rs10865710 polymorphism in PPARG promoter is associated with the severity of type 2 diabetes mellitus and coronary artery disease in a Chinese population.

BACKGROUND: Relationship between polymorphisms in peroxisome proliferator-activated receptor gamma (PPARG) and progression of type 2 diabetes mellitus (T2DM) and coronary artery disease (CAD) remains to be clarified. METHODS: 635 subjects were divided into T2DM, CAD, T2DM complicated with CAD (T2DM/CAD) and control groups according to diagnostic criteria. The rs10865710 and rs3856806 polymorphisms were genotyped, and the severity of T2DM and CAD was evaluated for all subjects. RESULTS: In patients with T2DM, G allele carriers of rs10865710 polymorphism had significantly higher levels of glucose, triglycerides, apolipoprotein B (ApoB) and lipoprotein (a) (Lp(a)) than non-carriers, T allele carriers of rs3856806 polymorphism had significantly higher levels of glucose, low-density lipoprotein cholesterol (LDL-C), ApoB and Lp(a) than non-carriers. In patients with CAD, G allele carriers of rs10865710 polymorphism had significantly higher levels of total cholesterol (TC), ApoB and Lp(a) than non-carriers, T allele carriers of rs3856806 polymorphism had significantly higher levels of body mass index, blood pressure, TC, LDL-C and ApoB than non-carriers. Patients with one or two G alleles of rs10865710 polymorphism had significantly higher levels of Gensini scores and more diseased coronary branches than those patients without CAD. The rs3856806 polymorphism was not associated with CAD severity, but it was found to be significantly associated with T2DM/CAD, T allele frequency was significantly higher in T2DM/CAD group than that in T2DM/CAD-free group. CONCLUSIONS: The rs10865710 and rs3856806 polymorphisms in PPARG are significantly associated with glucose levels in patients with T2DM. The rs10865710 polymorphism is significantly associated with the severity of CAD, which is possibly mediated by hyperlipidaemia and hyperglycaemia.

Soluble expression, purification, and characterization of active recombinant human tissue plasminogen activator by auto-induction in E. coli.

BACKGROUND: Human tissue plasminogen activator (tPA) belongs to the serine protease family. It converts plasminogen into plasmin and is used clinically to treat thrombosis. Human tPA is composed of 527 amino acids residues and contains 17 disulfide bonds. Escherichia coli has been used only rarely for the efficient production of recombinant tPA. However, the functional expression of full-length tPA that contains multiple disulfide bonds on an industrial scale remains challenging. Here, we describe the soluble expression and characterization of full-length tPA by auto-induction in E. coli. RESULTS: We achieved optimal levels of gene expression, minimized negative effects related to the production of heterologous proteins, and optimized cytoplasmic yields. Three different E. coli strains, BL21 (DE3), Rosetta, and Origami 2, could express tPA using an auto-induction mechanism. In addition, similar yields of recombinant protein were produced at temperatures of 33, 35, and 37°C. The E. coli strain origami 2 could increase disulfide bond formation in cytoplasmic tPA and produce purified soluble recombinant protein (~0.9 mg/l medium). The full-length tPA was monomeric in solution, and fibrin plate assays confirmed that the recombinant tPA displayed serine protease activity. CONCLUSIONS: This is the first report that describes the heterologous expression of correctly folded active full-length tPA. This could provide valuable information for using prokaryotic auto-induction expression systems to produce tPA at industrial and pharmaceutical levels without in vitro refolding during the production step.

Nuclear expression of CDK4 correlates with disease progression and poor prognosis in human nasopharyngeal carcinoma.

