Effect of tert-butyl acetate and ethyl butyrate on the dissolution of gallbladder cholesterol stones / 中华肝胆外科杂志

Objective:To compare the effect and biotoxicity of tert-butyl acetate (TBA) and ethyl butyrate (EB) on stone dissolution in vitro.Methods:Ten gallstone samples from patients with multiple gallbladder stones were selected and the cholesterol content was analyzed by HPLC. Stone dissolution tests of TBA and EB were performed on cholesterol gallstone in vitro, and the weight of stone at each time point was recorded, meanwhile, methyl tert-butyl ether (MTBE) was used as the control. The inhibitory effects of MTBE, TBA and EB on proliferation of human normal liver cell line LO2 were analyzed by cell proliferation inhibition assay. Flow cytometry was used to analyze the effects of MTBE, TBA and EB on the early and late apoptosis of LO2 cells, and the changes of reactive oxygen species level in LO2 cells were also analyzed.Results:Of the 10 gallbladder gallstones, 6 were cholesterol gallstones and 4 were non-cholesterol gallstones. Stone dissolution experiment showed that the remaining stones of MTBE, TBA and EB groups were (47.83±3.84)%, (58.12±4.53)% and (75.75±4.61)% 30 minutes later. The remaining stones were (18.38±6.47)%, (33.82±6.22)% and (56.38±3.91)% 90 minutes later. MTBE had the best stone dissolution effect in vitro, the stone dissolution effect of TBA was slightly weaker than MTBE, and the stone dissolution effect of EB was relatively weak in all ( P<0.05). The cell proliferation inhibition experiment showed that the cell viability of the control group, MTBE group and TBA group were (100.00±4.46)%, (96.79±4.32)% and (93.72±3.51)%, respectively, and there were no significant differences among the three groups ( P>0.05). However, the cell viability of EB group (87.57±5.29)% was lower than the above three groups, and the differences were statistically significant ( P<0.001). The early apoptosis and late apoptosis of the control group were (1.67±0.15)% and (1.27±0.06)%, respectively. EB induced early apoptosis (15.90±0.53)% ( P<0.001) and late apoptosis (5.13±0.76)% ( P<0.05). However, MTBE and TBA had no significant effect on cell apoptosis ( P>0.05). Compared with control group, MTBE, TBA and EB all significantly inhibited the level of reactive oxygen species ( P<0.05), and the inhibitory effect of EB was the most obvious. Conclusions:TBA has good stone dissolution effect and biosafety for gallbladder cholesterol stones in vitro, while EB has relatively poor performance. TBA is a potential drug for gallstone dissolution.

Melatonin alleviates alcoholic liver disease via EGFR-BRG1-TERT axis regulation

Chronic alcohol consumption causes liver steatosis, cell death, and inflammation. Melatonin (MLT) is reported to alleviate alcoholic liver disease (ALD)-induced injury. However, its direct regulating targets in hepatocytes are not fully understood. In the current study, a cell-based screening model and a chronic ethanol-fed mice ALD model were used to test the protective mechanisms of MLT. MLT ameliorated ethanol-induced hepatocyte injury in both cell and animal models (optimal doses of 10 μmol/L and 5 mg/kg, respectively), including lowered liver steatosis, cell death, and inflammation. RNA-seq analysis and loss-of-function studies in AML-12 cells revealed that telomerase reverse transcriptase (TERT) was a key downstream effector of MLT. Biophysical assay found that epidermal growth factor receptor (EGFR) on the hepatocyte surface was a direct binding and regulating target of MLT. Liver specific knock-down of Tert or Egfr in the ALD mice model impaired MLT-mediated liver protection, partly through the regulation of nuclear brahma-related gene-1 (BRG1). Long-term administration (90 days) of MLT in healthy mice did not cause evident adverse effect. In conclusion, MLT is an efficacious and safe agent for ALD alleviation. Its direct regulating target in hepatocytes is EGFR and downstream BRG1-TERT axis. MLT might be used as a complimentary agent for alcoholics.

