In silico and in vitro evaluation of the anti-virulence potential of patuletin, a natural methoxy flavone, against Pseudomonas aeruginosa.

This study aimed to investigate the potential of patuletin, a rare natural flavonoid, as a virulence and LasR inhibitor against Pseudomonas aeruginosa. Various computational studies were utilized to explore the binding of Patuletin and LasR at a molecular level. Molecular docking revealed that Patuletin strongly interacted with the active pocket of LasR, with a high binding affinity value of -20.96 kcal/mol. Further molecular dynamics simulations, molecular mechanics generalized Born surface area (MM/GBSA), protein-ligand interaction profile (PLIP), and essential dynamics analyses confirmed the stability of the patuletin-LasR complex, and no significant structural changes were observed in the LasR protein upon binding. Key amino acids involved in binding were identified, along with a free energy value of -26.9 kcal/mol. In vitro assays were performed to assess patuletin's effects on P. aeruginosa. At a sub-inhibitory concentration (1/4 MIC), patuletin significantly reduced biofilm formation by 48% and 42%, decreased pyocyanin production by 24% and 14%, and decreased proteolytic activities by 42% and 20% in P. aeruginosa isolate ATCC 27853 (PA27853) and P. aeruginosa clinical isolate (PA1), respectively. In summary, this study demonstrated that patuletin effectively inhibited LasR activity in silico and attenuated virulence factors in vitro, including biofilm formation, pyocyanin production, and proteolytic activity. These findings suggest that patuletin holds promise as a potential therapeutic agent in combination with antibiotics to combat antibiotic-tolerant P. aeruginosa infections.

Anti-virulence potential of patuletin, a natural flavone, against Staphylococcus aureus: In vitro and In silico investigations.

Staphylococcus aureus is a highly prevalent and aggressive human pathogen causing a wide range of infections. This study aimed to explore the potential of Patuletin, a rare natural flavone, as an anti-virulence agent against S. aureus. At a sub-inhibitory concentration (1/4 MIC), Patuletin notably reduced biofilm formation by 27 % and 23 %, and decreased staphyloxanthin production by 53 % and 46 % in Staphylococcus aureus isolate SA25923 and clinical isolate SA1, respectively. In order to gain a more comprehensive understanding of the in vitro findings, several in silico analyses were conducted. Initially, a 3D-flexible alignment study demonstrated a favorable structural similarity between Patuletin and B70, the co-crystallized ligand of CrtM, an enzyme that plays a pivotal role in the biosynthesis of staphyloxanthin. Molecular docking highlighted the strong binding of Patuletin to the active site of CrtM, with a high affinity of -20.95 kcal/mol. Subsequent 200 ns molecular dynamics simulations, along with MM-GBSA, ProLIF, PLIP, and PCAT analyses, affirmed the stability of the Patuletin-CrtM complex, revealing no significant changes in CrtM's structure upon binding. Key amino acids crucial for binding were also identified. Collectively, this study showcased the effective inhibition of CrtM activity by Patuletin in silico and its attenuation of key virulence factors in vitro, including biofilm formation and staphyloxanthin production. These findings hint at Patuletin's potential as a valuable therapeutic agent, especially in combination with antibiotics, to counter antibiotic-resistant Staphylococcus aureus infections.

Crippling of Klebsiella pneumoniae virulence by metformin, N-acetylcysteine and secnidazole.

