Objective To investigate the role of BRAFV600E mutation in diagnosis of thyroid nodules when it is inconsonant with cytological results. Methods This study included 9837 patients who underwent US-FNA. We mainly analyzed 239 cases with benign or indeterminate cytology, but having a detection of BRAFV600E muta-tion. BRAFV600E mutation analysis was performed using a Amplification Refractory Mutation System Polymerase Chain Reaction. Results In 93 nodules with benign cytology results but positive BRAFV600E mutation, 84 nodules were malignant. Based on the results, US-FNA combined with BRAFV600E mutation analysis will improve sensitivity (Se=94.03％)and negative predictive value (NPV=2.69％) of the thyroid nodules diagnosis than using US-FNA alone(Se=71.03％, NPV=20.76％). Conclusion BRAFV600E mutation analysis is an important tool in the diagnosis of PTC with high sensitivity and NPV. When facing patients with benign or indeterminate cytology but positive BRAFV600E mutation, thyroidectomy should be considered.

Objective: To investigate the role of BRAF^V600E mutation in diagnosis of thyroid nodules when it is inconsonant with cytological results. Methods: This study included 9837 patients who underwent US-FNA. We mainly analyzed 239 cases with benign or indeterminate cytology, but having a detection of BRAF^V600E mutation. BRAF^V600E mutation analysis was performed using a Amplification Refractory Mutation System Polymerase Chain Reaction. Results: In 93 nodules with benign cytology results but positive BRAF^V600E mutation, 84 nodules were malignant. Based on the results, US-FNA combined with BRAF^V600E mutation analysis will improve sensitivity (Se=94.03%) and negative predictive value (NPV=2.69%) of the thyroid nodules diagnosis than using US-FNA alone (Se=71.03%, NPV=20.76%) . Conclusion BRAF^V600E mutation analysis is an important tool in the diagnosis of PTC with high sensitivity and NPV. When facing patients with benign or indeterminate cytology but positive BRAF^V600E mutation, thyroidectomy should be considered.

On the basis of successful cloning the full length hemagglulinin (HA) and neuramidinase (NA) gene and sequence analysis of influenza virus H1N1, part of the gene was ligated into pMETA. Expression vectors pMETA/HA (52-1 557 bp) and pMETA/NA (121-1 263 bp) were constructed and expressed in pMAD16 induced by methanol. Recombinant protein was purified through Ni2+ affinity chromatography. Western blotting and ELISA were used to determine the antigenic activity of the recombinant protein. SDS-PAGE showed that the recombinant capsid gene could be overexpressed in Pichia methanolica. ELISA and Western blotting showed that the recombinant protein had antigenicity.
Cloning, Molecular ; Genetic Vectors ; genetics ; Hemagglutinins ; biosynthesis ; genetics ; Influenza A Virus, H1N1 Subtype ; genetics ; Neuraminidase ; biosynthesis ; genetics ; Pichia ; genetics ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics ; immunology

OBJECTIVETo evaluate the association between very low density lipoprotein cholesterol (VLDL-C) and cholesterol absorption and synthesis markers in patients with moderate and high risk of coronary heart disease.

METHODSA total 363 statin-naïve patients with moderate and high risk of coronary heart disease were consecutively recruited from two hospitals in Shanxi and Henan provinces between October 2008 and June 2009. A standard questionnaire and physical examination were performed at baseline. Atorvastatin (20 mg/day) was administered to patients for 4 weeks. Venous blood samples after an overnight fast were collected before and after treatment for measuring VLDL-C and cholesterol absorption and synthesis markers. In qualitative analyses, the baseline level of cholesterol absorption and synthesis markers and their reduction after atorvastatin treatment were categorized into 3 tertile groups.

