Synthesis, biological assessment, and computational investigations of nifedipine and monastrol analogues as anti-leishmanial major and anti-microbial agents.

Leishmaniasis includes a range of parasitic diseases caused by numerous types of the protozoan kinetoplastid parasite. Fungal and bacterial pathogens have led to infectious illnesses causing some main public health problem in current years. A series of dihydropyridine and tetrahydropyrimidine derivatives having fluoro, bromo, and nitro substituents at para-phenyl ring on C4 of dihydropyridine and tetrahydropyrimidine rings were synthesized. Then, anti-leishmanial and antimicrobial potencies of compounds were assessed. All compounds were synthesized via Hantzsch and Biginelli reactions. All derivatives were evaluated for their anti-leishmanial and antimicrobial activities. Moreover, docking and molecular dynamics simulation calculations of the compounds in PRT1 binding site were performed to report the results of anti-leishmanial and antimicrobial activities. Compounds 4a and 4b showed the highest anti-amastigote and anti-promastigote activities. Compound 4a revealed the highest antimicrobial activity against E. coli, P. aeruginosa, and C. albicans strains. In addition, compound 4c showed the highest activity against S. aureus. The fluoro, bromo, and nitro substituents in para-position of phenyl group at C4 of dihydropyridine and tetrahydropyrimidine moieties as well as the bulk and length of the chain linking to the ester moieties are essential for anti-leishmanial and anti-microbial activities of these derivatives. Low cytotoxicity was shown by most of derivatives against macrophages. The molecular docking studies were in agreement with in vitro assay. Moreover, hydrogen binds, RMSF, RMSD, and Rg, strongly showed the steady binding of 4a and 4b compounds in PRT1 active site.

The Zic family homologue Odd-paired regulates Alk expression in Drosophila.

The Anaplastic Lymphoma Kinase (Alk) receptor tyrosine kinase (RTK) plays a critical role in the specification of founder cells (FCs) in the Drosophila visceral mesoderm (VM) during embryogenesis. Reporter gene and CRISPR/Cas9 deletion analysis reveals enhancer regions in and upstream of the Alk locus that influence tissue-specific expression in the amnioserosa (AS), the VM and the epidermis. By performing high throughput yeast one-hybrid screens (Y1H) with a library of Drosophila transcription factors (TFs) we identify Odd-paired (Opa), the Drosophila homologue of the vertebrate Zic family of TFs, as a novel regulator of embryonic Alk expression. Further characterization identifies evolutionarily conserved Opa-binding cis-regulatory motifs in one of the Alk associated enhancer elements. Employing Alk reporter lines as well as CRISPR/Cas9-mediated removal of regulatory elements in the Alk locus, we show modulation of Alk expression by Opa in the embryonic AS, epidermis and VM. In addition, we identify enhancer elements that integrate input from additional TFs, such as Binou (Bin) and Bagpipe (Bap), to regulate VM expression of Alk in a combinatorial manner. Taken together, our data show that the Opa zinc finger TF is a novel regulator of embryonic Alk expression.

Jeb/Alk signalling regulates the Lame duck GLI family transcription factor in the Drosophila visceral mesoderm.

The Jelly belly (Jeb)/Anaplastic Lymphoma Kinase (Alk) signalling pathway regulates myoblast fusion in the circular visceral mesoderm (VM) of Drosophila embryos via specification of founder cells. However, only a limited number of target molecules for this pathway are described. We have investigated the role of the Lame Duck (Lmd) transcription factor in VM development in relationship to Jeb/Alk signal transduction. We show that Alk signalling negatively regulates Lmd activity post-transcriptionally through the MEK/MAPK (ERK) cascade resulting in a relocalisation of Lmd protein from the nucleus to cytoplasm. It has previously been shown that downregulation of Lmd protein is necessary for the correct specification of founder cells. In the visceral mesoderm of lmd mutant embryos, fusion-competent myoblasts seem to be converted to 'founder-like' cells that are still able to build a gut musculature even in the absence of fusion. The ability of Alk signalling to downregulate Lmd protein requires the N-terminal 140 amino acids, as a Lmd(141-866) mutant remains nuclear in the presence of active ALK and is able to drive robust expression of the Lmd downstream target Vrp1 in the developing VM. Our results suggest that Lmd is a target of Jeb/Alk signalling in the VM of Drosophila embryos.

Plasminogen is a master regulator and a potential drug candidate for the healing of radiation wounds.

