Nanofat lysate ameliorates pain and cartilage degradation of osteoarthritis through activation of TGF-β–Smad2/3 signaling of chondrocytes

Introduction: Nanofat is an effective cell therapy for osteoarthritis (OA). However, it has clinical limitations due to its short half-life. We developed Nanofat lysate (NFL) to overcome the defect of Nanofat and explore its anti-OA efficacy and mechanism.Methods: Monoiodoacetate (MIA) was employed to establish rat OA model. For pain assessment, paw withdrawal latency (PWL) and thermal withdrawal latency (TWL) were evaluated. Degeneration of cartilage was observed by histopathological and immunohistochemical examination. Primary chondrocytes were treated with TNF-α to establish the cellular model of OA. MTT, wound healing, and transwell assays were performed to assess effects of NFL on chondrocytes. RNA-seq, qPCR and Western blot assays were conducted to clarify the mechanism of NFL.Results and Discussion: The animal data showed that PWL and TWL values, Mankin’s and OARSI scorings, and the Col2 expression in cartilage were significantly improved in the NFL-treated OA rats. The cellular data showed that NFL significantly improved the proliferation, wound healing, and migration of chondrocytes. The molecular data showed that NFL significantly restored the TNF-α-altered anabolic markers (Sox9, Col2 and ACAN) and catabolic markers (IL6 and Mmp13). The RNA-seq identified that TGF-β-Smad2/3 signaling pathway mediated the efficacy of NFL, which was verified by qPCR and Western blot that NFL significantly restored the abnormal expressions of TGFβR2, phosphorylated-Smad2, phosphorylated-Smad2/3, Col2, Mmp13 and Mmp3. After long-term storage, NFL exerted similar effects as its fresh type, indicating its advantage of storability. In sum, NFL was developed as a new therapeutic approach and its anti-OA efficacy and mechanism that mediated by TGF-β-Smad2/3 signaling was determined for the first time. Besides, the storability of NFL provided a substantial advantage than other living cell-based therapies

Urine Metabolomics Profiling of Lumbar Disc Herniation and its Traditional Chinese Medicine Subtypes in Patients Through Gas Chromatography Coupled With Mass Spectrometry

Lumbar disc herniation (LDH) possesses complex pathogenesis, which has not been well elucidated yet. To date, specific or early diagnosis of LDH remains unavailable, resulting in missed opportunity for effective treatment. According to Traditional Chinese medicine (TCM) theory, LDH can be divided into two subtypes (reality syndrome and deficiency syndrome). The purpose of this study was to analyze the metabolic disorders of LDH and its TCM subtypes and screen out potential biomarkers for LDH diagnosis. Gas chromatography coupled with mass spectrometry (GC-MS) was applied to test the urine samples from 66 participants (30 healthy volunteers, 18 LDH patients with deficiency syndrome and 18 patients with reality syndrome). PCA analysis showed a distinct separation tendency between the healthy subjects and LDH patients but no obvious separation between the different syndromes (reality syndrome and deficiency syndrome) of LDH patients. As a result, 23 metabolites were identified significantly altered in the LDH patients, as compared with the healthy subjects. The altered metabolites belong to amino acid metabolism, nucleic acid metabolism, carbohydrate metabolism, and vitamin metabolism, which are related to osteoporosis and inflammation. Our results indicate metabolic disorders of LDH and thereby propose a group of metabolic biomarkers for potential application in early diagnosis of LDH in clinic, which provide a reasonable explanation for the pathogenesis of LDH

Intra-Articular Injection of Fructus Ligustri Lucidi Extract Attenuates Pain Behavior and Cartilage Degeneration in Mono-Iodoacetate Induced Osteoarthritic Rats