RESULTS(1) Of 363 patients, 283 patients with mean age of (55.43±9.01)years old with complete data were finally analyzed. The median level of baseline VLDL-C was 1.06 (0.65, 1.86) mmol/L. The median level of baseline cholesterol absorption marker (Campesterol) and cholesterol synthesis marker (Lathosterol) was 6.01 (3.78, 9.45) mg/L and 13.46 (8.30, 21.07) mg/L, respectively. (2) Partial correlation analysis and multiple regression showed the baseline level of VLDL-C was positively correlated with Campesterol (r=0.153, P<0.05) but not with Lathosterol(r=0.182, P=0.173). Furthermore, baseline VLDL-C level significantly increased with tertile of the baseline level of Campesterol in the qualitative analyses(P for trend=0.035). (3) Mean reduction in VLDL-C levels was 38.0% after 4 weeks atorvastatin treatment. VLDL-C reduction was positively correlated with Campesterol reduction (r=0.331, P<0.001). VLDL-C reduction significantly increased with the tertile of Campesterol reduction (P for trend=0.032). But this trend was not observed between VLDL-C level and Lathosterol (P for trend=0.798).

CONCLUSIONThe level of VLDL-C was closely related to cholesterol absorption marker, and further studies are needed to validate if inhibitor of cholesterol absorption (for example by Ezetimibe) could bring about more effective VLDL-C lowering effect in this patient cohort.

Atorvastatin Calcium ; Biomarkers ; Cholesterol ; analogs & derivatives ; Cholesterol, LDL ; Cholesterol, VLDL ; Coronary Artery Disease ; Ezetimibe ; Humans ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; Phytosterols ; Risk Factors

The biophysical properties of the extracellular matrix (ECM) dictate tissue-specific cell behaviour. In the skeleton system, bone shows the potential to adapt its architecture and contexture to environmental rigidity via the bone remodelling process, which involves chondrocytes, osteoblasts, osteoclasts, osteocytes and even peripheral bone marrow-derived stem/stromal cells (BMSCs). In the current study, we generated stiff (~1 014 ± 56) kPa, Young's modulus) and soft (~46 ± 11) kPa silicon-based elastomer polydimethylsiloxane (PDMS) substrates by mixing curing agent into oligomeric base at 1:5 and 1:45 ratios, respectively, and investigated the influence of substrate stiffness on the cell behaviours by characterizing cell spreading area, cell cytoskeleton and cell adhesion capacity. The results showed that the cell spreading areas of chondrocytes, osteoblasts, osteoclasts, osteocytes and BMSCs were all reduced in the soft substrate relative to those in the stiff substrate. F-actin staining confirmed that the cytoskeleton was also changed in the soft group compared to that in the stiff group. Vinculin in focal adhesion plaques was significantly decreased in response to soft substrate compared to stiff substrate. This study establishes the potential correlation between microenvironmental mechanics and the skeletal system, and the results regarding changes in cell spreading area, cytoskeleton and cell adhesion further indicate the important role of biomechanics in the cell-matrix interaction.
Actins ; Cell Adhesion ; Elastic Modulus ; Focal Adhesions ; physiology ; Humans ; Vinculin ; analysis ; metabolism

OBJECTIVETo investigate the antibacterial activity of silver sulfadiazine (SD-Ag), mupirocin, and clotrimazole used alone or in combination against methicillin-resistant Staphylococcus aureus (MRSA) isolated from burn wounds.

METHODSEighteen MRSA isolates from wound excretion of 18 burn patients hospitalized in our unit from July to December 2011 were collected continuously and non-repetitively. (1) Minimum inhibitory concentration (MIC), 50% MIC (MIC50), and 90% MIC (MIC90) of SD-Ag, mupirocin, and clotrimazole used alone, those of SD-Ag and mupirocin used in combination, and those of SD-Ag, mupirocin, and clotrimazole used in combination to MRSA were determined by checkerboard agar dilution method. (2) Fractional inhibitory concentration (FIC) index was calculated to determine the combined effect of SD-Ag plus mupirocin, and SD-Ag plus mupirocin and clotrimazole. Synergy with FIC index less than or equal to 0.5 or additivity with FIC index more than 0.5 and less than or equal to 1.0 was regarded as effective, and indifference with FIC index more than 1.0 and less than or equal to 4.0 or antagonism with FIC index more than 4.0 was regarded as ineffective. The effective ratio was compared with overall ratio (assumed as 0) by unilateral binomial distribution test.

