Design, synthesis, and biological evaluation of thiazole/thiadiazole carboxamide scaffold-based derivatives as potential c-Met kinase inhibitors for cancer treatment.

As part of our continuous efforts to discover novel c-Met inhibitors as antitumor agents, four series of thiazole/thiadiazole carboxamide-derived analogues were designed, synthesised, and evaluated for the in vitro activity against c-Met and four human cancer cell lines. After five cycles of optimisation on structure-activity relationship, compound 51am was found to be the most promising inhibitor in both biochemical and cellular assays. Moreover, 51am exhibited potency against several c-Met mutants. Mechanistically, 51am not only induced cell cycle arrest and apoptosis in MKN-45 cells but also inhibited c-Met phosphorylation in the cell and cell-free systems. It also exhibited a good pharmacokinetic profile in BALB/c mice. Furthermore, the binding mode of 51am with both c-Met and VEGFR-2 provided novel insights for the discovery of selective c-Met inhibitors. Taken together, these results indicate that 51am could be an antitumor candidate meriting further development.

[Determination of bisphenols in sediment by accelerated solvent extraction and solid-phase extraction purification coupled with ultra performance liquid chromatography-tandem mass spectrometry].

Bisphenols are endocrine disruptors that are characterized with bioaccumulation, persistence, and estrogenic activity. Even low contents of bisphenols can exert adverse effects on human health and the ecological environment. Herein, a method combining accelerated solvent extraction and solid-phase extraction purification with ultra performance liquid chromatography-tandem mass spectrometry was developed for the accurate detection of bisphenol A (BPA), bisphenol B (BPB), bisphenol F (BPF), bisphenol S (BPS), bisphenol Z (BPZ), bisphenol AF (BPAF), and bisphenol AP (BPAP) in sediments. The mass spectrometric parameters of the seven bisphenols were optimized, and the response values, separation effects, and chromatographic peak shapes of the target compounds were compared under three different mobile phase conditions. The sediment samples were pretreated by accelerated solvent extraction, and orthogonal tests were used to optimize the extraction solvent, extraction temperature, and cycle number. The results showed that the use of 0.05% (v/v) ammonia and acetonitrile as the mobile phase for gradient elution could rapidly separate the seven bisphenols on an Acquity UPLC BEH C18 column (100 mm×2.1 mm, 1.7 µm). The gradient program was as follows: 0-2 min, 60%A; 2-6 min, 60%A-40%A; 6-6.5 min, 40%A; 6.5-7 min, 40%A-60%A; 7-8 min, 60%A. Orthogonal experiments indicated that the optimal extraction conditions were as follows: extraction solvent of acetonitrile, extraction temperature of 100 ℃, and cycle number of three. The seven bisphenols showed good linearity in the range of 1.0-200 µg/L, with correlation coefficients (r2) greater than 0.999, and the limits of detection were 0.01-0.3 ng/g. The recoveries for the seven bisphenols ranged from 74.9% to 102.8% at three spiking levels (2.0, 10, 20 ng/g), with relative standard deviations ranging from 6.2% to 10.3%. The established method was applied to detect the seven bisphenols in sediment samples collected from Luoma Lake and its inflow rivers. BPA, BPB, BPF, BPS, and BPAF were detected in the sediments of the lake, and BPA, BPF, and BPS were detected in the sediments of its inflow rivers. The detection frequency of BPA and BPF was 100%, and the contents of these bisphenols in the sediment were 11.9-38.0 ng/g and 11.0-27.3 ng/g, respectively. The developed method is simple, rapid with high accuracy and precision, and is suitable for the determination of the seven bisphenols in sediment.

AMPK inhibitor BML-275 induces neuroprotection through decreasing cyt c and AIF expression after transient brain ischemia.

