Human Fibroblast-like Cells Research Articles

Toxicological Assessment of Biodegradable Poli-ε-Caprolactone Polymer Composite Materials Containing Hydroxyapatite, Bioglass, and Chitosan as Potential Biomaterials for Bone Regeneration Scaffolds.

Polycaprolactone (PCL) is a biodegradable polyester that might be used in tissue engineering to obtain scaffolds for bone reconstruction using 3D-printing technologies. New material compositions based on PCL, with improved physicochemical properties and excellent biocompatibility, would improve its applicability in bone regeneration. The aim of this study was to assess the potential toxic effects of PCL-based composite materials containing 5% hydroxyapatite (PCL/SHAP), 5% bioglass (PCL/BIO), or 5% chitosan (PCL/CH) on MG-63 human fibroblast-like cells in vitro. Material tests were carried out using X-ray diffraction, differential thermal analysis/thermal gravimetry, BET specific surface analysis, and scanning electron microscopy. The effect of the biomaterials on the MG-63 cells was then assessed based on toxicity tests using indirect and direct contact methods. The analysis showed that the tested biomaterials did not significantly affect cell morphology, viability, proliferation, or migration. We concluded that biodegradable PCL-based scaffolds may be suitable for tissue scaffold production, and the addition of bioglass improves the growth of cultured cells.

Intron retention in PI-PLC γ1 mRNA as a key mechanism affecting MMP expression in human primary fibroblast-like synovial cells.

The intron retention (IR) is a phenomenon utilized by cells to allow diverse fates at the same mRNA, leading to a different pattern of synthesis of the same protein. In this study, we analyzed the modulation of phosphoinositide-specific phospholipase C (PI-PLC) enzymes by Harpagophytum procumbensextract (HPE) in synoviocytes from joins of osteoarthritis(OA) patients. In some samples, the PI-PLC γ1 isoform mature mRNA showed the IR and, in these synoviocytes, the HPE treatment increased the phenomenon. Moreover, we highlighted that as a consequence of IR, a lower amount of PI-PLC γ1 was produced. The decrease of PI-PLC γ1 was associated with the decrease of metalloprotease-3(MMP-3),and MMP-13, and ADAMTS-5 after HPE treatment. The altered expression of MMPs is a hallmark of the onset and progression of OA, thus substances able to decrease their expression are very desirable. The interesting outcomes of this study are that 35% of analyzed synovial tissues showed the IR phenomenon in the PI-PLC γ1 mRNAand that the HPE treatment increased this phenomenon. For the first time, we found that the decrease of PI-PLC γ1 protein in synoviocytes interferes withMMP production, thus affecting the pathways involved in the MMP expression. This finding was validated by the silencing of PI-PLC γ1 in synoviocytes where the IR phenomenon was not present. Our results shed new light on the biochemical mechanisms involved in the degrading enzyme production in the joint of OA patients, suggesting a new therapeutic target and highlighting the importance of personalized medicine.

Anti-rheumatic property and physiological safety of KMU-11342 in in vitro and in vivo models

Rheumatoid arthritis (RA) is a chronic, systemic inflammatory disorder characterized by joint destruction due to synovial hypertrophy and the infiltration of inflammatory cells. Despite substantial progress in RA treatment, challenges persist, including suboptimal treatment responses and adverse effects associated with current therapies. This study investigates the anti-rheumatic capabilities of the newly identified multi-protein kinase inhibitor, KMU-11342, aiming to develop innovative agents targeting RA. In this study, we synthesized the novel multi-protein kinase inhibitor KMU-11342, based on indolin-2-one. We assessed its cardiac electrophysiological safety using the Langendorff system in rat hearts and evaluated its toxicity in zebrafish in vivo. Additionally, we examined the anti-rheumatic effects of KMU-11342 on human rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS), THP-1 cells, and osteoclastogenesis in RAW264.7 cells. KMU-11342 demonstrated the ability to inhibit LPS-induced chemokine inhibition and the upregulation of pro-inflammatory cytokines, cyclooxygenase-2, inducible nitric oxide synthase, p-IKKα/β, p-NF-κB p65, and the nuclear translocation of NF-κB p65 in RA-FLS. It effectively suppressed the upregulation of NLR family pyrin domain containing 3 (NLRP3) and caspase-1 cleavage. Furthermore, KMU-11342 hindered the activation of osteoclast differentiation factors such as RANKL-induced TRAP, cathepsin K, NFATc-1, and c-Fos in RAW264.7 cells. KMU-11342 mitigates LPS-mediated inflammatory responses in THP-1 cells by inhibiting the activation of NLRP3 inflammasome. Notably, KMU-11342 exhibited minimal cytotoxicity in vivo and electrophysiological cardiotoxicity ex vivo. Consequently, KMU-11342 holds promise for development as a therapeutic agent in RA treatment.

