ABSTRACT
In-situ Fourier transform infrared (FT-IR) spectroscopy has been recognized as an important technology for online monitoring of chemical reactions. However, analysis of the real-time IR data for identification and quantification of uncertain reactants or intermediates is often ambiguous and difficult. Here, we propose an analysis algorithm based on reaction kinetic modeling and the chemometric method of partial least squares (PLS) to comprehensively and quantitatively study reaction processes. Concentration profiles and apparent kinetic parameters can be simultaneously calculated from the spectral data, without the demand of complicated analysis on characteristic absorbance peaks or tedious sampling efforts for multivariate modeling. Paal-Knorr reactions and glyoxylic acid synthesis reactions were selected as typical reactions to validate the algorithm. A lack of fit of the Paal-Knorr reaction spectra was less than 2.5% at various conditions, and the absolute errors between the predicted values and HPLC measurement of glyoxylic acid synthesis were less than 6% during the reaction process. Moreover, the reaction kinetic models extracted from FT-IR data were used to simulate reaction processes and optimize the conditions in order to maximize product yields, which proved that this analysis method could be used for process optimization.
Subject(s)
Least-Squares Analysis , Spectroscopy, Fourier Transform Infrared/methods , Kinetics , Fourier AnalysisABSTRACT
Osteoarthritis (OA) is a major cause of joint pain and disability, resulting in large socioeconomic costs worldwide. Isofraxidin (ISO), a bioactive coumarin compound isolated from the functional foods Siberian ginseng and Apium graveolens, exerts anti-inflammatory effects in a variety of diseases. However, no studies have reported the protective effects of ISO against OA development. Accordingly, this study aimed to assess the therapeutic effect of ISO in human OA chondrocytes, and in a mouse model of OA induced by destabilisation of the medial meniscus (DMM). In vitro, lipopolysaccharide (LPS)-induced overproduction of nitric oxide (NO), prostaglandin E2 (PGE2), tumour necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) was decreased by ISO pre-treatment. Furthermore, ISO attenuated the increased expression of inflammatory enzymes, including inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), in response to LPS stimulation. Meanwhile, LPS-induced extracellular matrix (ECM) degradation was also reversed by ISO treatment. Mechanistically, ISO competitively inhibited Toll-like receptor 4 (TLR4)/myeloid differentiation protein-2 (MD-2) complex formation, and thus TLR4/nuclear factor kappa B (NF-κB) signalling cascades. In vivo, ISO treatment not only prevented the calcification and erosion of cartilage, as well as the thickening of subchondral bone, but also reduced the serum levels of inflammatory cytokines in the mouse OA model. Taken together, these data suggest that ISO has potential in the treatment of OA.
Subject(s)
Coumarins/pharmacology , Lymphocyte Antigen 96/metabolism , Osteoarthritis/drug therapy , Toll-Like Receptor 4/metabolism , Aged , Animals , Cartilage/drug effects , Cartilage/metabolism , Cell Differentiation/drug effects , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , Chondrocytes/drug effects , Chondrocytes/metabolism , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Dinoprostone/genetics , Dinoprostone/metabolism , Disease Models, Animal , Extracellular Matrix/genetics , Extracellular Matrix/metabolism , Female , Humans , Interleukin-6/genetics , Interleukin-6/metabolism , Lipopolysaccharides/toxicity , Lymphocyte Antigen 96/genetics , Male , Mice , Middle Aged , NF-kappa B/genetics , NF-kappa B/metabolism , Nitric Oxide/metabolism , Nitric Oxide Synthase Type II/genetics , Nitric Oxide Synthase Type II/metabolism , Osteoarthritis/genetics , Toll-Like Receptor 4/genetics , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolismABSTRACT
The Masquelet's induced membrane technique for repairing bone defects has been demonstrated to be a promising treatment strategy. Previous studies have shown that the vessel density of induced membrane is decreased in the late stage of membrane formation, which consequently disrupts the bone healing process. However, relatively little is known about certain mechanisms of vessel degeneration in the induced membrane tissue and whether promotion of angiogenesis in induced membranes can improve bone regeneration. Here, we showed that the Delta-like ligand 4/ Notch homolog 1 (Dll4/Notch1) pathway was relatively activated in the late stage of induced membrane, especially at the subcutaneous site. Then, DAPT, a classical γ-secretase inhibitor, was applied to specifically inhibit Notch1 activation, followed by up-regulation of vascular endothelial growth factor receptor 2 (VEGFR2) and CD31 expression. DAPT-modified induced membranes were further confirmed to contribute to bone regeneration after autogenous bone grafting. Finally, in vitro experiments revealed that knocking down Notch1 contributed to the functional improvement of endothelial progenitor cells (EPCs) and that DAPT-treated induced membrane tissue was more favorable for angiogenesis of EPCs compared with the vehicle group. In conclusion, the present findings demonstrate that Dll4/Notch1 signaling is negatively associated with the vessel density of induced membrane. Pharmacological inhibition of Notch1 attenuated the vessel degeneration of induced membrane both in vitro and in vivo, which consequently improved bone formation at the bone defect site and graft resorption at the subcutaneous site.
Subject(s)
Fractures, Bone/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Membrane Proteins/metabolism , Neovascularization, Physiologic , Orthopedic Procedures , Receptor, Notch1/metabolism , Animals , Fractures, Bone/pathology , Fractures, Bone/surgery , Male , Rats , Rats, Sprague-DawleyABSTRACT
Osteoarthritis (OA), which is characterized by progressive degradation of the articular cartilage, is the most prevalent form of human arthritis. Accumulating evidence has shown that polydatin (PD) exerts special biological functions in a variety of diseases. However, whether it protects against OA development has remained unknown. Here, we investigated the anti-inflammatory and chondroprotective effects of PD on interleukin (IL)-1ß-induced human osteoarthritic chondrocytes and in the surgical destabilization of medial meniscus mouse (DMM) OA models. In vitro, PD treatment completely suppressed the over-production of pro-inflammatory mediators, including prostaglandin E2 (PGE2), tumor necrosis factor-α (TNF-α), nitric oxide (NO), cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and IL-6 in IL-1ß-induced human OA chondrocytes. Moreover, PD exerted a suppressive effect on the expression of matrix-degrading proteases, including matrix metalloproteinase 13 (MMP13) and thrombospondin motifs 5 (ADAMTS-5), which leads to the degradation of the extracellular matrix (ECM). Meanwhile, specific inhibition of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) level by short-interfering RNA (siRNA) strongly reversed the anti-inflammatory and chondroprotective effects of PD in human OA chondrocytes. The protective effects of PD were also observed in vivo. In conclusion, our studies demonstrate that PD holds novel therapeutic potential for the development of OA.