Around 95% of cancer patients undergoing radiotherapy experience cutaneous side effects, and some develop radiation wounds or fibrosis. Currently, there is no effective treatment for these indications. We show here that plasminogen administration enhanced the healing of radiation wounds via pleiotropic effects on gene expression. Using RNA sequencing, we found that plasminogen downregulated the expression of genes in the TLR, TNF, WNT, MAPK, and TGF-ß signaling pathways, and enhanced the anti-inflammatory effect of arachidonic acid, leading to significantly decreased inflammation and improved remodeling of granulation tissue compared with placebo treatment. In addition, plasminogen induced metabolic changes, including decreased glycolysis. Importantly, many of the factors downregulated by plasminogen are pro-fibrotic. Therefore, in radiation wounds with excessive inflammation, plasminogen is able to enhance and redirect the healing process, such that it more closely resembles physiological healing with significantly reduced risk for developing fibrosis. This makes plasminogen an attractive drug candidate for the treatment of radiation wounds in cancer patients.

Plasminogen activation is required for the development of radiation-induced dermatitis.

Skin damage caused by radiation therapy (radiodermatitis) is a severe side effect of radiotherapy in cancer patients, and there is currently a lack of effective strategies to prevent or treat such skin damage. In this work, we show with several lines of evidence that plasminogen, a pro-inflammatory factor, is key for the development of radiodermatitis. After skin irradiation in wild-type (plg+/+) mice, the plasminogen level increased in the irradiated area, leading to severe skin damage such as ulcer formation. However, plasminogen-deficient (plg-/-) mice and mice lacking plasminogen activators were mostly resistant to radiodermatitis. Moreover, treatment with a plasminogen inhibitor, tranexamic acid, decreased radiodermatitis in plg+/+ mice and prevented radiodermatitis in plg+/- mice. Together with studies at the molecular level, we report that plasmin is required for the induction of inflammation after irradiation that leads to radiodermatitis, and we propose that inhibition of plasminogen activation can be a novel treatment strategy to reduce and prevent the occurrence of radiodermatitis in patients.

Plasminogen is a critical regulator of cutaneous wound healing.

Wound healing is a complicated biological process that consist of partially overlapping inflammatory, proliferation and tissue remodelling phases. A successful wound healing depends on a proper activation and subsequent termination of the inflammatory phase. The failure to terminate the inflammation halts the completion of wound healing and is a known reason for formation of chronic wounds. Previous studies have shown that wound closure is delayed in plasminogen-deficient mice, and a role for plasminogen in dissection of extracellular matrix was suggested. However, our finding that plasminogen is transported to the wound by inflammatory cells early during the healing process, where it potentiates inflammation, indicates that plasminogen may also have other roles in the wound healing process. Here we report that plasminogen-deficient mice have extensive fibrin and neutrophil depositions in the wounded area long after re-epithelialisation, indicating inefficient debridement and chronic inflammation. Delayed formation of granulation tissue suggests that fibroblast function is impaired in the absence of plasminogen. Therefore, in addition to its role in the activation of inflammation, plasminogen is also crucial for subsequent steps, including resolution of inflammation and activation of the proliferation phase. Importantly, supplementation of plasminogen-deficient mice with human plasminogen leads to a restored healing process that is comparable to that in wild-type mice. Besides of being an activator of the inflammatory phase during wound healing, plasminogen is also required for the subsequent termination of inflammation. Based on these results, we propose that plasminogen may be an important future therapeutic agent for wound treatment.

Structural Insights into Polymorphic ABO Glycan Binding by Helicobacter pylori.

The Helicobacter pylori adhesin BabA binds mucosal ABO/Le(b) blood group (bg) carbohydrates. BabA facilitates bacterial attachment to gastric surfaces, increasing strain virulence and forming a recognized risk factor for peptic ulcers and gastric cancer. High sequence variation causes BabA functional diversity, but the underlying structural-molecular determinants are unknown. We generated X-ray structures of representative BabA isoforms that reveal a polymorphic, three-pronged Le(b) binding site. Two diversity loops, DL1 and DL2, provide adaptive control to binding affinity, notably ABO versus O bg preference. H. pylori strains can switch bg preference with single DL1 amino acid substitutions, and can coexpress functionally divergent BabA isoforms. The anchor point for receptor binding is the embrace of an ABO fucose residue by a disulfide-clasped loop, which is inactivated by reduction. Treatment with the redox-active pharmaceutic N-acetylcysteine lowers gastric mucosal neutrophil infiltration in H. pylori-infected Le(b)-expressing mice, providing perspectives on possible H. pylori eradication therapies.