Stroke is a major public health problem with an imperative need for a more effective and tolerated therapy. Neuroprotective therapy may be an effective therapeutic intervention for stroke. The morbidity and mortality of stroke-induced secondary brain injury is mainly caused by neuronal apoptosis, which can be executed in a caspase-dependent or apoptosis inducing factor (AIF)-dependent manner. As apoptosis is an energy-dependent process with a relative time delay, abnormal energy metabolism could be a significant and fundamental pathophysiological basis of stroke. To our knowledge, convincible evidences that AMPK inhibition exerts neuroprotection in cerebral ischemia injury via anti-apoptosis remain to be investigated. Accordingly, the aims of this study were to investigate the protective effects of AMPK inhibitor BML-275 on cerebral ischemic/reperfusion (I/R) injury and to elucidate the underlying mechanisms. Cerebral ischemia was induced by transient middle cerebral artery occlusion (tMCAO) in male C57BL/6 mice. The therapeutic effects of BML-275 were evaluated by infarct sizes, neurological scores and the proportion of apoptotic neurons after 24 h of reperfusion. The cell apoptosis markers cyt c and AIF were also evaluated. The results showed that intraperitoneally administration of BML-275 alleviate the cerebral infarction, neurological deficit and neuronal apoptosis induced by MCAO. BML-275 simultaneously induces anti-apoptosis and decreases the expression of cyt c and AIF. This study supports the hypothesis that anti-apoptosis is one of potential neuroprotective strategies for the treatment of stroke.

Study on the application of oral digital design in aesthetic restoration of anterior teeth of cleft lip/palate patients.

OBJECTIVES: A study was conducted to investigate the clinical effects of oral digital design on the aesthetic restoration of anterior teeth of cleft lip/palate patients. METHODS: Nine adult cleft lip/palate patients who need aesthetic restoration of anterior teeth were recruited. Digital information of patients' dental arches, the surrounding soft tissue and face were captured by digital camera and scanner. The aesthetic analysis and design were conducted using keynote and 3shape software and were demonstrated to the patients. The optimized treatment plan was ensured by communicating with the patients. Digital wax-up models were exported and printed into resin diagnostic models, which were then utilized in the treatment process to guide the doctors and the technicians in tooth preparation and in making the final restorations, respectively. The adhesive procedure was completed after satisfactory try-in. Aesthetics assessment was conducted in accordance with the anterior esthetic evaluation form. The scores of patient's satisfaction were recorded on a questionnaire containing six items of aesthetic index and doctor-patient communication. Patients were interviewed and examined after 1, 3, 6, and 12 months, respectively, and the clinical effects of restorations were evaluated. RESULTS: All nine patients had satisfactory clinical results. The aesthetic defects of the patients were effectively addressed. All treatments met the requirements of the preoperative digital designs. The patients' scores were all above 90 on the satisfaction scale. At 12 months after the operation, the clinical effects of restorations of all cases achieved A class in each evaluation indicator. CONCLUSIONS: For cleft lip/palate patients with esthetic defect in the anterior teeth, the digital design plays an important role in optimizing the treatment plan and guides the whole treatment process. This design can help clinicians achieve predictable satisfactory aesthetic results.

[Impact assessment on enamel remineralization after orthodontic treatment with casein phosphopeptide calcium phosphate complex].

PURPOSE: To compare the effect on enamel demineralization following fluoride rinse or casein phosphopeptide calcium phosphate complex (CPP-ACP) after fixed appliance orthodontic treatment. METHODS: The study population consisted of 21 post-orthodontic patients (13 females, 8 males, 84 affected teeth) with white spot lesions (WSL). They were divided into 3 groups with 28 affected teeth in each group. Participants in the control group were brushed with fluoride toothpaste twice a day. Participants in the fluoride group were instructed to rinse the mouth with 20mL 0.01% sodium fluoride rinse in addition to brushing twice a day. Participants in CPP-ACP group were instructed to use tooth moss after brushing their teeth twice a day for 6 months. SPSS 17.0 software package was used to analyze the data. RESULTS: Within 6 months after orthodontic treatment, white spot lesions areas of the three groups caused by enamel demineralization were all reduced in different degrees, and the differences of success rate were significant among three groups (P<0.05). CPP-ACP group achieved the highest success rate (51%) than the other group, the fluoride group (44%) and the control group (42%). CONCLUSIONS: Brushing teeth, fluoride rinse and CPP-ACP have certain effect on remineralization of demineralized teeth in 6 months after orthodontic treatment. Compared with brushing and fluoride rinse, CPP-ACP can reduce the area of enamel demineralization more effectively.