Fluidic shear stress alters clathrin dynamics and vesicle formation in endothelial cells

Endothelial cells (ECs) experience a variety of highly dynamic mechanical stresses. Among others, cyclic stretch and increased plasma membrane tension inhibit clathrin-mediated endocytosis (CME) in non-ECs. It remains elusive how ECs maintain CME in these biophysically unfavorable conditions. Previously, we have used simultaneous two-wavelength axial ratiometry (STAR) microscopy to show that endocytic dynamics are similar between statically cultured human umbilical vein endothelial cells (HUVECs) and fibroblast-like Cos-7 cells. Here, we asked whether biophysical stresses generated by blood flow influence CME. We used our data processing platform—DrSTAR—to examine if clathrin dynamics are altered in HUVECs after experiencing fluidic shear stress (FSS). We found that HUVECs cultivated under a physiological level of FSS had increased clathrin dynamics compared with static controls. FSS increased both clathrin-coated vesicle formation and nonproductive flat clathrin lattices by 2.3-fold and 1.9-fold, respectively. The curvature-positive events had significantly delayed curvature initiation relative to clathrin recruitment in flow-stimulated cells, highlighting a shift toward flat-to-curved clathrin transitions in vesicle formation. Overall, our findings indicate that clathrin dynamics and clathrin-coated vesicle formation can be modulated by the local physiological environment and represent an important regulatory mechanism.

Changes in the mitochondrial function of fibroblast-like cells exposed to copper oxide nanoparticles

Introduction. Mitochondria are targets for almost all types of damaging agents, including toxins and oxidative stress. There is no doubt that possible effects on mitochondria should be taken into account in a comprehensive assessment of metal toxicity. Our objective was to establish changes in the mitochondrial function under the effect of copper oxide nanoparticles in vitro.
 Material and methods. A monolayer culture of human lung fibroblast-like cells of the FLECH-104 line were exposed to copper oxide nanoparticles (CuO NPs) 21±4 nm in size, final concentrations of which in the media were 25, 50, and 100 μg/mL. We measured the rate of oxygen consumption by the cells and its changes under the influence of modulators, such as oligomycin, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone, and rotenone combined with antimycin A. We estimated parameters of mitochondrial function and the bioenergetics index.
 Results. At the concentration of 100 μg/mL in the incubation medium, CuO NPs induced changes in the culture of fibroblast-like cells that impeded further assessment of the mitochondrial function. At the lower concentrations of 25 and 50 μg/mL, we observed a dose-dependent trend toward a decrease in ATP-related respiration and bioenergetics index. It is noteworthy that the concentration of 25 μg/mL increased the maximum and reserve respiratory capacity of the cells, which was probably related to the dual effect of copper as a toxicant and an essential element.
 Limitations of the study. The study was conducted using only one cell line and three concentrations of CuO nanoparticles suspended in the culture medium.
 Conclusion. We established that copper oxide nanoparticles, when added to the incubation medium, have a multidirectional effect on the mitochondrial function of fibroblast-like cells potentially attributed to biotic properties of this metal.

Exploring the supplementary potential of all-trans retinoic acid with methotrexate in rheumatoid arthritis: modulation of synovial cell apoptosis and autophagy.

