleolin both stabilize K-Ras levels on the plasma membrane, but NPM concurrently increases the clustered fraction of GTP-K-Ras. The increase in nanoclustered GTP-K-Ras in turn enhances signal gain in the MAPK pathway. In summary these results reveal novel extranucleolar functions for NPM and nucleolin as regulators of K-Ras nanocluster formation and activation of the MAPK pathway. The study also identifies a new class of K-Ras nanocluster regulator that operates independently of the structural scaffold galectin-3

The clinical utility and costs of whole-genome sequencing to detect cancer susceptibility variants—a multi-site prospective cohort study

Abstract Background Many families and individuals do not meet criteria for a known hereditary cancer syndrome but display unusual clusters of cancers. These families may carry pathogenic variants in cancer predisposition genes and be at higher risk for developing cancer. Methods This multi-centre prospective study recruited 195 cancer-affected participants suspected to have a hereditary cancer syndrome for whom previous clinical targeted genetic testing was either not informative or not available. To identify pathogenic disease-causing variants explaining participant presentation, germline whole-genome sequencing (WGS) and a comprehensive cancer virtual gene panel analysis were undertaken. Results Pathogenic variants consistent with the presenting cancer(s) were identified in 5.1% (10/195) of participants and pathogenic variants considered secondary findings with potential risk management implications were identified in another 9.7% (19/195) of participants. Health economic analysis estimated the marginal cost per case with an actionable variant was significantly lower for upfront WGS with virtual panel ($8744AUD) compared to standard testing followed by WGS ($24,894AUD). Financial analysis suggests that national adoption of diagnostic WGS testing would require a ninefold increase in government annual expenditure compared to conventional testing. Conclusions These findings make a case for replacing conventional testing with WGS to deliver clinically important benefits for cancer patients and families. The uptake of such an approach will depend on the perspectives of different payers on affordability