INTRODUCTION: The emergence of multidrug-resistant Klebsiella pneumoniae in hospitals represents a serious threat to public health. Infections caused by Klebsiella pneumoniae are widespread in healthcare institutions, mainly pneumonia, bloodstream infections, and infections affecting neonates in intensive care units; so, it is necessary to combat this pathogen with new strategies. Targeting virulence factors necessary to induce host damage and disease is a new paradigm for antimicrobial therapy with several potential benefits that could lead to decreased resistance. BACKGROUND: The influence of metformin, N-acetylcysteine, and secnidazole on Klebsiella pneumoniae virulence factors production was tested. The production of Klebsiella pneumoniae virulence factors such as biofilm formation, urease, proteases, hemolysins, and tolerance to oxidative stress was evaluated phenotypically using sub-inhibitory concentration (1/8 MIC) of metformin, N-acetylcysteine, and secnidazole. For more confirmation, qRT-PCR was used to assess the relative expression level of rmpA, wcaG, fimH-1, mrkD, ureA, and khe genes regulating virulence factors production. RESULTS: Metformin, N-acetylcysteine, and secnidazole were all found to have a powerful inhibitory effect on the production of virulence factors phenotypically. Our results showed a significant reduction in the expression level of rmpA, wcaG, fimH-1, mrkD, ureA, and khe genes. Furthermore, the tested drugs were investigated in vivo to inform their ability to protect mice against Klebsiella pneumoniae pathogenesis. CONCLUSIONS: Metformin, N-acetylcysteine, and secnidazole inhibited the virulence of Klebsiella pneumoniae. Besides combating resistant Klebsiella pneumoniae, the tested drugs could also serve as an adjuvant to traditional antibiotics.

Immobilization of ZnO-TiO2 Nanocomposite into Polyimidazolium Amphiphilic Chitosan Film, Targeting Improving Its Antimicrobial and Antibiofilm Applications.

This study presents a green protocol for the fabrication of a multifunctional smart nanobiocomposite (NBC) (ZnO-PIACSB-TiO2) for secure antimicrobial and antibiofilm applications. First, shrimp shells were upgraded to a polyimidazolium amphiphilic chitosan Schiff base (PIACSB) through a series of physicochemical processes. After that, the PIACSB was used as an encapsulating and coating agent to manufacture a hybrid NBC in situ by co-encapsulating ZnONPs and TiO2NPs. The physicochemical and visual characteristics of the new NBC were investigated by spectral, microscopic, electrical, and thermal methods. The antimicrobial indices revealed that the newly synthesized, PIACSB-coated TiO2-ZnO nanocomposite is an exciting antibiotic due to its amazing antimicrobial activity (MIC/MBC→0.34/0.68 µg/mL, 0.20/0.40 µg/mL, and 0.15/0.30 µg/mL working against S. aureus, E. coli, and P. aeruginosa, respectively) and antifungal capabilities. Additionally, ZnO-PIACSB-TiO2 is a potential fighter of bacterial biofilms, with the results being superior to those of the positive control (Cipro), which worked against S. aureus (only 8.7% ± 1.9 biofilm growth), E. coli (only 1.4% ± 1.1 biofilm growth), and P. aeruginosa (only 0.85% ± 1.3 biofilm growth). Meanwhile, the NBC exhibits excellent biocompatibility, as evidenced by its IC50 values against both L929 and HSF (135 and 143 µg/mL), which are significantly higher than those of the MIC doses (0.24-24.85 µg/mL) that work against all tested microbes, as well as the uncoated nanocomposite (IC50 = 19.36 ± 2.04 and 23.48 ± 1.56 µg/mL). These findings imply that the new PIACSB-coated nanocomposite film may offer promising multifunctional food packaging additives to address the customer demand for safe, eco-friendly food products with outstanding antimicrobial and antibiofilm capabilities.

Involvement of the SAGA and TFIID coactivator complexes in transcriptional dysregulation caused by the separation of core and tail Mediator modules.

