Peripheral nerve-derived Sema3A promotes osteogenic differentiation of mesenchymal stem cells through the Wnt/ß-catenin/Nrp1 positive feedback loop.

Sensory nerves play a crucial role in maintaining bone homeostasis by releasing Semaphorin 3A (Sema3A). However, the specific mechanism of Sema3A in regulation of bone marrow mesenchymal stem cells (BMMSCs) during bone remodelling remains unclear. The tibial denervation model was used and the denervated tibia exhibited significantly lower mass as compared to sham operated bones. In vitro, BMMSCs cocultured with dorsal root ganglion cells (DRGs) or stimulated by Sema3A could promote osteogenic differentiation through the Wnt/ß-catenin/Nrp1 positive feedback loop, and the enhancement of osteogenic activity could be inhibited by SM345431 (Sema3A-specific inhibitor). In addition, Sema3A-stimulated BMMSCs or intravenous injection of Sema3A could promote new bone formation in vivo. To sum up, the coregulation of bone remodelling is due to the ageing of BMMSCs and increased osteoclast activity. Furthermore, the sensory neurotransmitter Sema3A promotes osteogenic differentiation of BMMSCs via Wnt/ß-catenin/Nrp1 positive feedback loop, thus promoting osteogenesis in vivo and in vitro.

Naringin activates semaphorin 3A to ameliorate TGF-ß-induced endothelial-to-mesenchymal transition related to atrial fibrillation.

The loss of semaphorin 3A (Sema3A), which is related to endothelial-to-mesenchymal transition (EndMT) in atrial fibrosis, is implicated in the pathogenesis of atrial fibrillation (AF). To explore the mechanisms by which EndMT affects atrial fibrosis and assess the potential of a Sema3A activator (naringin) to prevent atrial fibrosis by targeting transforming growth factor-beta (TGF-ß)-induced EndMT, we used human atria, isolated human atrial endocardial endothelial cells (AEECs), and used transgenic mice expressing TGF-ß specifically in cardiac tissues (TGF-ß transgenic mice). We evaluated an EndMT marker (Twist), a proliferation marker (proliferating cell nuclear antigen; PCNA), and an endothelial cell (EC) marker (CD31) through triple immunohistochemistry and confirmed that both EndMT and EC proliferation contribute to atrial endocardial fibrosis during AF in TGF-ß transgenic mice and AF patient tissue sections. Additionally, we investigated the impact of naringin on EndMT and EC proliferation in AEECs and atrial fibroblasts. Naringin exhibited an antiproliferative effect, to which AEECs were more responsive. Subsequently, we downregulated Sema3A in AEECs using small interfering RNA to clarify a correlation between the reduction in Sema3A and the elevation of EndMT markers. Naringin treatment induced the expression of Sema3A and a concurrent decrease in EndMT markers. Furthermore, naringin administration ameliorated AF and endocardial fibrosis in TGF-ß transgenic mice by stimulating Sema3A expression, inhibiting EndMT markers, reducing atrial fibrosis, and lowering AF vulnerability. This suggests therapeutic potential for naringin in AF treatment.

CircZNF609 sponges miR-135b to up-regulate SEMA3A expression to alleviate ox-LDL-induced atherosclerosis.

The initiation and progression of atherosclerotic plaque caused by abnormal lipid metabolism is one of the main causes of atherosclerosis (AS). Lipid droplet accumulation has become a novel research pointcut for AS treatment in recent years. In AS patients, miR-135b level was up-regulated relative to the normal cases, which showed negative correlations with the levels of Semaphorin 3A (SEMA3A) and circZNF609, separately. The U937-derived macrophages were cultured with ox-LDL to establish AS models in vitro. After that, the lipid accumulation, inflammation, mitochondrial dysfunction and cell death were evaluated by ORO, ELISA, RT-qPCR, western blot, JC-1 and FCM assays respectively. Transfection of the circZNF609 expression vector notably declined lipid accumulation, attenuated inflammation, reduced mitochondrial dysfunction and inhibited cell death in ox-LDL-stimulated cells. The direct binding of miR-135b to circZNF609 in vitro was confirmed using RIP assay, and SEMA3A expression was up-regulated by circZNF609 overexpression. After manipulating the endogenous expressions of circZNF609, miR-135b and SEMA3A, the above damages in ox-LDL-stimulated cells were rescued by inhibition of miR-135b expression and overexpression of circZNF609 or SEMA3A. Besides, the AS mice model was built to demonstrate the excessive lipid accumulation, increasing inflammation and cell death in AS pathogenesis according to the results of HE staining, ELISA and IHC assays, while these damages were reversed after overexpression of circZNF609 or SEMA3A. In AS models, overexpressed circZNF609 prevents the AS progression through depleting miR-135b expression and subsequent up-regulation of SEMA3A expression to overwhelm lipid accumulation, mitochondrial dysfunction and cell death.