Immunohistochemical and mutational status of telomerase reverse transcriptase in conjunctival squamous cell carcinoma

Purpose: Mutations in human telomerase reverse transcriptase (TERT) are associated with increased telomerase activity in cutaneous melanomas. Conjunctival squamous cell carcinoma, also referred to as ocular surface squamous cell carcinoma, is cancer on the surface of the eye. Recent studies have identified UV signature mutations in TERT promoters in ocular melanoma and ocular surface squamous neoplasia. However, its immunohistochemical status has not been reported in ocular surface squamous cell carcinoma. This study aimed to explore the immunohistochemical and mutational status of TERT in ocular surface SCC. Methods: The immunohistochemical expression of TERT and mutational status of TERT promoter was evaluated in 19 ocular surface squamous cell carcinoma cases. Conjunctival melanoma tissue was used as a positive control. Results: The cytoplasmic overexpression of TERT was detected in 11/19 (57%), and TERT promoter mutations were identified in 6/19 (31%) of ocular surface squamous cell carcinoma. Out of these, 66% had a C228T mutation, and 33% had a C250T mutation. The TERT expression was found to be associated with a high (?T3) AJCC category (P = 0.023), and TERT immunoexpression was significantly correlated with reduced disease?free survival (P = 0.024, log?rank analysis) in ocular surface squamous cell carcinoma patients. Conclusion: The present study demonstrates that TERT promoter mutations with UV signatures are frequent in ocular surface squamous cell carcinoma. The increased expression of TERT could be of biological significance in aggressive ocular surface squamous cell carcinoma.

Protective effect of tert-butylhydroquinone on retinal morphology and function in early diabetic rats and its mechanism / 中华实验眼科杂志

Objective:To investigate the protective effects of an antioxidant tert-butylhydroquinone (tBHQ) on the morphology and function of retina in early-stage experimental diabetic rats, and to explore the mechanism of its protective effect.Methods:Forty-five healthy SD rats of clean degree were randomized into normal control group, diabetes model group and tBHQ intervention group, with 15 rats in each group according to a random number table.The diabetes model was established via a single intraperitoneal injection of streptozotocin (STZ) in diabetes model group and tBHQ intervention group.Normal control group was intraperitoneally administered with an equal-volume injection of sodium citrate buffer.Rats in the tBHQ intervention group maintained a diet with 1% tBHQ for 2 weeks before the STZ injection, and the other two groups were fed with normal rat food only.Blood from tail vein was collected to assay the blood glucose level at 72 hours, 2 weeks and 4 weeks following modeling.Rat electroretinogram (ERG) was detected at 4 weeks after modeling.Morphological changes of rat retina were observed by hematoxylin and eosin staining.The apoptosis of retinal cells in different layers was detected by TUNEL assay.The expression of protein kinase B (Akt), p-Akt, endothelial nitric oxide synthase (eNOS) and p-eNOS was detected by Western blot.Müller cell line rMC-1 cells cultured in vitro were divided into 5 groups, including normal control group (72-hour culturing in normal medium), mannitol control group (72-hour culturing in medium containing 5.5 mmol/L glucose and 24.5 mmol/L mannitol), high glucose group (72-hour culturing in high-glucose medium), tBHQ intervention group (24-hour culturing in normal-glucose medium containing 5 μmol/L tBHQ, 72-hour culturing in high-glucose medium containing 5 μmol/L tBHQ), and phosphoinositide 3-kinase (PI3K) inhibitor group (6-hour culturing in normal medium containing 5 μmol/L LY294002, 24-hour culturing in normal-glucose medium containing 5 μmol/L LY294002 and 5 μmol/L tBHQ, 72-hour culturing in high-glucose medium containing 5 μmol/L LY294002 and 5 μmol/L tBHQ). The expression of Akt, p-Akt, eNOS and p-eNOS in the cells was detected by western blot.The use and care of animals complied with Regulations for the Administration of Laboratory Animals in Southwest Medical University.The study protocol was approved by the Animal Ethics Committee of Southwest Medical University (No.201711189). Results:The blood glucose level at 72 hours, 2 weeks and 4 weeks after modeling was higher in diabetic model group than tBHQ intervention group and normal control group (all at P<0.01). Four weeks after modeling, the scotopic ERG a-wave and b-wave amplitudes of diabetic model group were lower than those of normal control group and tBHQ intervention group (all at P<0.05). With edema and thickening of inner plexiform layer, thinning of inner nuclear layer and outer nuclear layer, as well as loosely arrangement and disorder of retinal layers, the number of retinal ganglion cells was decreased in diabetic model group in comparison with normal control group, all of which were improved in tBHQ intervention group in comparison with diabetic model group.There were more apoptotic retinal cells in diabetic model group than normal control group and tBHQ intervention group (both at P<0.05), which mainly existed in the outer nuclear layer.The relative expressions of p-Akt/Akt and p-eNOS/eNOS in rat retina of normal control group, diabetic model group and tBHQ intervention group were 0.76±0.11 and 0.83±0.06, 0.52±0.10 and 0.52±0.08, 1.14±0.31 and 1.03±0.13, respectively.The relative expressions of p-Akt/Akt and p-eNOS/eNOS in diabetic model group were lower than those of normal control group and tBHQ intervention group (all at P<0.01). The relative expressions of p-Akt/Akt and p-eNOS/eNOS in normal glucose group, mannitol control group, high glucose group, tBHQ intervention group and PI3K inhibitor group were 0.95±0.38 and 0.86±0.11, 0.94±0.27 and 0.74±0.29, 0.33±0.25 and 0.45±0.29, 1.32±0.37 and 1.28±0.22, 0.24±0.09 and 0.73±0.29, respectively.The relative expressions of p-Akt/Akt and p-eNOS/eNOS were significantly lower in high glucose group than those in normal glucose group and tBHQ intervention group (all at P<0.05), which were significantly lower in PI3K inhibitor group compared with tBHQ intervention group (both at P<0.01). Conclusions:tBHQ has protective effects on the morphology and function of retina in early diabetic rats, and the mechanism may be related to the activation of Akt/eNOS signaling pathway.

