Porphyromonas gingivalis Impairs Oral Epithelial Barrier through Targeting GRHL2.

Oral mucosa provides the first line of defense against a diverse array of environmental and microbial irritants by forming the barrier of epithelial cells interconnected by multiprotein tight junctions (TJ), adherens junctions, desmosomes, and gap junction complexes. Grainyhead-like 2 (GRHL2), an epithelial-specific transcription factor, may play a role in the formation of the mucosal epithelial barrier, as it regulates the expression of the junction proteins. The current study investigated the role of GRHL2 in the Porphyromonas gingivalis (Pg)-induced impairment of epithelial barrier functions. Exposure of human oral keratinocytes (HOK-16B and OKF6 cells) to Pg or Pg-derived lipopolysaccharides (Pg LPSs) led to rapid loss of endogenous GRHL2 and the junction proteins (e.g., zonula occludens, E-cadherin, claudins, and occludin). GRHL2 directly regulated the expression levels of the junction proteins and the epithelial permeability for small molecules (e.g., dextrans and Pg bacteria). To explore the functional role of GRHL2 in oral mucosal barrier, we used a Grhl2 conditional knockout (KO) mouse model, which allows for epithelial tissue-specific Grhl2 KO in an inducible manner. Grhl2 KO impaired the expression of the junction proteins at the junctional epithelium and increased the alveolar bone loss in the ligature-induced periodontitis model. Fluorescence in situ hybridization revealed increased epithelial penetration of oral bacteria in Grhl2 KO mice compared with the wild-type mice. Also, blood loadings of oral bacteria (e.g., Bacteroides, Bacillus, Firmicutes, ß-proteobacteria, and Spirochetes) were significantly elevated in Grhl2 KO mice compared to the wild-type littermates. These data indicate that Pg bacteria may enhance paracellular penetration through oral mucosa in part by targeting the expression of GRHL2 in the oral epithelial cells, which then impairs the epithelial barrier by inhibition of junction protein expression, resulting in increased alveolar tissue destruction and systemic bacteremia.

Clopidogrel may decrease the risk of post-stroke infection after ischaemic stroke.

BACKGROUND AND PURPOSE: The P2Y12 receptor, a well-known factor in the platelet activation pathway, plays a role in thrombosis as well as systemic inflammation. Clopidogrel, a prototype P2Y12 receptor antagonist, reportedly decreases inflammation and systemic infection. The aim of this study was to evaluate whether clopidogrel use decreases the risk of post-stroke infection following ischaemic stroke. METHODS: A total of 1643 patients with acute ischaemic stroke (within 7 days after onset) were included for analysis between March 2010 and December 2015. Patients were categorized into two groups (clopidogrel users versus clopidogrel non-users), and clinical characteristics and risks of post-stroke infection were compared between the two groups. The inverse probability of treatment weighting using propensity scores for baseline imbalance adjustments was applied. RESULTS: Of the included patients (mean age 67.7 years; men 60.6%), 670 (40.8%) patients were clopidogrel users and 164 (10.0%) patients had post-stroke infection. The proportion of patients with post-stroke infection was significantly lower in clopidogrel users compared to clopidogrel non-users (6.7% vs. 12.2%, P ≤ 0.001). Moreover, clopidogrel users were less likely to be admitted to the intensive care unit (13.3% vs. 35.3%, P = 0.006). A multivariate analysis with inverse probability of treatment weighting revealed that clopidogrel users exhibited a lower risk of post-stroke infection (odds ratio 0.56, 95% confidence interval 0.42-0.75) and intensive care unit admission (odds ratio 0.34, 95% confidence interval 0.22-0.53). CONCLUSIONS: The study suggested that clopidogrel users exhibit a lower risk of infection and develop less severe infections after ischaemic stroke. Further prospective studies are needed.

Effects of ultrasonic instrumentation on enamel surfaces with various defects.