Fructus Ligustri Lucidi (FLL) has been widely used as a traditional Chinese medicine (TCM) for treating soreness and weakness of waist and knees. It has potential for treating OA owing to its kidney-tonifying activity with bone-strengthening effects, but there is so far no report of its anti-OA effect. This study established a rat OA model by intra-articular (IA) injection of mono-iodoacetate (1.5 mg) and weekly treated by IA administration of FLL at 100 μg/mL for 4 weeks. Thermal withdrawal latency, mechanical withdrawal threshold, and spontaneous activity were tested for evaluation of pain behavior, and histopathological (HE, SO, and ABH staining) and immunohistochemical (Col2, Col10, and MMP13) analyses were conducted for observation of cartilage degradation. In vitro effect of FLL on chondrocytes was evaluated by MTT assay and qPCR analysis. Moreover, HPLC analysis was performed to determine its chemoprofile. The pain behavioral data showed that FLL attenuated joint pain hypersensitivity by increasing thresholds of mechanical allodynia and thermal hyperalgesia as well as spontaneous activity. The histopathological result showed that FLL reversed OA cartilage degradation by protecting chondrocytes and extracellular matrix in cartilage, and the immunohistochemical analysis revealed its molecular actions on protein expressions of MMP13, Col2, and Col10 in cartilage. The MTT assay showed its proliferative effects on chondrocytes, and qPCR assay clarified its mechanism associated with gene expressions of Mmp13, Col2, Col10, Adamts5, Aggrecan, and Runx2 in TNF-α treated chondrocytes. Our results revealed an anti-OA effect of FLL on pain behavior and cartilage degradation in OA rats and clarified a molecular mechanism in association with the suppression of chondrocyte hypertrophy and catabolism. IA FLL can be regarded as novel and promising option for OA therapy

Tonic Herbs and Herbal Mixtures in Chinese Medicine

This review results from a PubMed-based data-mining of scientific literature concerning typical tonic herbs and formulas of Chinese herbal medicine and their application principle under the theory of CM. We have focused on two aspects of tonic activity: body tonification against qi气-, blood血-, yin阴- and yang阳-deficiency, respectively, and organ (Zang 脏and Fu腑) tonification against heart-, liver-, spleen/stomach-, lung-, and kidney-deficiency, respectively. Body-tonifying herbs are: Astragalus Membranaceus Radix (Huang-qi黄芪) and Panax Ginseng Radix (Ren-shen人参) for qi-tonification; Angelicae Sinensis Radix (Dang-gui当归) and Rehmannia Glutinosa Radix Preparata (Shu-di熟地) for blood-tonification; Ophiopogon Japonicus Radix (Mai-dong麦冬) and Scrophularia Ningpoensis Radix (Xuan-shen玄参) for yin-deficiency; Myristica Fragrans Semen (Rou-dou-kou肉豆蔻) and Psoralea Corylifolia Fructus (Bu-gu-zhi补骨脂) for yang-deficiency. The corresponding CM formulas are: Bu-zhong-yi-qi decoction (补中益气汤) for qi-tonification; Si-wu decoction (四物汤) for blood-tonification; Zeng-ye decoction (增液汤) for yin-tonification; Si-shen pill (四神丸) for yang-tonification. Organ-tonifying herbs are: Glycyrrhizae Uralensis Radix Preparata (Zhi-gan-cao炙甘草) and Rehmannia Glutinosa Radix (Di-huang地黄) for heart-tonification; Lycium Barbarum Fructus (Gou-qi-zi枸杞子) and Angelicae Sinensis Radix (Dang-gui当归) for liver-tonification; Panax Ginseng Radix (Ren-shen人参) and Atractylodis Macrocephala Rhizoma (Bai-zhu白术) for spleen/stomach-tonification; Panax Ginseng Radix (Ren-shen人参) and Astragalus Membranaceus Radix (Huang-qi黄芪) for lung-tonification; Cornus Officinalis Fructus (Shan-zhu-yu山茱萸) and Dioscorea Opposite Rhizoma (Shan-yao山药) for kidney-tonification. The corresponding CM formulas are: Zhi-gan-cao decoction (炙甘草汤) for heart-tonification; Yi-guan decoction (一贯煎) for liver-tonification; Shen-ling-bai-zhu powder (参苓白术散) for spleen/stomach-tonification; Bu-fei decoction (补肺汤) for lung-tonification; Liu-wei-di-huang pill (六味地黄丸) for kidney tonification. These herbs and formulas were described regarding their efficacy, reasonable use and unreasonable abuse specific to different patients with different symptoms. The scientific investigation on efficacy and safety of Chinese formulas will propel the acceptance and spread of TCM in the western world for the sake of patients worldwide

Verbascoside attenuates experimental varicocele-induced damage to testes and sperm levels through up-regulation of the hypothalamus-pituitary-gonadal (HPG) axis

