Geographical distribution of MTHFR C677T gene polymorphisms among the reproductive-age women in Chinese Han populations: based on migration.

BACKGROUND: Methylenetetrahydrofolate reductase (MTHFR) is essential for the metabolism of folic acid and homocysteine. The MTHFR C677T polymorphism is associated with several disorders. Our study aims to explore the geographical distributions of the MTHFR C677T polymorphism of women in China and how migration affected the polymorphism in Suzhou. METHODS: A total of 7188 women of reproductive age were recruited in Suzhou of the study. Subjects were classified according to their native places after data extraction. MTHFR C677T gene polymorphisms were detected by quantitative PCR with genomic DNA isolated from blood samples. RESULTS: The frequencies of the 677T allele and 677TT genotype were higher in northern China than that in southern China and decreased in geographical gradients from north to south. The frequencies were considerably higher in the migrant population than that in the indigenous population of Suzhou. The migrant population have gradually changed the prevalence in Suzhou. CONCLUSIONS: Our study suggested that the prevalence of MTHFR C677T polymorphisms among women varied across different geographical regions in Chinese Han populations. The 677T allele frequencies of the northern populations were significantly higher than those of the southern populations. The migrant population gradually changed the prevalence of the MTHFR C677T polymorphism in Suzhou.

The role of NPY2R/NFATc1/DYRK1A regulatory axis in sebaceous glands for sebum synthesis.

BACKGROUND: Sebaceous glands (SGs) synthesize and secret sebum to protect and moisturize the dermal system via the complicated endocrine modulation. Dysfunction of SG are usually implicated in a number of dermal and inflammatory diseases. However, the molecular mechanism behind the differentiation, development and proliferation of SGs is far away to fully understand. METHODS: Herein, the rat volar and mammary tissues with abundant SGs from female SD rats with (post-natal day (PND)-35) and without puberty onset (PND-25) were arrested, and conducted RNA sequencing. The protein complex of Neuropeptide Y receptor Y2 (NPY2R)/NPY5R/Nuclear factor of activated T cells 1 (NFATc1) was performed by immunoprecipitation, mass spectrum and gel filtration. Genome-wide occupancy of NFATc1 was measured by chromatin immunoprecipitation sequencing. Target proteins' expression and localization was detected by western blot and immunofluorescence. RESULTS: NPY2R gene was significantly up-regulated in volar and mammary SGs of PND-25. A special protein complex of NPY2R/NPY5R/NFATc1 in PND-25. NFATc1 was dephosphorylated and activated, then localized into nucleus to exert as a transcription factor in volar SGs of PND-35. NFATc1 was especially binding at enhancer regions to facilitate the distal SG and sebum related genes' transcription. Dual specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A) contributed to NFATc1 phosphorylation in PND-25, and inactivated of DYRK1A resulted in NFATc1 dephosphorylation and nuclear localization in PND-35. CONCLUSIONS: Our findings unmask the new role of NPY2R/NFATc1/DYRK1A in pubertal SG, and are of benefit to advanced understanding the molecular mechanism of SGs' function after puberty, and provide some theoretical basis for the treatment of acne vulgaris from the perspective of hormone regulation.

Investigation of three-rooted deciduous mandibular second molars in a Chinese population using cone-beam computed tomography.