OBJECTIVE: The aim of this study was to analyse the enamel damage caused by ultrasonic scaling of teeth with various enamel conditions that are difficult to identify by visual inspection, such as enamel cracks, early caries and resin restorations. METHODS: In total, 120 tooth surfaces were divided into 4 experimental groups using a quantitative light-induced fluorescence-digital system: sound enamel group, enamel cracks group, early caries group and resin restoration group. A skilled dental hygienist performed ultrasonic scaling under a standardized set of conditions: a ≤ 15° angle between the scaler tip and tooth surface and 40-80 g of lateral pressure at the rate of 12 times/10 s. Following scaling, the depth of enamel damage was measured using a surface profilometer and observed using scanning electron microscopy (SEM). RESULTS: The damage depth was the greatest in the enamel cracks group (37.63 ± 34.42 µm), followed by the early caries group (26.81 ± 8.67 µm), resin restoration group (19.63 ± 6.73 µm) and the sound enamel group (17.00 ± 5.66 µm). The damage depth was significantly deeper in the enamel cracks and early caries groups than in the sound enamel group (P < .05). SEM clearly revealed enamel loss in the enamel cracks, early caries and resin restoration groups. CONCLUSIONS: The results of this study suggest that ultrasonic scaling can cause further damage to teeth with enamel cracks, early caries and resin restorations. Therefore, accurate identification of tooth conditions and calculus before the initiation of ultrasonic scaling is necessary to minimize damage.

Human Papillomavirus 16 E6 Induces FoxM1B in Oral Keratinocytes through GRHL2.

High-risk human papillomavirus (HPV) is a major risk factor for oral and pharyngeal cancers (OPCs), yet the detailed mechanisms by which HPV promotes OPCs are not understood. Forkhead box M1B (FoxM1B) is an oncogene essential for cell cycle progression and tumorigenesis, and it is aberrantly overexpressed in many tumors. We previously showed that FoxM1B was the putative target of an epithelial-specific transcription factor, Grainyhead-like 2 (GRHL2). In the current study, we demonstrate that HPV type 16 (HPV-16) E6 induces FoxM1B in human oral keratinocytes (HOKs) and tonsillar epithelial cells (TECs) in part through GRHL2. FoxM1B was barely detectable in cultured normal human oral keratinocytes (NHOKs) and progressively increased in immortalized HOKs harboring HPV-16 genome (HOK-16B) and tumorigenic HOK-16B/BaP-T cells. Retroviral expression of HPV-16 E6 and/or E7 in NHOKs, TECs, and hypopharyngeal carcinoma cells (FaDu) revealed induction of FoxM1B and GRHL2 by the E6 protein but not E7. Both GRHL2 and FoxM1B were strongly induced in the epidermis of HPV-16 E6 transgenic mice and HPV+ oral squamous cell carcinomas. Ectopic expression of FoxM1B led to acquisition of transformed phenotype in HOK-16B cells. Loss of FoxM1B by lentiviral short hairpin RNA vector or chemical inhibitor led to elimination of tumorigenic characteristics of HOK-16B/BaP-T cells. Luciferase reporter assay revealed that GRHL2 directly bound and regulated the FoxM1B gene promoter activity. Using epithelial-specific Grhl2 conditional knockout mice, we exposed wild-type (WT) and Grhl2 KO mice to 4-nitroquinolin 1-oxide (4-NQO), which led to induction of FoxM1B in the tongue tissues and rampant oral tumor development in the WT mice. However, 4-NQO exposure failed to induce tongue tumors or induction of FoxM1B expression in Grhl2 KO mice. Collectively, these results indicate that HPV-16 induces FoxM1B in part through GRHL2 transcriptional activity and that elevated FoxM1B level is required for oropharyngeal cancer development.

Clastic cells are absent around the root surface in pulp-exposed periapical periodontitis lesions in mice.

INTRODUCTION: Clastic cells, originating from the monocyte-macrophage lineage, resorb mineralized tissues. In periapical periodontitis, alveolar bone around the tooth apex becomes resorbed; however, the roots of the teeth are often left intact by yet unknown mechanisms. Here, we examined the status of clastic cells in a periapical periodontitis model in mice. METHODS: Periapical periodontitis was induced by performing pulp exposure on the maxillary first molar. The contralateral maxillary first molar was used as a control. The maxillae were harvested, fixed, and subjected to µCT scanning and three-dimensional volumetric analysis. TRAP staining was performed, and osteoclasts were quantified. Immunohistochemical staining was performed for RANKL, OPG, and F4/80, a marker for macrophages. RESULTS: At the apex of the tooth, pulp exposure resulted in periapical radiolucency with mineralized tissues at the surrounding bone surfaces but not on the root surfaces. Histologically, clastic cells were present on the bone surfaces but absent around the root surfaces. Expression of F4/80 and RANKL was not found at close proximity to the root surfaces, but OPG was globally expressed. CONCLUSION: The absence of clastic cells around the root surface of pulp-exposed teeth, in part, is associated with the lack of macrophages and RANKL expression.