Expression of semaphorin-3A in the joint and role in osteoarthritis.

Osteoarthritis (OA) is characterised by the deterioration of cartilage in the joints and pain. We hypothesise that semaphorin-3A (sema-3A), a chemorepellent for sensory nerves, plays a role in joint degradation and pain. We used the mechanical joint loading (MJL) model of OA to investigate sema-3A expression in the joint and examine its association with the development of OA and pain. We also analyse its effect on chondrocyte differentiation using the ATDC5 cell line. We demonstrate that sema-3A is present in most tissues in the healthy joint and its expression increases in highly innervated tissues, such as cruciate ligaments, synovial lining and subchondral bone, in loaded compared to nonloaded control joints. In contrast, sema-3A expression in cartilage was decreased in the severe OA induced by the application of high loads. There was a significant increase in circulating sema-3A, 6 weeks after MJL compared to the nonloaded mice. mRNA for sema-3A and its receptor Plexin A1 were upregulated in the dorsal root ganglia of mice submitted to MJL. These increases were supressed by zoledronate, an inhibitor of bone pain. Sema-3A was expressed at all stages of Chondrocyte maturation and, when added exogenously, stimulated expression of markers of chondrocyte differentiation. This indicates that sema-3A could affect joint tissues distinctively during the development of OA. In highly innervated joint tissues, sema-3A could control innervation and/or induce pain-associated neuronal changes. In cartilage, sema-3A could favour its degeneration by modifying chondrocyte differentiation.

CD72-semaphorin3A axis: A new regulatory pathway in systemic lupus erythematosus.

CD72 is a regulatory co-receptor on B cells, with a role in the pathogenesis of systemic lupus erythematosus (SLE) in both human and animal models. Semaphorin3A (sema3A) is a secreted member of the semaphorin family that can reconstruct B cells' regulatory functions by upregulating IL-10 expression and inhibiting the pro-inflammatory activity of B and T cells in autoimmune diseases. The aim of our present study was to identify a new ligand for CD72, namely sema3A, and exploring the signal transduction pathways following its ligation in B cells. We established that CD72 functions as sema3A binding and signal-transducing receptor. These functions of CD72 are independent of neuropilin-1 (NRP-1) (the known sema3A receptor). We discovered that sema3A induces the phosphorylation of CD72 on tyrosine residues and the association of CD72 with SHP-1 and SHP-2. In addition, the binding of sema3A to CD72 on B cells inhibits the phosphorylation of STAT-4 and HDAC-1 and induces the phosphorylation of p38-MAPK and PKC-theta in B-cells derived B-lymphoblastoid (BLCL) cells, and in primary B-cells isolated from either healthy donors or SLE patients. We concluded that sema3A is a functional regulatory ligand for CD72 on B cells. The sema3A-CD72 axis is a crucial regulatory pathway in the pathogenesis of autoimmune and inflammatory diseases namely SLE, and modulation of this pathway may have a potential therapeutic value for autoimmune diseases.

The role of Semaphorin 3A in oral diseases.

