TGFßR-1/ALK5 inhibitor RepSox induces enteric glia-to-neuron transition and influences gastrointestinal mobility in adult mice.

Promoting adult neurogenesis in the enteric nervous system (ENS) may be a potential therapeutic approach to cure enteric neuropathies. Enteric glial cells (EGCs) are the most abundant glial cells in the ENS. Accumulating evidence suggests that EGCs can be a complementary source to supply new neurons during adult neurogenesis in the ENS. In the brain, astrocytes have been intensively studied for their neuronal conversion properties, and small molecules have been successfully used to induce the astrocyte-to-neuron transition. However, research on glia-to-neuron conversion in the ENS is still lacking. In this study, we used GFAP-Cre:Rosa-tdTomato mice to trace glia-to-neuron transdifferentiation in the ENS in vivo and in vitro. We showed that GFAP promoter-driven tdTomato exclusively labelled EGCs and was a suitable marker to trace EGCs and their progeny cells in the ENS of adult mice. Interestingly, we discovered that RepSox or other ALK5 inhibitors alone induced efficient transdifferentiation of EGCs into neurons in vitro. Knockdown of ALK5 further confirmed that the TGFßR-1/ALK5 signalling pathway played an essential role in the transition of EGCs to neurons. RepSox-induced neurons were Calbindin- and nNOS-positive and displayed typical neuronal electrophysiological properties. Finally, we showed that administration of RepSox (3, 10 mg· kg-1 ·d-1, i.g.) for 2 weeks significantly promoted the conversion of EGCs to neurons in the ENS and influenced gastrointestinal motility in adult mice. This study provides a method for efficiently converting adult mouse EGCs into neurons by small-molecule compounds, which might be a promising therapeutic strategy for gastrointestinal neuropathy.

Neural cell adhesion molecule regulates chondrocyte hypertrophy in chondrogenic differentiation and experimental osteoarthritis.

Chondrocyte hypertrophy-like change is an important pathological process of osteoarthritis (OA), but the mechanism remains largely unknown. Neural cell adhesion molecule (NCAM) is highly expressed and involved in the chondrocyte differentiation of mesenchymal stem cells (MSCs). In this study, we found that NCAM deficiency accelerates chondrocyte hypertrophy in articular cartilage and growth plate of OA mice. NCAM deficiency leads to hypertrophic chondrocyte differentiation in both murine MSCs and chondrogenic cells, in which extracellular signal-regulated kinase (ERK) signaling plays an important role. Moreover, NCAM expression is downregulated in an interleukin-1ß-stimulated OA cellular model and monosodium iodoacetate-induced OA rats. Overexpression of NCAM substantially inhibits hypertrophic differentiation in the OA cellular model. In conclusion, NCAM could inhibit hypertrophic chondrocyte differentiation of MSCs by inhibiting ERK signaling and reduce chondrocyte hypertrophy in experimental OA model, suggesting the potential utility of NCAM as a novel therapeutic target for alleviating chondrocyte hypertrophy of OA.

Fengshi Gutong Capsule Attenuates Osteoarthritis by Inhibiting MAPK, NF-κB, AP-1, and Akt Pathways.

Background and purpose: Fengshi Gutong capsule (FSGTC), a traditional herbal formula, has been used clinically in China for the treatment of arthritis. However, the mechanism underlying the therapeutic effects of FSGTC on osteoarthritis (OA) has not been elucidated. The present study investigated the function and mechanisms of FSGTC in rat OA model and interleukin (IL)-1ß-stimulated synovial cells. Materials and methods: Rat OA model was established by intra-articular injection containing 4% papain. IL-1ß-induced SW982 cells were used as an OA cell model. Safranin-O-Fast green (S-O) and hematoxylin-eosin (HE) stainings were used to observe the changes in cartilage morphology. Enzyme-linked immunosorbent assay (ELISA) and real-time quantitative PCR (qPCR) detected the expression of inflammatory cytokines. In addition, molecular mechanisms were analyzed by Western blot in the OA cell model. Results: FSGTC treatment significantly relieved the degeneration of cartilage and reduced the contents of tumor necrosis factor-α (TNF-α) and IL-6 in the serum in papain-induced OA rats. FSGTC also reduced the protein and mRNA levels of IL-6 and IL-8 in IL-1ß-stimulated SW982 cells. Moreover, it inhibited the phosphorylation levels of ERK (extracellular signal-related kinase), JNK (c-Jun N-terminal kinase), p38, Akt (protein kinase B), and c-Jun. It also decreased the extent of IκBα degradation and p65 protein translocation into the nucleus. Conclusion: The current data confirmed the protective effects of FSGTC in the rat and OA cell models. The results suggested that FSGTC reduced the production of inflammatory mediators via restraining the activation of mitogen-activated protein kinases (MAPK), nuclear factor kappa B (NF-κB), activator protein-1 (AP-1), and Akt.

Anti-inflammatory effects of pitavastatin in interleukin-1ß-induced SW982 human synovial cells.

