Genistein-mediated inhibition of glycosaminoglycan synthesis, which corrects storage in cells of patients suffering from mucopolysaccharidoses, acts by influencing an epidermal growth factor-dependent pathway

<p>Abstract</p> <p>Background</p> <p>Mucopolysaccharidoses (MPS) are inherited metabolic disorders caused by mutations leading to dysfunction of one of enzymes involved in degradation of glycosaminoglycans (GAGs). Due to their impaired degradation, GAGs accumulate in cells of patients, which results in dysfunction of tissues and organs. Substrate reduction therapy is one of potential treatment of these diseases. It was demonstrated previously that genistein (4', 5, 7-trihydroxyisoflavone) inhibits synthesis and reduces levels of GAGs in cultures of fibroblasts of MPS patients. Recent pilot clinical study indicated that such a therapy may be effective in MPS III (Sanfilippo syndrome).</p> <p>Methods</p> <p>To learn on details of the molecular mechanism of genistein-mediated inhibition of GAG synthesis, efficiency of this process was studied by measuring of incorporation of labeled sulfate, storage of GAGs in lysosomes was estimated by using electron microscopic techniques, and efficiency of phosphorylation of epidermal growth factor (EGF) receptor was determined by using an ELISA-based assay with fluorogenic substrates.</p> <p>Results</p> <p>Effects of genistein on inhibition of GAG synthesis and accumulation in fibroblasts from patients suffering from various MPS types were abolished in the presence of an excess of EGF, and were partially reversed by an increased concentration of genistein. No such effects were observed when an excess of 17β-estradiol was used instead of EGF. Moreover, EGF-mediated stimulation of phsophorylation of the EGF receptor was impaired in the presence of genistein in both wild-type and MPS fibroblasts.</p> <p>Conclusion</p> <p>The results presented in this report indicate that the mechanism of genistein-mediated inhibition of GAG synthesis operates through epidermal growth factor (EGF)-dependent pathway.</p

Heat Treatment Effect on Eu 3+

Glass systems of 73TeO2-4BaO-3Bi2O3-2Eu2O3-xAg (in molar ratio where x = 0, 1, 2, and 3) compositions have been successfully synthesized. Silver nanoparticles were obtained with the employment of heat treatment (HT) procedure executed at 350°C. Glass transition temperatures of different compositions have been determined through DSC measurements. XRD results presented characteristic amorphous halo indicating lack of long range order in the samples. FTIR structural studies revealed that glass matrix is mainly composed of TeO3 and TeO4 species and is stable after different applied heat treatment times. X-ray photoelectron spectroscopy (XPS) measurements confirmed that in selected samples part of Ag ions changed oxidation state to form Ag0 species. TEM measurements revealed nanoparticles of size in the range of 20–40 nm. UV-vis absorption results demonstrated characteristic transitions of Eu3+ ions. Additionally, UV-vis spectra of samples heat-treated for 6, 12, 24, and 48 hours presented bands related to silver nanoparticles. Photoluminescence (PL) studies have been performed with excitation wavelength of λexc=395 nm. Obtained spectra exhibited peaks due to 5D0-7FJ (where J=2,3,4) and 5D1-7FJ (where J=1,2,3) transitions of Eu3+. Moreover, luminescence measurement indicated enhancement of rare earth ions emissions in several of the annealed samples. Increase of emission intensity of about 35% has been observed