BACKGROUND: To investigate the anatomic features of three-rooted deciduous mandibular second molars (DMSMs) in Chinese children by using cone-beam computed tomography (CBCT). METHODS: A total of 247 CBCT scans of Chinese children were selected and retrospectively analyzed. The occurrence, gender and side predilection of three-rooted DMSMs were examined. The pattern of concurrence of bilateral three-rooted DMSMs, and concurrence of three-rooted DMSM and three-rooted permanent mandibular first molar (PMFM) was analyzed by the concurrence rate and Spearman's rank correlation test. The geometric parameters of the disto-buccal (DB) and disto-lingual (DL) roots, including the vertical root length, level and angle of distal root furcation, angle of root curvature (by Schneider technique) and the spreading angle, were measured and compared to the three-rooted PMFMs (n = 42) from 100 randomly selected adult subjects. RESULTS: The occurrence of three-rooted DMSMs was 24.0% (54/225) calculated by individual, and 18.6% (88/472) by tooth. A significant right-side predilection was detected (23.0% vs 14.2%, p < 0.05), while gender predilection was not detected (p > 0.05). The bilateral concurrence rate was 49.0%, and Spearman's correlation test indicated a significant relationship between the antimetric teeth (rho = 0.609, p < 0.01); whereas a weak but significant co-relationship was detected between the three-rooted DMSM and three-rooted PMFM (right side: concurrence rate = 31.6%, rho = 0.325, p < 0.01; left side: concurrence rate = 23.0%, rho = 0.260, p < 0.01). The length of DL roots in the DMSMs was 7.4 ± 1.5 mm, and the curvature angle was 16.4 ± 11.3 degrees, which was significantly (both p < 0.01) lower than that of the three-rooted PMFMs (root length = 11.0 ± 1.3 mm; degrees of curvature = 34.2 ± 16.1 degrees), whereas the spreading angle of the DL root in DMSMs (34.6 ± 8.4 degrees) was significantly (p < 0.01) greater than in the PMFMs (26.8 ± 6.5 degrees). CONCLUSIONS: Three-rooted DMSMs have a high occurrence rate in the Chinese children with a right-side predilection, and they have a weak but statistically significant correlation with three-rooted PMFMs. The DL roots of DMSMs are shorter, less curved, and spreading more widely as compared with those in the three-rooted PMFMs.

Assessment of the presence of a second mesiobuccal canal in maxillary first molars according to the location of the main mesiobuccal canal-a micro-computed tomographic study.

OBJECTIVES: To investigate the root canal morphology of mesiobuccal (MB) roots in maxillary first molars, and to assess the presence of a second mesiobuccal canal (MB2) according to the location of the main MB canal. MATERIALS AND METHODS: A total of 72 extracted permanent maxillary first molars were collected from dental clinics and were scanned with micro-CT and reconstructed three-dimensionally. The root canal systems were recorded according to Vertucci's classification, and the occurrence of accessory canals was also recorded. The root canal dimensions were measured at the coronal (furcation plane), middle, and apical root levels. The long (D) and short (d) diameters as well as the palatal (P) and buccal (B) distances from the center of the first mesiobuccal canal (MB1) to the root surface were measured, and the ratios of D/d and P/B were calculated. Receiver operating characteristic (ROC) curve analysis was used to evaluate the diagnostic accuracy of using the ratio of P/B for predicting the presence of an MB2 canal. The best cut-off point was determined according to the sensitivity and specificity. RESULTS: The MB roots most frequently had a type 2-2 root canal with an incidence of 37.5% (27/72), followed by the type 1-1 (23.6%, 17/72) and type 2-1 (16.7%, 12/72) canal forms. Type 1-2 canals were detected only in 5 molars (6.9%), and type 2-1-2 canals were detected in 6 molars (8.3%). The other 5 cases included 1 case of type 1-2-1 canal and 4 cases of triple canals. MB2 canals were detected in 76.4% (55/72) of the total sample teeth. The incidence of accessory canals was 56.9% (41/72). The mean ratio of D/d was generally "greatest to least": coronal level > middle level > apical level for different root levels and MB single > MB1 > MB2 for different canals, which reflected a trend from a flat to a circular cross-sectional shape. ROC curve analysis showed that at the coronal and middle root levels, areas under the ROC curve (AUC) were greater than 0.99 (P < 0.01), and the best cut-off point was 1.58 and 1.55, respectively; at the apical level, the AUC was 0.94 (P < 0.01), and the best cut-off point was 1.77. CONCLUSIONS: The MB2 canals may be present in the MB roots of maxillary first molars with a high occurrence rate at various levels, and the P/B ratio of the MB1 is a good index for predicting the presence of an MB2. However, since all the sample teeth were collected from a Chinese population, clinicians have to be cautious while trying to apply the conclusions on teeth of other ethnic populations. CLINICAL RELEVANCE: By calculating the P/B ratio, an index reflecting the buccal deviation of the MB1, clinicians can predict the presence of an invisible MB2 in cone-beam computed tomography images with inadequate resolution.

Increased Expression of Ubiquitin-Specific Protease 4 Participates in Neuronal Apoptosis After Intracerebral Hemorrhage in Adult Rats.