AIMS: The purpose of this study was to examine the correlation between nuclear expression of cyclin-dependent kinase 4 (CDK4) and clinicopathological data in nasopharyngeal carcinoma (NPC), including patient survival. METHODS AND RESULTS: Using real-time PCR and immunohistochemistry, the expression of CDK4 was examined in NPC and nasopharyngeal (NP) tissues. We observed that mRNA expression of CDK4 was elevated significantly in NPC tissues compared to NP tissues. Further, we found that CDK4 protein was expressed in both the nucleus and cytoplasm. Nuclear expression of CDK4 was correlated positively with clinical stage (P = 0.048), but not associated with other clinical features. Patients with tumours showing nuclear expression of CDK4 had poorer overall survival rates than those without nuclear tumour expression of CDK4. Nuclear expression of CDK4 was associated inversely with survival time for NPC patients in stages T1-2, stages N2-3 and clinical stages III-IV, and after treatment with radiotherapy or chemotherapy. Nuclear expression of CDK4 was an independent and unfavourable prognostic factor for patients with NPC. CONCLUSIONS: Our findings suggest that nuclear expression of CDK4 is a potential marker for the progression and poor prognosis of NPC.

Knocking down CDK4 mediates the elevation of let-7c suppressing cell growth in nasopharyngeal carcinoma.

BACKGROUND: CDK4 is a protein kinase in the CDK family important for G1/S phase cell cycle progression. However, the roles and molecular mechanisms of CDK4 triggering nasopharynx carcinogenesis are still unclear. METHODS: Lentiviral-vector mediated shRNA was used to suppress CDK4 expression and examine its molecular mechanisms. Using immunohistochemistry, we analyzed CDK4 protein expression in clinicopathologically characterized nasopharyngeal carcinoma (NPC) cases and nasopharyngeal tissues (NPs). Survival curves were plotted by the Kaplan-Meier method and compared using the log-rank test. RESULTS: In this investigation, we knocked down CDK4 expression and observed that NPC cell growth and cell cycle progression were significantly blocked by suppressing expression of CCND1, CDK6, and E2F1 as well as elevated p21 expression. Further, we found that reduced CDK4 expression elevated the expression of let-7c, a tumor-suppressive miRNA modulated by E2F1. We found that let-7c was markedly downregulated in NPC tissues compared to NPs and suppressed cell growth and cell cycle progression by modulating p15/p16/CDK4/E2F1 pathway. Finally, CDK4 protein was observed to be overexpressed in NPC tissues and could be considered an unfavorable prognosis factor for NPC patients although its independent prognostic value did not reach statistical significance (p = 0.087). CONCLUSIONS: Our results demonstrated that overexpressed CDK4 is an unfavorable prognostic factor which suppresses the expression of tumor suppressive-factor let-7c through p21/CCND1/CDK6/E2F1 signaling, and inhibits cell proliferation by p15/p16/CDK4/E2F1 feedback signaling in NPC.

[Expression of ABCG2 and p-glycoprotein in residual breast cancer tissue after chemotherapy and their correlation with epithelial-mesenchymal transition].

OBJECTIVE: To explore the expression of breast cancer resistance protein (ABCG2), p-glycoprotein (P-gp) in residual breast cancer tissue after chemotherapy and their correlation with epithelial mesenchymal transition (EMT). METHODS: Seventy-six cases of breast cancer were collected. The expression of ABCG2, P-gp and EMT markers E-cadherin and vimentin in residual breast cancer tissue after chemotherapy was detected by immunohistochemistry (EnVision method). MCF7 cells were divided into three group:untreated control group, positive control (TGF-ß1 induced) group and drug surviving cells (DSC) group (selected viable MCF7 cells after docetaxel and epirubicin treatment). The expression of EMT markers E-cadherin and vimentin was detected by immunofluorescence. The mRNA and protein expression of ABCG2, P-gp and EMT markers were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot, respectively. RESULTS: Compared with breast cancer tissue before chemotherapy, ABCG2, P-gp and vimentin protein were highly expressed in residual breast cancer tissue after chemotherapy. The expression of ABCG2 and P-gp correlated positively with vimentin protein (r1=0.97, P1=0.000; r2=0.83, P2=0.001) and negatively with E-cadherin protein (r3=-0.55, P3=0.010; r4=-0.43, P4=0.020) expression. RT-PCR results showed that ABCG2, P-gp and vimentin mRNA were highly expressed in residual breast cancer tissue after chemotherapy. The expression of ABCG2 and P-gp mRNA correlated positively with vimentin mRNA (r1=0.99, r2=0.96, P<0.05) but negatively with E-cadherin protein (r3=-0.99, r4=-0.98, P<0.05); Western blot showed that ABCG2, P-gp and vimentin protein were highly expressed in residual breast cancer tissue after chemotherapy. The expression of ABCG2 and P-gp protein correlated positively with vimentin protein (r1=0.98, r2=0.89, P<0.05) and negatively with E-cadherin protein (r3=-0.47, r4=-0.33, P<0.05). CONCLUSIONS: The expression of resistance-associated proteins in the residual breast cancer tissue after chemotherapy is significantly correlated with EMT. The expression of EMT profile may be one of important mechanisms for multidrug resistance in breast cancer.

