Microfluidic Droplet Digital PCR Is a Powerful Tool for Detection of BRAF and TERT Mutations in Papillary Thyroid Carcinomas.

We examined the utility of microfluidic digital PCR (dPCR) for detection of BRAF and TERT mutations in thyroid tumors. DNA extracted from 100 thyroid tumors (10 follicular adenomas, 10 follicular cancers, 5 medullary cancers, and 75 papillary thyroid cancer (PTC) were used for detection of BRAF and TERT mutations. Digital PCRs were performed using rare mutation SNP genotyping assays on QuantStudio 3D platform. In PTCs, BRAFV600E was detected by dPCR and Sanger sequencing in 42/75 (56%) and in 37/75 (49%), respectively. BRAFV600E was not detected in other tumors. The ratio of mutant/total BRAF alleles varied from 4.7% to 47.5%. These ratios were higher in classical PTCs (27.1%) as compared to follicular variant PTCs (9.4%) p = 0.001. In PTCs with and without metastases, the ratios of mutant/total BRAF alleles were 27.6% and 18.4%, respectively, (p = 0.03). In metastatic lesions percentages of mutant/total BRAF alleles were similar to those detected in primary tumors. TERTC228T and TERTC250T were found in two and one cases, respectively, and these tumors concomitantly harbored BRAFV600E. These tumors exhibited gross extra-thyroidal extension, metastases to lymph nodes, and pulmonary metastases (one case). Our results showed that dPCR allows quantitative assessment of druggable targets in PTCs and could be helpful in a molecular-based stratification of prognosis in patients with thyroid cancer.

Identification and characterization of novel Helicobacter pylori apo-fur-regulated target genes.

In Helicobacter pylori, the ferric uptake regulator (Fur) has evolved additional regulatory functions not seen in other bacteria; it can repress and activate different groups of genes in both its iron-bound and apo forms. Because little is understood about the process of apo-Fur repression and because only two apo-Fur-repressed genes (pfr and sodB) have previously been identified, we sought to expand our understanding of this type of regulation. Utilizing published genomic studies, we selected three potential new apo-Fur-regulated gene targets: serB, hydA, and the cytochrome c553 gene. Transcriptional analyses confirmed Fur-dependent repression of these genes in the absence of iron, as well as derepression in the absence of Fur. Binding studies showed that apo-Fur directly interacted with the suspected hydA and cytochrome c553 promoters but not that of serB, which was subsequently shown to be cotranscribed with pfr; apo-Fur-dependent regulation occurred at the pfr promoter. Alignments of apo-regulated promoter regions revealed a conserved, 6-bp consensus sequence (AAATGA). DNase I footprinting showed that this sequence lies within the protected regions of the pfr and hydA promoters. Moreover, mutation of the sequence in the pfr promoter abrogated Fur binding and DNase protection. Likewise, fluorescence anisotropy studies and binding studies with mutated consensus sequences showed that the sequence was important for apo-Fur binding to the pfr promoter. Together these studies expand the known apo-Fur regulon in H. pylori and characterize the first reported apo-Fur box sequence.

Experimental study of Ti-24Nb-4Zr-7.9Sn alloy after treated with micro-arc oxidation in vivo / 华西口腔医学杂志

<p><b>OBJECTIVE</b>To study the biocompatibility of Ti-24Nb-4Zr-7.9Sn (TNZS) ahoy treated with micro-arc oxidation (MAO).</p><p><b>METHODS</b>The tibia bones of New Zealand rabbits were used to build the animal model. TNZS and MAO-TNZS samples were implanted into one side of tibia, pure titanium samples were implanted into the other side as control. After 4 and 26 weeks, radiographs and HE staining technique was used to observe the dynamic remodeling process of bone-implant interface.</p><p><b>RESULTS</b>As the cure time increased, it was showed well biocompatibility of all implants. X-ray indicated that there was no permeable area produced around the three different materials at each time point. The density of bone matrix and arrangement of bone trabecula was almost the same as in the host bone. It was revealed by histological examination that the MAO-TNZS greatly prompted the bonding ability between implant and surrounding hard tissues. Four weeks after implantation, fine attachment was found at the bone-implant interface of all the implants and the fibrous tissue at the interface was gradually remodeled to form new bone. Twenty-six weeks later, MAO-TNZS showed that a biological fixation was created between bone and oxidation layer, while a layer of fibers formed between non-coated TNZS and titanium implants surrounding bone.</p><p><b>CONCLUSION</b>The Ti-24Nb-4Zr-7.9Sn after treated with micro-arc oxidation shows good biocompatibility and can stimulate the bone growth in the bone-implant region, which provides support for clinical usage tests of TNZS alloy as implant after treated with micro-arc oxidation.</p>