Prognostic value of mutations in isocitrate dehydrogenase 1 (IDH1) and reverse telomerase transcriptase (TERT ) in Argentine patients` gliomas / Valor pronóstico de mutaciones en Isocitrato dehidrogenasa1 (IDH1) y Telomerasa transcriptasa reversa (TERT ) en gliomas de pacientes argentinos

Background and aim: Gliomas are the most common primary brain tumors, classified according to their histopathological and genetic features. Tumorigenesis depends on alterations in different genes. The aim of this study was the identification of mutations in IDH1 and TERT genes in gliomas of Argentine patients and to correlate them with clinical features and prognosis. Methods: DNA was isolated from 19 biopsies with different glioma grades matched with blood samples. IDH1 and TERT mutations were studied by PCR amplifica-tion and sequencing. Results: Six out of seven patients with low-grade glioma (grade II) harbor IDH1 mutations, mainly without tumor growth and overall survival of more than 12 months. Eleven out of twelve patients with high-grade gliomas (grade III/IV) showed wild type IDH1, mainly with tumor growth and shorter survival than low-grade gliomas. Mutated TERT promoter was present in 5 out of 11 high-grade gliomas, showing the prevalence of polymorphic C allele. In 1 out of 5 low-grade gliomas with a predominance of T allele. TERT and IDH1 mutations were mutually exclusive in most gliomas. Conclusions: Our results show that genetic tests provided a more accurate prognosis than histopathological analysis. The evolution of gliomas can be predicted primarily by the mutational status of IDH1 and secondarily by other markers, such as TERT mutational status

Antecedentes y objetivo: los gliomas son los tumores cerebrales primarios más comunes y se clasifican según sus características histopatológicas y genéticas. La tumorigénesis depende de alteraciones en diferentes genes. El objetivo de este estudio fue identificar mutaciones en los genes IDH1 y TERT en gliomas de pacientes argentinos y correlacionarlos con la evolución clínica. Métodos: se obtu-vieron 19 muestras pareadas de ADN de gliomas y de la sangre. Las mutaciones en IDH1 y TERT se analizaron por PCR y secuenciación. Resultados: la IDH1 mutada se encontró en 6 de los 7 gliomas de bajo grado (grado II), mayormente sin crecimiento tumoral y una sobrevida mayor de 12 meses. La IDH1 salvaje estaba presente en 11 de los 12 gliomas de alto grado (grado III y IV) mayormente con crecimiento tumoral y menor sobrevida que los tumores de bajo grado. Las mutaciones en el promotor del gen TERT se observaron en 5 de los 11 gliomas de alto grado, con la prevalencia de alelo polimórfico C, en cambio, en gliomas de bajo grado TERT mutado estaba presente en 1 de los 5 gliomas con predominio del alelo T. Las mutaciones en IDH1 y TERT fueron mutuamente excluyentes en la mayoría de los gliomas. Conclusiones: el análisis genético provee un pronóstico más certero que el análisis histopatológico. Nuestros resulta-dos muestran que la evolución de gliomas puede predecirse primariamente por el estado mutacional de IDH1 y secundariamente por mutaciones en otros marcadores tales como el TERT