Loss of cytoplasmic KLF4 expression is correlated with the progression and poor prognosis of nasopharyngeal carcinoma.

AIMS: To examine, in nasopharyngeal carcinoma (NPC), the correlation of Krüppel-like factor 4 (KLF4) expression with clinicopathological features including patient prognosis. METHODS AND RESULTS: Using real-time PCR and immunohistochemistry, expression of KLF4 mRNA and protein was examined in NPC and nasopharyngeal tissues. The relationship of KLF4 expression levels with clinical features and prognosis of NPC patients was analysed. mRNA expression was markedly lower in NPC than in the nasopharyngeal tissues. Using immunohistochemistry, staining for KLF4 protein was found in the nuclei and cytoplasm of nasopharyngeal and malignant epithelial cells, but decreased cytoplasmic expression was observed in atypical hyperplasia and NPC samples compared to normal and squamous epithelium samples (P < 0.001). In addition, levels of cytoplasmic KLF4 protein were correlated inversely with the nodal (N) status (TNM classification; P = 0.002) and overall clinical stage (P < 0.001) of NPC patients. Patients with NPC showing lower cytoplasmic KLF4 expression had a significantly shorter overall survival time than those with high NPC KLF4 expression. Multivariate analysis suggested that the level of KLF4 expression was an independent prognostic indicator (P = 0.008) for NPC survival. CONCLUSION: Low levels of cytoplasmic KLF4 expression are a potentially unfavourable prognostic factor for patients with NPC.

Proteomic features of potential tumor suppressor NESG1 in nasopharyngeal carcinoma.

We previously defined the recently revised NESG1 gene as a potential tumor suppressor in nasopharyngeal carcinoma (NPC). Here, we further used proteomics technology to globally examine NESG1-controlled proteins in NPC cells. Twenty-six proteins were found to be deregulated by NESG1 using proteomics analysis while enolase 1 (alpha) (ENO1), heat shock protein 90 kDa beta (Grp94), member 1 (HSP90B1), and cathepsin D (CTSD) proteins were differentially expressed by Western blot. Interestingly, a-enolase (ENO1), an overexpressed gene in NPC, was confirmed as a NESG1-regulated protein in NPC cells. Overexpressed ENO1 not only restored cell proliferation and cell-cycle progression, but also antagonized the regulation of NESG1 to cell-cycle regulators p21 and CCNA1 expression as well as induced the expression of C-Myc, pRB, and E2F1 in NESG1-ovexpressed NPC cells. Real-time PCR and immunohistochemistry analysis showed that NESG1 expression is negatively correlated with ENO1 expression in NPC tissues. Our observations suggest that ENO1 downregulation plays an important role in NESG1-induced growth inhibition of NPC cancer cells.

Nuclear p27 expression confers a favorable outcome for nasopharyngeal carcinoma patients.

OBJECTIVE: The purpose of the present study is to explore the correlation between nuclear expression of cyclin-dependent kinase inhibitor 1B (p27) and clinicopathologic features in nasopharyngeal carcinoma (NPC), including patient survival. METHODS: Immunohistochemistry was used to examine the expression of p27 in 130 primary NPC tissues. The relationship between the levels of p27 expression and clinicopathologic characteristics was analyzed. Survival curves were plotted using the Kaplan-Meier method and compared using the log-rank test. The significance of various survival variables was analyzed using multivariate Cox proportional hazards model. RESULTS: p27 was expressed in both nuclear and cytoplasmic compartments. Nuclear expression of p27 was inversely correlated with T classification and clinical stage. Patients with nuclear p27 expression had better overall survival rates than those without nuclear expression of p27. Further, we observed that nuclear expression of p27 was positively associated with survival time of NPC patients not only in N0-1 and M0 classifications but also in radiotherapy and chemotherapy treatment groups. Finally, we found that nuclear expression of p27 was not an independent prognostic factor for patients with NPC. CONCLUSIONS: Our findings hint that nuclear expression of p27 is a potentially favorable factor in the progression and prognosis of NPC.