Semaphorin 3A (SEMA3A), also referred to as H-Sema III, is a molecule with significant biological importance in regulating physiological and pathological processes. However, its role in oral diseases, particularly its association with inflammatory immunity and alveolar bone remodeling defects, remains poorly understood. This comprehensive review article aims to elucidate the recent advances in understanding SEMA3A in the oral system, encompassing nerve formation, periodontitis, pulpitis, apical periodontitis, and oral squamous cell carcinoma. Notably, we explore its novel function in inflammatory immunomodulation and alveolar bone formation during oral infectious diseases. By doing so, this review enhances our comprehension of SEMA3A's role in oral biology and opens up possibilities for modulatory approaches and potential treatments in oral diseases.

Assessment of semaphorin 3A and semaphorin 7A levels in primary Sjogren's syndrome.

Sjogren's syndrome (SS) is a chronic autoimmune connective tissue disease. Varying rates of system involvements may be seen in the course of the disease. Semaphorins has multifunctions in several physiological and pathological processes such as immune system regulation. The association of Semaphorin 3A (Sema3A) and Semaphorin 7A (Sema7A), which are immune semaphorins, with autoimmune diseases is interesting for researchers. We aimed to compare serum Sema3A and Sema7A levels between primary SS and control subjects, and investigated Sema3A and Sema7A levels in disease subgroups and associated system involvements. 50 consecutive primary SS patients and 40 healthy subjects followed in the Rheumatology clinic of Cumhuriyet University Medical Faculty between 2017 and 2018 were included in the study. Inclusion criteria of patients were diagnosis of primary SS according to the 2016 ACR/EULAR classification criteria. Serum Sema3A and Sema7A levels were measured by commercial ELISA kit. Both groups were similar in terms of age, gender, and body mass index. Serum Sema3A and Sema7A levels were significantly lower in SS than in the controls (p = 0.001 and p = 0.005, respectively). Serum Sema3A levels were significantly lower in patients with renal involvement than in patients without (p = 0.03). Sema3A and Sema7A may play a role in the etiopathogenesis of SS and may be a potential serological marker for the diagnosis of SS and may be a target for treatment.

Evaluation of serum semaphorin 3A and interleukin 6 levels in patients with pseudoexfoliation syndrome.

PURPOSE: To evaluate serum semaphorin 3A (Sema3A) and interleukin 6 (IL-6) levels in pseudoexfoliation syndrome (PXS) patients to determine whether these mediators play a role in the systemic manifestations of PXS. METHODS: This prospective case-control study included 70 patients divided into PXS (n = 30) and a control group (n = 40). Serum Sema3A and IL-6 levels were analyzed using the enzyme-linked immunosorbent assay. RESULTS: The PXS group had a statistically higher IL-6 level [3.6(0.64-100) pg/mL], compared to the control group [2.1(0.41-39.93) pg/mL] (p < 0.05). On the other hand, the Sema3A level of the PXS group was lower at [21.55(13.2-67.5) ng/mL] compared to the control group at [29.05(11.5-103.3) ng/mL] (p < 0.05). In the PXS group, there was no correlation between the participants' IL-6 values and Sema3A, age, and body mass index (BMI) (r = 0.153, 0.000, - 0.103, respectively, all, p > 0.05), and between Sema3A values and age and BMI values (r = 0.048, - 0.133, respectively, all, p > 0.05). In the control group, there was no correlation between the participants' IL-6 values and Sema3A, age, and BMI values (r = 0.138, - 0.001, - 0.145, respectively, all, p > 0.05) and between the Sema3A and age and BMI values (r = - 0.078, - 0.281, respectively, all, p > 0.05). CONCLUSIONS: Decreased levels of the anti-inflammatory mediator Sema3A and increased levels of inflammatory mediator IL-6 detected in PXS suggest that these molecules may play a role in systemic manifestations of this syndrome, such as inflammation, atherosclerosis, heart arrhythmia, and Alzheimer's disease.

Inhibition of semaphorin-3a alleviates lipopolysaccharide-induced vascular injury.