Effect of Small-molecule GLP- 1 Receptor Agonist in Ovariectomized Mice with Osteoporosis / 中国药房

OBJECTIVE：To ex plore the improvement effects of small-molecule glucagon-like peptide- 1 receptor（GLP-1R） agonists（6，7-dichloro-2-methylsulfonyl-3-N-tert-butylaminoquinoxaline，DMB）on osteoporosis model mice. METHODS ：C57BL/ 6J mice were randomly divided into sham operation group ，model group ，positive control group （17β-estradiol，10 μg/kg）and DMB group （1 mg/kg），with 10 mice in each group. The sham operation group received bilateral laparotomy without ovariectomy ， and the other groups received bilateral ovariectomy to reproduce the osteoporosis model. At 4th week after operation ，sham operation group and model group were given constant volume of florence oil intragastrically ；administration groups were given relevant medicine intragastrically ，once a day ，for consecutive 8 weeks. After last medication ，Micro-CT was used to detect the femur microstructure ，bone mineral density （BMD），bone mineral content （BMC）and bone morphometric parameters [bone tissue volume fraction （BV/TV），trabecular thickness （Tb.Th），trabecular number （TB.N）and trabecular spacing （Tb.Sp）]. ELISA assay was used to detect the expression of bone formation indexes [bone specific alkaline phosphatase （BALP），osteoprotegerin（OPG）] and bone resorption indexes [tartrate resistant acid phosphatase （TRAP）and C-terminal cross-linked peptide of type Ⅰ collagen （CTX-Ⅰ）]. Three-point bending test was used to detect biomechanical parameters （maximum load ，stiffness，stress and Young ’s modulus）of femur of mice in each group. RESULTS ：In sham operation group ，trabeculae were numerous ，thick，complete in morphological structure ，well-organized and reticular ；bone Δ 基金项目：国家自然科学基金资助项目（No.81402931）；陕西省 重点研发计划项目（No.2017SF-261）；西安医学院 2016年博士科研启 microstructure and bone mass were normal. Compared with 动基金项目（No.2016DOC27） sham operation group ， the number of trabecular bone ＊讲师，博士。研究方向 ：分子药理学 。E-mail：zhouying209@ decreased，the thickness became thinner ，twisted or broken ， 163.com and the trabecular space increased ；the bone microstructure ·284· China Pharmacy 2021Vol. 32 No. 3 中国药房 2021年第32卷第3期 deteriorated and the bone m ass decreased ；BMD，BMC，BV/TV，Tb.Th，Tb.N and biomechanical parameters of femur were decreased significantly ，while Tb.Sp and serum levels of BLAP ，OPG，TRAP and CTX- Ⅰ were increased significantly （P＜0.05 or P＜0.01）. Compared with model group ，the bone mass of positive control group and DMB group were increased ，while the number，thickness and shape of trabecular bone partially recovered ；BMD，BMC，BV/TV，Tb.Th，Tb.N and biomechanical parameters of femur were increased significantly （P＜0.05 or P＜0.01）；Tb.Sp，serum levels of BLAP and OPG were increased significantly （P＜0.01）， while the levels of TRAP and CTX- Ⅰ were decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS：DMB can improve osteoporosis by increasing bone mass ，improving bone microstructure ，increasing the expression of bone formation related indexes and biomechanical parameters and decreasing bone resorption related indexes.

Tert-butanol application and related research progress in biomedicine / 药学学报

italic>Tert-butanol is an organic solvent, widely used in the medical field and chemical industry. It could be characterized by high crystallization temperature and vapor pressure. It could be easily sublimed and removed during the freeze-drying process. This review mainly describes the use of tert-butanol in the lyophilized formulations of poorly soluble drugs, the lyophilization solvent of porous structure productions, and as an ice crystal growth guider. In addition, the application of tert-butanol in nano drugs and aerogels has also been reviewed, as well as the current research progress in its quality and safety.

