Alteration/deficiency in activation-3 (Ada3) plays a critical role in maintaining genomic stability

Cell cycle regulation and DNA repair following damage are essential for maintaining genome integrity. DNA damage activates checkpoints in order to repair damaged DNA prior to exit to the next phase of cell cycle. Recently, we have shown the role of Ada3, a component of various histone acetyltransferase complexes, in cell cycle regulation, and loss of Ada3 results in mouse embryonic lethality. Here, we used adenovirus-Cre-mediated Ada3 deletion in Ada3^fl/fl mouse embryonic fibroblasts (MEFs) to assess the role of Ada3 in DNA damage response following exposure to ionizing radiation (IR). We report that Ada3 depletion was associated with increased levels of phospho-ATM (pATM), γH2AX, phospho-53BP1 (p53BP1) and phospho-RAD51 (pRAD51) in untreated cells; however, radiation response was intact in Ada3^−/− cells. Notably, Ada3^−/− cells exhibited a significant delay in disappearance of DNA damage foci for several critical proteins involved in the DNA repair process. Significantly, loss of Ada3 led to enhanced chromosomal aberrations, such as chromosome breaks, fragments, deletions and translocations, which further increased upon DNA damage. Notably, the total numbers of aberrations were more clearly observed in S-phase, as compared with G₁ or G₂ phases of cell cycle with IR. Lastly, comparison of DNA damage in Ada3^fl/fl and Ada3^−/− cells confirmed higher residual DNA damage in Ada3^−/− cells, underscoring a critical role of Ada3 in the DNA repair process. Taken together, these findings provide evidence for a novel role for Ada3 in maintenance of the DNA repair process and genomic stability.     

IL-3 and TNFα increase Thymic Stromal Lymphopoietin Receptor (TSLPR) expression on eosinophils and enhance TSLP-stimulated degranulation

Background

Thymic stromal lymphopoietin (TSLP) and eosinophils are prominent components of allergic inflammation. Therefore, we sought to determine whether TSLP could activate eosinophils, focusing on measuring the regulation of TSLPR expression on eosinophils and degranulation in response to TSLP, as well as other eosinophil activation responses.

Methods

Eosinophil mRNA expression of TSLPR and IL-7R?? was examined by real-time quantitative PCR of human eosinophils treated with TNF?? and IL-5 family cytokines, and TSLPR surface expression on eosinophils was analyzed by flow cytometry. Eosinophils were stimulated with TSLP (with and without pre-activation with TNF?? and IL-3) and evaluated for release of eosinophil derived neurotoxin (EDN), phosphorylation of STAT5, and survival by trypan blue exclusion. A blocking antibody for TSLPR was used to confirm the specificity of TSLP mediated signaling on eosinophil degranulation.

Results

Eosinophil expression of cell surface TSLPR and TSLPR mRNA was upregulated by stimulation with TNF?? and IL-3. TSLP stimulation resulted in release of EDN, phosphorylation of STAT5 as well as promotion of viability and survival. TSLP-stimulated eosinophil degranulation was inhibited by a functional blocking antibody to TSLPR. Pre-activation of eosinophils with TNF?? and IL-3 promoted eosinophil degranulation at lower concentrations of TSLP stimulation.

Conclusions

This study demonstrates that eosinophils are activated by TSLP and that eosinophil degranulation in response to TSLP may be enhanced on exposure to cytokines present in allergic inflammation, indicating that the eosinophil has the capacity to participate in TSLP-driven allergic responses.     

Novel diarylheptanoids as inhibitors of TNF-α production

Synthesis and anti-inflammatory activity of novel diarylheptanoids [5-hydroxy-1-phenyl-7-(pyridin-3-yl)-heptan-3-ones and 1-phenyl-7-(pyridin-3-yl)hept-4-en-3-ones] as inhibitors of tumor necrosis factor-α (TNF-α) production is described in the present article. The key reactions involve the formation of a β-hydroxyketone by the reaction of substituted 4-phenyl butan-2-ones with pyridine-3-carboxaldehyde in presence of LDA and the subsequent dehydration of the same to obtain the α,β-unsaturated ketones. Compounds 4i, 5b, 5d, and 5g significantly inhibit lipopolysaccharide (LPS)-induced TNF-α production from human peripheral blood mononuclear cells in a dose-dependent manner. Of note, the in vitro TNF-α inhibition potential of 5b and 5d is comparable to that of curcumin (a naturally occurring diarylheptanoid). Most importantly, oral administration of 4i, 5b, 5d, and 5g (each at 100 mg/kg) but not curcumin (at 100 mg/kg) significantly inhibits LPS-induced TNF-α production in BALB/c mice. Collectively, our findings indicate that these compounds may have potential therapeutic implications for TNF-α-mediated auto-immune/inflammatory disorders.     