Regulation of RNA polymerase II transcription requires the concerted efforts of several multisubunit coactivator complexes, which interact with the RNA polymerase II preinitiation complex to stimulate transcription. We previously showed that separation of the Mediator core from Mediator's tail module results in modest overactivation of genes annotated as highly dependent on TFIID for expression. However, it is unclear if other coactivators are involved in this phenomenon. Here, we show that the overactivation of certain genes by Mediator core/tail separation is blunted by disruption of the Spt-Ada-Gcn5-Acetyl transferase complex through the removal of its structural Spt20 subunit, though this downregulation does not appear to completely depend on reduced Spt-Ada-Gcn5-Acetyl transferase association with the genome. Consistent with the enrichment of TFIID-dependent genes among genes overactivated by Mediator core/tail separation, depletion of the essential TFIID subunit Taf13 suppressed the overactivation of these genes when Med16 was simultaneously removed. As with Spt-Ada-Gcn5-Acetyl transferase, this effect did not appear to be fully dependent on the reduced genomic association of TFIID. Given that the observed changes in gene expression could not be clearly linked to alterations in Spt-Ada-Gcn5-Acetyl transferase or TFIID occupancy, our data may suggest that the Mediator core/tail connection is important for the modulation of Spt-Ada-Gcn5-Acetyl transferase and/or TFIID conformation and/or function at target genes.

Attenuating the virulence of the resistant superbug Staphylococcus aureus bacteria isolated from neonatal sepsis by ascorbic acid, dexamethasone, and sodium bicarbonate.

BACKGROUND: Infections affecting neonates caused by Staphylococcus aureus are widespread in healthcare facilities; hence, novel strategies are needed to fight this pathogen. In this study, we aimed to investigate the effectiveness of the FDA-approved medications ascorbic acid, dexamethasone, and sodium bicarbonate to reduce the virulence of the resistant Staphylococcus aureus bacteria that causes neonatal sepsis and seek out suitable alternatives to the problem of multi-drug resistance. METHODS: Tested drugs were assessed phenotypically and genotypically for their effects on virulence factors and virulence-encoding genes in Staphylococcus aureus. Furthermore, drugs were tested in vivo for their ability to reduce Staphylococcus aureus pathogenesis. RESULTS: Sub-inhibitory concentrations (1/8 MIC) of ascorbic acid, dexamethasone, and sodium bicarbonate reduced the production of Staphylococcus aureus virulence factors, including biofilm formation, staphyloxanthin, proteases, and hemolysin production, as well as resistance to oxidative stress. At the molecular level, qRT-PCR was used to assess the relative expression levels of crtM, sigB, sarA, agrA, hla, fnbA, and icaA genes regulating virulence factors production and showed a significant reduction in the relative expression levels of all the tested genes. CONCLUSIONS: The current findings reveal that ascorbic acid, dexamethasone, and sodium bicarbonate have strong anti-virulence effects against Staphylococcus aureus. Thus, suggesting that they might be used as adjuvants to treat infections caused by Staphylococcus aureus in combination with conventional antimicrobials or as alternative therapies.

Structure based design and synthesis of 3-(7-nitro-3-oxo-3,4-dihydroquinoxalin-2-yl)propanehydrazide derivatives as novel bacterial DNA-gyrase inhibitors: In-vitro, In-vivo, In-silico and SAR studies.