The imbalance between apoptosis and proliferation in fibroblast-like synoviocytes (FLSs) plays a key role in the pathogenesis of rheumatoid arthritis (RA). This study aims to investigate the potential of all-trans retinoic acid (ATRA) as a supplementary therapeutic agent alongside methotrexate (MTX) for RA, by examining its ability to inhibit synovial cell proliferation and enhance apoptosis through the ROS-JNK signalling pathway. The viability, apoptosis, and autophagy levels of human rheumatoid arthritis fibroblast-like synovial cells (HFLS-RA) were evaluated, while ROS generation was measured through the DCFH-DA fluorescence microplate assay. Western blotting was used to analyse the expression levels of JNK signalling pathway-related proteins. To assess therapeutic potential in vivo, a collagen-induced arthritis (CIA) model was established in Wistar rats. Small doses of MTX did not significantly affect the viability of HFLS-RAs or induce apoptosis. However, when ATRA was added to the treatment, the therapy markedly inhibited cell proliferation and induced apoptosis and excessive autophagy. Mechanistically, ATRA activated the ROS/JNK signalling pathway in HFLS-RAs. ROS scavengers and JNK inhibitors significantly attenuated ATRA-induced apoptosis and autophagy. In vivo, the combination therapy demonstrated a remarkable enhancement of the anti-arthritic efficacy in CIA rats. The ability of ATRA to inhibit proliferation in RA FLSs through autophagy and apoptosis underscores its potential as a supplementary therapeutic agent alongside MTX for RA, particularly when compared to the limited impact of MTX on these processes. This combined strategy holds promise for enhancing therapeutic outcomes and warrants further investigation in the management of RA.

Plasminogen Receptors Promote Lipoprotein(a) Uptake by Enhancing Surface Binding and Facilitating Macropinocytosis.

High levels of Lp(a) (lipoprotein(a)) are associated with multiple forms of cardiovascular disease. Lp(a) consists of an apoB100-containing particle attached to the plasminogen homologue apo(a). The pathways for Lp(a) clearance are not well understood. We previously discovered that the plasminogen receptor PlgRKT (plasminogen receptor with a C-terminal lysine) promoted Lp(a) uptake in liver cells. Here, we aimed to further define the role of PlgRKT and to investigate the role of 2 other plasminogen receptors, annexin A2 and S100A10 (S100 calcium-binding protein A10) in the endocytosis of Lp(a). Human hepatocellular carcinoma (HepG2) cells and haploid human fibroblast-like (HAP1) cells were used for overexpression and knockout of plasminogen receptors. The uptake of Lp(a), LDL (low-density lipoprotein), apo(a), and endocytic cargos was visualized and quantified by confocal microscopy and Western blotting. The uptake of both Lp(a) and apo(a), but not LDL, was significantly increased in HepG2 and HAP1 cells overexpressing PlgRKT, annexin A2, or S100A10. Conversely, Lp(a) and apo(a), but not LDL, uptake was significantly reduced in HAP1 cells in which PlgRKT and S100A10 were knocked out. Surface binding studies in HepG2 cells showed that overexpression of PlgRKT, but not annexin A2 or S100A10, increased Lp(a) and apo(a) plasma membrane binding. Annexin A2 and S100A10, on the other hand, appeared to regulate macropinocytosis with both proteins significantly increasing the uptake of the macropinocytosis marker dextran when overexpressed in HepG2 and HAP1 cells and knockout of S100A10 significantly reducing dextran uptake. Bringing these observations together, we tested the effect of a PI3K (phosphoinositide-3-kinase) inhibitor, known to inhibit macropinocytosis, on Lp(a) uptake. Results showed a concentration-dependent reduction confirming that Lp(a) uptake was indeed mediated by macropinocytosis. These findings uncover a novel pathway for Lp(a) endocytosis involving multiple plasminogen receptors that enhance surface binding and stimulate macropinocytosis of Lp(a). Although the findings were produced in cell culture models that have limitations, they could have clinical relevance since drugs that inhibit macropinocytosis are in clinical use, that is, the PI3K inhibitors for cancer therapy and some antidepressant compounds.

Effects of a co-stimulation with S-PRG filler eluate and muramyl dipeptide (MDP) on matrix metalloproteinase-1 production by human dental pulp fibroblast-like cells.