Tumor suppressor PDCD4 modulates miR-184-mediated direct suppression of C-MYC and BCL2 blocking cell growth and survival in nasopharyngeal carcinoma.

Programmed cell death 4 (PDCD4), a novel tumor suppressor, inhibits cell proliferation, migration and invasion as well as promotes cell apoptosis in tumors. However, the molecular mechanism of its tumor-suppressive function remains largely unknown in tumors including nasopharyngeal carcinoma (NPC). In this study, downregulated PDCD4 expression was significantly associated with the status of NPC progression and poor prognosis. PDCD4 markedly suppressed the ability of cell proliferation and cell survival by modulating C-MYC-controlled cell cycle and BCL-2-mediated mitochondrion apoptosis resistance signals, and oncogenic transcription factor C-JUN in NPC. Furthermore, miR-184, a tumor-suppressive miRNA modulated by PDCD4 directly targeting BCL2 and C-MYC, participated in PDCD4-mediated suppression of cell proliferation and survival in NPC. Further, we found that PDCD4 decreased the binding of C-Jun to the AP-1 element on the miR-184 promoter regions by PI3K/AKT/JNK/C-Jun pathway and stimulated miR-184 expression. In clinical fresh specimens, reduced PDCD4 mRNA level was positively correlated with miR-184 expression in NPC. Our studies are the first to demonstrate that PDCD4 as tumor suppressor regulated miR-184-mediated direct targeting of BCL2 and C-MYC via PI3K/AKT and JNK/C-Jun pathway attenuating cell proliferation and survival in NPC.

[Normal anatomy of aditus of antrum and antrum on high-resolution CT and three-dimensional reconstruction].

OBJECTIVE: To study the normal anatomy of aditus of antrum and antrum on the high-resolution CT (HRCT) and the three-dimensional reconstruction, testing the normal range. And comparison was carried out according to the age, sex and side. METHOD: Ninety cases were randomly selected without ear lesions. Scanning were taken in sagittal, transverse and coronal planes on HRCT respectively. The structure of aditus and antrum was displayed by three-dimensional reconstruction. The left-right distances and up-down distances, antero-posterior distances were measured and analyzed. RESULT: The image of antrum varied with age, while aditus remained constant on the HRCT and the three-dimensional reconstruction. The average of left-right distance of aditus was (5.19 +/- 1.39) mm, and the average of up-down distance of aditus was (5.74 +/- 1.16) mm. The average of left-right distance of antrum was (8.27 +/- 1.41) mm (<6 years old) and (5.41 +/- 1.32) mm (> or = 6 years old). The average of up-down distance of antrum was (11.78 +/- 1.65) mm (<6 years old) and (9.91 +/- 2.04) mm (> or = 6 years old). The average of antero-posterior distance of antrum was (12.25 +/- 1.23) mm (<6 years old) and (10.05 +/- 1.69) mm (> or = 6 years old). No statistically significant differences were seen in left-right distance of aditus by age, sex and side (P > 0.05). Significant differences in up-down distance of aditus was found between male and female, and the distance in male was greater than that in female (P < 0.05). Statistically significant differences were seen in left-right distance and up-down distance, antero-posterior distance of antrum by age (P < 0.05), but no statistically significant differences by sex or side (P > 0.05). CONCLUSION: Imaging of aditus ad antrum is relatively constant in the normal persons, while the aditus is more diverse. Significant gender differences were seen in up-down distance. There were significant differences in left-right distance, up-down distance, and antero-posterior distance of aditus among all age groups.

[Clinical analysis of plasma cell granuloma occurred in head and neck].