Antimicrobial resistance (AMR) is one of the critical challenges that have been encountered over the past years. On the other hand, bacterial DNA gyrase is regarded as one of the most outstanding biological targets that quinolones can extensively inhibit, improving AMR. Hence, a novel series of 3-(7-nitro-3-oxo-3,4-dihydroquinoxalin-2-yl)propanehydrazide derivatives (3-6j) were designed and synthesized employing the quinoxaline-2-one scaffold and relying on the pharmacophoric features experienced by the quinolone antibiotic; ciprofloxacin. The antibacterial activity of the synthesized compounds was assessed via in-vitro approaches using eight different Gram-positive and Gram-negative bacterial species. Most of the synthesized compounds revealed eligible antibacterial activities. In particular, compounds 6d and 6e displayed promising antibacterial activity among the investigated compounds. For example, compounds 6d and 6e displayed MIC values of 9.40 and 9.00 µM, respectively, regarding S. aureus, and 4.70 and 4.50 µM, respectively, regarding S. pneumonia in comparison to ciprofloxacin (12.07 µM). The cytotoxicity of compounds 6d and 6e were performed on normal human WI-38 cell lines with IC50 values of 288.69 and 227.64 µM, respectively assuring their safety and selectivity. Besides, DNA gyrase inhibition assay of compounds 6d and 6e was carried out in comparison to ciprofloxacin, and interestingly, compounds 6d and 6e disclosed promising IC50 values of 0.242 and 0.177 µM, respectively, whereas ciprofloxacin displayed an IC50 value of 0.768 µM, assuring the proposed mechanism of action for the afforded compounds. Consequently, compounds 6d and 6e were further assessed via in-vivo approaches by evaluating blood counts, liver and kidney functions, and histopathological examination. Both compounds were found to be safer on the liver and kidney than the reference ciprofloxacin. Moreover, in-silico molecular docking studies were established and revealed reasonable binding affinities for all afforded compounds, particularly compound 6d which exhibited a binding score of -7.51 kcal/mol, surpassing the reference ciprofloxacin (-7.29 kcal/mol) with better anticipated stability at the DNA gyrase binding pocket. Moreover, ADME studies were conducted, disclosing an eligible bioavailability score of >0.55 for all afforded compounds, and reasonable GIT absorption without passing the blood brain barrier was attained for most investigated compounds, ensuring their efficacy and safety. Lastly, a structure activity relationship study for the synthesized compounds was established and unveiled that not only the main pharmacophores required for DNA gyrase inhibition are enough for exerting promising antimicrobial activities, but also derivatization with diverse aryl/hetero aryl aldehydes is essential for their enhanced antimicrobial potential.

In a search for potential drug candidates for combating COVID-19: computational study revealed salvianolic acid B as a potential therapeutic targeting 3CLpro and spike proteins.

The global prevalence of COVID-19 disease and the overwhelming increase in death toll urge scientists to discover new effective drugs. Although the drug discovery process is a challenging and time-consuming, fortunately, the plant kingdom was found to have many active therapeutics possessing broad-spectrum antiviral activity including those candidates active against severe acute respiratory syndrome coronaviruses (SARS-CoV). Herein, nine traditional Chinese medicinal plant constituents from different origins (Glycyrrhizin 1, Lycorine 2, Puerarin 3, Daidzein 4, Daidzin 5, Salvianolic acid B 6, Dihydrotanshinone I 7, Tanshinone I 8, Tanshinone IIa 9) previously reported to exhibit antiviral activity against SARS-CoV were virtually screened in silico (molecular docking) as potential inhibitors of SARS-CoV-2 target proteins. The tested medicinal plant compounds were in silico screened for their activity against two key SARS-CoV-2 target proteins; 3CLpro, and Spike binding-domain proteins. Among the tested medicinal plant compounds, Salvianolic acid B 6 (Sal-B) showed promising binding affinities against the two specified SARS-CoV-2 target proteins compared to the reference standards used. Hence molecular dynamics simulations followed by calculating the free-binding energy were carried out for Sal-B providing information on its affinity, stability, and thermodynamic behavior within the two SARS-CoV-2 target proteins as well as key ligand-protein binding aspects. Besides, the quantum mechanical calculations showed that Sal-B can adopt different conformations due to the existence of various rotatable bonds. Therefore, the enhanced antiviral activity of Sal-B among other studied compounds can be also attributed to the structural flexibility of Sal-B. Our study gives an explanation of the structure activity relationship required for targeting SARS-CoV-2 3CLpro and Spike proteins and also facilitates the future design and synthesis of new potential drugs exhibiting better affinity and specificity. Besides, an ADME study was carried out on screened compounds and reference controls revealing their pharmacokinetics properties.Communicated by Ramaswamy H. Sarma.

In vitro and computational insights revealing the potential inhibitory effect of Tanshinone IIA against influenza A virus.