Alleviating vascular injury improves the prognosis of atherosclerosis. Semaphorin-3a (Sema3A) is a special membrane-associated secreted protein with various biological properties, like pro-inflammation, anti-tumor and et al. This study aims to investigate the effects of inhibition of Sema3A on lipopolysaccharide (LPS)-induced vascular injury in mice. The mice were randomized into three groups: control, LPS, and LPS + siRNA. Mice in the combined group were given siRNA through fast tail vein injection, then LPS was injected intraperitoneally 7 days later, finally the mice were euthanized 24 h later. Vascular function and structure were assessed by vascular injury biomarkers and relevant stainings. LPS-induced vascular dysfunction and pathological injury were substantially improved by inhibition of Sema3A. Western blot and quantitative real-time polymerase chain reaction assays were used for investigating molecular pathways. The relevant proteins of vascular endothelial cells activation, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), increased after LPS stimulation, while these effects were reversed by inhibition of Sema3A. The levels of inflammatory cytokines (IL-1ß, IL-6 and NLRP3) were upregulated after LPS stimulation, however, inhibition of Sema3A reversed it through NF-κB and MAPKs signaling pathways involvement. Moreover, inhibition of Sema3A alleviated LPS-induced oxidative stress, evidenced by a decrease in total reactive oxygen species and an increase in antioxidant protein of SOD-1. The results showed that inhibition of Sema3A protects against LPS-induced vascular injury by suppressing vascular endothelial cells activation, vascular inflammation, and vascular oxidative stress, implying that inhibition of Sema3A might be used as a therapeutic strategy for septic vascular injury or atherosclerosis.

Sema3A accelerates bone formation during distraction osteogenesis in mice.

Distraction osteogenesis (DO) is a bone regeneration technique used to treat maxillofacial disorders, fracture nonunion, and large bone defects. It is well known for its amazing regenerative potential, but an extended consolidation period limits its clinical use. The interaction between the nervous system and bone regeneration has attracted great attention in recent years. Sema3A is a key axonal chemorepellent which has been proved to have bone-protective effects. In this article, we try to improve DO by local administration of Sema3A and explore the possible mechanisms. Forty wildtype, male, adult mice were divided into two groups after tibia osteotomy surgery. Sema3A or Saline was daily injected transcutaneous into the center of the distraction zone during the consolidation period. Micro-CT images were taken at 4, 6,8 and 10 weeks post-surgery; vascular density and biomechanical testing were performed at 10 weeks post-surgery. We also set up in vitro vessel growth assay to evaluate the effect of Sema3A on angiogenesis. Compared with the Saline group, Sema3A treatment significantly accelerated bone regeneration, improved angiogenesis and callus' biomechanical strength. At 10 weeks post-surgery, compared with the Saline group, the BV/TV, BMD, TMD increased by about 23%, 22%, 18% respectively, vascular density increased by about 49% in the Sema3A group. Histological images and western-blot showed decreased expression of VEGF-A and increased expression of Ang-1 at 4 weeks post-surgery in the Sema3A group. In vitro, Sema3A suppressed VEGF-induced angiogenesis but had little effect on Ang-induced angiogenesis. Conclusion: Sema3A could accelerate bone regeneration and improve angiogenesis during DO.

The Responsiveness of Thymic Stromal Cells to semaphorin-3A.

BACKGROUND: The thymus is responsible for thymocyte differentiation into immunocompetent T lymphocytes. Different cell types in the thymic microenvironment actively cooperate in this process, interacting with the developing thymocytes through soluble factors, extracellular matrix (ECM) molecules, and receptors. In addition, this microenvironment can be influenced by several factors, such as semaphorin-3A (Sema3A), which is a multifunctional protein involved in cell migration. We evaluated the Sema3A effects on the cellular parameters and functional features of thymic stromal cells. METHODS: Thymic stromal cells were obtained by enzymatic digestion of the murine thymus. These cells were treated with Sema3A and evaluated as follows: cell morphology by scanning electron microscope, F-actin cytoskeleton and deposition of ECM molecules by fluorescence microscopy, and adhesion assays with freshly obtained thymocytes. RESULTS: The obtained thymic stroma was composed of 67% of thymic epithelial cells (TECs), and 90% of the TECs were positive for the Sema3A receptor neuropilin-1. These cells secreted CXCL12, IL-7 and extended thymocyte survival. Sema3A changed the morphology of thymic stromal cells and promoted F-actin reorganization. In addition, the fibronectin fibers were reoriented, and the laminin production was increased in Sema3A-treated thymic stromal cells. In the adhesion assays, there was an increase in the number of adhered thymocytes when thymic stromal cells were pretreated with Sema3A. CONCLUSION: Our data strongly suggest the active participation of Sema3A in thymic physiology, highlighting its role as an immunomodulatory molecule. This may provide important knowledge for understanding the interactions of thymic cells.