[Effect of HBXIP on biological function and PI3K/Akt signaling pathway of adenoid cystic carcinoma cell line ACC-M].

PURPOSE: To study the effect of hepatitis B virus X protein binding protein (HBXIP) on proliferation, migration and invasion of adenoid cystic carcinoma cell line ACC-M, and the possible mechanism of PI3K/Akt signaling pathway. METHODS: HBXIP plasmid was transfected into ACC-M. The cells were divided into experimental group (transfected with plasmid pEGFP-N1-HBXIP) control group (non-transfected group) and blank control group (vector group, pEGFP-N1). RT-PCR was used to detect the expression HBXIP in ACC-M; MTT assay, transwell chamber experiments and scratches over the proliferation of HBXIP were utilized individually to evaluate the influence of HBXIP on ACC-M expression, migration and invasion; Western blotting was used to detect the protein expression of Akt, p-Akt, PI3K, p-PI3K and S100A4 after overexpression of HBXIP. Statistical analysis was performed using SPSS 18.0 software package. RESULTS: MTT results showed that the number of surviving cells of experimental group was significantly higher than the control group (P<0.05); Scratch test results showed that the cell mobility of the experimental group was significantly higher than the control group (P<0.01); Transwell chamber experiments showed that the number of cell invasion of the experimental group was significantly higher than the control group (P<0.01); Western blotting results showed that compared with the control group, the expression of p-Akt, p-PI3K and S100A4 in the experimental group with overexpressed HBXIP was relatively increased. CONCLUSIONS: Overexpression of HBXIP gene promotes ACC-M proliferation, invasion and migration. Further, ACC-M proliferation, invasion and migration may be promoted by increased Akt, PI3K phosphorylation and S100A4 protein expression.

[DNA repair gene polymorphisms in ERCC4 rs6498486 and ERCC5 rs751402 and risk of salivary gland tumors].

PURPOSE: To investigate the association between ERCC4 and ERCC5 polymorphisms and risk of salivary gland tumors. METHODS: Case-control study was carried out in 133 cases of histologically confirmed salivary gland tumors and 142 age and gender matched healthy control cases. Polymorphisms of ERCC4 rs6498486 and ERCC5 rs751402 were detected by PCR-RFLP. Multiple factors logistic regression analysis was conducted to investigate the association between gene polymorphisms and risk of salivary gland tumors using SPSS 18.0 software package. RESULTS: The genotype distribution of each polymorphism was found to be of Hardy-Weinberg equilibrium in the study. ERCC5 rs751402 TT genotype was associated with risk of salivary gland tumors (TT vs. CC+CT: OR=0.45, 95% CI: 0.21-0.98, P=0.046). No significant association was found in ERCC4 rs6498486. CONCLUSIONS: The results suggest that ERCC5 rs751402 polymorphism may be associated with risk of salivary gland tumors.

[Expression and significance of pericytes and TGF-ß in infantile parotid hemangioma].

PURPOSE: To study the expression and significance of pericytes and TGF-ß in infantile parotid hemangioma. METHODS: The expressions of pericytes and TGF-ß at protein level were examined in 76 cases of infant parotid hemangioma by strep avidin-biotin complex (SABC) immunohistochemical technique. All statistical analysis were performed using SPSS 13.0 statistical package. The relationship between the expression of pericytes and TGF-ß and clinical phase of hemangioma was analyzed by using Chi-square test. Kappa test was used to determine the relationship between pericytes and TGF-ß expression. RESULTS: The rates of positive expression of pericytes were 86.7%(13/15), 45.5%(10/22) and 51.3%(20/39) in early, middle and advanced stage of hemangioma, respectively. The rates of positive expression of TGF-ß were 33.3%(5/15), 40.9%(9/22) and 76.9%(30/39) in early, middle and advanced stage of hemangioma, respectively. There was significantly close correlation between the level of pericytes and clinical phase of hemangioma, as well as between TGF-ß expression and clinical phase of hemangioma(P<0.05). CONCLUSION: Pericytes and TGF-ß may significantly contribute to the proliferation of infantile parotid hemangioma.

