Locally Delivered Metabolite Derivative Promotes Bone Regeneration in Aged Mice.

Repair of large bone defects is still a major challenge, especially for the aged population. One alternative to address this issue is using the biomaterial-mediated bone morphogenetic protein 2 (BMP2) delivery technique, although high-dose BMP2 can cause serious concerns. α-Ketoglutarate (AKG) is a key intermediate in the tricarboxylic acid cycle and emerging as an intriguing antiaging molecule to extend the life/health span in different organisms. While one recent study indicates that the dietary AKG could significantly reduce bone loss and improve bone anabolism in aged mice, the therapeutic potential of AKG for bone regeneration has not been studied so far. Moreover, the poor cell permeability, large dose requirement, and long-term systemic administration of AKG hinder its applications in clinics and cellular mechanism studies. Dimethyl α-ketoglutarate (DMAKG) is a cell-permeable derivative of AKG with promising potential, although its role in osteogenesis is still elusive. Therefore, we aim to study the potential roles of DMAKG for bone regeneration using both in vitro cell culture and in vivo aged mouse models. Compared to AKG, our data indicated that DMAKG could more effectively improve osteoblastic differentiation. In addition, DMAKG significantly reduced adipogenic differentiation and improved osteogenic differentiation of a mouse multipotential mesenchymal stem cell line. Importantly, our result indicated that DMAKG significantly promoted BMP2-induced osteoblastic differentiation and mineralization in vitro. Moreover, DMAKG could not only significantly mitigate lipopolysaccharide (LPS)-stimulated inflammation in macrophages but also largely rescue LPS-inhibited osteoblastic differentiation. Consistently, our in vivo study demonstrated that gelatin scaffold-mediated local release of DMAKG significantly promoted BMP2-induced bone regeneration in aged mice, which is compromised by chronic inflammation and high adipogenesis. Overall, we, for the first time, report that locally delivered metabolite derivative, DMAKG, could improve BMP2-induced bone regeneration in aged mice. Our study suggests DMAKG has a promising therapeutic potential for bone regeneration through modulating local inflammation and stem cell differentiation.

High Molecular Weight Poly(glutamic acid) to Improve BMP2-Induced Osteogenic Differentiation.

FDA-approved bone morphogenetic protein 2 (BMP2) has serious side effects due to the super high dose requirement. Heparin is one of the most well-studied sulfated polymers to stabilize BMP2 and improve its functionality. However, the clinical use of heparin is questionable because of its undesired anticoagulant activity. Recent studies suggest that poly(glutamic acid) (pGlu) has the potential to improve BMP2 bioactivity with less safety concerns; however, the knowledge on pGlu's contribution remains largely unknown. Therefore, we aimed to study the role of pGlu in BMP2-induced osteogenesis and its potential application in bone tissue engineering. Our data, for the first time, indicated that both low (L-pGlu) and high molecular weight pGlu (H-pGlu) were able to significantly improve the BMP2-induced early osteoblastic differentiation marker (ALP) in MC3T3-E1 preosteoblasts. Importantly, the matrix mineralization was more rapidly enhanced by H-pGlu compared to L-pGlu. Additionally, our data indicated that only α-H-pGlu could significantly improve BMP2's activity, whereas γ-H-pGlu failed to do so. Moreover, both gene expression and mineralization data demonstrated that α-H-pGlu enabled a single dose of BMP2 to induce a high level of osteoblastic differentiation without multiple doses of BMP2. To study the potential application of pGlu in tissue engineering, we incorporated the H-pGlu+BMP2 nanocomplexes into the collagen hydrogel with significantly elevated osteoblastic differentiation. Furthermore, H-pGlu-coated 3D porous gelatin and chitosan scaffolds significantly enhanced osteogenic differentiation through enabling sustained release of BMP2. Thus, our findings suggest that H-pGlu is a promising new alternative with great potential for bone tissue engineering applications.

Development and assessment of machine learning models for predicting recurrence risk after endovascular treatment in patients with intracranial aneurysms.

Intracranial aneurysms (IAs) remain a major public health concern and endovascular treatment (EVT) has become a major tool for managing IAs. However, the recurrence rate of IAs after EVT is relatively high, which may lead to the risk for aneurysm re-rupture and re-bleed. Thus, we aimed to develop and assess prediction models based on machine learning (ML) algorithms to predict recurrence risk among patients with IAs after EVT in 6 months. Patient population included patients with IAs after EVT between January 2016 and August 2019 in Hunan Provincial People's Hospital, and an adaptive synthetic (ADASYN) sampling approach was applied for the entire imbalanced dataset. We developed five ML models and assessed the models. In addition, we used SHapley Additive exPlanations (SHAP) and local interpretable model-agnostic explanation (LIME) algorithms to determine the importance of the selected features and interpret the ML models. A total of 425 IAs were enrolled into this study, and 66 (15.5%) of which recurred in 6 months. Among the five ML models, gradient boosting decision tree (GBDT) model performed best. The area under curve (AUC) of the GBDT model on the testing set was 0.842 (sensitivity: 81.2%; specificity: 70.4%). Our study firstly demonstrated that ML-based models can serve as a reliable tool for predicting recurrence risk in patients with IAs after EVT in 6 months, and the GBDT model showed the optimal prediction performance.

Fluorometric determination of mercury(II) via a graphene oxide-based assay using exonuclease III-assisted signal amplification and thymidine-Hg(II)-thymidine interaction.