OBJECTIVE: To discuss the diagnosis and treatment of plasma cell granuloma in the head and neck region. METHOD: Retrospective analysis of 3 cases of plasma cell granuloma occurred in head and neck. One case treated with endoscopic surgery and nasal and oral steroids after operation. Two cases treated with Caldwell-Luc style resection, followed by oral steroids and radiotherapy, one of the two cases combined with chemotherapy simultaneously. RESULT: The patient done with endoscopic surgery followed up for 5 years, there was no tumor recurrence in the nasal cavity and maxillary sinus openings and sinus lining. Paranasal sinus CT was examed on the patient done with partial resection and radiotherapy 2 years after operation, no residual tumor proliferation. The third patient underwent paranasal sinus CT examination 3 years after operation, and no limitation of mouth opening was found. CONCLUSION: The incidence rate of Plasma cell granuloma occured in head and neck is low. It is an inflammatory disease, but its physical signs and imaging findings are similar to malignant tumors. Because of the invasion feature, surgical resection should be the first choice, and if combined with radiotherapy and chemotherapy, the prognosis could be better.

[Mastoidectomy and ventilation tube placement for refractory secretory otitis media].

OBJECTIVE: To study the mastoidectomy and ventilation tube placement for refractory secretory otitis media of clinical efficacy and mechanism. METHOD: Retrospective analysis of 22 patients (33 ears) in refractory secretory otitis media, all patients treated by ventilation tube placement have 3 or more than 3 times, but not significantly alleviate the symptoms. Mastoid surgery and ventilation tube placement were basic surgical management. Tympanic membrane ventilation tube was pulled out at 3 to 6 months. RESULT: Twenty-two patients perceived improvement of hearing after surgery, ear fullness disappeared; tympanic membrane was gray, no significant tympanic membrane mobility is limited; 33 ears conductive hearing loss, air-bone gap(13.54 +/- 4.86) dB; after 29 ears tympanograms showed A-type, 4 ears for C-type, 30 ears appear ipsilateral acoustic reflex. CONCLUSION: For 3 or more than 3 times repeated ventilation tube insertion, patients more than 2 years of refractory secretory otitis media were treated with mastoidectomy and ventilation tube placement, it was satisfied that ears lesions were cleaned and expanded middle ear and mastoid air cell volume, good drainage of the tympanic membrane ventilation tube.

[The expressions of alpha-enolase in the nasopharyngeal cancer tissue].

OBJECTIVE: To study the meaning of expressions of ENO1 in the nasopharyngeal cancer tissue. METHOD: By using SP immunohistochemical methods. RESULT: The positive expressions for ENO1 were 60.0% in the nasopharyngeal cancer tissues,significantly higher than that (27.5%, 11/40) of in inflammatory nasopharyngeal mucosa (P<0.05). The positive expression rate of ENO1 protein in stage T1 + T2 + T3 was higher than that of in patients with T4 (chi2 = 11.424, P<0.05). But there was no significant relationship between expression of ENO1 and lymph node metastasis and postradiotherapy distant metastasis (P>0.05). CONCLUSION: The results show that ENO1 over expression may play a important role in cell proliferation and canceration of nasopharyngeal cancer.

[Mastoid surgery for secretory otitis media with mixed hearing loss].

OBJECTIVE: To analyze the therapeutic effect of mastoid surgery for secretory otitis media with mixed hearing loss. METHODS: A retrospective analysis was conducted of the data from 26 cases (43 ears) of secretory otitis media with bone conduction hearing loss collected from 2001 to 2008. Thirty-two ears were treated with mastoid surgery and myringotomy with insertion of ventilation tubes. All the patients received medications after the operation. RESULTS: All the patients showed obvious improvement after mastoid surgery. The average pure tone of air conduction hearing threshold was about 25 dB after the surgery, with the average pure tone of bone conduction hearing threshold of about 15 dB. The patients were followed up for 1-2 years during which no significant change in hearing was recorded, and no middle ear effusion in the tympanic cavity was found after removal of the ventilation tubes. CONCLUSION: Persistent secretory otitis media can be associated with mixed hearing loss, and mastoid surgery can significantly enhance the hearing level to produce positive therapeutic effects.