Ubiquitinating enzymes catalyze protein ubiquitination, a reversible process countered by deubiquitinating enzyme (DUB) action. Ubiquitin-specific protease 4 (USP4) is a member of the ubiquitin-specific protease (USP) family of DUBs that has a role in spliceosome regulation. In the present study, we demonstrated that USP4 may be involved in neuronal apoptosis in the processes of intracerebral hemorrhage (ICH). We obtained a significant up-regulation of USP4 in neurons adjacent to the hematoma following ICH by the results of Western blot, immunohistochemistry, and immunofluorescence. Increasing USP4 level was found to be accompanied by the up-regulation of active caspase-3, γH2AX, Bax, and decreased expression of Bcl-2. In addition, USP4 co-localized well with γH2AX in the nucleus in the ICH model and hemin-induced apoptosis model. Moreover, in vitro study, knocking down USP4 by USP4-specific siRNA in PC12 cells reduced active caspase-3 expression. All these results above suggested that USP4 may be involved in neuronal apoptosis after ICH.

Regulation of Glioma Cells Migration by DYRK2.

Dual-specificity tyrosine-regulated kinase 2 (DYRK2), a protein kinase that phosphorylates its substrates on serine/threonine, is expressed in numerous human tumors, but little is known about its role in the pathophysiology of glioma. In this study, we made an effort to explore the expression and function in human glioma. Western blot and immunohistochemistry analysis were performed to investigate the expression of DYRK2 protein in glioma tissues in 84 patients. Wound healing and transwell assay were carried out to determine the cell migration ability. We showed that the level of DYRK2 was significantly decreased in high-grade glioma tissues compared with low-grade tissues. In addition, the expression level of DYRK2 was positively correlated with glioma pathological grade and E-cadherin expression. Kaplane-Meier analysis revealed that low expression of DYRK2 was related to poor prognosis of glioma patients. Furthermore, wound healing and transwell assay revealed that DYRK2 could suppress cell migration and affect the expression levels of E-cadherin and vimentin through PI3K/AKT/GSK3ß signaling pathway. Taken together, our results implied that DYRK2 could serve as a promising prognostic biomarker as well as a potential therapeutical target of glioma.

CD109 Mediates Cell Survival in Hepatocellular Carcinoma Cells.

BACKGROUND: Hepatocellular carcinoma (HCC) accounts for 75-80 % of primary liver cancer, and usually arises after years of liver disease. Thus it is important to understand the molecular mechanisms which drive or mediate the development of HCC. AIM: In this work, we examined whether CD109 was associated with a poor prognosis in HCC and explored possible underlying mechanisms. METHODS: We examined the CD109 and Ki67 expression levels in 97 patients with HCC using immunohistochemistry. CD109 levels in HCC cells were down-regulated by shRNA transfection. The cycle progression and cell proliferation status of HCC cells were evaluated by flow cytometry and CCK-8 assay. The effect of CD109 on proliferation and apoptosis was investigated by western blot and TUNEL activity assays. RESULTS: The CD109 protein was up-regulated in HCC tissue compared with adjacent noncancerous tissue. CD109 expression levels in the 97 patients with HCC were positively correlated with histological grade. Univariate and multivariate survival analysis revealed that CD109 was a significant predictor of overall survival among HCC patients. CD109 shRNA knockdown delayed the G1-S phase transition, abrogated cell proliferation, and increased cell apoptosis. Furthermore, CD109 impaired TGF-ß/Smad signaling through control of p-smad2. CONCLUSIONS: CD109 promoted HCC proliferation and predicted poor prognosis. In addition, CD109 expression was associated with anti-apoptosis in HCC cells.

[The role of ROS/JNK/caspase 3 axis in apoptosis induction by menthol-favored electronic cigarette liquid in human periodontal ligament stem cells].