The present study investigated the effects of a co-stimulation with surface reaction-type pre-reacted glass-ionomer (S-PRG) filler eluate and muramyl dipeptide (MDP) on matrix metalloproteinase (MMP)-1 production by human dental pulp fibroblast-like cells (hDPFs). S-PRG filler eluate contains 6 ions (F, Na, Al, B, Sr, and Si) released from S-PRG filler. Each S-PRG filler eluate and MDP stimulation enhanced MMP-1 production by hDPFs. The co-stimulation with S-PRG filler eluate and MDP enhanced MMP-1 production more than the MDP stimulation alone. A similar stimulation induced the phosphorylation of ERK 1/2. The increased secretion of MMP-1 and enhanced phosphorylation of ERK 1/2 by the co-stimulation with S-PRG filler eluate and MDP were suppressed by the selective and potent CaSR antagonist NPS 2143. Since strontium binds to CaSR, these results suggest that the enhanced production of MMP-1 by the co-stimulation with S-PRG filler eluate and MDP was due to the effects of strontium.

DHA Induces Cell Death through the Production of ROS and the Upregulation of CHOP in Fibroblast-like Synovial Cells from Human Rheumatoid Arthritis Patients.

Rheumatoid arthritis (RA) is an inflammatory disease marked by a massive proliferation of synovial cells in the joints. In this study, we investigated the pro-apoptotic effects of docosahexaenoic acid (DHA) in human fibroblast-like synovial cells from RA patients (RA-FLS). An in vitro study using MH7A cells showed that DHA treatment induced caspase-8-dependent apoptosis in a dose-dependent manner and reduced the TNF-α-mediated induction of MMP-9 and IL-1β. DHA also induced the phosphorylation of eIF2α, the expression of the ER stress markers ATF4 and C/EBP homologous protein (CHOP), and death receptor 5 (DR5). The knockdown of CHOP or DR5 increased cell viability and reduced apoptosis in DHA-treated cells. Furthermore, the knockdown of CHOP reduced DHA-mediated DR5 expression, while the overexpression of CHOP increased DR5 expression. We also found that DHA treatment induced the accumulation of reactive oxygen species (ROS), and pretreatment with the anti-oxidant Tiron effectively abrogated not only the expression of CHOP and DR5, but also DHA-induced apoptosis. Under this condition, cell viability was increased, while PARP-1 cleavage and caspase-8 activation were reduced. All the findings were reproduced in human primary synovial cells obtained from RA patients. These results suggest that the DHA-mediated induction of ROS and CHOP induced apoptosis through the upregulation of DR5 in RA-FLSs, and that CHOP could be used as a therapy for RA.

Cotransplantation of marginal mass allogeneic islets with 3D culture-derived adult human skin cells improves glycemia in diabetic mice.

Islet transplantation represents a therapeutic option for type 1 diabetes (T1D). Long-term viability of transplanted islets requires improvement. Mesenchymal stromal cells (MSCs) have been proposed as adjuvants for islet transplantation facilitating grafting and functionality. Stem cell aggregation provides physiological interactions between cells and enhances the in situ concentration of modulators of inflammation and immunity. We established a hanging-drop culture of adult human skin fibroblast-like cells as spheroids, and skin spheroid-derived cells (SphCs) were characterized. We assessed the potential of SphCs in improving islet functionality by cotransplantation with a marginal mass of allogeneic islets in an experimental diabetic mouse model and characterized the secretome of SphCs by mass spectrometry-based proteomics. SphCs were characterized as multipotent progenitors and their coculture with anti-CD3 stimulated mouse splenocytes decreased CD4+ T cell proliferation with skewed cytokine secretion through an increase in the Th2/Th1 ratio profile. SphCs-conditioned media attenuated apoptosis of islets induced by cytokine challenge in vitro and importantly, intratesticular SphCs administration did not show tumorigenicity in immune-deficient mice. Moreover, SphCs improved glycemic control when cotransplanted with a marginal mass of allogeneic islets in a diabetic mouse model without pharmacological immunosuppression. SphCs' protein secretome differed from its paired skin fibroblast-like counterpart in containing 70% of up- and downregulated proteins and biological processes that overall positively influenced islets such as cytoprotection, cellular stress, metabolism, and survival. In summary, SphCs improved the performance of transplanted allogeneic islets in an experimental T1D model, without pharmacological immunosuppression. Future research is warranted to identify SphCs-secreted factors responsible for islets' endurance.