Seasonal human influenza is a serious respiratory infection caused by influenza viruses that can be found all over the world. Type A influenza is a contagious viral infection that, if left untreated, can lead to life-threatening consequences. Fortunately, the plant kingdom has many potent medicines with broad-spectrum antiviral activity. Herein, six plant constituents, namely Tanshinone IIA 1, Carnosic acid 2, Rosmarinic acid 3, Glycyrrhetinic acid 4, Baicalein 5, and Salvianolic acid B 6, were screened for their antiviral activities against H1N1 virus using in vitro and in silico approaches. Hence, their anti-influenza activities were tested in vitro to determine inhibitory concentration 50 (IC50) values after measuring their CC50 values using MTT assay on MDCK cells. Interestingly, Tanshinone IIA (TAN) 1 was the most promising member with CC50 = 9.678 µg/ml. Moreover, the plaque reduction assay carried on TAN 1 revealed promising viral inhibition percentages of 97.9%, 95.8%, 94.4%, and 91.7% using concentrations 0.05 µg/µl, 0.025 µg/µl, 0.0125 µg/µl, and 0.006 µg/µl, respectively. Furthermore, in silico molecular docking disclosed the superior affinities of Salvianolic acid B (SAL) 6 towards both surface glycoproteins of influenza A virus (namely, hemagglutinin (HA) and neuraminidase (NA)). The docked complexes of both SAL and TAN inside HA and NA receptor pockets were selected for 100 ns MD simulations followed by MM-GBSA binding free energy calculation to confirm the docking results and give more insights regarding the stability of both compounds inside influenza mentioned receptors, respectively. The selection criteria of the previously mentioned complexes were based on the fact that SAL showed the highest docking scores on both viral HA and NA glycoproteins whereas TAN achieved the best inhibitory activity on the other hand. Finally, we urge more advanced preclinical and clinical research, particularly for TAN, which could be used to treat the human influenza A virus effectively.

Genome-Wide Profiling of Protein-DNA Interactions with Chromatin Endogenous Cleavage and High-Throughput Sequencing (ChEC-Seq ).

Interactions between regulatory proteins and specific genomic regions are critical for all chromatin-based processes, including transcription, DNA replication, and DNA repair. Genome-wide mapping of such interactions is most commonly performed with chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq), but a number of orthogonal methods employing targeted enzymatic activity have also been introduced. We previously described a genome-wide implementation of chromatin endogenous cleavage (ChEC-Seq), wherein a protein of interest is fused to micrococcal nuclease (MNase) to enable targeted, calcium-dependent genomic cleavage. Here, we describe the ChEC-Seq protocol for use in budding yeast though it can be used in other organisms in conjunction with appropriate methods for introduction of an MNase fusion protein.

Connection of core and tail Mediator modules restrains transcription from TFIID-dependent promoters.

The Mediator coactivator complex is divided into four modules: head, middle, tail, and kinase. Deletion of the architectural subunit Med16 separates core Mediator (cMed), comprising the head, middle, and scaffold (Med14), from the tail. However, the direct global effects of tail/cMed disconnection are unclear. We find that rapid depletion of Med16 downregulates genes that require the SAGA complex for full expression, consistent with their reported tail dependence, but also moderately overactivates TFIID-dependent genes in a manner partly dependent on the separated tail, which remains associated with upstream activating sequences. Suppression of TBP dynamics via removal of the Mot1 ATPase partially restores normal transcriptional activity to Med16-depleted cells, suggesting that cMed/tail separation results in an imbalance in the levels of PIC formation at SAGA-requiring and TFIID-dependent genes. We propose that the preferential regulation of SAGA-requiring genes by tailed Mediator helps maintain a proper balance of transcription between these genes and those more dependent on TFIID.

Alterations in skin microbiome mediated by radiotherapy and their potential roles in the prognosis of radiotherapy-induced dermatitis: a pilot study.