The utility of intra-operative frozen section for the evaluation of microscopic extrathyroidal extension in papillary thyroid carcinoma.

OBJECTIVES: This study was designed to evaluate the usefulness of intra-operative frozen section for the evaluation of microscopic extrathyroidal extension (ETE) in papillary thyroid carcinoma (PTC). DESIGN: Retrospective cohort study. SETTING: Dong-A University Medical Center, Busan, Korea. PARTICIPANTS: Three hundred and sixty-four patients who underwent thyroid surgery from January 2000 to December 2010 with PTC confined to one unilateral lobe as diagnosed using preoperative ultrasonography were enrolled. MAIN OUTCOME MEASURES: The patients who had microscopic ETE on frozen section were classified into "group A," and those who did not have microscopic ETE on frozen section were classified into "group B." Clinicopathologic factors including age, gender, size of the tumour, extent of operation, ETE, multifocality, bilaterality, lymph node metastasis and recurrence were compared between the two groups. RESULTS: Of the 364 patients enrolled, ETE was confirmed in 100 patients (group A, 27.5%) on frozen biopsy. The nodule size in group A (0.94±0.87 cm) was larger than that in group B (0.86±0.79 cm) (P=.042). In group A, 15 patients (15%) showed multifocality and 11 patients (14.47%) showed bilaterality. In group B, 37 patients (14.02%) showed multifocality and seven patients (43.35%) showed bilaterality. They did not differ significantly between the two groups (P=.811, P=.182). There was a higher frequency of lymph node metastases in group A (52/86, 60.47%) than in group B (7/16, 43.75%, P=.214). Recurrence was observed in only two patients who had received thyroid lobectomy as the initial surgery in group A. CONCLUSIONS: Intra-operative frozen biopsy can be a useful method for identifying the microscopic ETE. During the surgery, it can also help the surgeon to decide the optimal extent of surgery and the need for central compartment neck dissection in PTC patients.

Effects of Bioactive Compounds on Odontogenic Differentiation and Mineralization.

Direct pulp capping involves the placement of dental materials directly onto vital pulp tissues after deep caries removal to stimulate the regeneration of reparative dentin. This physical barrier will serve as a "biological seal" between these materials and the pulp tissue. Although numerous direct pulp capping materials are available, the use of small bioactive compounds that can potently stimulate and expedite reparative dentin formation is still underexplored. Here, the authors compared and evaluated the pro-osteogenic and pro-odontogenic effects of 4 small bioactive compounds- phenamil (Phen), purmorphamine (Pur), genistein (Gen), and metformin (Met). The authors found that these compounds at noncytotoxic concentrations induced differentiation and mineralization of preosteoblastic MC3T3-E1 cells and preodontoblastic dental pulp stem cells (DPSCs) in a dose-dependent manner. Among them, Phen consistently and potently induced differentiation and mineralization in vitro. A single treatment with Phen was sufficient to enhance the mineralization potential of DPSCs in vitro. More importantly, Phen-treated DPSCs showed enhanced odontogenic differentiation and mineralization in vivo. Our study suggests that these small bioactive compounds merit further study for their potential clinical use as pulp capping materials.

Atypia of undetermined significance on thyroid fine needle aspiration - risk factors for malignancy.

OBJECTIVES: This study is designed to determine the clinical predictors of malignancy in the atypia of undetermined significance (AUS) category resulted from thyroid fine needle aspiration (FNA). DESIGN: Retrospective cohort study. SETTING: Dong-A University Medical Center, Busan, Korea. PARTICIPANTS: Sixty-two patients who underwent thyroid surgery from January 2010 to December 2013, following a diagnosis of AUS from preoperative thyroid FNA. MAIN OUTCOME MEASURES: We investigated the age, gender, maximum size and site of the nodules, ultrasonographic findings, cytological features, BRAF gene mutation, surgical method, number of AUS on repeated FNA and final pathologic results. RESULTS: Forty-one of sixty-two patients underwent total thyroidectomy and the rest had lobectomy. The final pathologic results were 41 malignancies and 21 benign diseases. Nodules less than 1.5 cm, ultrasonographic findings suggestive of malignancy were risk factors for malignancy on univariated analysis (P < 0.001). Multivariated analysis showed that nodules less than 1.5 cm, ultrasonographic findings suggestive of malignancy and more than 2 results of atypia from repeated FNAs were significant risk factors for malignancy (P < 0.001). A BRAF gene mutation analysis was performed in 38 patients, and 13 patients had the mutation. All patients with the BRAF gene mutation had been diagnosed with papillary thyroid cancer (P > 0.05). CONCLUSIONS: We recommend close observation or diagnostic surgery in patients with nodules <1.5 cm and with two or more malignant ultrasound feature and a BRAF mutation, or with two or more AUS findings on repeated FNAs.