Effect of Xanthii Fructus alcohol extract on proliferation and apoptosis of HFLS-RA and its mechanism

Xanthii fructus (XF) is the dried and mature fruit of Xanthium sibiricum Patr. It has the effects of anti-inflammatory, antioxidant and anti-arthritic. Rheumatoid arthritis (RA) is the most common inflammatory disorder and often leads to disability. However, there are few studies on the treatment of RA by XF and the specific mechanism of treatment has not been clarified. This study was designed to explore the effects of proliferation and apoptosis by XF on human fibroblast-like synovial-RA (HFLS-RA) cells and investigate its mechanism. The cell proliferation ability was detected by MTS assay. Hoechst 33,342 staining was used to detect apoptosis, and the apoptosis rate was detected by flow cytometry. The expression levels of NF-κB p65 and β-catenin were detected by Western Blotting. MTS, Hoechst 33,342, flow cytometry analysis showed that the alcohol extract of XF inhibited human fibroblast-like synovial-RA cells proliferation and promoted apoptosis in a dose-dependent manner. Western Blotting experiment showed that the extract of XF could reduce the expression levels of NF-κB p65 and β-catenin. The extract of XF has a significant therapeutic effect on RA in vitro by regulating NF-κB signaling pathway and Wnt/β-catenin signaling pathway. Our research will help to clarify the potential pharmacological mechanism of XF on RA and provide experimental basis for the application of XF in clinical treatment.

Direct RNA sequencing of respiratory syncytial virus infected human cells generates a detailed overview of RSV polycistronic mRNA and transcript abundance.

To characterize species of viral mRNA transcripts generated during respiratory syncytial virus (RSV) infection, human fibroblast-like MRC-5 lung cells were infected with subgroup A RSV for 6, 16 and 24 hours. In addition, we characterised the viral transcriptome in infected Calu-3 lung epithelial cells at 48 hours post infection. Total RNA was harvested and polyadenylated mRNA was enriched and sequenced by direct RNA sequencing using an Oxford nanopore device. This platform yielded over 450,000 direct mRNA transcript reads which were mapped to the viral genome and analysed to determine the relative mRNA levels of viral genes using our in-house ORF-centric pipeline. We examined the frequency of polycistronic readthrough mRNAs were generated and assessed the length of the polyadenylated tails for each group of transcripts. We show a general but non-linear decline in gene transcript abundance across the viral genome, as predicted by the model of RSV gene transcription. However, the decline in transcript abundance is not uniform. The polyadenylate tails generated by the viral polymerase are similar in length to those generated by the host polyadenylation machinery and broadly declined in length for most transcripts as the infection progressed. Finally, we observed that the steady state abundance of transcripts with very short polyadenylate tails less than 20 nucleotides is less for N, SH and G transcripts in both cell lines compared to NS1, NS2, P, M, F and M2 which may reflect differences in mRNA stability and/or translation rates within and between the cell lines.

Exercise Regulates the Lactate Receptor HCAR1 and ERK1/2-PI3K/Akt Pathways to Promote Cerebral Angiogenesis.

We aimed to explore the ole and mechanism of lactate receptor (HCAR1) in the angiogenesis of leptomeningeal fibroblast-like cells. Human brain fibroblast-like cells were selected and some cells were deactivated, analyzed and compared with HCAR1 mRNA and protein expressions in deactivated/normal cells. HCAR1-/- mice and wild type (WT) mice were selected and divided into WT, WT exercise, HCAE1 KO and HCAE1 KO exercise groups, with 10 mice for each group. HCAR1mRNA and expression levels of proteins in fibroblast-like cells, mRNA and expression levels of proteins in Collagen IV, phosphatidylinositol trihydroxykinase (PI3K), serine threonine kinase (AKT) and extracellular signal-regulated kinases 1 and 2 (ERK1/2) in hippocampus were compared, and the microvessel density (MVD) and diameter were calculated. mRNA and expression levels of proteins in Collagen IV, PI3K, AKT, ERK1/2 and MVD in hippocampus were significantly higher in the WT exercise group than those in the WT group, microvessel diameter was significantly lower than that in the WT group (P<0.05). mRNA and expression levels of proteins in Collagen IV, PI3K, AKT, ERK1/2 and MVD in hippocampus in the HCAR1 KO and HCAR1 KO exercise groups were significantly lower than those in the WT group, microvessel diameter was higher than that in the WT group (P<0.05). Compared with the HCAR1 KO exercise group, the changes of mRNA in Collagen IV, PI3K, AKT, ERK1/2 and microvascular were not significant. Exercise can promote cerebral angiogenesis through the activation of the lactate receptor HCAR1 and the ERK1/2-PI3K/Akt signaling pathways.