PURPOSE: To explore the cytotoxic effect of a menthol-favored E-liquid on human periodontal ligament stem cells (hPDLSCs), as well as the underlying mechanism of electronic cigarette (E-cig)-induced cell apoptosis. METHODS: PDLSCs were isolated and cultured from periodontal ligament tissues of healthy premolars extracted for orthodontic reasons. Cells in passage 3 were used to detect the surface markers of stem cells by flow cytometry. Then the cells were exposed to different doses of menthol-favored E-liquid (at 59 mg/L nicotine concentration) in the culture median (the final nicotine concentrations were 0.1 µg/mL, 1.0 µg/mL, 10 µg/mL, 50 µg/mL, 0.1 mg/mL, 0.2 mg/mL and 0.5 mg/mL, respectively) for different period of times (24, 48 and 72 h). The cell viability was analyzed by CCK-8 assay. Cell apoptosis was evaluated by flow cytometry (7-AAD and Annexin V staining) and TUNEL assay. Reactive oxygen species (ROS) production was detected with fluorescence probe DCFH-DA by confocal microscopy and flow cytometry. The protein expression levels associated with ROS/JNK/caspase 3 axis(p-JNK, JNK, c-Jun, p-c-Jun, Bcl-2, Bax and cleaved-caspase 3) were analyzed by Western blot. Immunocytofluorescense staining was applied to evaluate the expression level of p-JNK. After addition of NAC, a ROS scavenger, and MAPK/JNK specific blocker SP600125, their effects on E-cig-induced cell apoptosis were evaluated. Statistical analysis was performed with Graph Pad 5.0 software package. RESULTS: Human PDLSCs were successfully isolated and cultured and flow cytometry assay showed the mesenchymal stem cell surface biomarkers (CD73, CD90 and CD105) were positively expressed. CCK8 assay indicated cell viability was significantly(P＜0.001) different among all concentration groups at various time points (24, 48 or 72 h), and the difference in apoptosis rate among all concentration groups was also statistically significant (P＜0.001). After exposure to E-liquid with nicotine concentration ≥50 µg/mL, cell viability was significantly reduced, and the proportion of apoptotic cells and the cellular ROS level was significantly increased in a dose-dependent manner as compared with the control group(0.0 mg/mL). Western blot assay showed E-cig exposure could promote MAPK/JNK phosphorylation in a dose-dependent and time-dependent manner. Either NAC or SP600125 could partially rescue the E-cig-induced cell apoptosis via reversing up-regulation of p-JNK and cleaved caspase 3. CONCLUSIONS: ROS/JNK/caspase 3 axis is involved in menthol-favored E-liquid-induced apoptosis of hPDLSCs.

In situ tissue profile of rat trigeminal nerve in trigeminal neuralgia using spatial transcriptome sequencing.

BACKGROUND: Trigeminal neuralgia (TN) is the most common neuropathic disorder in the maxillofacial region. The etiology and pathogenesis of TN have not been clearly determined to date, although there are many hypotheses. OBJECTIVE: The goal of this study was to investigate the interactions between different types of cells in TN, particularly the impact and intrinsic mechanism of demyelination on the trigeminal ganglion, and to identify new important target genes and regulatory pathways in TN. METHODS: TN rat models were prepared by trigeminal root compression, and trigeminal nerve tissues were isolated for spatial transcriptome sequencing. The gene expression matrix was reduced dimensionally by PCA and presented by UMAP. Gene function annotation was analyzed by Metascape. The progression of certain clusters and the developmental pseudotime were analyzed using the Monocle package. Modules of the gene coexpression network between different groups were analyzed based on weighted gene coexpression network analysis and assigned AddModuleScore values. The intercellular communication of genes in these networks via ligand-receptor interactions was analyzed using CellPhoneDB analysis. RESULTS: The results suggested that the trigeminal ganglion could affect Schwann cell demyelination and remyelination responses through many ligand-receptor interactions, while the effect of Schwann cells on the trigeminal ganglion was much weaker. Additionally, ferroptosis may be involved in the demyelination of Schwann cells. CONCLUSIONS: This study provides spatial transcriptomics sequencing data on TN, reveals new markers, and redefines the relationship between the ganglion and myelin sheath, providing a theoretical basis and supporting data for future mechanistic research and drug development.

Transcriptome characterization of human gingival mesenchymal and periodontal ligament stem cells in response to electronic-cigarettes.