Epigenetic gene regulation by histone demethylases: emerging role in oncogenesis and inflammation.

Histone N-terminal tails of nucleosomes are the sites of complex regulation of gene expression through post-translational modifications. Among these modifications, histone methylation had long been associated with permanent gene inactivation until the discovery of Lys-specific demethylase (LSD1), which is responsible for dynamic gene regulation. There are more than 30 members of the Lys demethylase (KDM) family, and with exception of LSD1 and LSD2, all other KDMs possess the Jumonji C (JmjC) domain exhibiting demethylase activity and require unique cofactors, for example, Fe(II) and α-ketoglutarate. These cofactors have been targeted when devising KDM inhibitors, which may yield therapeutic benefit. KDMs and their counterpart Lys methyltransferases (KMTs) regulate multiple biological processes, including oncogenesis and inflammation. KDMs' functional interactions with retinoblastoma (Rb) and E2 factor (E2F) target promoters illustrate their regulatory role in cell cycle progression and oncogenesis. Recent findings also demonstrate the control of inflammation and immune functions by KDMs, such as KDM6B that regulates the pro-inflammatory gene expression and CD4+ T helper (Th) cell lineage determination. This review will highlight the mechanisms by which KDMs and KMTs regulate the target gene expression and how epigenetic mechanisms may be applied to our understanding of oral inflammation.

The Role of ORAI1 in the Odontogenic Differentiation of Human Dental Pulp Stem Cells.

Pulp capping, or placing dental materials directly onto the vital pulp tissues of affected teeth, is a dental procedure that aims to regenerate reparative dentin. Several pulp capping materials are clinically being used, and calcium ion (Ca(2+)) released from these materials is known to mediate reparative dentin formation. ORAI1 is an essential pore subunit of store-operated Ca(2+) entry (SOCE), which is a major Ca(2+) influx pathway in most nonexcitable cells. Here, we evaluated the role of ORAI1 in mediating the odontogenic differentiation and mineralization of dental pulp stem cells (DPSCs). During the odontogenic differentiation of DPSCs, the expression of ORAI1 increased in a time-dependent manner. DPSCs knocked down with ORAI1 shRNA (DPSC/ORAI1sh) or overexpressed with dominant negative mutant ORAI1(E106Q) (DPSC/E106Q) exhibited the inhibition of Ca(2+) influx and suppression of odontogenic differentiation and mineralization as demonstrated by alkaline phosphatase (ALP) activity/staining as well as alizarin red S staining when compared with DPSCs of their respective control groups (DPSC/CTLsh and DPSC/CTL). The gene expression for odontogenic differentiation markers such as osteocalcin, bone sialoprotein, and dentin matrix protein 1 (DMP1) was also suppressed. When DPSC/CTL or DPSC/E106Q cells were subcutaneously transplanted into nude mice, DPSC/CTL cells induced mineralized tissue formation with significant increases in ALP and DMP1 staining in vivo, whereas DPSC/E106Q cells did not. Collectively, our data showed that ORAI1 plays critical roles in the odontogenic differentiation and mineralization of DPSCs by regulating Ca(2+) influx and that ORAI1 may be a therapeutic target to enhance reparative dentin formation.

Pulp-dentin Regeneration: Current State and Future Prospects.

The goal of regenerative endodontics is to reinstate normal pulp function in necrotic and infected teeth that would result in reestablishment of protective functions, including innate pulp immunity, pulp repair through mineralization, and pulp sensibility. In the unique microenvironment of the dental pulp, the triad of tissue engineering would require infection control, biomaterials, and stem cells. Although revascularization is successful in resolving apical periodontitis, multiple studies suggest that it alone does not support pulp-dentin regeneration. More recently, cell-based approaches in endodontic regeneration based on pulpal mesenchymal stem cells (MSCs) have demonstrated promising results in terms of pulp-dentin regeneration in vivo through autologous transplantation. Although pulpal regeneration requires the cell-based approach, several challenges in clinical translation must be overcome-including aging-associated phenotypic changes in pulpal MSCs, availability of tissue sources, and safety and regulation involved with expansion of MSCs in laboratories. Allotransplantation of MSCs may alleviate some of these obstacles, although the long-term stability of MSCs and efficacy in pulp-dentin regeneration demand further investigation. For an alternative source of MSCs, our laboratory developed induced MSCs (iMSCs) from primary human keratinocytes through epithelial-mesenchymal transition by modulating the epithelial plasticity genes. Initially, we showed that overexpression of ΔNp63α, a major isoform of the p63 gene, led to epithelial-mesenchymal transition and acquisition of stem characteristics. More recently, iMSCs were generated by transient knockdown of all p63 isoforms through siRNA, further simplifying the protocol and resolving the potential safety issues of viral vectors. These cells may be useful for patients who lack tissue sources for endogenous MSCs. Further research will elucidate the level of potency of these iMSCs and assess their transdifferentiation capacities into functional odontoblasts when transplanted into the root canal microenvironment.