A highly sensitive and selective fluorometric method is described for determination of mercury(II). It is based on (a) the use of graphene oxide (GO) acting as a quencher of the fluoresence of the carboxy-fluorescein (FAM), and (b) of Hg(II)-triggered cleavage of the newly formed nucleic acid sequences harbored blunt 3'-hydroxyl termini by exonuclease III (Exo III) that leads to signal amplification. Two DNA probes are used, viz. a capture probe (CP) and a help probe; HP) that is partially complementary. In the absence of Hg(II), the FAM-labeled hairpin (signal probe, SP) is adsorbed onto the surface of GO via π-stacking interactions. CP blocks the release of the HP for binding to SP. This results in quenching of the green fluorescence of the label. Upon addition of Hg(II), the linear structure of CP is converted to a hairpin structure due to the formation of thymidine-Hg(II)-thymidine duplexes. HP is released from the CP/HP hybrids, and this causes SP to be released from from GO and fluorescence to be recovered. The signal is strongly amplified by using Exo III-assisted targeting and recycling of HP. Hence, Hg(II) can be detected via the strong increase in fluorescence. The method has a linear response in the 0.1 to 30 nM Hg(II) concentration range and a 10 pM detection limit. It was applied to the determination of Hg(II) in three (spiked) Chinese medicines. Graphical abstract Schematic representation of fluorescence sensing strategy for Hg2+ by using graphene oxide as a quencher and exonuclease III-assisted signal amplification.

[Effects of crocin on hippocampus rapid kindling epilepsy in mice].

Objective: To investigate the effect of crocin on the progression and generalized seizure of temporal lobe epilepsy in mice. Methods: Hippocampus rapid kindling model was established in C57BL/6J mice. The effects of crocin on seizure stage, afterdischarge duration (ADD), number of stimulation in each stage and final state, the incidence of generalized seizure (GS), average seizure stage and ADD were observed. Results: Crocin (20 mg/kg) significantly retarded behavioral seizure stages ( P<0.05) and shortened cumulative ADD ( P<0.01) during hippocampus rapid kindling acquisition in mice compared with vehicle group. Meanwhile, number of stimulations in stage 1-2 was significantly increased ( P<0.05) and the incidence of fully kindled animals was significantly decreased ( P<0.01). However, 10 or 50 mg/kg crocin showed no significant effect on the above indexes (all P>0.05). Crocin (100 or 200 mg/kg) significantly decreased the incidence of GS (all P<0.01) and reduced average seizure stages (all P<0.01) in fully-kindled mice compared with vehicle group; Fifty mg/kg crocin only reduced average seizure stages ( P<0.05). Conclusion: Low-dose crocin can retard the progression in hippocampus rapid kindling acquisition in mice, while high-dose crocin relieves the GS in fully-kindled mice, which suggests that crocin may be a potential anti-epileptic compound.

Rapid and undamaged analysis of crude and processed Radix Scrophulariae by Fourier transform infrared spectroscopy coupled with soft independent modeling of class analogy.

OBJECTIVE: The main objective of this work is to determine the feasibility of identification of crude and processed Radix Scrophulariae using the Fourier transform infrared spectroscopy couple with soft independent modeling of class analogy (FT-IR-SIMCA). MATERIALS AND METHODS: A total of 50 different crude Radix Scrophulariae was used to product processed ones. The spectra were acquired by FT-IR spectroscopy using a diffuse reflectance fiber optic probe. For the multivariate analysis, SIMCA was used. Results showed that FT-IR-SIMCA was useful to discriminate the processed Radix Scrophulariae samples from crude samples. These samples could be successfully classified by SIMCA. RESULTS: In all cases, the recognition and rejection rates were 97.8% and 100%, respectively. When testing with the blind sample that was picked out from the chosen samples, the accuracy was up to 90%. CONCLUSION: It means that the methodology is capable of accurately separating processed Radix Scrophulariae from crude samples.

Analysis of the influence of processing of stir-baking with glycyrrhizae on the main components of Euodiae Fructus by high-performance liquid chromatography with diode array detector.

Euodiae Fructus is one of the most commonly used Chinese herbs in China. Specifically, the crude Euodiae Fructus and its processed products of Gancao Zhi Pin are used clinically for the treatment of different diseases. In order to improve the quality control standard and evaluate the crude and processed Euodiae Fructus, in this study, a simple and sensitive high-performance liquid chromatography-diode array detector method was developed for the simultaneous determination of five major compounds in Euodiae Fructus. The results indicated that the five components had significant linear relation (r(2) ≥ 0.9997) between the peak area and the injected concentration. The average recoveries of the five components were in the range from 97.38% to 102.56%. Overall intra- and inter-day variations were less than 1.36%. The developed method can be applied to the intrinsic quality control of crude and processed Euodiae Fructus.

[Effect of nano-liposome sustained elemene in inducing cell apoptosis of C6 glioma].

OBJECTIVE: To study the effect of nano-liposome sustained elemene in inducing cell apoptosis of C6 glioma and to explore its influence on the expression of caspase-3 gene. METHODS: C6 glioma cells were cultured in medium with the same amount of nano-liposome sustained elemene and common elemene respectively, also in blank medium for control. The status of cell apoptosis was determined by flow cytometry at 0, 48 h and 72 h, and the expression of Caspase-3 protein was measured simultaneously by immunohistochemistry assay. RESULTS: Marked apoptosis presented in cells cultured in the medium with nano-liposome sustained elemene or common elemene at 48 h and 72 h, with the apoptotic rate significantly higher than that in the control. At the same time, Caspase-3 protein expression raised significantly in cells cultured in medium with either kinds of elemene, showing significant difference when compared with that in the control. CONCLUSION: Elemene has significant apoptosis promoting and Caspase-3 protein expression inducing effect on C6 glioma cells, which could be facilitated by nano-liposome bearing.