Synthetic genistein derivatives as modulators of glycosaminoglycan synthesis

Background: Mucopolysaccharidoses (MPS) are severe metabolic disorders caused by 26 accumulation of undegraded glycosaminoglycans (GAGs) in lysosomes due to defects in certain 27 lysosomal hydrolases. Substrate reduction therapy (SRT) has been proposed as one of potential 28 treatment procedures of MPS. Importantly, small molecules used in such a therapy might 29 potentially cross the blood-brain barrier (BBB) and improve neurological status of patients, as 30 reported for a natural isoflavone, 5, 7-dihydroxy-3- (4-hydroxyphenyl)-4H-1-benzopyran-4-one, 31 also known as genistein. Although genistein is able to cross BBB to some extent, its delivery to 32 the central nervous system is still relatively poor (below 10% efficiency). Thus, we aimed to 33 develop a set of synthetically modified genistein molecules and characterize physicochemical as 34 well as biological properties of these compounds. Methods: Following parameters were 35 determined for the tested synthetic derivatives of genistein: cytotoxicity, effects on cell 36 proliferation, kinetics of GAG synthesis, effects on epidermal growth factor (EGF) receptor’s 37 tyrosine kinase activity, effects on lysosomal storage, potential ability to cross BBB. Results: We 38 observed that some synthetic derivatives inhibited GAG synthesis similarly to, or more 39 efficiently than, genistein and were able to reduce lysosomal storage in MPS III fibroblasts. The 40 tested compounds were generally of low cytotoxicity and had minor effects on cell proliferation. 41 Moreover, synthetic derivatives of genistein revealed higher lipophilicity (assessed in silico) than 42 the natural isoflavone. Conclusion: Some compounds tested in this study might be promising 43 candidates for further studies on therapeutic agents in MPS types with neurological symptoms

Coupling of transcription and replication machineries in λ DNA replication initiation: evidence for direct interaction of Escherichia coli RNA polymerase and the λO protein

Transcription proceeding downstream of the λ phage replication origin was previously shown to support initial steps of the λ primosome assembly in vitro and to regulate frequency and directionality of λ DNA replication in vivo. In this report, the data are presented indicating that the RNA polymerase β subunit makes a direct contact with the λO protein, a replication initiator of λ phage. These results suggest that the role of RNA polymerase during the initiation of λ phage DNA replication may be more complex than solely influencing DNA topology. Results demonstrated in this study also show that gyrase supercoiling activity stimulates the formation of a complex between λO and RNA polymerase, suggesting that the introduction of negative supercoils by DNA gyrase, besides lowering the energy required for DNA strand separation, may play an additional role in modeling protein–protein interactions at early steps of DNA replication initiation

Biodiversity of bacteriophages: morphological and biological properties of a large group of phages isolated from urban sewage

A large scale analysis presented in this article focuses on biological and physiological variety of bacteriophages. A collection of 83 bacteriophages, isolated from urban sewage and able to propagate in cells of different bacterial hosts, has been obtained (60 infecting Escherichia coli, 10 infecting Pseudomonas aeruginosa, 4 infecting Salmonella enterica, 3 infecting Staphylococcus sciuri, and 6 infecting Enterococcus faecalis). High biological diversity of the collection is indicated by its characteristics, both morphological (electron microscopic analyses) and biological (host range, plaque size and morphology, growth at various temperatures, thermal inactivation, sensitivity to low and high pH, sensitivity to osmotic stress, survivability upon treatment with organic solvents and detergents), and further supported by hierarchical cluster analysis. By the end of the research no larger collection of phages from a single environmental source investigated by these means had been found. The finding was confirmed by whole genome analysis of 7 selected bacteriophages. Moreover, particular bacteriophages revealed unusual biological features, like the ability to form plaques at low temperature (4 °C), resist high temperature (62 °C or 95 °C) or survive in the presence of an organic solvents (ethanol, acetone, DMSO, chloroform) or detergent (SDS, CTAB, sarkosyl) making them potentially interesting in the context of biotechnological applications

Experimental Plant Biology: Why Not?! Genistein: a natural isoflavone with a potential for treatment of genetic diseases