Gingival crevicular fluid lipocalin-2 and semaphorin3A in stage III periodontitis: Non-surgical periodontal treatment effects.

BACKGROUND AND OBJECTIVE: Identification of biomarkers to assess individual risk and monitor periodontal health status is important. Research on lipocalin-2 (LCN2) and semaphorin3A (Sema3A) is lacking. This study aimed to evaluate gingival crevicular fluid (GCF) LCN2, Sema3A, and tumor necrosis factor-α (TNF-α) levels in periodontally healthy (H), gingivitis (G), and periodontitis (P) patients, and their changes following non-surgical periodontal therapy. METHODS: Sixty systemically healthy and non-smoker participants, diagnosed as periodontally healthy, gingivitis, and stage III grade C periodontitis, were recruited (n = 20/group). Clinical periodontal parameters were recorded and GCF samples were obtained at baseline from all groups; for group P, these were repeated one and three months following non-surgical periodontal treatment. GCF LCN2, Sema3A, and TNF-α levels were evaluated with enzyme-linked immunosorbent assay. RESULTS: GCF LCN2, Sema3A, and TNF-α total amounts were significantly higher in disease groups than group H (p < .001). Between P and G groups, only TNF-α levels were significantly different (p < .001). Non-surgical periodontal therapy resulted in significant improvement of all clinical parameters and significant decreases of GCF LCN2 and TNF-α levels, at both time points, compared with baseline (p < .001). Sema3A levels remained unchanged following treatment (p > .05). LCN2 and TNF-α levels were significantly positively correlated with clinical parameters. LCN2 (AUC [area under the curve] = 0.94) and TNF-α (AUC = 0.98) levels were similarly accurate in differentiating between periodontal disease (whether G or P) and healthy controls. CONCLUSIONS: LCN2 and TNF-α levels in GCF are correlated with clinical parameters and could prove useful as non-invasive screening tools for periodontitis.

Semaphorin 3A protects against alveolar bone loss during orthodontic tooth movement in mice with periodontitis.

OBJECTIVE: This study investigated the effect of local semaphorin 3A (Sema3A) administration on alveolar bone loss during OTM in a mouse model of periodontitis. BACKGROUND: Orthodontic tooth movement (OTM) for patients with periodontal disease is known to increase the risk of exacerbating alveolar bone loss due to inflammation of the periodontal tissue. However, its mechanism of action and prevention remains unclear. METHODS: Mice (male 7-8 weeks old, C57BL/6J, n = 12) were divided into six groups: untreated group (control), without OTM and recovered from induced periodontitis (RP), with OTM and administered PBS or Sema3A to the gingiva after induced periodontitis (VehPO, SemaPO), with OTM and administered PBS or Sema3A to the gingiva without periodontitis induction (VehNO, SemaNO). Samples were collected on 14 days, and bone loss, histological analysis, cytokine production level, and tooth movement were assessed. Cultured human periodontal ligament (hPDL) cells were stimulated with lipopolysaccharide (LPS) and compressive force (CF), and mRNA expression levels of Sema3A and its receptors were analyzed. RESULTS: The bone loss was significantly lower in the SemaPO group than in the VehPO group. The number of TRAP-positive cells in the SemaPO group was significantly lower than that in the VehPO group and was at the same level as that in the control group. The receptor activator of nuclear factor (NF)-kB-ligand/osteoprotegerin (RANKL/OPG) ratio and the levels of proinflammatory cytokines, including interleukin (IL)-1ß, IL-6, IL-17, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ, in the gingival tissues were significantly lower in the SemaPO group than in the VehPO group. Additionally, Sema3A mRNA expression in hPDL cells was significantly decreased by co-stimulation with LPS and CF compared with that in the control group. Finally, the distance moved (dist.) and the mesial tipping angle (θ) was significantly smaller in the SemaPO group than in the VehPO group and was not significantly different from that of VehNO. CONCLUSION: Pathological alveolar bone loss exacerbated by OTM in periodontitis might be prevented by local administration of Sema3A without inhibiting OTM.