Discovery of thiosemicarbazone derivatives as effective New Delhi metallo-β-lactamase-1 (NDM-1) inhibitors against NDM-1 producing clinical isolates

@#New Delhi metallo-β-lactamase-1 (NDM-1) is capable of hydrolyzing nearly all β-lactam antibiotics, posing an emerging threat to public health. There are currently less effective treatment options for treating NDM-1 positive “superbug”, and no promising NDM-1 inhibitors were used in clinical practice. In this study, structure–activity relationship based on thiosemicarbazone derivatives was systematically characterized and their potential activities combined with meropenem (MEM) were evaluated. Compounds 19bg and 19bh exhibited excellent activity against 10 NDM-positive isolate clinical isolates in reversing MEM resistance. Further studies demonstrated compounds 19bg and 19bh were uncompetitive NDM-1 inhibitors with Ki = 0.63 and 0.44 μmol/L, respectively. Molecular docking speculated that compounds 19bg and 19bh were most likely to bind in the allosteric pocket which would affect the catalytic effect of NDM-1 on the substrate meropenem. Toxicity evaluation experiment showed that no hemolysis activities even at concentrations of 1000 mg/mL against red blood cells. In vivo experimental results showed combination of MEM and compound 19bh was markedly effective in treating infections caused by NDM-1 positive strain and prolonging the survival time of sepsis mice. Our finding showed that compound 19bh might be a promising lead in developing new inhibitor to treat NDM-1 producing superbug.

Facile synthesis of insulin fusion derivatives through sortase A ligation

Insulin derivatives such as insulin detemir and insulin degludec are U.S. Food and Drug Administration (FDA)-approved long-acting insulin currently used by millions of people with diabetes. These derivatives are modified in C-terminal B29 lysine to retain insulin bioactivity. New and efficient methods for facile synthesis of insulin derivatives may lead to new discovery of therapeutic insulin. Herein, we report a new method using sortase A (SrtA)-mediated ligation for the synthesis of insulin derivatives with high efficiency and functional group tolerance in the C-terminal B chain. This new insulin molecule (Ins-SA) with an SrtA-recognizing motif can be conjugated to diverse groups with N-terminal oligoglycines to generate new insulin derivatives. We further demonstrated that a new insulin derivative synthesized by this SrtA-mediated ligation shows strong cellular and

Clinical significance of co-mutation of BRAF V600E and TERT promoter in diagnosis and treatment of papillary thyroid carcinoma / 中华内分泌外科杂志

BRAF V600E mutation is a common gene mutation in papillary thyroid carcinoma (PTC) , which is associated with the occurrence, development, and prognosis of PTC. TERT promoter mutations are rare in PTC, but PTC with its mutations is more aggressive and has a worse prognosis. Clinically, a small number of PTC patients have both BRAF V600E mutation and TERT promoter mutations. This article provides a brief overview of the co-mutation of BRAF V600E and TERT promoter and the occurrence and development, clinical diagnosis, surgical strategies, postoperative adjuvant treatment, and prognosis of PTC.

Biosynthetic L-tert-leucine using Escherichia coli co-expressing a novel NADH-dependent leucine dehydrogenase and a formate dehydrogenase

BACKGROUND: L-tert-Leucine has been widely used in pharmaceutical, chemical, and other industries as a vital chiral intermediate. Compared with chemical methods, enzymatic methods to produce L-tert-leucine have unparalleled advantages. Previously, we found a novel leucine dehydrogenase from the halophilic thermophile Laceyella sacchari (LsLeuDH) that showed good thermostability and great potential for the synthesis of L-tertleucine in the preliminary study. Hence, we manage to use the LsLeuDH coupling with a formate dehydrogenase from Candida boidinii (CbFDH) in the biosynthesis of L-tert-leucine through reductive amination in the present study. RESULT: The double-plasmid recombinant strain exhibited higher conversion than the single-plasmid recombinant strain when resting cells cultivated in shake flask for 22 h were used. Under the optimized conditions, the double-plasmid recombinant E. coli BL21 (pETDute-FDH-LDH, pACYCDute-FDH) transformed 1 mol·L-1 trimethylpyruvate (TMP) completely into L-tert-leucine with greater than 99.9% ee within 8 h. CONCLUSIONS: The LsLeuDH showed great ability to biosynthesize L-tert-leucine. In addition, it provided a new option for the biosynthesis of L-tert-leucine.