Grainyhead-like 2 (GRHL2) inhibits keratinocyte differentiation through epigenetic mechanism.

We recently identified Grainyhead-like 2 (GRHL2), a mammalian homolog of Grainyhead in Drosophila, to be a novel transcription factor that regulates hTERT gene expression and enhances proliferation of normal human epidermal keratinocytes (NHEK). In the current study, we show that GRHL2 impairs keratinocyte differentiation through transcriptional inhibition of the genes clustered at the epidermal differentiation complex (EDC), located at chromosome 1q21. Gene expression profiling and subsequent in vitro assays revealed consistent downregulation of EDC genes, for example, IVL, KRT1, FLG, LCEs, and SPRRs, in NHEK expressing exogenous GRHL2. In vivo binding assay by chromatin immunoprecipitation revealed GRHL2 association at the promoter regions of its target genes, many of which belong to EDC. Exogenous GRHL2 expression also inhibited recruitment of histone demethylase Jmjd3 to the EDC gene promoters and enhanced the level of histone 3 Lys 27 trimethylation enrichment at these promoters. Survey of GRHL2 expression in human skin tissues demonstrated enhanced protein and mRNA levels in chronic skin lesions with impaired keratinocyte differentiation, for example, atopic dermatitis and psoriasis, compared with normal epidermis. These data indicate that GRHL2 impairs epidermal differentiation by inhibiting EDC gene expression through epigenetic mechanisms and support its role in the hyperproliferative skin diseases.

Bacterial diversity on different surfaces in urban freshwater.

Microbial loads in freshwater systems have important implications in biogeochemical cycling in urban environments. Immersed surfaces in freshwaters provide surfaces for bacterial attachment and growth. Microorganisms that adhere initially to these surfaces play a critical role in biofilm formation and sustenance. Currently, there is little understanding on the type of organisms that initially adhere to different surfaces in urban canals. In this study, water from an urban stormwater canal was employed to allow bacteria to attach to different surfaces in a flowcell apparatus and understand the differences and changes in bacterial community structure. Bacterial communities were highly diverse on different surfaces as indicated by Jaccard's indices of 0.14-0.56. Bacteria on aluminium were the most diverse and on Plexiglas the least. Bacterial communities were highly dynamic in the early attachment phase and it changed by 59% between 3 and 6 h on aluminium. Specificity of attachment to surfaces was observed for some bacteria. Judicious use of materials in urban aquatic environment would help mitigate microbial load in urban waters.

Bisphosphonates induce senescence in normal human oral keratinocytes.

Bisphosphonate-related osteonecrosis of the jaw (BRONJ) commonly occurs in individuals receiving bisphosphonates (BPs) with clinical manifestations of the exposed necrotic bone. Although defective wound healing of soft tissue is frequently, if not always, observed in BRONJ, the effects of BPs on oral soft tissue or cells remain unknown. To investigate the effects of BPs on cells of oral mucosal tissue, we studied the effect of pamidronate (PAM), one of the BPs most commonly administered to cancer patients, on the phenotypes of normal human oral keratinocytes (NHOK) and fibroblasts (NHOF). When exposed to PAM at 10 µM, NHOK, not NHOF, underwent senescence: NHOK overexpressed senescence-associated ß-galactosidase (SA-ß-Gal), p16INK4A, IL-6, and IL-8. When exposed to a higher level (50 µM) of PAM, NHOK maintained senescent phenotypes, but NHOF underwent apoptosis. PAM-induced senescence in NHOK is mediated, in part, via geranylgeranylation of the mevalonate pathway. Our in vitro 3D oral mucosal tissue construction studies further demonstrated that PAM induced senescence and impaired re-epithelialization of oral mucosa. Analysis of these data indicates that premature senescence of oral mucosal cells and subsequent defective soft-tissue wound healing might be partly responsible for the development of BRONJ in individuals receiving PAM or other BPs.