Genetics of melanocytic nevi

Melanocytic nevi are a benign clonal proliferation of cells expressing the melanocytic phenotype, with heterogeneous clinical and molecular characteristics. In this review, we discuss the genetics of nevi by salient nevi subtypes: congenital melanocytic nevi, acquired melanocytic nevi, blue nevi, and Spitz nevi. While the molecular etiology of nevi has been less thoroughly studied than melanoma, it is clear that nevi and melanoma share common driver mutations. Acquired melanocytic nevi harbor oncogenic mutations in BRAF, which is the predominant oncogene associated with melanoma. Congenital melanocytic nevi and blue nevi frequently harbor NRAS mutations and GNAQ mutations, respectively, while Spitz and atypical Spitz tumors often exhibit HRAS and kinase rearrangements. These initial ‘driver’ mutations are thought to trigger the establishment of benign nevi. After this initial phase of the cell proliferation, a senescence program is executed, causing termination of nevi growth. Only upon the emergence of additional tumorigenic alterations, which may provide an escape from oncogene‐induced senescence, can malignant progression occur. Here, we review the current literature on the pathobiology and genetics of nevi in the hope that additional studies of nevi promise to inform our understanding of the transition from benign neoplasm to malignancy.     

Socio-Economic Disparities in Use of Family Planning Methods among Pakistani Women: Findings from Pakistan Demographic and Health Surveys

BackgroundSeveral developing countries like Pakistan step into Sustainable Development Goals period with crucial maternal and child health needs that need to be addressed for improving health outcomes among people. We aim to explore existent socio-economic disparities in use of family planning methods (FPM) among Pakistani women, and compare any such inequalities between the years 2006 and 2013.SettingPakistan Demographic and Health Surveys (PDHS) 2006–7 (n = 9177) and the most recent 2012–13(n = 13558) data were used to conduct secondary analysis. Participants were ever married women aged between 15 and 49 years. Socio-economic status was assessed by the education level and wealth index. Inequalities were measured through Odds Ratio (OR), Relative Index of inequality (RII), and Slope index of inequality (SII) on non-use of FPM.ResultsAlthough the prevalence of FPM use has increased over time (28% in 2006 versus 54% in 2013), the socio-economic inequalities persistently exist. Comparing results of PDHS 2006 with PDHS 2013, education related absolute inequalities among urban dwellers increased from -0.41 (95% CI -0.67, -0.13, p-value < 0.01) to -0.83 (95% CI -1.02, -0.63, p-value < 0.01); and increased from -0.93 (95% CI -1.21, -0.64, p-value < 0.01) to -0.98 (95% CI -1.20, -0.76, p-value < 0.01) among rural dwellers. Similarly wealth related absolute inequalities are also existent.ConclusionsAlthough the FPM use has increased over time, but it is important to note that socio-economic gap in use of FPM persists. Such differences have disadvantaged the poor and the illiterate. Family planning programs may target the disadvantaged subgroups for ensuring well-being of women and children in Pakistan.     

Lyotropic Liquid Crystalline Nanoparticles of Amphotericin B: Implication of Phytantriol and Glyceryl Monooleate on Bioavailability Enhancement

Implication of different dietary specific lipids such as phytantriol (PT) and glyceryl monooleate (GMO) on enhancing the oral bioavailability of amphotericin B (AmB) was examined. Liquid crystalline nanoparticles (LCNPs) were prepared using hydrotrope method, followed by in vitro characterization, Caco-2 cell monolayer uptake, and in vivo pharmacokinetic and toxicity evaluation. Optimized AmB-LCNPs displayed small particle size (<?210 nm) with a narrow distribution (~?0.2), sustained drug release and high gastrointestinal stability, and reduced hemolytic toxicity. PLCNPs presented slower release, i.e., ~?80% as compared to ~?90% release in case of GLCNPs after 120 h. Significantly higher uptake in Caco-2 monolayer substantiated the role of LCNPs in increasing the intestinal permeability followed by increased drug titer in plasma. Pharmacokinetic studies demonstrated potential of PT in enhancing the bioavailability (approximately sixfold) w.r.t. of its native counterpart with reduced nephrotoxicity as presented by reduced nephrotoxicity biomarkers and histology studies. These studies established usefulness of PLCNPs over GLCNPs and plain drug. It can be concluded that acid-resistant lipid, PT, can be utilized efficiently as an alternate lipid for the preparation of LCNPs to enhance bioavailability and to reduce nephrotoxicity of the drug as compared to other frequently used lipid, i.e., GMO.