Green approach toward protection of secondary amine in pyrazole nucleus by PEG-400 and Boc catalyst

The objective of the present study is the protection of secondary amine in the substituted pyrazole derivative withthe help of a green catalyst to facilitate the synthesis of anticancer compounds. Di-tert-butyl dicarbonate (Boc) hasbeen used as a protecting agent with various catalysts such as Polyethylene glycols-400, Dimethyl aminopyridine,and N, N-Diisopropyl ethylamine. In this study, five different synthetic methods have been applied, but success hasbeen achieved in only two. The very first method has been reported for the protection of secondary amine in pyrazole,associated with these catalysts. These synthetic methods had given a good yield and fewer side products, and it also agreen approach toward synthetic chemistry.

Relationship between BRAF V600E, TERT promoter mutations and the clinicopathological features of papillary thyroid carcinoma / 中国肿瘤临床

Objective: To explore the relationship between TERT promoter mutations and BRAF V600E as well as their coexistence with the clinicopathological features of papillary thyroid cancer (PTC). Methods: A total of 728 patients enrolled in Shanxi Cancer Hospital from December 2014 to December 2016 with PTC were retrospectively analyzed. We reviewed and analyzed the clinical results, pathology records, ultrasound results, and BRAF V600E and TERT status. Results: BRAF V600E mutations were found in 42.3% (308 of 728) of patients, and TERT C228T and C250T promoter mutations were found in 10.7% (78 of 728) and 0.5% (4 of 728) of patients, respectively. The TERT promoter mutation was significantly associated with old age (P=0.034), extrathyroidal invasion (P=0.026), large tumor size (P=0.028), cervical lymph node metastasis (P=0.012), distant metastasis (P=0.001), advanced disease stages (P<0.001), histological type (P=0.003) and recurrence (P=0.002). The BRAF V600E mutation was significantly associated with extrathyroidal invasion (P= 0.001), large tumor size (P<0.001), Hashimoto thyroiditis (P<0.001), distant metastasis (P=0.010), advanced disease stages (P=0.009) and recurrence (P=0.001). The coexistence of BRAF V600E and TERT promoter mutations was particularly associated with high-risk clinicopathological features, such as old age (P=0.024), extrathyroidal invasion (P=0.022), Hashimoto thyroiditis (P=0.005), the cervical lymph node (P=0.018), and advanced disease stages (P=0.002). Conclusions: Our study demonstrates that BRAF V600E and TERT promoter mutations play a significant role in the aggressiveness of PTC, particularly when the two mutations coexist. The results reveal the significant role of these mutations in the treatment and prognosis prediction of PTC.

Synthesis, in-vitro antioxidant activity and in-silico prediction of drug-likeness properties of a novel compound: 4-(3,5-Di-tert-butyl- 4-hydroxybenzylidene)-3-methylisoxazol-5(4H)-one

A simple and eco-friendly method has been used for the synthesis of novel compound 4-(3,5-di-tert-butyl-4-hydroxybenzylidene)-3-methylisoxazol-5(4H)-one. The synthesized compound was characterized by its physical andspectral data. This compound was screened for in-vitro antioxidant activity by 1,1-diphenyl-2-picrylhydrazyl (DPPH)and nitric oxide free radical scavenging assays at 100 µM concentration. The DPPH scavenging ability of evaluatedcompound was superior to the synthetic antioxidant butylated hydroxytoluene (BHT) and comparable to the standardascorbic acid. The nitric oxide scavenging ability of the compound was also found greater than the BHT. The insilico prediction of molecular properties and bioactivity scores of synthesized compound and BHT were carried outusing Molinspiration Cheminformatics online software. The study revealed that the predicted compounds obeyedLipinski’s rule of five and showed good intestinal absorption and membrane permeability indicating the drug-likenessproperties of predicted compounds. Finally, the in silico study identified the title compound as a nuclear receptorligand compared to BHT.

