Staphylococcus aureus (SAU) stands as the prevailing pathogen in post-traumatic infections, with the emergence of antibiotic resistance presenting formidable treatment hurdles. The pressing need is ...to explore novel antibiotics to address this challenge. ShangKeHuangShui (SKHS), a patented traditional Chinese herbal formula, has gained widespread use in averting post-traumatic infections, but its biological effects remain incomplete understanding. This study’s primary objective was to delve into the antibacterial properties, potential antibacterial compounds within SKHS, and their associated molecular targets. In vitro SKHS antibacterial assays demonstrated that the minimum inhibitory concentration (MIC) was 8.625 mg/mL and the minimum bactericide concentration (MBC) was 17.25 mg/mL. Proteomic analysis based on tandem mass tag (TMT) showed significant changes in the expression level of 246 proteins in SKHS treated group compared to control group, with 79 proteins upregulated and 167 proteins downregulated (>1.5-fold, p < 0.05). Subsequently, thirteen target proteins related to various biological processes and multiple metabolic pathways were selected to conduct parallel reaction monitoring (PRM) and molecular docking screen. In protein tyrosine phosphatase PtpA (ptpA) docking screening, phellodendrine and obacunone can bind to ptpA with the binding energy of − 8.4 and − 8.3 kcal/mol, respectively. This suggests their potential impact on antibacterial activity by modulating the two-component system of SAU. The discovery lays a groundwork for future research endeavors for exploring new antibacterial candidates and elucidating specific active chemical components within SKHS that match target proteins. Further investigations are imperative to unveil the biological effects of these monomers and their potential synergistic actions.
Shang-Ke-Huang-Shui (SKHS) is a classic traditional Chinese medicine formula originally from the southern China city of Foshan. It has been widely used in the treatment of osteoarthritis (OA) but ...underlying molecular mechanisms remain unclear.
Recently, activation of C-X-C chemokine receptor type 4 (CXCR4) signaling has been reported to induce cartilage degradation in OA patients; therefore, inhibition of CXCR4 signaling has becoming a promising approach for OA treatment. The aim of this study was to validate the cartilage protective effect of SKHS and test whether the anti-OA effects of SKHS depend on its inhibition on CXCR4 signaling. Additionally, CXCR4 antagonist in SKHS should be identified and its anti-OA activity should also be tested in vitro and in vivo.
The anti-OA effects of SKHS and the newly identified CXCR4 antagonist was evaluated by monosodium iodoacetate (MIA)-induced rats. The articular cartilage surface was examined by hematoxylin and eosin (H&E) staining and Safranin O-Fast Green (S–F) staining whereas the subchondral bone was examined by micro-CT. CXCR4 antagonist screenings were conducted by molecular docking and calcium response assay. The CXCR4 antagonist was characterized by UPLC/MS/MS. The bulk RNA-Seq was conducted to identify CXCR4-mediated signaling pathway. The expression of ADAMTS4,5 was tested by qPCR and Western blot.
SKHS protected rats from MIA-induced cartilage degradation and subchondral bone damage. SKHS also inhibited CXCL12-indcued ADAMTS4,5 overexpression in chondrocytes through inhibiting Akt pathway. Coptisine has been identified as the most potent CXCR4 antagonist in SKHS. Coptisine reduced CXCL12-induced ADAMTS4,5 overexpression in chondrocytes. Furthermore, in MIA-induced OA model, the repaired cartilage and subchondral bone were observed in the coptisine-treated rats.
We first report here that the traditional Chinese medicine formula SKHS and its predominate phytochemical coptisine significantly alleviated cartilage degradation as well as subchondral bone damage through inhibiting CXCR4-mediated ADAMTS4,5 overexpression. Together, our work has provided an important insight of the molecular mechanism of SKHS and coptisine for their treatment of OA.
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•Shang-Ke-Huang-Shui (SKHS), a traditional Chinese medicine formula, protects rats from MIA-induced cartilage degradation.•Coptisine has been identified as the most potent CXCR4 antagonist in SKHS.•Coptisine inhibits ADAMTS4,5 overexpression in chondrocytes through inhibiting Akt signaling pathway.•Coptisine alleviates symptoms of OA and reduces ADAMTS4,5 expression in MIA-induced rats.