Context Verbascoside (VB), which is found in many medicinal plant families, exhibits biological activities in various diseases. However, its effects on varicocele (VCL)-induced damage remain unknown. Objective To investigate the effects and mechanism of VB on experimental rats with varicocele (VCL)-induced damage. Materials and methods Sixty sexually mature male Sprague-Dawley (SD) rats were divided into six groups (n = 10): control, control-sham, VCL-vehicle (normal saline), and VCL + VB groups (50, 100, and 200 mg/kg/day, intraperitoneally). After 4 weeks of VB treatment, all animals were sacrificed, and the body and testicular weight, sperm quality parameters, histopathology, antioxidant status, and hormone levels were tested. The levels of gonadotropin-releasing hormone (GnRH) and gonadotropin-inhibitory hormone in the hypothalamus were detected by western blot. Results Compared with the VCL-vehicle group (41.14%), administration of VB significantly increased the sperm viability (59.29, 65.45, 84.93%). VB groups showed higher Johnson’s score (3.57 ± 0.15, 4.71 ± 0.26, 7.93 ± 0.37) than VCL-vehicle group (2.72 ± 0.24). Antioxidant status and hormone levels alterations were also observed. Meanwhile, the mean number of apoptotic tubules (8.15 ± 0.96, 6.61 ± 1.05, 2.17 ± 0.08) and apoptotic index showed a marked decrease. Compared with the VCL-vehicle group (0.21 ± 0.09), the VB groups (0.36 ± 0.07, 0.42 ± 0.06, 0.88 ± 0.10) showed considerable increases in GnRH. Discussion and conclusions VB has protective effects on reproductive organs and VB may be therapeutically useful in the treatment of varicocele through up-regulation of the HPG axis

Selection of safe artemisinin derivatives using a machine learning-based cardiotoxicity platform and in vitro and in vivo validation

The majority of drug candidates fails the approval phase due to unwanted toxicities and side effects. Establishment of an effective toxicity prediction platform is of utmost importance, to increase the efficiency of the drug discovery process. For this purpose, we developed a toxicity prediction platform with machine-learning strategies. Cardiotoxicity prediction was performed by establishing a model with five parameters (arrhythmia, cardiac failure, heart block, hypertension, myocardial infarction) and additional toxicity predictions such as hepatotoxicity, reproductive toxicity, mutagenicity, and tumorigenicity are performed by using Data Warrior and Pro-Tox-II software. As a case study, we selected artemisinin derivatives to evaluate the platform and to provide a list of safe artemisinin derivatives. Artemisinin from Artemisia annua was described first as an anti-malarial compound and later its anticancer properties were discovered. Here, random forest feature selection algorithm was used for the establishment of cardiotoxicity models. High AUC scores above 0.830 were achieved for all five cardiotoxicity indications. Using a chemical library of 374 artemisinin derivatives as a case study, 7 compounds (deoxydihydro-artemisinin, 3-hydroxy-deoxy-dihydroartemisinin, 3-desoxy-dihydroartemisinin, dihydroartemisinin-furano acetate-d3, deoxyartemisinin, artemisinin G, artemisinin B) passed the toxicity filtering process for hepatotoxicity, mutagenicity, tumorigenicity, and reproductive toxicity in addition to cardiotoxicity. Experimental validation with the cardiomyocyte cell line AC16 supported the findings from the in silico cardiotoxicity model predictions. Transcriptomic profiling of AC16 cells upon artemisinin B treatment revealed a similar gene expression profile as that of the control compound, dexrazoxane. In vivo experiments with a Zebrafish model further substantiated the in silico and in vitro data, as only slight cardiotoxicity in picomolar range was observed. In conclusion, our machine-learning approach combined with in vitro and in vivo experimentation represents a suitable method to predict cardiotoxicity of drug candidates

In Vivo Cardiotoxicity Induced by Sodium Aescinate in Zebrafish Larvae

Sodium aescinate (SA) is a widely-applied triterpene saponin product derived from horse chestnut seeds, possessing vasoactive and organ-protective activities with oral or injection administration in the clinic. To date, no toxicity or adverse events in SA have been reported, by using routine models (in vivo or in vitro), which are insufficient to predict all aspects of its pharmacological and toxicological actions. In this study, taking advantage of transparent zebrafish larvae (Danio rerio), we evaluated cardiovascular toxicity of SA at doses of 1/10 MNLC, 1/3 MNLC, MNLC and LC10 by yolk sac microinjection. The qualitative and quantitative cardiotoxicity in zebrafish was assessed at 48 h post-SA treatment, using specific phenotypic endpoints: heart rate, heart rhythm, heart malformation, pericardial edema, circulation abnormalities, thrombosis and hemorrhage. The results showed that SA at 1/10 MNLC and above doses could induce obvious cardiac and pericardial malformations, whilst 1/3 MNLC and above doses could induce significant cardiac malfunctions (heart rate and circulation decrease/absence), as compared to untreated or vehicle-treated control groups. Such cardiotoxic manifestations occurred in more than 50% to 100% of all zebrafish treated with SA at MNLC and LC10. Our findings have uncovered the potential cardiotoxicity of SA for the first time, suggesting more attention to the risk of its clinical application. Such a time- and cost-saving zebrafish cardiotoxicity assay is very valid and reliable for rapid prediction of compound toxicity during drug research and development

