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The aim of this study was to compare the functional characteristics of mesenchymal stromal cells (MSCs) from a sheep model of traumatic temporomandibular joint (TMJ) fibrous and bony ankylosis. A sheep model of bilateral TMJ trauma-induced fibrous ankylosis on one side and bony ankylosis on the contralateral side was used. MSCs from fibrous ankylosed callus (FA-MSCs) or bony ankylosed callus (BA-MSCs) at weeks 1, 2, 4, and 8 after surgery were isolated and cultured. MSCs derived from the bone marrow of the mandibular condyle (BM-MSCs) were used as controls. The MSCs from the different sources were characterized morphologically, phenotypically, and functionally. Adherence and trilineage differentiation potential were presented in the ovine MSCs. These cell populations highly positively expressed MSC-associated specific markers, namely CD29, CD44, and CD166, but lacked CD31 and CD45 expressions. The BA-MSCs had higher clonogenic and proliferative potentials than the FA-MSCs. The BA-MSCs also showed higher osteogenic and chondrogenic potentials, but lower adipogenic capacity than the FA-MSCs. In addition, the BA-MSCs demonstrated higher chondrogenic, but lower osteogenic capacity than the BM-MSCs. Our study suggests that inhibition of the osteogenic and chondrogenic differentiations of MSCs might be a promising strategy for preventing bony ankylosis in the future.

High-throughput gene expression profiling has recently emerged as a promising technique that provides insight into cancer subtype classification and improved prediction of prognoses. Immune/inflammatory-related mRNAs may potentially enrich genes to allow researchers to better illustrate cancer microenvironments. Oral cavity squamous cell carcinoma (OC-SCC) exhibits high morbidity and poor prognosis compared to that of other types of head and neck squamous cell carcinoma (HNSCC), and these differences may be partially due to differences within the tumor microenvironments. Based on this, we designed an immune-related signature to improve the prognostic prediction of OC-SCC. A cohort of 314 OC-SCC samples possessing whole genome expression data that were sourced from The Cancer Genome Atlas (TCGA) database was included for discovery. The GSE41613 database was used for validation. A risk score was established using immune/inflammatory signatures acquired from the training dataset. Principal components analysis, GO analysis, and gene set enrichment analysis were used to explore the bioinformatic implications. When grouped by the dichotomized risk score based on the signature, this classifier could successfully discriminate patients with distinct prognoses within the training and validation cohorts (P<0.05 in both cohorts) and within different clinicopathological subgroups. Similar somatic mutation patterns were observed between high and low risk score groups, and different copy number variation patterns were also identified. Further bioinformatic analyses suggested that the lower risk score group was significantly correlated with immune/inflammatory-related biological processes, while the higher risk score group was highly associated with cell cycle-related processes. The analysis indicated that the risk score was a robust predictor of patient survival, and its functional annotation was well established. Therefore, this bioinformatic-based immune-related signature suggested that the microenvironment of OC-SCC could distinguish among patients with different underlying biological processes and clinical outcomes, and the use of this signature may shed light on future OC-SCC classification and therapeutic design.

BACKGROUND The type of traumatic temporomandibular joint (TMJ) ankylosis depends on the degree of severity of TMJ trauma. Here, we performed comprehensive differential molecular profiling between TMJ fibrous and bony ankylosis. MATERIAL AND METHODS Six sheep were used and a bilateral different degree of TMJ trauma was performed to induce fibrous ankylosis in one side and bony ankylosis in the other side. The ankylosed calluses were harvested at days 14 and 28 postoperatively and analyzed by Affymetrix OviGene-1_0-ST microarrays. DAVID was used to perform the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis for the different expression genes (DEGs). The DEGs were also typed into protein-protein interaction (PPI) networks to get the interaction data. Ten DEGs, including 7 hub genes from PPI analysis, were confirmed by real-time PCR. RESULTS We found 90 and 323 DEGs at least 2-fold at days 14 and 28, respectively. At day 14, bony ankylosis showed upregulated DEGs, such as TLR8, SYK, NFKBIA, PTPRC, CD86, ITGAM, and ITGAL, indicating a stronger immune and inflammatory response and cell adhesion, while genes associated with anti-adhesion (PRG4) and inhibition of osteoblast differentiation (SFRP1) had higher expression in fibrous ankylosis. At day 28, bony ankylosis showed increased biological process related to new bone formation, while fibrous ankylosis was characterized by a prolonged immune and inflammatory reaction. CONCLUSIONS This study provides a differential gene expression profile between TMJ fibrous and bony ankylosis. Further study of these key genes may provide new ideas for future treatment of TMJ bony ankylosis.

Introduction The aim of this experimental study was to investigate, under the premise of discectomy, whether damage to either the fibrous layer of the condyle or that of the glenoid fossa, could induce temporomanibular joint (TMJ) ankylosis. And if not, which of the fibrous layer was more important in the genesis of TMJ ankylosis. Materials and methods Bilateral TMJ surgery was performed in 6 growing Xiao-wei Han sheep. Disk and condylar fibrous layer removal (DCFLR) was performed on the left TMJ, and disk and glenoid fibrous layer removal (DGFLR) was performed on the right TMJ. All animals were sacrificed at 3 months postoperatively. The TMJ complexes were examined by histological evaluation. Results Partial fibrous ankylosis was achieved on the DCFLR side in the 6 sheep at 3 months after surgery. On the DGFLR side, pathologic characteristics of TMJ osteoarthritis could be seen; however, no evidence of ankylosis was observed. The scores of TMJ ankylosis for the DGFLR side were significantly lower than those for the DCFLR side (P Conclusion This study demonstrated that removal of the condylar fibrous layer, not the glenoid fibrous layer, combined with discectomy could lead to traumatic TMJ ankylosis.