Semaphorin 3A attenuates the hypoxia suppression of osteogenesis in periodontal ligament stem cells.

OBJECTIVE AND BACKGROUND: The occurrence and development of periodontitis are closely related to hypoxia of the periodontal microenvironment. Periodontal ligament stem cells (PDLSCs) are considered to have potential to regenerate periodontal tissues. Semaphorin 3A (Sema3A) plays an essential role in promoting osteogenesis. However, the effect of Sema3A on osteogenesis of PDLSCs under hypoxia remains unclear. The aim of this study was to investigate the effect of Sema3A on osteogenesis of PDLSCs under hypoxia. METHODS: Isolated PDLSCs were identified using flow cytometry. Adipogenic differentiation potential was identified by oil red O staining. Osteogenesis was measured using Alizarin Red S staining and ALP staining. Intracellular hypoxia was induced using cobalt chloride (CoCl2 ). The expression level of hypoxia-inducible factor-1α (HIF-1α) was detected via ELISA. Expression of osteogenic markers and Sema3A was analyzed using western blot and real-time PCR. RESULTS: The proliferation and osteogenesis of PDLSCs were markedly inhibited with increased concentrations of CoCl2 . Under the treatment with a low concentration of CoCl2 , expression of related osteogenic markers and Sema3A decreased in a time-dependent manner. ARS and ALP staining results also showed that osteogenic calcification decreased under hypoxia. Apigenin, an inhibitor of HIF-1α, effectively up-regulated expression of Sema3A and osteogenic markers with CoCl2 treatment. Moreover, exogenous Sema3A significantly increased the expression of osteogenesis-related markers and mineralization of PDLSCs according to ALP and ARS staining with CoCl2 treatment. CONCLUSIONS: Hypoxia markedly inhibited osteogenesis of PDLSCs. Sema3A explicitly attenuated the hypoxia suppression of osteogenesis in PDLSCs.

Semaphorin 3A in the Immune System: Twenty Years of Study.

Semaphorin 3A is a secreted glycoprotein, which was originally identified as axon guidance factor in the neuronal system, but it also possesses immunoregulatory properties. Here, the effect of semaphorin 3A on T-lymphocytes, myeloid dendritic cells and macrophages is systematically analyzed on the bases of all publications available in the literature for 20 years. Expression of semaphorin 3A receptors - neuropilin-1 and plexins A - in these cells is described in details. The data obtained on human and murine cells is described comparatively. A comprehensive overview of the interaction of semaphorin 3A with mononuclear phagocyte system is presented for the first time. Semaphorin 3A signaling mostly results in changes of the cytoskeletal machinery and cellular morphology that regulate pathways involved in migration, adhesion, and cell-cell cooperation of immune cells. Accumulating evidence indicates that this factor is crucially involved in various phases of immune responses, including initiation phase, antigen presentation, effector T cell function, inflammation phase, macrophage activation, and polarization. In recent years, interest in this field has increased significantly because semaphorin 3A is associated with many human diseases and therefore can be used as a target for their treatment. Its involvement in the immune responses is important to study, because semaphorin 3A and its receptors turn to be a promising new therapeutic tools to be applied in many autoimmune, allergic, and oncology diseases.

Semaphorin3A promotes osseointegration of titanium implants in osteoporotic rabbits.