[Expression of ERK and nm23-H1 in tongue squamous cell carcinoma and its relation with invasion and metastasis].

PURPOSE: To reveal the relations between expression of extracellular signal-regulated kinase (ERK) and nm23-H1 and tumor invasion and metastasis in tongue squamous cell carcinoma (TSCC). METHODS: The expression of ERK and nm23-H1 at protein level was examined in 74 cases of TSCC by strep avidin-biotin complex (SABC) immunohistochemical technique. SPSS10.0 software package was used for data analysis. RESULTS: The expression of ERK was positively related to TNM clinical stage and cervical lymph node metastasis. The expression of nm23-H1 was negatively related to TNM clinical stage and cervical lymph node metastasis. The expression of ERK had a negative correlation with the expression of nm23-H1. TSCC cases with ERK(+)/nm23-H1(-) revealed significant higher tendency to cervical lymph node metastasis than those with ERK(-)/nm23-H1(+). CONCLUSION: The expression of ERK and nm23-H1 is correlated with invasion and cervical lymph node metastasis in TSCC.

[Expression of MMP-2 during differentiation of vascular endothelial cell from bone marrow mesenchymal stem cell in porcine].

PURPOSE: To investigate the expression of MMP-2 in porcine bone marrow mesenchymal stem cells (MSCs) in the process of differentiation into vascular endothelial cells in vitro. METHODS: Porcine MSCs were isolated from bone marrow and grown in vitro. At 3-, 7-day after VEGF induction, immunohistochemical and flow cytometrical examinations were carried out to detect the staining of FVIII. The expression of MMP-2 was analyzed by Western blot. RESULTS: At 3-, 7-day after VEGF induction, the morphology of the cells gradually changed. The cells gradually showed a ball-like appearance at 3-day. They connected with adjacent cells and formed tube-like structure after 7 days. The factor VIII-positive cells could be observed at 3-day after VEGF induction. The positive cells increased and joined to neighboring FVIII-positive cells, formed tube-like structures at 7-day after VEGF treatment. MSCs maintained in control medium stained negative. FACS analysis showed that untreated MSCs were negative for FVIII. The number of FVIII positive cells was not significantly increased at 3-day after induction (3%). But at 7-day after induction, FVIII-positive cells were significantly increased (13%). The expression of MMP-2 was increased as early as 1 hour post VEGF treatment. CONCLUSION: Porcine MSCs have the potential to differentiate into vascular endothelial cells in vitro and this process involves MMP-2.

[Repairing of peri-implant bone defect with platelet-rich plasma and platelet-rich plasma/ osteoinduction active material composite: an experimental study in dogs].

PURPOSE: To investigate the effect of platelet-rich plasma (PRP) and osteoinduction active material(OAM) compounded with PRP in the repair of bone defects of peri-implant and evaluate the feasibility of PRP as a material for repair of peri-implant bone defects. METHODS: Four Beagle dogs underwent extraction of the mandibular first, second and fourth premolars to create edentulous regions. Three months later, titanium implants were inserted and bone defects around implants were created. The bone defects were repaired with PRP/OAM compounds in experimental group A, repaired with PRP/ tricalcium phosphate in experimental group B, and repaired with tricalcium phosphate in the control group. The animals were sacrificed at the end of 8-week and 16-week, respectively. The specimens were observed with histological examination and energy disperse analysis of Ca(2+). The data were analyzed with SPSS13.0 software package for one-way ANOVA. RESULTS: At 8-week, in experimental group A, some parts of implants were directly contacted with new bone. Some new bones formed in the experimental group B. There was fibrous connection between the bone and implant in the control group. At 16-week, there was mature Haversian system in the experimental group A. There was good osseointegration between the bone and implant in the experimental groups but only was fibrous connection in the control group. At 8-and 16-week, there was significant difference between there groups in the percentage of Ca(2+) (P<0.05). CONCLUSIONS: PRP and PRP/OAM composite can effectively accelerate the regeneration of peri-implant bone defects and improve the osseointegration in the interface between the implant and bone.