The potential toxicities and threats of electronic cigarettes (E-cigs) on periodontal health remain elusive. Gingival mesenchymal stem cells (GMSCs) and periodontal ligament stem cells (PDLSCs) contribute to cell differentiation and regeneration for periodontium as well as inflammatory modulation. However, the effects of E-cig exposure on periodontal tissues, particularly GMSCs and PDLSCs, and the underlying epigenetic mechanisms remain largely unknown. In this study, we conducted RNA-seq analysis to examine the transcriptome of human GMSCs and PDLSCs exposed to four types of E-cigs (aerosol and liquid with tobacco and menthol flavor) and conventional tobacco smoke in vitro. Our results showed that E-cig exposure primarily impacted the immunoregulation and inflammatory responses to pathogenic microorganisms in GMSCs, and the microenvironment, differentiation and response to corticosteroid in PDLSCs, which were significantly different from the damage effects caused by tobacco smoke. Additionally, we discovered a large number of differentially expressed non-coding RNAs among the different E-cig exposure methods and flavors. We also noticed that in GMSCs, CXCL2 was especially down-regulated by E-cig aerosol exposure whereas up-regulated by E-liquid exposure compared to control. Of note, the enhancer elements near CXCL2 and other genes located at Chromosome 4 contributed to the transcription activity of these genes, and KDM6B was remarkably elevated in response to E-liquid exposure. Lastly, we conducted ChIP-seq analysis to confirm that the elevated gene transcription by E-liquids was due to the weakened H3K27me3 at genome-wide enhancer elements in GMSCs, but not at promoter regions. Taken together, our results characterized the diverse gene expression profiles of GMSCs and PDLSCs in response to E-cigs with different exposure methods and flavors in vitro, and indicated a novel mechanism of KDM6B-mediated H3K27me3 on enhancers for gene transcription regulation. Our data could be served as a resource for emphasizing the understanding of E-cigs in periodontal health.

Genomic distribution of signal transducer and activator of transcription (STAT) family in colorectal cancer.

JAK/STAT pathway has been widely acknowledged in the development of human cancers. However, the role of different phosphorylated STAT proteins translocating into nucleus in transcription activation of target genes is not fully understood. In present research, ChIP-seq was carried on to investigate the genome-wide distribution of the activated STAT1, STAT2, STAT3, STAT5 and STAT6 in colorectal cancer HCT-116 cells. Our observations indicated that the homodimers rather than heterodimers of STAT protein predominantly occupied on genomic DNA. STAT3 accounted for the largest proportion among all STAT proteins HCT-116 cells. Furthermore, the biased binding motif targeted by different STAT homodimers suggested the distinct biological functions. Here, we noticed that NR5A2 was a specific co-activator of STAT3 by DNA motif analysis. Co-IP assay determined that NR5A2 indeed interacted with STAT3 homodimer rather than other homodimers or heterodimers. NR5A2 knockdown resulted in a reduced binding affinity of STAT3 homodimer in the original regions. Taken together, we characterize the genome-wide landscape of activated STAT proteins, and reveal the differences of binding patterns as well as the target genes and associated functions between homodimer and heterodimer of STAT proteins in HCT-116 cells. We also present some new findings and possible mechanisms regarding the role of NR5A2 on STAT3 in CRC. Our findings may provide new insights into the design of STAT inhibitors to treat CRC and other diseases.

Transcriptome-scale spatial gene expression in rat arcuate nucleus during puberty.

BACKGROUND: A variety of neurons in hypothalamus undergo a complicated regulation on transcription activity of multiple genes for hypothalamic-pituitary-gonadal axis activation during pubertal development. Identification of puberty-associated cell composition and characterization of the unique transcriptional signatures across different cells are beneficial to isolation of specific neurons and advanced understanding of their functions. METHODS: The hypothalamus of female Sprague-Dawley rats in postnatal day-25, 35 and 45 were used to define the dynamic spatial atlas of gene expression in the arcuate nucleus (ARC) by 10× Genomics Visium platform. A surface protein expressed selectively by kisspeptin neurons was used to sort neurons by flow cytometric assay in vitro. The transcriptome of the isolated cells was examined using Smart sequencing. RESULTS: Four subclusters of neurons with similar gene expression signatures in ARC were identified. Only one subcluster showed the robust expression of Kiss1, which could be isolated by a unique membrane surface biomarker Solute carrier family 18 member A3 (SLC18A3). Moreover, genes in different subclusters presenting three expression modules distinctly functioned in each pubertal stage. Different types of cells representing distinct functions on glial or neuron differentiation, hormone secretion as well as estradiol response precisely affect and coordinate with each other, resulting in a complicated regulatory network for hypothalamic-pituitary-gonadal axis initiation and modulation. CONCLUSION: Our data revealed a comprehensive transcriptomic overview of ARC within different pubertal stages, which could serve as a valuable resource for the study of puberty and sexual development disorders.

[Effect of high-concentration fluoride on apoptosis of human periodontal ligament stem cells].