Abstract Genistein [4 ,5,7-trihydroxyisoflavone or 5,7-dihydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-one] is a natural isoflavone occurring in many plants known to possess various biological activities, ranging from phyto-oestrogenic to antioxidative actions. Recent studies indicated that this isoflavone can also be considered as a drug for as yet untreatable genetic diseases. In the present review, we discuss a plausible use of genistein in treatment of two genetic disorders: CF (cystic fibrosis) and MPS (mucopolysaccharidosis). Although various biological actions of genistein are employed in these two cases, in vitro studies, tests on animal models and pilot clinical trials suggest that this plant-derived compound might be a real hope for patients suffering from severe inherited disorders with relatively complicated pathomechanisms, including those affecting the central nervous system

Experimental Plant Biology: Why Not?! Genistein: a natural isoflavone with a potential for treatment of genetic diseases

Abstract Genistein [4 ,5,7-trihydroxyisoflavone or 5,7-dihydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-one] is a natural isoflavone occurring in many plants known to possess various biological activities, ranging from phyto-oestrogenic to antioxidative actions. Recent studies indicated that this isoflavone can also be considered as a drug for as yet untreatable genetic diseases. In the present review, we discuss a plausible use of genistein in treatment of two genetic disorders: CF (cystic fibrosis) and MPS (mucopolysaccharidosis). Although various biological actions of genistein are employed in these two cases, in vitro studies, tests on animal models and pilot clinical trials suggest that this plant-derived compound might be a real hope for patients suffering from severe inherited disorders with relatively complicated pathomechanisms, including those affecting the central nervous system

Effects of flavonoids on glycosaminoglycan synthesis: implications for substrate reduction therapy in Sanfilippo disease and other mucopolysaccharidoses

Sanfilippo disease (mucopolysaccharidosis type III, MPS III) is a severe metabolic disorder caused by accumulation of heparan sulfate (HS), one of glycosaminoglycans (GAGs), due to a genetic defect resulting in a deficiency of GAG hydrolysis. This disorder is characterized as the most severe neurological form of MPS, revealing rapid deterioration of brain functions. Among therapeutic approaches for MPS III, one of the most promising appears to be the substrate reduction therapy (SRT). Genistein (5, 7-dihydroxy-3- (4-hydroxyphenyl)-4H-1-benzopyran-4-one) is an isoflavone that has been used in SRT for MPS III. In this report, we tested effects of other flavonoids (apigenin, daidzein, kaempferol and naringenin) on GAG synthesis. Their cytotoxicity and anti-proliferation features were also tested. We found that daidzein and kaempferol inhibited GAG synthesis significantly. Moreover, these compounds were able to reduce lysosomal storage in MPS IIIA fibroblasts. Interestingly, although genistein is believed to inhibit GAG synthesis by blocking the tyrosine kinase activity of the epidermal growth factor receptor, we found that effects of other flavonoids were not due to this mechanism. In fact, combinations of various flavonoids resulted in significantly more effective inhibition of GAG synthesis than the use of any of these compounds alone. These results, together with results published recently by others, suggest that combination of flavonoids can be considered as a method for improvement of efficiency of SRT for MPS III

A dual promoter system regulating λ DNA replication initiation

Transcription and DNA replication are tightly regulated to ensure coordination of gene expression with growth conditions and faithful transmission of genetic material to progeny. A large body of evidence has accumulated, indicating that encounters between protein machineries carrying out DNA and RNA synthesis occur in vivo and may have important regulatory consequences. This feature may be exacerbated in the case of compact genomes, like the one of bacteriophage λ, used in our study. Transcription that starts at the rightward pR promoter and proceeds through the λ origin of replication and downstream of it was proven to stimulate the initiation of λ DNA replication. Here, we demonstrate that the activity of a convergently oriented pO promoter decreases the efficiency of transcription starting from pR. Our results show, however, that a lack of the functional pO promoter negatively influences λ phage and λ-derived plasmid replication. We present data, suggesting that this effect is evoked by the enhanced level of the pR-driven transcription, occurring in the presence of the defective pO, which may result in the impeded formation of the replication initiation complex. Our data suggest that the cross talk between the two promoters regulates λ DNA replication and coordinates transcription and replication processes