Radiotherapy-induced dermatitis (RID) is an inflammatory cutaneous disorder that is acquired as an adverse effect of undergoing radiotherapy. Skin microbiome dysbiosis has been linked to the outcomes of several dermatological diseases. To explore the skin microbiota of RID and deduce their underlying impact on the outcome of RID, cutaneous microbiomes of 78 RID patients and 20 healthy subjects were characterized by sequencing V1-V3 regions of 16S rRNA gene. In total, a significantly apparent reduction in bacterial diversity was detected in microbiomes of RID in comparison to controls. Overall, the raised Proteobacteria/ Firmicutes ratio was significantly linked to delayed recovery or tendency toward the permanence of RID (Kruskal Wallis: P = 2.66 × 10-4). Moreover, applying enterotyping on our samples stratified microbiomes into A, B, and C dermotypes. Dermotype C included overrepresentation of Pseudomonas, Staphylococcus and Stenotrophomonas and was markedly associated with delayed healing of RID. Strikingly, coexistence of diabetes mellitus and RID was remarkably correlated with a significant overrepresentation of Klebsiella or Pseudomonas and Staphylococcus. Metabolic abilities of skin microbiome could support their potential roles in the pathogenesis of RID. Cutaneous microbiome profiling at the early stages of RID could be indicative of prospective clinical outcomes and maybe a helpful guide for personalized therapy.

Architectural Mediator subunits are differentially essential for global transcription in Saccharomyces cerevisiae.

Mediator is a modular coactivator complex involved in the transcription of the majority of RNA polymerase II-regulated genes. However, the degrees to which individual core subunits of Mediator contribute to its activity have been unclear. Here, we investigate the contribution of two essential architectural subunits of Mediator to transcription in Saccharomyces cerevisiae. We show that acute depletion of the main complex scaffold Med14 or the head module nucleator Med17 is lethal and results in global transcriptional downregulation, though Med17 removal has a markedly greater negative effect. Consistent with this, Med17 depletion impairs preinitiation complex (PIC) assembly to a greater extent than Med14 removal. Co-depletion of Med14 and Med17 reduced transcription and TFIIB promoter occupancy similarly to Med17 ablation alone, indicating that the contributions of Med14 and Med17 to Mediator function are not additive. We propose that, while the structural integrity of complete Mediator and the head module are both important for PIC assembly and transcription, the head module plays a greater role in this process and is thus the key functional module of Mediator in this regard.

Zinc oxide nanoparticles inhibits quorum sensing and virulence in Pseudomonas aeruginosa.

BACKGROUND: Quorum sensing inhibitionis an advanced strategy that aims to interfere with bacterial cell-to-cell communication systems (quorum sensing), which regulate virulence factors production in Pseudomonas aeruginosa, in order to overcome the globalcrisis of antimicrobial resistance. OBJECTIVES: Study the potential quorum sensing inhibitory effect of Zinc oxide (ZnO)nanoparticlesin Pseudomonas aeruginosa and the impact on production of virulence factors. METHODS: Quorum sensing inhibitory effect of ZnO was evaluated by assessing its ability to reducePseudomonas aeruginosa virulence factors production; rhamnolipids, pyocyanin, pyoverdin, hemolysins, elastase and proteases. Furthermore, qRT-PCR was performed to determine ZnO inhibitory effect onQS-regulatory geneslasI, lasR, rhlI, rhlR, pqsA and pqsR that control virulence factors secretion. Moreover, mice survival test was conducted to investigate the influence of ZnO on Pseudomonas aeruginosa-induced mortality in vivo. RESULTS: ZnO revealed a statistically significant reduction in the production of QS-controlled virulence factors rhamnolipids, pyocyanin, pyoverdin, hemolysins, elastase and proteases. Furthermore, ZnO exhibited a significant decrease in the relative expression of QS-regulatory geneslasI, lasR, rhlI, rhlR, pqsA and pqsR. Additionally, ZnO significantly reduced the pathogenesis of Pseudomonas aeruginosa in vivo. CONCLUSION: ZnO nanoparticles can be used as a quorum sensing inhibitor in Pseudomonas aeruginosa infections either as an adjuvant or alternative to conventional antimicrobials.

Repositioning secnidazole as a novel virulence factors attenuating agent in Pseudomonas aeruginosa.