Chloroquine, a prototype anti-malaria drug, has been reported to possess anti-inflammatory effects. Moreover, chloroquine pretreatment could improve DNA damage repair. It is therefore reasonable to hypothesize that chloroquine pretreatment could attenuate ischemia/reperfusion injury in the brain. Considering the fact that chloroquine could also improve glucose metabolism, we speculated that the potential effects of chloroquine on ischemia/reperfusion injury might be particularly pronounced in diabetic mice. In this study, chloroquine pretreatment protected neurons from Oxygen Glucose Deprivation (OGD) induced cytotoxicity and apoptosis. In vivo, Ob/ob mice and wildtype (WT) mice were pretreated with chloroquine for 3 weeks. Then, ischemic stroke was induced by 60 min Middle Cerebral Artery Occlusion (MCAO). We found that chloroquine pretreatment normalized blood glucose in diabetic ob/ob mice, and reduced cerebral damage after ischemic stroke especially for diabetic mice. In addition, chloroquine pretreatment reduced High-mobility group box 1 (HMGB1) content in the cerebrospinal fluid (CSF) and serum and lowered myeloperoxidase (MPO) activity and inflammatory cytokines gene expression both in the ob/ob diabetic mice and WT mice. Moreover, harmful DNA damage-signaling responses, including PARP activation and p53 activation, were also attenuated by chloroquine pretreatment in these two kinds of mice. In conclusion, chloroquine pretreatment could reduce cerebral damage after ischemic stroke especially in diabetic mice through multiple mechanisms, which include reducing neural cell DNA injury, restoring euglycemia and anti-inflammatory effects. The findings may provide potential for the development of chloroquine in the prevention and treatment of stroke in diabetic high-risk patients.

Paroxysmal kinesigenic dyskinesia (PKD) is a heterogeneous movement disorder characterized by recurrent dyskinesia attacks triggered by sudden movement. PRRT2 has been identified as the first causative gene of PKD. However, it is only responsible for approximately half of affected individuals, indicating that other loci are most likely involved in the etiology of this disorder. To explore the underlying causative gene of PRRT2-negative PKD, we used a combination strategy including linkage analysis, whole-exome sequencing and copy number variations analysis to detect the genetic variants within a family with PKD. We identified a linkage locus on chromosome 12 (12p13.32-12p12.3) and detected a novel heterozygous mutation c.956 T>G (p.319 L>R) in the potassium voltage-gated channel subfamily A member 1, KCNA1. Whole-exome sequencing in another 58 Chinese patients with PKD who lacked mutations in PRRT2 revealed another novel mutation in the KCNA1 gene [c.765 C>A (p.255 N>K)] within another family. Biochemical analysis revealed that the L319R mutant accelerated protein degradation via the proteasome pathway and disrupted membrane expression of the Kv1.1 channel. Electrophysiological examinations in transfected HEK293 cells showed that both the L319R and N255K mutants resulted in reduced potassium currents and respective altered gating properties, with a dominant negative effect on the Kv1.1 wild-type channel. Our study suggests that these mutations in KCNA1 cause the Kv1.1 channel dysfunction, which leads to familial PKD. The current study further extended the genotypic spectrum of this disorder, indicating that Kv1.1 channel dysfunction maybe one of the underlying defects in PKD.

To investigate the inhibition effects of fragile histidine triad (FHIT) gene on the malignant growth of A549 cell line.A mammalian expression vector PEGFP-FHIT was constructed and transfected into the A549 cell line by lipofectamine. Then the transfected cell line was screened by G418. Individual G418-resistant colonies were isolated by limited dilution. The monoclonal transfected cell line was screened by RT-PCR and immunochemical staining. The inhibition growth efficacy of extraneous FHIT was evaluated by clonogenic survival assay, flow cytometry and heteroplastic transplant on nude mice.Presence of extraneous FHIT gene in FHIT-A549 cell was proved by RT-PCR. Immunochemical stain demonstrated that the expression of extraneous FHIT protein was positive in FHIT-A549 cell and negative in PEGFP-A549 cell and A549 cell. The clonal formation rate of FHIT-A549 (2.6%) was significantly lower than that of A549 cell (50.1%) and PEGFP-A549 cell (53.6%, P0.01). FHIT-A549 cell (95.8%) was blocked in G(2) phage. Tumorigenicity of A549 cells in nude mice was greatly inhibited by expression of ectogenous FHIT gene. The weight of tumor was significantly lower in FHIT-A549 cell (0.04 +/- 0.03) than in A549 cell (0.24 +/- 0.11) and PEGFP-A549 cell (0.25 +/- 0.07, P0.01).Reintroduction of the expression of ectogeneous FHIT gene can obviously suppress the proliferation and tumorigenicity in human lung cancer cell line A549 and induce apoptosis. The data demonstrate oncosuppressive properties of FHIT gene.