The present study shows the basis for the anti-inflammatory effects of pitavastatin in interleukin (IL)-1ß-induced human synovial cells. The SW982 cells were pretreated with pitavastatin at different concentrations (5µM and 10µM), followed by IL-1ß (10ng/mL) stimulation. The results showed that pitavastatin inhibited the expression of inflammatory mediators IL-6 and IL-8. Furthermore, pitavastatin inhibited the phosphorylation of p38, extracellular signal-related kinase (ERK), c-jun N-terminal kinase (JNK) and protein kinase B (Akt). It also suppressed the degradation of I kappa B alpha and blocked p65 translocation into the nucleus. These findings suggest that the mechanism underlying the inhibitory effects of pitavastatin on IL-1ß-induced IL-6 and IL-8 release might be mediated by the suppression of mitogen-activated protein kinase (MAPK), Akt, and nuclear factor-κB (NF-κB) signaling pathways. These results may also indicate that pitavastatin may be potentially utilized as an effective therapeutic agent for the treatment of osteoarthritis.

Hydroxysafflor yellow A inhibits IL-1ß-induced release of IL-6, IL-8, and MMP-1 via suppression of ERK, NF-κB and AP-1 signaling in SW982 human synovial cells.

Hydroxysafflor yellow A (HSYA), the main active ingredient in medical and edible dual purpose plant safflower, is reported to have multiple bioactivities. In the present study, the anti-inflammatory effects of HSYA and the underlying mechanisms were investigated in interleukin (IL)-1ß-induced SW982 human synovial cells. The cells were pretreated with HSYA at various concentrations (2.5, 10 and 40 µM) followed by IL-1ß (10 ng mL-1) stimulation. HSYA significantly inhibited the expression of IL-6, IL-8 and matrix metalloproteinase (MMP)-1 in IL-1ß-stimulated SW982 cells. HSYA also inhibited the phosphorylation of extracellular signal-regulated kinase (ERK), p65 and c-Jun. It also suppressed the degradation of IκBα and blocked p65 translocation into the nucleus. These results indicate that the inhibitory effects of HSYA on IL-1ß-induced IL-6, IL-8 and MMP-1 release might be mediated via suppression of ERK, nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) signaling pathways. The present data support the potential role of HSYA as an effective therapeutic agent in osteoarthritis.

Thrombospondin-4 Promotes Neuronal Differentiation of NG2 Cells via the ERK/MAPK Pathway.

NG2-expressing neural progenitors can produce neurons in the central nervous system, providing a potential cell resource of therapy for neurological disorders. However, the mechanism underlying neuronal differentiation of NG2 cells remains largely unknown. In this report, we found that a thrombospondin (TSP) family member, TSP4, is involved in the neuronal differentiation of NG2 cells. When TSP4 was overexpressed, NG2 cells underwent spontaneous neuronal differentiation, as demonstrated by the induction of various neuronal differentiation markers such as NeuN, Tuj1, and NF200, at the messenger RNA and protein levels. In contrast, TSP4 silencing had an opposite effect on the expression of neuronal differentiation markers in NG2 cells. Next, the signaling pathway responsible for TSP4-mediated NG2 cell differentiation was investigated. We found that ERK but not p38 and AKT signaling was affected by TSP4 overexpression. Furthermore, when ERK signaling was blocked by the inhibitor U0126, the neuronal marker expression of NG2 cells was substantially increased. Together, these findings suggested that TSP4 promoted neuronal differentiation of NG2 cells by inhibiting ERK/MAPK signaling, revealing a novel role of TSP4 in cell fate specification of NG2 cells.

[Inhibitory effect of oleanolic acid on inflammatory response in IL-1ß-stimulated human synovial sarcoma SW982 cells].

To study the role of oleanolic acid on interleukin (IL)-1ß-stimulated expression of inflammatory cytokines, and to explore its anti-inflammatory mechanism in SW982 cells, the toxicity of oleanolic acid on SW982 cells was detected by MTT; effects of different concentrations of oleanolic acid(5, 10, 20 µmol·L(-1)) on the expression of inflammatory factors IL-6, IL-8 and matrix metalloproteinase-1 (MMP-1) was tested at protein and m RNA levels. The study was performed in IL-1ß-stimulated SW982 cells together with enzyme-linked immunosorbent assay (ELISA) and real-time fluorescence quantitative PCR (real-time PCR) methods; the influence of oleanolic acid on the phosphorylation of mitogen-activated protein kinase (MAPK), phosphatidyl inositol-3-kinase/Akt (PI3K/Akt) and nuclear transcription factor-κB (NF-κB) signaling pathways related protein was analyzed by Western blot. Results showed that different concentrations of oleanolic acid(≤40 µmol·L(-1)) were almost non-toxicity to SW982 cells; oleanolic acid significantly inhibited the expression of inflammatory factors in a dose-dependent manner; oleanolic acid restrained extracellular signal-related kinase (ERK), p38, c-jun N-terminal kinase (JNK) and Akt protein phosphorylation and IκB-α protein degradation obviously. The inhibition effect of oleanolic acid on inflammatory factors stimulated by IL-1ß may be worked through MAPK, PI3K/Akt and NF-κB signaling pathways.