PURPOSE: To explore the effects of high-concentration fluoride(F) on apoptosis of human periodontal ligament stem cells (PDLSCs). METHODS: PDLSCs were isolated from periodontal ligament tissues of extracted third molars, and treated with different concentrations (0-40 ppm F) of NaF for indicated period of time. CCK-8 assay was performed to detect cell viability. After stained with Annexin V-PI and JC-1, cell apoptosis and mitochondrial membrane potential were analyzed by flow cytometry. Immunofluorescence staining and confocal microscopic assay were used to detect the protein expression level of cyt-c, cleaved-caspase-9 and -3. The mRNA level of caspase -9 and -3 were examined by RT-PCR. The protein expression level of total and phosphate-ERK, JNK and p38 were analyzed by Western blot. SPSS 13.0 software package was used for statistical analysis. RESULTS: Fluoride treatment inhibited cell viability (CCK-8 assay) and induced apoptosis of PDLSCs (Annexin V-PI staining) in a dose- and time-dependent manner. Immunofluorescence assay showed that fluoride with a dose ≥10 ppm significantly induced release of cyt-c from the mitochondria to cytosol, and up-regulation of expression of cleaved-caspase -9 and -3. RT-PCR confirmed that the mRNA level of caspase-9 and -3 increased with the dose of fluoride. Western blot assay confirmed that fluoride induced up-regulation of p-ERK, but not that of p-JNK and p-p38, and specifically blocking ERK pathway with U0126 could partially rescue the fluoride-induced cell apoptosis. CONCLUSIONS: High concentrations of fluoride induces apoptosis of PDLSCs via intrinsic mitochondrial pathway, and phosphation of MAPK/ERK is involved in the F-induce cell apoptosis.

The role of N-glycosylation of CD200-CD200R1 interaction in classical microglial activation.

BACKGROUND: Microglial inflammatory activation is the common feature of the central nervous system (CNS) diseases. Microglia can be activated and particularly polarized toward a dual role in the injured CNS. The CD200 receptor 1 (CD200R1) inhibits inflammatory microglia activation as illustrated by studies. Publications show abnormal activation of microglia secondary to the deficient inhibit of CD200-CD200R interaction. In the present study, we established a neuronal-microglia co-culture system to investigate the association between CD200R1 engagement and classical microglial activation. We analyzed the glycosylation of CD200R1 and the CD200 binding. Secretion of pro-inflammatory cytokines were measured. RESULTS: CD200R1 was N-glycosylated at Asparagine 44 (Asn44, N44). Mutation of this site disrupted CD200-CD200R1 interaction and up-regulated the expression of cytokines iNOS, CD86, IL-1ß and TNF-α. CONCLUSION: N44 of CD200R1 is a significant binding site for CD200-CD200R1 interaction and play a critical role in the maintenance of microglia. The N-glycosylation of CD200R1 could serve as a therapeutic agent for CNS inflammation.

Root and root canal variations of the human maxillary and mandibular third molars in a Chinese population: A micro-computed tomographic study.

OBJECTIVES: To investigate the anatomical variations of the root and root canal configuration of the human third molars. DESIGNS: A total of 130 maxillary and 130 mandibular third molars were collected from a native Chinese population. All teeth were scanned by micro-computed tomography. After 3D reconstruction, the root and canal morphology of each tooth was examined both qualitatively and quantitatively. RESULTS: For maxillary molars, a single fused root (67 cases, 51.5%) and a single root canal system (64 cases, 49.2%) was most common root/canal form; the typical three-rooted molars were detected only in 33 cases (25.4%), and the secondary MB canals were detected only in 9 molars (6.9%). For mandibular molars, 62 teeth were single-rooted (47.7%) and 42 had a single root canal system (32.3%); 20 singled-rooted and 60 double-rooted molars exhibited independent mesial and distal root canal systems (61.5%), and the type 1-1 canal was the most common configuration for mesial (57 cases) and distal (81 cases) root canal systems. C-shaped canals were detected in 11 maxillary and 36 mandibular single-rooted molars. The mean root surface area, root and crown volume of mandibular third molars were significantly higher than the maxillary third molars (P < 0.01). CONCLUSION: The root canal system of the third molars may exhibit several anatomic variations. Whereas in most of cases, the degree of the canal differentiation was at a low level, and the canal form was not complicate.

Upregulated expression of Nucleostemin/GNL3 is associated with poor prognosis and Sorafenib Resistance in Hepatocellular Carcinoma.