Abstract Rosiglitazone (Rosi) is a drug in the thiazolidinedione class for treatment of type 2 diabetes mellitus (T2DM), which binds and activates PPARγ nuclear receptor in fat cells, sensitizing ...them to insulin. Despite proven antidiabetic efficacy, Rosi therapy may be associated with trabecular bone loss and an increased risk of fractures. To examine the potential side effects of Rosi treatment on bone formation, we delivered Rosi to mice using a combined model of distraction osteogenesis (DO) and type 2 diabetes mellitus (T2DM). DO provides a unique method to isolate the sequence of intramembranous bone formation, an important component of both fracture healing and bone homeostasis. Four groups of n = 6 mice were used to compare the effects of Rosi on bone formation and cellular composition in both diabetic (Avy/a strain) and non-diabetic mice (a/a strain). New bone formation was examined by high resolution radiographs, micro-computed tomography, and histology. Precursor cells in the distraction gap were quantitated using immunohistochemical stains for proliferating cell nuclear antigen. Committed osteoblasts and adipocytes in the gap were identified and quantitated by immunostaining for osteocalcin and FABP4/aP2, respectively. The diabetic model developed obesity, hyperglycemia, hyperinsulinemia and insulin resistance, while the control littermates remained lean, normoglycemic and insulin sensitive. Rosi treatment decreased levels of non-fasted glucose and insulin and improved insulin sensitivity in the Avy /a mice, but had no effect in a/a mice, indicating antidiabetic efficacy of Rosi at the tested dose. Despite the diabetic, obese mice having twice the number of fat cells in their marrow than the non-diabetic mice, bone formation using DO was not adversely affected by the diabetes itself. However, Rosi treatment significantly diminished intramembranous endosteal bone formation, while increasing adipogenesis in and adjacent to the distraction gap up to 3.5- to 3.8-fold in both diabetic and non-diabetic models. This effect was independent of the anti-diabetic therapeutic response. These results raise the question of whether osteoblast precursors are inhibited in their development or actually converted to adipocytic phenotypes, possibly via marrow fat PPARγ nuclear receptor.
Correspondence to Dr Yan Wang; yan.wang@siat.ac.cn ; Dr Qian Xie; qian.xie@szu.edu.cn Obesity is recognised as a risk factor for triggering rheumatoid arthritis (RA), and it can worsen joint ...deformities1 and diminish the quality of life in patients with RA.2 The reduction of body weight in obese individuals is believed to alleviate RA symptoms.3 Body mass index (BMI) serves as the primary standard for evaluating obesity.4 An increase in BMI by 1 SD notably elevates the incidence rate of RA, suggesting a causal link between higher BMI and an increased risk of developing RA.5 The association between BMI and obesity is straightforward, as a higher BMI typically indicates a greater risk of obesity.4 Obesity is clinically defined as having a BMI of 30 kg/m2 or greater.4 Here, we established collagen-induced arthritis (CIA) models in mice using both regular and high-fat diets (HFDs) to see if HFD can induce severe RA symptoms in mice. Furthermore, the HFD sham group displayed significantly higher levels of cholesterol (CHO) and high-density lipoprotein (HDL) compared with the regular diet sham group, whereas the HFD CIA group exhibited similar serum levels of CHO, HDL, low-density lipoprotein and triglycerides compared with the regular diet CIA group (figure 1C). (C) Cholesterol (CHO), high-density lipoprotein (HDL), low-density lipoprotein (LDL) and triglyceride (TG) levels were measured at the end of the experiment.
Background
Intervertebral disc degeneration (IDD) is a significant cause of low back pain and poses a significant public health concern. Genetic factors play a crucial role in IDD, highlighting the ...need for a better understanding of the underlying mechanisms.
Aim
The aim of this study was to identify potential IDD‐related biomarkers using a comprehensive bioinformatics approach and validate them in vitro.
Materials and Methods
In this study, we employed several analytical approaches to identify the key genes involved in IDD. We utilized weighted gene coexpression network analysis (WGCNA), MCODE, LASSO algorithms, and ROC curves to identify the key genes. Additionally, immune infiltrating analysis and a single‐cell sequencing dataset were utilized to further explore the characteristics of the key genes. Finally, we conducted in vitro experiments on human disc tissues to validate the significance of these key genes in IDD.
Results
we obtained gene expression profiles from the GEO database (GSE23130 and GSE15227) and identified 1015 DEGs associated with IDD. Using WGCNA, we identified the blue module as significantly related to IDD. Among the DEGs, we identified 47 hub genes that overlapped with the genes in the blue module, based on criteria of |logFC| ≥ 2.0 and p.adj <0.05. Further analysis using both MCODE and LASSO algorithms enabled us to identify five key genes, of which CKAP4 and SSR1 were validated by GSE70362, demonstrating significant diagnostic value for IDD. Additionally, immune infiltrating analysis revealed that monocytes were significantly correlated with the two key genes. We also analyzed a single‐cell sequencing dataset, GSE199866, which showed that both CKAP4 and SSR1 were highly expressed in fibrocartilage chondrocytes. Finally, we validated our findings in vitro by performing real time polymerase chain reaction (RT‐PCR) and immunohistochemistry (IHC) on 30 human disc samples. Our results showed that CKAP4 and SSR1 were upregulated in degenerated disc samples. Taken together, our findings suggest that CKAP4 and SSR1 have the potential to serve as disease biomarkers for IDD.