Regulation of the hTERT promoter activity by MSH2, the hnRNPs K and D, and GRHL2 in human oral squamous cell carcinoma cells.

Higher expression of human telomerase reverse transcriptase (hTERT) and subsequent activation of telomerase occur during cellular immortalization and are maintained in cancer cells. To understand the mode of hTERT expression in cancer cells, we identified cancer-specific trans-regulatory proteins that interact with the hTERT promoter, using the promoter magnetic precipitation assay coupled with mass spectrometry. The identified proteins include MutS homolog 2 (MSH2), heterogeneous nuclear ribonucleoprotein (hnRNP) D, hnRNP K and grainyhead-like 2 (GRHL2). We noticed a higher expression of these proteins in human oral squamous cell carcinoma (OSCC) cells than in normal cells, which do not exhibit telomerase activity. Knockdown of MSH2, hnRNP D and GRHL2 resulted in a notable reduction of the hTERT promoter activity in tested cancer cells. Silencing of the above genes resulted in a significant reduction of the telomerase activity in OSCC cells. Interestingly, among the four identified genes, silencing of GRHL2 was essential in reducing telomerase activity and viability of tested cancer cells. These results suggest a possible role of GRHL2 in telomerase activation during cellular immortalization.

The Antimicrobial Effect of Silver Ion Impregnation into Endodontic Sealer against Streptococcus mutans.

Pulpal and periradicular diseases are primarily caused by bacterial invasion of the root canal system as a result of caries progression. The presence of residual bacteria at the time of root canal completion (obturation) is associated with significantly higher rate of treatment failure. Re-infection of obturated root canals can be potentially prevented by enhancing the antibacterial activities of root canal obturation materials. We evaluated, in an in vitro model, the antimicrobial efficacy of silver ions added to a common endodontic sealer. For that purpose we performed growth inhibition studies and bacterial viability tests. We measured the zone of inhibition, optical density and performed confocal laser scanning microscopy. Our results show that the silver ions enhance the antimicrobial activity of the root canal sealer against Streptococcus mutans. This study approach may hold promise for studying other biologically based therapies and therefore increasing the success rate of routine orthograde root canal treatment.

Interleukin-6 induces proliferation in adult spinal cord-derived neural progenitors via the JAK2/STAT3 pathway with EGF-induced MAPK phosphorylation.

INTRODUCTION: In a previous study, we observed cell proliferation 3 days after spinal cord injury, and levels of interleukin-6 (IL-6) and epidermal growth factor (EGF) had significantly increased in the region of the injury. OBJECTIVES: The purpose of the new study described here was to evaluate the roles of IL-6 and EGF after traumatic damage to the spinal cord having isolated neural progenitor cells (NPC) from adult mice. METHODS AND RESULTS: Evidence provided by the trypan blue dye exclusion assay, 5-bromodeoxyuridine immunoreactivity and Western blot analysis indicated that IL-6 and EGF induced proliferation of these spinal cord-derived NPCs via phosphorylation of Janus-activated kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) and mitogen-activated protein kinases (MAPK), respectively. Combined treatment with IL-6 and EGF accelerated proliferation of cells synergistically and phosphorylation of STAT3 and extracellular signal-regulated kinase 1/2 (Erk1/2). Furthermore, AG490 and AG1478, JAK2 inhibitor and EGFR inhibitor, respectively, prevented the IL-6- and EGF-induced proliferation of the cells. Interestingly, IL-6-activated MAPKs but EGF did not influence JAK2/STAT3 activation; AG490 specifically inhibited IL-6-induced Erk1/2 phosphorylation without affecting IL-6-induced phosphorylation of Raf and MEK1/2. These results indicate that IL-6 is directly involved in Erk1/2 activation via JAK2 and that Erk1/2 provides a signal bridge between the IL-6-induced JAK2/STAT3 pathway and EGF-induced MAPK pathway. CONCLUSIONS: Our study is the first demonstration of IL-6- and EGF-stimulated proliferation of spinal cord progenitor cells via JAK2/STAT3 and MAPK signalling pathways. These pathways play key roles in repopulation and regeneration of spinal cord tissue after injury. It may represent novel therapeutic targets for pharmacological intervention in central nervous system disease, including spinal cord injury.