Nucleostemin (NS)/GNL3 protein has been recently documented to be a nucleolar protein that was abundantly expressed in stem cells and cancer cells. Herein, we showed that NS was upregulated in HCC tissues and the expression of NS was inversely correlated with that of p53. Overexpression of NS predicted significantly worsened prognosis in HCC patients, suggesting that NS might serve as a prognostic marker of HCC. In addition, we found that depletion of NS sensitized HCC cells to sorafenib-induced apoptosis. Moreover, we found that the mechanism underlying NS-mediated sorafenib resistance involved dysregulated expression of p53, and downstream Bax and Bcl-2 proteins. NS interacted with p53 in HCC cells. Depletion of NS increased the expression of p53 and Bax, whereas impaired the level of cellular Bcl-2. Interference of NS enhanced the cytotoxic effects of sorafenib in HCC cells. Furthermore, ectopic expression of NS impaired the apoptosis of HCC cells following sorafenib exposure. Therefore, NS may contribute to sorafenib resistance in HCC cells through the modulation of p53 pathway and Bcl-2 proteins. These findings indicated that the combination of silencing NS expression and sorafenib treatment is a promising therapeutic strategy in treatment of HCC.

USP11, Deubiquitinating Enzyme, Associated with Neuronal Apoptosis Following Intracerebral Hemorrhage.

Protein ubiquitination is a dynamic two-way process that can be reversed or regulated by deubiquitinating enzymes (DUB). USP11, located on the X chromosome, 6 is a member of USP subclass of the DUB family. Here, we demonstrate that USP11 may be involved in neuronal apoptosis in the processes of intracerebral hemorrhage (ICH). From the results of Western blot, immunohistochemistry, and immunofluorescence, we obtained a significant up-regulation of USP11 in neurons adjacent to the hematoma following ICH. Increasing USP11 level was found to be accompanied by the up-regulation of active caspase-3, Fas receptor (Fas), Fas ligand (FasL), and active caspase-8. Besides, USP11 co-localized well with active caspase-3 in neurons, indicating its potential role in neuronal apoptosis. What is more, knocking down USP11 by RNA-interference in PC12 cells reduced active caspase-3 expression. Thus, USP11 may play a role in promoting the brain secondary damage following ICH.

Silencing of CtBP1 suppresses the migration in human glioma cells.

Carboxyl-terminal binding protein 1 (CtBP1), up-regulated in various types of human cancers, has been functionally associated with proliferation, anti-apoptosis, and EMT in vitro studies. However, the functional significance of CtBP1 in the pathophysiology of glioma remains unknown. In the present study, we showed the expression of CtBP1 was markedly higher in glioma tissues compared with normal brain tissues by Western blot analysis. Immunohistochemical analysis revealed that CtBP1 mainly localized in the nucleus of glioma cells. Statistical analysis suggested the upregulation of CtBP1 was considerably correlated with the WHO grade (P < 0.05) and those patients with high CtBP1 levels exhibited shorter survival time (P < 0.01). Silencing CtBP1 by short hairpin RNAi caused an inhibition of cell migration. Moreover, knockdown of CtBP1 increases E-cadherin expression and decreases vimentin expression. These data uncovered that CtBP1 protein is a valuable marker of glioma pathogenic process and that CtBP1 can serve as a novel prognostic marker for glioma therapy.

High VRK1 expression contributes to cell proliferation and survival in hepatocellular carcinoma.

VRK1 is a member of the vaccinia-related kinase (VRK) family of serine/threonine protein kinases, which is known to play multiple roles in cellular proliferation, cell cycle regulation and carcinogenesis. However, the expression and physiological significance of VRK1 in hepatocellular carcinoma (HCC) remain unclear. In this study, we aimed to investigate the potential role of VRK1 in the development and progression of HCC. Western blot and immunohistochemical analysis revealed that VRK1 was highly expressed in HCC tissues and cell lines, compared with adjacent nontumorous tissues and LO2 normal hepatocytes. Meanwhile, clinicopathological analysis showed that VRK1 was significantly associated with AJCC stage, Ki-67 and a poor prognosis in HCC specimens. Univariate and multivariate analysis showed that VRK1 could serve as an independent prognostic indicator of HCC patients' survival. Furthermore, we found that VRK1 was lowly expressed in serum-starved Huh7 cells, and was progressively increased after serum-refeeding. Finally, flow cytometry, CCK-8 and colony formation assay indicated that the depletion of VRK1 could retard cell cycle progression and reduce cells proliferation in HCC cells. On the basis of these findings, we conclude that VRK1 may be a candidate prognostic biomarker as well as a potential therapeutical target of HCC.