Study on the Therapeutic Effect of Lenalidomide on Hemophilic Arthropathy

To explore the effect of lenalidomide on human fibroblast-like synovial cells (HFLS) and the therapeutic efficacy on hemophilic arthropathy in hemophilia A mice model. In vitro, to remodel the inflammatory environment of synovial tissue after hemorrhage, ferric citrate and recombinant TNF-α were added into the cell culture medium of HFLS. Cell Counting Kit-8 (CCK-8), Enzyme-linked immunosorbent assay (ELISA), Quantitative Real-time PCR (RT-qPCR) and flow cytometry were employed for detection of the effects of lenalidomide on the proliferation ability, pro-inflammatory cytokines release and apoptosis of HFLS cells. In vivo, hemophilia arthropathy was remodeled in hemophilia A mice by induction of hemarthrosis. A series of doses of lenalidomide (0.1, 0.3 and 1.0 g/kg) was administrated intra-articularly. Tissues of knee joints were collected on the 14th day after administration, and the protective effect of lenalidomide on arthritis in hemophilia A mice were evaluated by RT-qPCR and histological grading. In vitro, compared with the untreated control group, lenalidomide could significantly inhibit the proliferation of HFLS cells (P<0.05), and the effect was the most significant when the concentration was 0.01 μmol/L (P<0.001). Compared with the control group, lenalidomide could significantly inhibit the expression levels of TNF-α, IL-1β, IL-6 and IFN-γ in HFLS cells (P<0.05). The flow cytometry results showed that lenalidomide could enhance the apoptotis of HFLS cells (P<0.05). The results of RT-qPCR showed that lenalidomide could significantly reduce the mRNA expression levels of TNF-α, IL-1β, IL-6,MCP-1 and VEGF in the joint tissues (P<0.05). Histological results showed that compared with the injured group, lenalidomide could significantly reduce the pathological sequela after hemarthrosis induction, e.g. synovial thickening and neo-angiogenesis in the synovium. The protection displayed a dose-response pattern roughly. In vitro, lenalidomide can inhibit the proliferation of HFLS cells, promote their apoptosis, and inhibit the expression of pro-inflammatory cytokines. In vivo, lenalidomide can significantly decrease the expression of pro-inflammatory cytokines in the joints of mice, and prevent the development of inflammation and neo-angiogenesis. The results provide a theoretical and experimental basis for the clinical application of lenalidomide in the treatment of hemophilic arthropathy.

Panlongqi Tablet (PLQT) Inhibits LPS-Induced Abnormal Proliferation of RA-HFLS by Regulating PI3K/ AKT and MAPK Signaling Pathways

Panlongqi Tablet (PLQT), a traditional Chinese formula, is effective in the clinical treatment of Rheumatoid Arthritis (RA) in China for several decades. However, the underlying mechanism of PLQT’s therapeutic effect remains unclear. The main chemical constituents of PLQT were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The human fibroblast-like synovial cells in rheumatoid arthritis (RA-HFLS) were stimulated with LPS (10 μg/mL) to establish a inflammatory model in vitro. The cells were then treated with PLQT-medicated serum to verify the protective effect of the herbal compound. MTT assay was used to detect the viability of RAHFLS and abnormal cell proliferation. The inflammatory factors levels including interleukin-1β (IL-1β), interleukin-6 (IL-6), and interleukin-17 (IL-17) in RAHFLS were measured by ELISA. In addition, the cell cycle progression was analyzed by flow cytometry and protein expression of Proliferating Cell Nuclear Antigen (PCNA) was detected using immunohistochemistry and western blot. The protein expression levels associated with PI3K/AKT and MAPK signaling pathways in RA-HFLS were detected by western blot analyses. Our results revealed that PLQT treatment effectively inhibited the abnormal proliferation of RA-HFLS and decreased the inflammatory factors levels. The proportion of cells in S phase was significantly increased with the treatment of PLQT. Meanwhile, the expression levels of PCNA, p-PI3K, p-AKT, p-mTOR, p-JNK/JNK, p-ERK/ ERK, and p-P38/P38 proteins from RA-HFLS were dramatically inhibited by PLQT treatment. PLQT inhibits LPS-induced abnormal proliferation of RAHFLS, which may be related to the down regulation of PCNA, and the inhibition of abnormally activated PI3K/AKT and MAPK signaling pathways.