Preoperative detection of TERT promoter and BRAFV600E mutations in papillary thyroid carcinoma in high-risk thyroid nodules

ABSTRACT Objectives: This observational study analyzed telomerase reverse transcriptase (pTERT) mutations in 45 fine-needle aspiration (FNA) specimens obtained from thyroid nodules followed by postoperatively confirmation of papillary thyroid cancer (PTC) diagnosis, examining their relationship with clinicopathologic aspects and the BRAFV600E mutation. Subjects and methods: Clinical information was collected from patients who presented to Ribeirao Preto University Hospital for surgical consultation regarding a thyroid nodule and who underwent molecular testing between January 2010 to October 2012. Tests included a DNA-based somatic detection of BRAFV600E and pTERT mutations. Results: We found coexistence of pTERTC228T and BRAFV600E mutations in 8.9% (4/45) of thyroid nodules. All nodules positive for pTERT mutations were BRAFV600E positives. There was a significant association between pTERTC228T/BRAFV600E with older age and advanced stage compared with the group negative for either mutation. Conclusions: This series provides evidence that FNA is a reliable method for preoperative diagnosis of high-risk thyroid nodules. pTERTC228T/BRAFV600E mutations could be a marker of poor prognosis. Its use as a personalized molecular medicine tool to individualize treatment decisions and follow-up design needs to be further studied.

Association between genetic polymorphisms of TERT with susceptibility of age-related macular degeneration / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To investigate the relationship between single nucleotide polymorphism(SNP)in the TERT gene and risk of age-related macular degeneration(ARMD).<p>METHODS: Seven reported SNPs in TERT were selected and genotyped using SEQUENOM MassArray technology in 191 ARMD patients and 197 healthy normal controls. The correlations of the alleles, genotypes and the susceptibility of ARMD were evaluated by Logistic regression analysis.<p>RESULTS: The frequencies of SNP rs10069690 CC, CT, TT genotypes in ARMD patients were 64.4%, 31.4%, 4.2% compared with 74.6%, 22.8%, 2.5% in normal controls, respectively. Genetic model analysis revealed that SNP rs10069690 CT+TT genotypes were significantly associated with an increased risk of ARMD patients(<i>OR</i>=1.63, 95% <i>CI</i>=1.05-2.53).<p>CONCLUSION: SNP rs10069690 in the TERT gene is associated with the risk of ARMD.

Effects of short-chain acyl-CoA dehydrogenase on human umbilical vein endothelial cell apoptosis / 中华危重病急救医学

Objective To observe the changes of short-chain acyl-CoA dehydrogenase (SCAD) expression on human umbilical vein endothelial cell (HUVEC) apoptosis and investigate its relationship with apoptosis. Methods The HUVEC was cultured normally for 2-3 days. The apoptotic model of HUVEC was established by tert-butyl hydrogen peroxide (tBHP). The HUVEC was treated by different concentrations of tBHP (0, 10, 20, 30, 40, 50 μmol/L) for 12 hours and different time (0, 3, 6, 9, 12 hours) with 50 μmol/L tBHP to establish the apoptotic model of HUVEC. The cell viability was detected by methyl thiazolyl tetrazolium (MTT), the mRNA expression of SCAD was determined by real-time polymerase chain reaction (PCR), the protein expression of SCAD was achieved by Western Blot. The best concentrate and time were determined to interfere the HUVEC to achieve the apoptotic model of HUVEC. The SCAD gene of HUVEC was knocked down by RNA interference sequence (siRNA274, siRNA414, siRNA679). The mRNA expression of SCAD, the protein expression of SCAD and the activity of SCAD enzyme were detected to achieve the best RNA interference sequence. The HUVEC was intervened by the best RNA interference sequence and tBHP. The cell activity and apoptosis rate, the enzyme activity of SCAD, the mRNA and protein expression of SCAD, the contents of reactive oxygen species (ROS), aderosine triphosphate (ATP) and free fatty acid (FFA) were detected to observe the effect of SCAD on apoptosis of HUVEC. Results ① The cell viability, the mRNA expression and the protein expression of SCAD were decreased gradually in a concentration and time dependent manner with the increase of tBHP concentration and the prolongation of intervention time. The decline was most significant in the group of the 50 μmol/L tBHP to interfere HUVEC for 12 hours. ② The siRNA679 transfection was the most significant in reducing SCAD mRNA and protein expressions among the three interference sequences (siRNA274, siRNA414, siRNA679). ③ Compare with blank control group, the cell viability was significantly decreased in the siRNA679 group (A value: 0.48±0.09 vs. 1.00±0.09, P < 0.01), the apoptotic rate of HUVEC was significantly increased [(29.96±2.09)% vs. (2.90±1.90)%, P < 0.01], the expression of SCAD mRNA and SCAD protein, the activity of SCAD enzyme and the content of ATP were significantly decreased [SCAD mRNA (2-ΔΔCt): 0.50±0.16 vs. 1.34±0.12, SCAD/α-Tubulin: 0.67±0.11 vs. 1.00±0.06, the activity of SCAD enzyme (kU/g): 0.38±0.04 vs. 0.53±0.04, the content of ATP (μmol/g): 0.14±0.02 vs. 0.19±0.01, all P < 0.05], the contents of FFA and ROS were significantly increased [FFA (nmol/g): 0.84±0.07 vs. 0.47±0.04, ROS (average fluorescence intensity): 647.5±23.7 vs. 434.2±46.5, both P < 0.01]. Meanwhile, SCAD siRNA treatment triggered the same apoptosis as HUVEC treated with tBHP. Conclusions Down-regulation of SCAD may play an important role in HUVEC apoptosis. Increase in the expression of SCAD may become an important part in intervening HUVEC apoptosis.