Long-term treatment with antibiotics gives rise to the evolution of multi-drug resistant bacteria which are hard to be treated. Virulence factors inhibitors depend on disarming of microbial pathogens through reducing expression of virulence factors, abolishing the pathogen capability to harm the host. In the present study, the influence of secnidazole on Pseudomonas aeruginosa virulence factors expression was characterized. Production of Pseudomonas aeruginosa virulence factors such as pyocyanin, pyoverdin, elastase, rhamnolipids, proteases and hemolysins was examined following treatment of bacteria with sub-inhibitory concentration of secnidazole. Interestingly, secnidazole showed a powerful inhibitory effect on Pseudomonas aeruginosa virulence factors. Our results were further confirmed using qRT-PCR showing that there was a significant decrease in the expression of quorum sensing genes; lasI, lasR, rhlI, rhlR, pqsA and pqsR that regulate expression of virulence factors in Pseudomonas aeruginosa. Moreover, in vivo experiment using mice as infection model showed that secnidazole-treated bacteria were less capable to kill mice as compared to untreated bacteria. Importantly, there was a significant reduction in mortality in mice injected with secnidazole-treated bacteria relative to mice inoculated with untreated bacteria. In summary, our data showed that secnidazole could play a role in attenuating Pseudomonas aeruginosa through reducing virulence factors production. Moreover, our data clearly suggest that secnidazole could be involved in the treatment of Pseudomonas aeruginosa infections in order to control infection and lower the development of bacterial resistance to antibiotics.

Mediator Is Essential for Small Nuclear and Nucleolar RNA Transcription in Yeast.

Eukaryotic RNA polymerase II (RNAPII) transcribes mRNA genes and non-protein-coding RNA (ncRNA) genes, including those encoding small nuclear and nucleolar RNAs (sn/snoRNAs). In metazoans, RNAPII transcription of sn/snoRNAs is facilitated by a number of specialized complexes, but no such complexes have been discovered in yeast. It has been proposed that yeast sn/snoRNA and mRNA expression relies on a set of common factors, but the extent to which regulators of mRNA genes function at yeast sn/snoRNA genes is unclear. Here, we investigated a potential role for the Mediator complex, essential for mRNA gene transcription, in sn/snoRNA gene transcription. We found that Mediator maps to sn/snoRNA gene regulatory regions and that rapid depletion of the essential structural subunit Med14 strongly reduces RNAPII and TFIIB occupancy as well as nascent transcription of sn/snoRNA genes. Deletion of Med3 and Med15, subunits of the activator-interacting Mediator tail module, does not affect Mediator recruitment to or RNAPII and TFIIB occupancy of sn/snoRNA genes. Our analyses suggest that Mediator promotes PIC formation and transcription at sn/snoRNA genes, expanding the role of this critical regulator beyond its known functions in mRNA gene transcription and demonstrating further mechanistic similarity between the transcription of mRNA and sn/snoRNA genes.

Diabetes-induced perturbations are subject to intergenerational transmission through maternal line

The hypothesis of fetal origins of adult disease states that early life events program the occurrence of significant adult diseases, including diabetes and obesity. Maternal diabetes is associated with general stress environment for developing fetus, and gestational diabetes is an independent risk factor for type 2 diabetes and metabolic syndrome in offspring. Intra-uterine fetal programming of fetal tissues exposes the offspring to increased risk of impaired glucose tolerance, type 2 diabetes, and cardiovascular disease. Here, we examined the transmission of maternal diabetes-induced fetal programming in second generation and compared maternal and paternal routes of intergenerational effects. We organized 40 Wistar rats into three groups, male offspring of diabetic mothers, female offspring of diabetic mothers, and offspring of control mothers. These groups were mated with normal healthy rats to assess the effect of grand-maternal diabetes on pregnancy outcome in F2 rats, as well as glucose-sensing parameters, insulin resistance, and glucose tolerance prenatally and postnatally. We found that F2 offspring of diabetic mothers had impaired glucose sensing, increased oxidative stress, insulin resistance, and impaired glucose tolerance, and these effects were more prominent in the F2 offspring of F1 female rats (F2-DF1F). We deduce that fetal programming of maternal diabetes is mostly transmitted through maternal line across two generations (AU)

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In Utero Nutritional Manipulation Provokes Dysregulated Adipocytokines Production in F1 Offspring in Rats.