Tectoridin exhibits anti-rheumatoid arthritis activity through the inhibition of the inflammatory response and the MAPK pathway in vivo and in vitro

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by inflammation infiltration of the synovial tissues and the fibroblast-like synoviocytes. Tectoridin is a botanical active ingredient with anti-inflammatory properties. In this study, the anti-arthritic effects of tectoridin and its mechanism of action are examined in TNF-α-induced human fibroblast-like synovial cells (HFLSs cells) and complete Freund's adjuvant (CFA)-stimulated arthritic mice. Arthritis progression was evaluated via bodyweight, hind paw swelling, organ index, and synovial pathology. IL-1β, IL-6 and other pro-inflammatory factors concentrations, and the expression of MAPK pathway proteins in HFLSs cells and arthritic mice were measured using ELISA and western blotting. Results showed that tectoridin significantly decreased the swelling of the paws and joints as well as the increased immune organ index within CFA-induced arthritic mice. Histopathological analysis showed that tectoridin alleviated the lesions of ankle joints and synovial tissues induced by CFA. Secretion of pro-inflammatory cytokines in TNF-α-induced HFLSs cells and CFA-stimulated arthritic mice were also abated by tectoridin. Similarly, the presence of tectoridin significantly inhibited the abnormal phosphorylation levels of ERK, JNK, and p38 in vivo and in vitro. All those results highlighted that tectoridin exhibits anti-arthritis effects by inhibiting MAPK-mediated inflammatory responses.

Anti-inflammatory effects of aucubin in cellular and animal models of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease. It is known that aucubin (AU) exerts anti-inflammatory activity, but its effects and mechanisms in RA are unclear. This study investigated the anti-inflammatory effects and mechanisms of AU in vivo and in vitro. Human fibroblast-like synoviocyte cells from patients with RA (HFLS-RA), RAW264.7 cells, and MC3T3-E1 cells were used to evaluate the effects of AU on migration, invasion, apoptosis, osteoclast differentiation and production. Immunofluorescence was used to observe nuclear translocation of nuclear factor (NF)-κB, the double luciferase reporter gene method was used to observe NF-κB-p65 activity in AU-treated MC3T3-E1 cells. RT-qPCR was used to measure expression of bone metabolism and inflammation-related genes, and western blot was used to measure bone metabolism and NF-κB protein expression levels. Collagen-induced arthritis (CIA) rat model was used for pharmacodynamics study. Arthritis indexes were measured in the ankle and knee, histological staining and Micro-computed tomography were performed on the ankle joints. Also, inflammatory factor gene expression and the levels of NF-κB-related proteins were detected as in vitro. AU effectively inhibited HFLS-RA cell migration and invasion, promoted apoptosis, and inhibited RAW264.7 cell differentiation into osteoclasts, as well as inhibited NF-κB-p65 activity in MC3T3-E1 cells. Notably, AU significantly reduced the gene expression levels of three cell-related inflammatory factors and bone metabolism factors, effectively inhibited the expression of p-Iκκα β, p-IκBα, and p-p65 proteins. In vivo, AU relieved joint inflammation, reduced related inflammatory factors, and inhibited NF-κB signaling. It could be used to treat RA-related synovial inflammation and bone destruction through the NF-κB pathway.