OBJECTIVE: In the present study, we intend to assess the function of Sema3A in osteointegration of titanium implants both in vivo and in vitro. MATERIAL AND METHODS: Briefly, Sema3A was transfected in HBMSCs cells to detect its effect on osteogenesis. Subsequently, an in vivo rabbit model was established. Eighteen female rabbits were randomly assigned into three groups (n=6), and rabbits in the two treatment groups (OVX groups) were subjected to bilateral ovariectomy, while those in the control group were treated with sham operation. Twelve weeks later, we first examined expression levels of Sema3A in rabbits of the three groups. Titanium implants were implanted in rabbit proximal tibia. Specifically, rabbits in sham group were implanted with Matrigel, while the remaining in the OVX experimental group (OVX+Sema3A group) and OVX group were implanted with Matrigel containing Sema3A adeno-associated virus or empty vector, respectively. RESULTS: Histomorphometry results uncovered that rabbits in the OVX+Sema3A group had a significantly higher BIC compared with those of the OVX group on the 12th week of post-implantation. And compared with the OVX group, the maximum push-out force increased by 89.4%, and the stiffness increased by 39.4%, the toughness increased by 63.8% in the OVX+Sema3A group at 12 weeks. CONCLUSION: Sema3A has a positive effect on promoting early osseointegration of titanium implants in osteoporotic rabbits. CLINICAL RELEVANCE: Our research found that Sema3A can improve the osteogenic ability of bone marrow stem cells and promotes osseointegration during osteoporosis.

Semaphorin 3A-Neuropilin-1 Signaling Modulates MMP13 Expression in Human Osteoarthritic Chondrocytes.

Osteoarthritis (OA) is a complex disorder of diarthrodial joints caused by multiple risk factors and is characterized by articular cartilage destruction as well as changes in other articular tissues. Semaphorin 3A (Sema3A), known to be a chemo-repellent for sensory nerve fibers, has recently been implicated in cartilage OA pathophysiology. We demonstrated that the expression of SEMA3A and its receptor neuropilin-1 (NRP1) are synchronously upregulated in chondrocytes isolated from knee cartilage of OA patients compared to non-OA control chondrocytes. In addition, we observed that during in vitro passaging of OA chondrocytes, the Nrp-1 level increases, whereas the Sema3A level decreases. In this study, we aimed to uncover how Sema3A-Nrp-1 signaling affects metabolism and viability of OA chondrocytes via siRNA-mediated inhibition of Nrp-1 expression. We observed a decreased proliferation rate and an increase in adhesion and senescence after Nrp-1 silencing. Moreover, MMP13 gene expression was reduced by approximately 75% in NRP1 knockdown OA chondrocytes, whereas MMP13 expression was induced by Sema3A treatment in control (nt siRNA) OA chondrocytes, accompanied by an impaired AKT phosphorylation. These findings suggest a potential catabolic function of Sema3A signaling in OA chondrocytes by inducing MMP13 expression and by compromising pro-survival AKT activation. We propose that targeting the Sema3A-Nrp-1 signaling axis might be an opportunity to interfere with OA pathogenesis and progression.

Semaporin3A inhibitor ameliorates renal fibrosis through the regulation of JNK signaling.

Renal fibrosis is the common pathological pathway in progressive renal diseases. In the present study, we analyzed the roles of semaphorin 3 A (SEMA3A) on renal fibrosis and the effect of SEMA3A inhibitor (SEMA3A-I) using a unilateral ureteral obstruction (UUO) mouse model. Expression of SEMA3A in the proximal tubulus and neuropilin-1, a recepor of SEMA3A, in fibloblast and tubular cells were increased in UUO kidneys. The expression of myofibroblast marker tenascin-C and fibronection as well as renal fibrosis were increased in UUO kidneys, all of which were ameliorated by SEMA3A-I. In addition, the JNK signaling pathway, known as the target of SEMA3A signaling, was activated in proximal tubular cells and fibroblast cells after UUO surgery, and SEMA3A-I significantly attenuated the activation. In vitro, treatments with SEMA3A as well as transforming growth factor-ß1 (TGF-ß1) in human proximal tubular cells lost epithelial cell characteristics, and SEMA3A-I significantly ameliorated this transformation. The JNK inhibitor SP600125 partially reversed SEMA3A and TGF-ß1-induced cell transformation, indicating that JNK signaling is involved in SEMA3A-induced renal fibrosis. In addition, treatment with SEMA3A in fibroblast cells activated expression of tenascin-C, collagen type I, and fibronection, indicating that SEMA3A may accelerate renal fibrosis through the activation of fibroblast cells. Analysis of human data revealed the positive correlation between urinary SEMA3A and urinary N-acetyl-ß-d-glucosaminidase, indicating the association between SEMA3A and tubular injury. In conclusion, SEMA3A signaling is involved in renal fibrosis through the JNK signaling pathway and SEMA3A-I might be a therapeutic option for protecting from renal fibrosis.NEW & NOTEWORTHY Renal fibrosis is the common pathological pathway in the progression of renal diseases. This study, using a unilateral ureteral obstruction (UUO) mouse model, indicated increased semaphorin3A (SEMA3A) signaling in renal tubular cells as well as fibroblast cells under UUO surgery, and SEMA3A inhibitor ameliorated UUO-induced renal fibrosis through the regulation of JNK signaling. The study proposes the potential therapeutic option of SEMA3A inhibitor to treat renal fibrosis.