Autologous Stem Cells Combined Core Decompression for Treatment of Avascular Necrosis of the Femoral Head: A Systematic Meta-Analysis

Objective. This study aims to systematically evaluate the efficacy and safety of core decompression combined transplantation of autologous bone marrow stem cells (CDBMSCs) for treatment of avascular necrosis of the femoral head (ANFH). Methods. Randomized controlled trials (RCTs) regarding effectiveness of core decompression combined transplantation of autologous bone marrow stem cells for treating ANFH were searched in 8 comprehensive databases prior to September 2016. The data analysis was performed by using the RevMan version 5.3. Results. A total of 11 studies with 507 participants were included. Results showed that CDBMSCs group was more effective than CD group in increasing Harris hip score, decreasing necrotic area of femoral head, collapse of femoral head, and conversion to total hip replacement incidence. In the subgroup analysis, the results did not change in different intervention measure substantially. In addition, the safety of CDBMSCs for ANFH is reliable. Conclusion. Based on the systematic review, our findings suggest that core decompression combined transplantation of autologous bone marrow stem cells appeared to be more efficacious in the treatment at early stages of ANFH

Detrimental alteration of mesenchymal stem cells by an articular inflammatory microenvironment results in deterioration of osteoarthritis

Abstract Background Articular injection of mesenchymal stem cells (MSCs) has been applied to treat knee osteoarthritis (kOA), but its clinical outcomes are controversial. This study investigated whether an articular inflammatory microenvironment (AIM) impacts MSC-based therapy in a rat model of kOA. Methods The biological change of MSCs and the functional change of MSCs on chondrocytes were evaluated under AIM. The key mediator and mechanism for the AIM impact on MSC therapy were explored via gain- and loss-of-function approaches. Results The results showed that MSCs exerted potent anti-kOA effects in vivo and in vitro, but that this therapy become chondrodestructive if a chronic AIM was present. Mechanistically, the overexpression of MMP13 in the injected MSCs via a MAPKs-AP1 signaling axis was revealed as the underlying mechanism for the detriment outcome. Conclusions This study thus clarifies recent clinical findings while also suggesting a means to overcome any detrimental effects of MSC-based therapy while improving its efficacy

Identification of Gedunin from a Phytochemical Depository as a Novel Multidrug Resistance-Bypassing Tubulin Inhibitor of Cancer Cells

The chemotherapy of tumors is frequently limited by the development of resistance and severe side effects. Phytochemicals may offer promising candidates to meet the urgent requirement for new anticancer drugs. We screened 69 phytochemicals, and focused on gedunin to analyze its molecular modes of action. Pearson test-base correlation analyses of the log10IC50 values of 55 tumor cell lines of the National Cancer Institute (NCI), USA, for gedunin with those of 91 standard anticancer agents revealed statistically significant relationships to all 10 tested microtubule inhibitors. Thus, we hypothesized that gedunin may be a novel microtubule inhibitor. Confocal microscopy, cell cycle measurements, and molecular docking in silico substantiated our assumption. Agglomerative cluster analyses and the heat map generation of proteomic data revealed a subset of 40 out of 3171 proteins, the expression of which significantly correlated with sensitivity or resistance for the NCI cell line panel to gedunin. This indicates the complexity of gedunin’s activity against cancer cells, underscoring the value of network pharmacological techniques for the investigation of the molecular modes of drug action. Finally, we correlated the transcriptome-wide mRNA expression of known drug resistance mechanism (ABC transporter, oncogenes, tumor suppressors) log10IC50 values for gedunin. We did not find significant correlations, indicating that gedunin’s anticancer activity might not be hampered by classical drug resistance mechanisms. In conclusion, gedunin is a novel microtubule-inhibiting drug candidate which is not involved in multidrug resistance mechanisms such as other clinically established mitotic spindle poisons