The progress of hepatocellular adenoma malignant transforming in molecule mechanism researches / 中华肝胆外科杂志

Hepatocellular adenoma is a rare type of benign tumor in the liver. It has high risk of rupture and low risk of malignant transform. Recently the incidence of hepatocellular adenoma malignant transforming has been increasing. The malignant progress of hepatocellular adenoma develop to hepatocellular carcinoma has the transition state. This course not only relyes on the CTNNB1 gene exon 3 mutations, but also depends on TERT gene promoter mutation. This article will elaborate the hepatocellular adenoma malignant transforming in molecule mechanism, pathological diagnosis and therapies.

Protective Effect and Mechanism of Tetramethylpyrazine Against t-BHP-induced PC12 Cell Injury / 中国实验方剂学杂志

Objective:To study the protective effect of tetramethylpyrazine (TMP) on PC12 cells induced by tert-butyl hydroperoxide (t-BHP) and the regulatory mechanism on signaling pathway of phosphatidylinositol-3-kinases (PI3K)/kinase B (Akt)/mammalian target of rapamycin(mTOR). Method:PC12 cells cultured in vitro were treated with t-BHP (200 μmol·L-1) for 6 h to establish a model of oxidative damage in PC12 cells. The experiment was divided into blank group, model group (200 μmol·L-1t-BHP), TMP group. PC12 cells were pretreated with TMP(25, 50, 100 μmol·L-1) for 12 h, and then treated with t-BHP for 6 h. The cell viability was detected by cell counting kit-8(CCK-8) method, and lactate dehydrogenase (LDH) leakage, malondialdehyde (MDA) content, superoxide dismutase (SOD) activity, reactive oxygen species (ROS) and glutathione peroxidase (GSH-Px) activity were detected by enzyme-linked immunosorbent assay (ELISA). Apoptosis was observed by annexin V-FITC/PI double staining. B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), total protein kinase B (Akt), and phosphorylated protein kinase B (p-Akt), mTOR and p-mTOR expressions were detected by Western blot. Result:The cell viability of PC12 cells treated with 200 μmol·L-1 t-BHP decreased to about 50%after 6 h. This condition was suitable for the establishment of oxidative damage model. Compared with the model group, TMP (25, 50, 100 μmol·L-1) pretreatment for 12 h significantly increased the survival rate of PC12 cells (PPPPPPP-1) pretreatment group increased significantly (PConclusion:Ligustrazine protects PC12 cell injury induced by t-BHP by activating PI3K/Akt/mTOR signaling pathway.

The progress of hepatocellular adenoma malignant transforming in molecule mechanism researches / 中华肝胆外科杂志

Hepatocellular adenoma is a rare type of benign tumor in the liver. It has high risk of rupture and low risk of malignant transform. Recently the incidence of hepatocellular adenoma malignant transforming has been increasing. The malignant progress of hepatocellular adenoma develop to hepatocellular carcinoma has the transition state. This course not only relyes on the CTNNB1 gene exon 3 mutations, but also depends on TERT gene promoter mutation. This article will elaborate the hepatocellular adenoma malignant transforming in molecule mechanism, pathological diagnosis and therapies.