Radiofrequency Ablation of Benign Thyroid Nodules and Recurrent Thyroid Cancers: Consensus Statement and Recommendations

Thermal ablation using radiofrequency is a new, minimally invasive modality employed as an alternative to surgery in patients with benign thyroid nodules and recurrent thyroid cancers. The Task Force Committee of the Korean Society of Thyroid Radiology has developed recommendations for the optimal use of radiofrequency ablation for thyroid nodules. These recommendations are based on a comprehensive analysis of the current literature, the results of multicenter studies, and expert consensus

The 5p15.33 Locus Is Associated with Risk of Lung Adenocarcinoma in Never-Smoking Females in Asia

Genome-wide association studies of lung cancer reported in populations of European background have identified three regions on chromosomes 5p15.33, 6p21.33, and 15q25 that have achieved genome-wide significance with p-values of 10−7 or lower. These studies have been performed primarily in cigarette smokers, raising the possibility that the observed associations could be related to tobacco use, lung carcinogenesis, or both. Since most women in Asia do not smoke, we conducted a genome-wide association study of lung adenocarcinoma in never-smoking females (584 cases, 585 controls) among Han Chinese in Taiwan and found that the most significant association was for rs2736100 on chromosome 5p15.33 (p = 1.30×10−11). This finding was independently replicated in seven studies from East Asia totaling 1,164 lung adenocarcinomas and 1,736 controls (p = 5.38×10−11). A pooled analysis achieved genome-wide significance for rs2736100. This SNP marker localizes to the CLPTM1L-TERT locus on chromosome 5p15.33 (p = 2.60×10−20, allelic risk = 1.54, 95% Confidence Interval (CI) 1.41–1.68). Risks for heterozygote and homozygote carriers of the minor allele were 1.62 (95% CI; 1.40–1.87), and 2.35 (95% CI: 1.95–2.83), respectively. In summary, our results show that genetic variation in the CLPTM1L-TERT locus of chromosome 5p15.33 is directly associated with the risk of lung cancer, most notably adenocarcinoma

Soluble expression and partial purification of recombinant human erythropoietin from E. coli

Human erythropoietin (hEpo) is an essential regulator of erythrocyte production that induces the division and differentiation of erythroid progenitor cells in the bone marrow into mature erythrocytes. It is widely used for the treatment of anemia resulting from chronic kidney disease, chemotherapy, and cancer-related therapies. Active hEpo, and hEpo analogs, have been purified primarily from mammalian cells, which has several disadvantages, including low yields and high production costs. Although an Escherichia coli (E. coli) expression system may provide economic production of therapeutic proteins, it has not been used for the production of recombinant hEpo (rhEpo) because it aggregates in inclusion bodies in the E. coli cytoplasm and is not modified post-translationally. We investigated the soluble overexpression of active rhEpo with various protein tags in E. coli, and found out that several tags increased the solubility of rhEpo. Among them maltose binding protein (MBP)-tagged rhEpo was purified using affinity and gel filtration columns. Non-denaturing electrophoresis and MALDI-TOF MS analysis demonstrated that the purified rhEpo had two intra-disulfide bonds identical to those of the native hEpo. An in vitro proliferation assay showed that rhEpo purified from E. coli had similar biological activity as rhEpo derived from CHO cells. Therefore, we report for the first time that active rhEpo was overexpressed as a soluble form in the cytoplasm of E. coli and purified it in simple purification steps. We hope that our results offer opportunities for progress in rhEpo therapeutics.ASTAR (Agency for Sci., Tech. and Research, S’pore

Cell proliferation assay of purified hGH in the Nb2-11 cell line.

<p>Dose-response proliferation curve of Nb2-11 cells by exposure to different concentrations of purified hGH and commercial hGH.•, commercial hGH; △, hGH from Trx-hGH; ◊, hGH from Trx-hGH in presence of DTT; □, hGH from MBP-hGH.</p

Schematic representation of domain structure and generation of the MBP-hGH construct.

<p>(A) Vector map of pHMGWA-hGH using the gateway cloning method. Expression of the fusion proteins in <i>E. coli</i> is controlled by the IPTG-inducible T7 promoter, with ampicillin as the selection marker. (B) Schematic structure of the seven fusion proteins His6-, Trx-, GST-, MBP-, NusA-, PDIb′a′-, and PDI-GH (total size). The arrows indicate the TEV protease cleavage site.</p

MALDI-TOF MS analysis of purified hGH from Trx-hGH in <i>E. coli</i> (no DTT during purification).

<p>(A) Tryptic peptide map of hGH (191 aa). (B) MALDI-TOF MS for purified hGH in reducing conditions. (C) MALDI-TOF MS for the purified hGH in non-reducing conditions. MALDI-TOF MS analysis of reduced and non-reduced hGH obtained from MBP-hGH and PDIb′a′-hGH showed similar results (data not shown).</p

Expression level and solubility of hGH with seven different tags.

<p>Expression level and solubility of hGH with seven different tags.</p

hGH purification from Trx-hGH expressed in <i>E. coli</i>.

<p>(A) Flowchart of the purification. (B) Trx-hGH was purified from <i>E. coli</i> with a combination of IMAC and gel filtration chromatography. M, molecular weight marker; lane 1, total cell protein before IPTG induction as a negative control; lane 2, total cell protein treated with IPTG; lane 3, soluble fraction after cell sonication; lane 4, Trx-hGH fusion protein purified using IMAC (37.4 kDa); lane 5, Trx tag cleavage with TEV protease: Trx (15.4 kDa) and hGH (22 kDa); lane 6, final purified hGH (22 kDa). Lane 5 shows that the Trx tag was almost completely cleaved. (C) Gel filtration chromatogram of PDIb′a′-hGH after second IMAC. hGH and oligomers were separated by their sizes. (D) Purity of final product hGH was evaluated by silver staining. M, molecular weight marker; hGH: final product in non-reducing conditions.</p