[Experimental study on osteoinduction active material in repairing the peri-implant bone defect].

PURPOSE: To explore the potential of using osteoinduction active material (OAM) for reconstruction of bone defects of peri-implant by animal experiment, and provide experimental evidence for clinical application. METHODS: Four Beagle dogs underwent extraction of the mandibular first and second premolar to create edentulous regions. 2 groups were divided randomly. Three months later, titanium implants were inserted and peri-implant bone defect models were established. The bone defects were repaired with OAM in the experimental group and repaired with tricalcium phosphate (TCP) in the control group. Two animals each were sacrificed at the end of 8 weeks and 16 weeks respectively. The specimens were evaluated with histological examination, scanning electric microscope, bone density examination, and energy disperse analysis of Ca(2+). The data was analyzed with SPSS13.0 software package for Student's t test. RESULTS: At 8-week,some parts of implants in the experimental group were directly contacted with new bone. There were fibers between the bone and implant in the control group. There was no significant difference between the experimental group and the control group in bone density (P>0.05). The percentage of Ca(2+) of the experimental group(22.16+/-3.33) was significantly greater than that of the control group(3.13+/-2.44) (P<0.05). At 16-week, there was good osseointegration between the bone and implant in the experimental group but no new bone formation in the control group. The bone density of the experimental group was significantly greater than that of the control group (P<0.05). The percentage of Ca(2+) of the experimental group(42.23+/-6.20) was significantly greater than that of the control group(10.40+/-3.12)(P<0.05). CONCLUSION: The results suggest that OAM can effectively accelerate the reconstruction of peri-implant bone defects and improve the osseointegration in the interface between the implant and bone.

[The expression and significance of ERK1, ERK2 gene in tongue squamous cell carcinoma].

PURPOSE: The aim of this study is to investigate the relationship between ERK1, ERK2 gene expression and tumor behavior such as invasion, metastasis in tongue squamous cell carcinoma (TSCC). METHODS: The expressions of ERK1, ERK2 at gene level were examined in 45 cases of TSCC by reverse transcriptase polymerase chain reaction(RT-PCR). Student's t test was used for data analysis using SPSS 12.0 software package. RESULTS: No correlation between the expression intensity of ERK gene and tumor pathological degree of TSCC.A significant relationship was observed between the expression intensity of ERK gene and T stage of TSCC. A significant relationship was observed between the expression intensity of ERK gene and lymphnode metastasis of TSCC. CONCLUSIONS: Activation of ERK gene may play a strong role in cervical lymph node metastasis of TSCC. Intensive expression of ERK may be an important presage marker in TSCC patient. Supported by Key Science and Technology Research Project of Liaoning Province (Grant No.2005225013-2).

[Expression and significance of PTEN protein in tongue squamous cell carcinoma].

PURPOSE: To study the expressions and clinical significance of PTEN protein in tongue squamous cell carcinoma (TSCC). METHODS: The expressions of PTEN at protein level was investigated in 74 cases of TSCC and 15 cases of normal peripheral tissues around cancer (as control) by strep avidin-biotin complex (SABC) immunohistochemical technique. Mann-Whitney U-test, Pearson's Chi-square test and Fisher exact test were used to analyze the data. RESULTS: The rates of the positive expression in normal tissue and TSCC of PTEN were 100% and 66.2% respectively (P<0.05). Pathological grade and T stage were not significantly related with the level of PTEN expression (P>0.05). There was significant correlation between the level of PTEN expression and cervical lymph node status (P<0.05). CONCLUSION: Expressions of PTEN is correlated with cervical lymphnode metastasis in tongue squamous cell carcinoma.