RESULTSThe MIC, MIC50, and MIC90 of SD-Ag, mupirocin, and clotrimazole used alone against 18 MRSA isolates were respectively 8, 8, 16 µg/mL; 2, 16, 64 µg/mL; 2, 2, 2 µg/mL. MIC of antimicrobial agents used in combination decreased from 3.1% to 50.0% as compared with that of individual agent used alone. Compared with those of single application of SD-Ag and mupirocin, MIC50 of SD-Ag and that of mupirocin both decreased 75.0%, and MIC90 of them decreased 87.5% when SD-Ag and mupirocin were used in combination. Compared with those of single application of SD-Ag, mupirocin, and clotrimazole, MIC50 of SD-Ag, mupirocin, and clotrimazole respectively decreased 75.0%, 87.5%, and 50.0%; MIC90 of them respectively decreased 87.5%, 96.9%, and 50.0% when SD-Ag, mupirocin, and clotrimazole were used in combination. Among the 18 MRSA isolates, the combined effect of SD-Ag and mupirocin was synergic in 9 isolates, additive in 7 isolates, indifferent in 2 isolates, and antagonistic in 0 isolate; the combined effect of SD-Ag, mupirocin, and clotrimazole was additive in 16 isolates, indifferent in 2 isolates, and antagonistic in 0 isolate. There were statistically significant differences between effective ratio and overall ratio of 18 MRSA isolates treated with combined antimicrobial agents (P values all above 0.01).

CONCLUSIONSFor burn wounds at middle and late stages infected with Staphylococcus aureus or Staphylococcus aureus and Fungus, low dose of SD-Ag or combination of above-mentioned antimicrobial agents can effectively control infection and decrease the adverse effect of antimicrobial agents on wound healing.

Adolescent ; Adult ; Aged ; Anti-Bacterial Agents ; administration & dosage ; adverse effects ; pharmacology ; Burns ; microbiology ; Child ; Child, Preschool ; Clotrimazole ; administration & dosage ; adverse effects ; pharmacology ; Drug Therapy, Combination ; Female ; Humans ; Infant ; Male ; Methicillin-Resistant Staphylococcus aureus ; drug effects ; isolation & purification ; Middle Aged ; Mupirocin ; administration & dosage ; adverse effects ; pharmacology ; Silver Sulfadiazine ; administration & dosage ; adverse effects ; pharmacology ; Young Adult

Programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) blocking therapy has become a major pillar of cancer immunotherapy. Compared with antibodies targeting, small-molecule checkpoint inhibitors which have favorable pharmacokinetics are urgently needed. Here we identified berberine (BBR), a proven anti-inflammation drug, as a negative regulator of PD-L1 from a set of traditional Chinese medicine (TCM) chemical monomers. BBR enhanced the sensitivity of tumour cells to co-cultured T-cells by decreasing the level of PD-L1 in cancer cells. In addition, BBR exerted its antitumor effect in Lewis tumor xenograft mice through enhancing tumor-infiltrating T-cell immunity and attenuating the activation of immunosuppressive myeloid-derived suppressor cells (MDSCs) and regulatory T-cells (Tregs). BBR triggered PD-L1 degradation through ubiquitin (Ub)/proteasome-dependent pathway. Remarkably, BBR selectively bound to the glutamic acid 76 of constitutive photomorphogenic-9 signalosome 5 (CSN5) and inhibited PD-1/PD-L1 axis through its deubiquitination activity, resulting in ubiquitination and degradation of PD-L1. Our data reveals a previously unrecognized antitumor mechanism of BBR, suggesting BBR is small-molecule immune checkpoint inhibitor for cancer treatment.

Using chemoproteomic techniques, we first identified EIF2AK2, eEF1A1, PRDX3 and VPS4B as direct targets of berberine (BBR) for its synergistically anti-inflammatory effects. Of them, BBR has the strongest affinity with EIF2AK2 via two ionic bonds, and regulates several key inflammatory pathways through EIF2AK2, indicating the dominant role of EIF2AK2. Also, BBR could subtly inhibit the dimerization of EIF2AK2, rather than its enzyme activity, to selectively modulate its downstream pathways including JNK, NF-κB, AKT and NLRP3, with an advantage of good safety profile. In EIF2AK2 gene knockdown mice, the inhibitory IL-1β, IL-6, IL-18 and TNF-α secretion of BBR was obviously attenuated, confirming an EIF2AK2-dependent anti-inflammatory efficacy. The results highlight the BBR's network mechanism on anti-inflammatory effects in which EIF2AK2 is a key target, and inhibition of EIF2AK2 dimerization has a potential to be a therapeutic strategy against inflammation-related disorders.