Decreased Presence of Mast Cells in the Bursa Premacularis of Proliferative Diabetic Retinopathy.

INTRODUCTION: We previously reported that the intravitreal activities of chymase and tryptase were more increased in the patients with macular hole (MH) and epiretinal membrane (ERM) than in those with proliferative diabetic retinopathy (PDR) and that the source of these serine proteases might be mast cells in the bursa premacularis (BPM). The purpose of this study was to compare the density of mast cells in BPM samples obtained from MH, ERM, and PDR patients. METHODS: BPM and vitreous core samples were first collected during vitrectomy from eyes afflicted with vitreoretinal diseases (MH: 6 eyes, ERM: 3 eyes, and PDR: 9 eyes), and then were stained with hematoxylin, toluidine blue, antibodies against chymase and tryptase, and a terminal deoxynucleotidyl transferase dUTP nick end labeling assay kit. RESULTS: Hematoxylin nuclear staining showed fewer positive-staining cells in the BPM samples obtained from PDR patients than in those obtained from MH and ERM patients. Toluidine blue staining of the BPM revealed metachromasia in the mast cells of the patients with MH and ERM, but not those of the patients with PDR. In addition, immunostaining using anti-chymase and anti-tryptase antibodies showed that the BPM samples were more intensely stained than the vitreous core samples from the patients with MH and ERM and that both tissue samples were poorly stained in the patients with PDR. The apoptotic cells were more frequently observed in the BPM samples from patients with MH than in those from patients with PDR. CONCLUSIONS: These findings indicated that lower activities of chymase and tryptase in the vitreous of PDR patients appeared to be attributable to the decreased presence of mast cells in the BPM. The lack of mast cells in the BPM might be related to the pathogenesis of PDR.

Human epicardium-derived cells reinforce cardiac sympathetic innervation.

RATIONALE: After cardiac damage, excessive neurite outgrowth (sympathetic hyperinnervation) can occur, which is related to ventricular arrhythmias/sudden cardiac death. Post-damage reactivation of epicardium causes epicardium-derived cells (EPDCs) to acquire a mesenchymal character, contributing to cardiac regeneration. Whether EPDCs also contribute to cardiac re/hyperinnervation, is unknown. AIM: To investigate whether mesenchymal EPDCs influence cardiac sympathetic innervation. METHODS AND RESULTS: Sympathetic ganglia were co-cultured with mesenchymal EPDCs and/or myocardium, and neurite outgrowth and sprouting density were assessed. Results showed a significant increase in neurite density and directional (i.e. towards myocardium) outgrowth when ganglia were co-cultured with a combination of EPDCs and myocardium, as compared to cultures with EPDCs or myocardium alone. In absence of myocardium, this outgrowth was not directional. Neurite differentiation of PC12 cells in conditioned medium confirmed these results via a paracrine effect, in accordance with expression of neurotrophic factors in myocardial explants co-cultured with EPDCs. Of interest, EPDCs increased the expression of nerve growth factor (NGF) in cultured, but not in fresh myocardium, possibly due to an "ischemic state" of cultured myocardium, supported by TUNEL and Hif1α expression. Cardiac tissues after myocardial infarction showed robust NGF expression in the infarcted, but not remote area. CONCLUSION: Neurite outgrowth and density increases significantly in the presence of EPDCs by a paracrine effect, indicating a new role for EPDCs in the occurrence of sympathetic re/hyperinnervation after cardiac damage.