Rabbit Anti-Human TBK1 monoclonal antibody, clone 20I26M24

beta-cell proliferation induction is a promising therapeutic strategy to restore beta-cell mass. By screening small molecules in a transgenic zebrafish model of type 1 diabetes, we identified inhibitors of noncanonical I kappa B kinases (IKKs), TANK-binding kinase 1 (TBK1) and I kappa B kinase epsilon (IKK epsilon), as enhancers of beta-cell regeneration. The most potent beta-cell regeneration enhancer was a cinnamic acid derivative (E)-3-(3-phenylbenzo[c] isoxazol-5-yl) acrylic acid (PIAA), which, acting through the cAMP-dependent protein kinase A (PKA), stimulated beta-cell-specific proliferation by increasing cyclic AMP (cAMP) levels and mechanistic target of rapamycin (mTOR) activity. A combination of PIAA and cilostamide, an inhibitor of beta-cell-enriched cAMP hydrolyzing enzyme phosphodiesterase (PDE) 3, enhanced beta-cell proliferation, whereas overexpression of PDE3 blunted the mitogenic effect of PIAA in zebrafish. PIAA augmented proliferation of INS-1 beta-cell and beta-cell in mammalian islets including human islets with elevation in cAMP levels and insulin secretion. PIAA improved glycemic control in streptozotocin (STZ)-induced diabetic mice with increases in beta-cell proliferation, beta-cell area, and insulin content in the pancreas. Collectively, these data reveal an evolutionarily conserved and critical role of TBK1/IKK epsilon suppression in expanding functional beta-cell mass.

Ethnopharmacological relevance: Flos Trollii is the dried flowers of Trollius chinensis. It has been used as a traditional herbal medicine for the treatment of upper respiratory tract infection, tonsillitis and pharyngitis in China for a long history. Veratric acid, vitexin, and trolline are the representative compounds of phenolic acids, flavonoids and alkaloids in this herbal medicine. All of these three compounds show antiviral activity which is related to the efficacy of Flos Trollii. Aim of the study: To investigate the anti-influenza A virus mechanism of the three representative compounds from the perspective of regulating TLRs signaling pathways, so as to understand the relevant efficacy of Flos Trollii. Materials and methods: Influenza A virus A/FM/1/47 (H1N1) and mouse peritoneal macrophages (RAW264.7) were used in the whole process of investigation. MTT assay was conducted to select the appropriate experimental concentrations of the three compounds on RAW264.7 cells. Western blot, RT-PCR, and ELISA assays were performed to determine the protein and mRNA expression of key factors and related inflammatory factors of TLRs signaling pathways. Griess method was employed to detect the production of NO. Results: The three representative compounds reduced the inflammatory factors including NO, IL-6, and TNF-alpha and enhanced the production of IFN-beta through dynamically regulating the TLRs 3, 4 and 7 pathways. Veratric acid significantly down-regulated the protein expression of TLR3 and IRF3 as well as the mRNA expression of TBK1 and TRIF. Vitexin significantly down-regulated the protein expression of TBK1 and IRF3 as well as the mRNA expression of TLR3, TBK1, TRIF and IRF3 while up-regulated the protein expression of TLR4 and IKK alpha. Trolline significantly down-regulated the protein expression of TLR7 whereas significantly up-regulated the protein expression of TLR4, IKK alpha and TAK1. Conclusions: The three representative compounds from Flos Trollii play their parts in anti-H1N1 viral effect through partially down-regulating TLRs 3 and 7 pathways and up-regulating TLR4 pathway. They counteract the inflammatory injury caused by excessive production of NO, IL-1, IL-6, and TNF-alpha induced by virus infection and enhance the production of IFN-beta so as to eliminate the virus.