Sodium propionate improves rheumatoid arthritis by inhibiting survivin mediated proliferation of fibroblast like synoviocytes by promoting miR-140-5p

Background Increased proliferation and impaired death of fibroblast-like synovial cells play an important role in the development of rheumatoid arthritis (RA). Survivin plays an important role in the prodromal stage and prognosis of RA and has been introduced as a biomarker of joint injury in RA patients. The purpose of this study was to explore whether propionate alleviates RA through miR-140-5p/survivin pathway. Methods The synovial tissues of RA patients were collected to detect the expression levels of miR-140-5p and survivin; normal human fibroblast-like synovial cells (HLSs) and RA fibroblast-like synovial cells (RA-FLSs) were cultured and treated with 10 mM of sodium propionate (SP), then the expressions of miR-140-5p and survivin, cell viability and apoptosis were detected; collagen induced arthritis (CIA) rat model was constructed and treated with SP, then the tissue inflammation level and the expression levels of miR-140-5p and Survivin were detected. Results The expression of miR-140-5p decreased in synovial tissues of RA patients and RA-FLSs cells, while the expression of survivin increased significantly in RA patients. SP promoted miR-140-5p expression and apoptosis in RA-FLSs cells and inhibited survivin expression and cell viability of RA-FLSs cells. In addition, miR-140-5p plays a protective role by targeting survivin. Importantly, in the CIA rat model, SP reduced joint inflammatory response, and the miR-140-5p inhibitor weakened the protective effect of SP. Conclusion SP can alleviate RA by promoting the expression of miR-140-5p and inhibiting the excessive proliferation and death impairment of RA-FLSs cells induced by survivin.

The various dose-dependent effect of selenium oxide and copper oxide nanoparticles in vitro and application of the hormesis paradigm

Introduction. In vitro studies on a culture of cardiomyocytes have shown that dose-response relationships could be monotonic for some effects and non-monotonic for others. In this work, we wanted to demonstrate that these features of the dose-response relationship are a general pattern. Materials and methods. In vitro experiments were conducted on the culture of human fibroblast-like cells FLECH-104. The cytotoxicity of spherical nanoparticles of selenium oxide (SeO-NP) and copper oxide (CuO-NP) was studied with an average diameter of 51 ± 14 nm and 21 ± 4 nm, respectively. Results. SeO-NP and CuO-NP were cytotoxic for human fibroblast-like cells, as judged by a decrease in ATP-dependent luminescence. In this case, the cytotoxicity of CuO-NP was somewhat more substantial than the SeO-NP one. Our experiment revealed doses that cause both cell hypertrophy and a decrease in the size of cells and nuclei. Discussion. We observed both monotonic and different variants of the non-monotonic dose-response relationship. For the latter, it was possible to construct adequate mathematical expressions based on the generalized hormesis paradigm that we had considered earlier concerning the CdS-NP and PbS-NP cytotoxicity for cardiomyocytes. Conclusion. The general rule is the variability of the dose-response dependence types manifested in different cytotoxic effects of nanoparticles.

Effect of Non-Thermal Atmospheric Pressure Plasma on Differentiation Potential of Human Deciduous Dental Pulp Fibroblast-like Cells

Human mesenchymal stem cells can differentiate into various cell types and are useful for applications in regenerative medicine. Previous studies indicated that dental pulp exfoliated from deciduous teeth is a valuable alternative for dental tissue engineering because it contains stem cells with a relatively high proliferation rate. For clinical application, it is necessary to rapidly obtain a sufficient number of cells in vitro and maintain their undifferentiated state; however, the abundance of stem cells in the dental pulp tissue is limited. Non-thermal atmospheric pressure plasma (NTAPP) has been applied in regenerative medicine because it activates cell proliferation. Here, we examined the effects of NTAPP to activate the proliferation of human deciduous dental pulp fibroblast-like cells (hDDPFs) in vitro. Compared with untreated cells, NTAPP increased cell proliferation by 1.3-fold, significantly upregulated well-known pluripotent genes for stemness (e.g., Oct4, Sox2, and Nanog), and activated the expression of stem cell-specific surface markers (e.g., CD105). Overall, NTAPP activated the proliferation of various mesodermal-derived human adult stem cells while maintaining their pluripotency and stemness. In conclusion, NTAPP is a potential tool to expand the population of various adult stem cells in vitro for medical applications.