[Effect of implant junction on bone growth by X-ray analysis].

OBJECTIVE: To explore the effects of mandibular implant junction on the growth of immature bone. METHODS: Eight Beagle dogs were randomly divided into three groups: control, unjunction and junction. Twelve implants were produced on the mandible of unjunction experimental group and junction experimental group. At three months after implanting, radiographic examination was performed. RESULTS: Three months after implanting, all implants were integrated with bone. None implants was mobile or got off. Radiographic examination demonstrated that the bone lose difference was no significant in junction and unjunction group. CONCLUSION: Mandibular implant connection wasn't effect on the growth of bone.

[Expressions and clinical significance of urokinase-type plasminogen activator (uPA) and uPA receptor (uPAR) in tongue squamous cell carcinoma].

PURPOSE: To study the expressions and clinical significance of urokinase-type plasminogen activator (uPA) and uPA receptor (uPAR) in tongue squamous cell carcinoma (TSCC). METHODS: The expressions of uPA and uPAR at protein level were examined in 74 cases of TSCC and 15 cases of normal peripheral tissues around cancer (as control) by strep avidin-biotin complex (SABC) immunohistochemical technique. The relationship between the expressions of uPA and uPAR was evaluated by the Spearman's rank correlation test. Mann-Whitney U-test was used to examine the association of these factors with conventional clinical pathological factors. Pearson's Chi-square test was used to analyze the relationship between uPA, uPAR and cervical lymph node status. RESULTS: The rates of the positive expression in TSCC of uPA and uPAR were 71.6% and 64.8% respectively, and a positive relationship between expression of uPA and uPAR was found (P<0.05). There was significant correlation between the level of uPA expression and TNM stage or cervical lymph node status, either between the level of uPAR expression and cervical lymph node status. Pathological grade was not significantly related to the level of uPA expression. Pathological grade and TNM stage were not significantly related to the level of uPAR expression. TSCC cases with concurrent positive expression of these proteins revealed significant higher tendency of cervical lymph node metastasis than those with concourse negative expression (P<0.05). CONCLUSIONS: The expression of uPA and uPAR increased in TSCC, and uPA, uPAR may contribute significantly to TSCC invasion and metastasis.

[Effect of implantation and implant connection on craniofacial bone growth in Beagle dogs].

PURPOSE: To study the effects of implantation and implant connection on Beagle craniofacial bone growth. METHODS: Eight Beagle dogs were randomly divided into two groups: unconnection group (4 dogs) and connection group (4 dogs). Two implant were implanted respectively in the lateral maxilla and zygomatic bone in each dog, the contra lateral side was used as normal control. Two implant was connected in the connection group 1 month later. Student's t test was used to analyze the data using SPSS11.5 software package. RESULTS: In the unconnection group, the distance increased between implants along with craniofacial bone growth; In the connection group, the distance fixed between implants along with craniofacial bone growth. The craniofacial values revealed no significant differences in the two groups (P > 0.05). CONCLUSION: The effect of implantation is localized and no inhibition in development of craniofacial bone is noted.

[Investigation of extracellular signal-regulated protein kinase and lymphnode metastasis in oral squamous cell carcinoma].

PURPOSE: To study the relationship between the expression of extracellular signal-regulated protein kinase (ERK) and the differentiation degree and lymphnode metastasis in oral squamous cell carcinoma. METHODS: Immunohistochemical staining was used in 41 cases of OSCC to investigate the expression of ERK protein. Student's t test were used for statistical analysis. RESULTS: A significant relationship was observed between the expression intensity of ERK protein and clinicopathological features of OSCC (P<0.01). A significant relationship was observed between the expression intensity of ERK protein and lymph node metastasis of OSCC(P<0.05). No correlation between the expression intensity of ERK protein and cell differentiation degree of OSCC (P>0.05) was found. CONCLUSIONS: Activation of ERK may play a strong role in the development of OSCC. Intensive expression of ERK protein may be an important presage marker in OSCC patients.