Background. Intrauterine environment plays a pivotal role in the origin of fatal diseases such as diabetes. Diabetes and obesity are associated with low-grade inflammatory state and dysregulated adipokines production. This study aims to investigate the effect of maternal obesity and malnutrition on adipokines production (adiponectin, leptin, and TNF-α) in F1 offspring in rats. Materials and Methods. Wistar rats were allocated in groups: F1 offspring of control mothers under control diet (CF1-CD) and under high-fat diet (CF1-HCD), F1 offspring of obese mothers under CD (OF1-CD) and under HCD (OF1-HCD), and F1 offspring of malnourished mothers under CD (MF1-CD) and under HCD (MF1-HCD). Every 5 weeks postnatally, blood samples were obtained for biochemical analysis. Results. At the end of the 30-week follow-up, OF1-HCD and MF1-HCD exhibited hyperinsulinemia, moderate dyslipidemia, insulin resistance, and impaired glucose homeostasis compared to CF1-CD and CF1-HCD. OF1-HCD and MF1-HCD demonstrated low serum levels of adiponectin and high levels of leptin compared to CF1-CD and CF1-HCD. OF1-CD, OF1-HCD, and MF1-HCD had elevated serum levels of TNF-α compared to CF1-CD and CF1-HCD (p < 0.05). Conclusion. Maternal nutritional manipulation predisposes the offspring to development of insulin resistance in their adult life, probably via instigating dysregulated adipokines production.

Maternal diabetes impairs oxidative and inflammatory response in murine placenta.

Placenta is the major exchange surface between mother and fetus and plays a pivotal role in fetal development. A better understanding of the mechanisms by which diabetes alters placental function may allow better management of diabetes pregnancies. In this study, we attempt to investigate the effect of diabetic milieu with and without malformation on placental function. In order to investigate the impact of diabetic pregnancy on oxidative stress, endothelial and vascular functions of placental tissue, we mated diabetic and non-diabetic female rats with normal male rats. At gestational day 17, we terminated pregnancy, assessed fetuses for malformations and isolated placenta for measurement of various parameters of placental function. Our results show that maternal diabetes induced a state of oxidative stress in placenta, which disrupts normal signaling, activating apoptosis, as well as perturbing endothelial and vascular placental functions. The coalescence of these insults on various levels of placental function could contribute to the pleiotropic nature of diabetes-induced placental stress.

Diabetes-induced perturbations are subject to intergenerational transmission through maternal line.

The hypothesis of fetal origins of adult disease states that early life events program the occurrence of significant adult diseases, including diabetes and obesity. Maternal diabetes is associated with general stress environment for developing fetus, and gestational diabetes is an independent risk factor for type 2 diabetes and metabolic syndrome in offspring. Intra-uterine fetal programming of fetal tissues exposes the offspring to increased risk of impaired glucose tolerance, type 2 diabetes, and cardiovascular disease. Here, we examined the transmission of maternal diabetes-induced fetal programming in second generation and compared maternal and paternal routes of intergenerational effects. We organized 40 Wistar rats into three groups, male offspring of diabetic mothers, female offspring of diabetic mothers, and offspring of control mothers. These groups were mated with normal healthy rats to assess the effect of grand-maternal diabetes on pregnancy outcome in F2 rats, as well as glucose-sensing parameters, insulin resistance, and glucose tolerance prenatally and postnatally. We found that F2 offspring of diabetic mothers had impaired glucose sensing, increased oxidative stress, insulin resistance, and impaired glucose tolerance, and these effects were more prominent in the F2 offspring of F1 female rats (F2-DF1F). We deduce that fetal programming of maternal diabetes is mostly transmitted through maternal line across two generations.