In this study, we identified 1015 differentially expressed genes (DEGs) between IDD samples and control samples from datasets of GSE23130 and GSE15227 for further WGCNA. Out of the 56 hub genes from the blue module with the highest gene significance, 47 overlapped with 508 genes screened from DEGs with | Log2FC | ≥2.0 and p.adj < 0.05. We then performed GO and KEGG pathway enrichment analysis on the 47 hub genes and used MCODE and LASSO algorithms to locate five key genes. We validated two of these genes, SSR1 and CKAP4, using ROC curves and demonstrated their potential diagnostic value in another dataset, GSE70362. To further analyze these genes, we conducted immunofiltration analysis using ssGSEA and cell subtype analysis on a single‐cell RNA‐seq dataset, GSE199866. Finally, we validated our findings in vitro by performing RT‐PCR and IHC on human disc samples.
Type 1 diabetes mellitus is associated with a number of disorders of skeletal health, conditions that rely, in part, on dynamic bone formation. A mouse model of distraction osteogenesis was used to ...study the consequences of streptozotocin-induced diabetes and insulin treatment on bone formation and osteoblastogenesis. In diabetic mice compared with control mice, new bone formation was decreased, and adipogenesis was increased in and around, respectively, the distraction gaps. Although insulin treatment restored bone formation to levels observed in nondiabetic control mice, it failed to significantly decrease adipogenesis. Molecular events altered during de novo bone formation in untreated type 1 diabetes mellitus, yet restored with insulin treatment were examined so as to clarify specific osteogenic genes that may contribute to diabetic bone disease. RNA from distraction gaps was analyzed by gene microarray and quantitative RT-PCR for osteogenic genes of interest. Runt-related transcription factor 2 (RUNX2), and several RUNX2 target genes, including matrix metalloproteinase-9, Akp2, integrin binding sialoprotein, Dmp1, Col1a2, Phex, Vdr, osteocalcin, and osterix, were all significantly down-regulated in the insulin-deficient, hyperglycemic diabetic animals; however, insulin treatment of diabetic animals significantly restored their expression. Expression of bone morphogenic protein-2, transcriptional coactivator with PDZ-binding motif, and TWIST2, all important regulators of RUNX2, were not impacted by the diabetic condition, suggesting that the defect in osteogenesis resides at the level of RUNX2 expression and its activity. Together, these data demonstrate that insulin and/or glycemic status can regulate osteogenesis in vivo, and systemic insulin therapy can, in large part, rescue the diabetic bone phenotype at the tissue and molecular level.
Bone Formation Is Impaired in a Model of Type 1 Diabetes
Kathryn M. Thrailkill 1 ,
Lichu Liu 2 ,
Elizabeth C. Wahl 2 ,
Robert C. Bunn 1 ,
Daniel S. Perrien 3 4 ,
Gael E. Cockrell 1 ,
Robert A. ...Skinner 4 5 ,
William R. Hogue 4 5 ,
Adam A. Carver 4 ,
John L. Fowlkes 1 ,
James Aronson 5 and
Charles K. Lumpkin , Jr. 1
1 Department of Pediatrics, University of Arkansas for Medical Sciences, Little Rock, Arkansas
2 Arkansas Children’s Hospital, Little Rock, Arkansas
3 Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, Arkansas
4 Center for Orthopedic Research, University of Arkansas for Medical Sciences, Little Rock, Arkansas
5 Department of Orthopedics, University of Arkansas for Medical Sciences, Little Rock, Arkansas
Address correspondence and reprint requests to Kathryn M. Thrailkill, MD, Arkansas Children’s Hospital, 800 Marshall St.,
Springer Building, Slot 512-6, Little Rock, AR 72202. E-mail: thrailkillkathrynm{at}uams.edu
Abstract
The effects of type 1 diabetes on de novo bone formation during tibial distraction osteogenesis (DO) and on intact trabecular
and cortical bone were studied using nonobese diabetic (NOD) mice and comparably aged nondiabetic NOD mice. Diabetic mice
received treatment with insulin, vehicle, or no treatment during a 14-day DO procedure. Distracted tibiae were analyzed radiographically,
histologically, and by microcomputed tomography (μCT). Contralateral tibiae were analyzed using μCT. Serum levels of insulin,
osteocalcin, and cross-linked C-telopeptide of type I collagen were measured. Total new bone in the DO gap was reduced histologically
( P ≤ 0.001) and radiographically ( P ≤ 0.05) in diabetic mice compared with nondiabetic mice but preserved by insulin treatment. Serum osteocalcin concentrations
were also reduced in diabetic mice ( P ≤ 0.001) and normalized with insulin treatment. Evaluation of the contralateral tibiae by μCT and mechanical testing demonstrated
reductions in trabecular bone volume and thickness, cortical thickness, cortical strength, and an increase in endosteal perimeter
in diabetic animals, which were prevented by insulin treatment. These studies demonstrate that bone formation during DO is
impaired in a model of type 1 diabetes and preserved by systemic insulin administration.
BMD, bone mineral density
DO, distraction osteogenesis
FIZ, fibrous interzone
μCT, microcomputed tomography
PMF, primary matrix front
ROI, region of interest
Footnotes
Accepted July 6, 2005.
Received March 31, 2005.
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