RET/PTC rearrangement in Hashimoto’s thyroiditis nodules: contribution to an ongoing debate.

An association between Hashimoto thyroiditis (HT) and thyroid carcinoma, has been postulated for decades, although definitive molecular links remain elusive. Activation of RET/PTC oncogene (typical of papillary thyroid carcinoma) has been reported in both non-neoplastic and neoplastic HT thyroid follicular cells (Tallini et al., 2006), but the issue is still controversial. To contribute new insight to this issue, we have investigated RET rearrangement on 207 fine needle aspiration biopsies on unselected nodules from 168 patients and 38 corresponding surgical samples. I-FISH with a homebrew-breakpart DNA probe was used. Normal tissue nuclei cutoff value (mean ± 3DS) was 3%. In addition, RET/PTC was investigated by RT-PCR. Coexistent overt autoimmune thyroid disease (AITD) was assessed by serological (antithyroid autoantibodies, ATA), clinical and echographic data. RET breakage (FISH+) was observed in 6.3% samples. RET/PTC was confirmed by RT-PCR (RT-PCR+) in 23% FISH+ nodules. RT-PCR+ corresponded to FISH+ value ≥ 6,8%. No correlation between RET rearrangement and ATA (5/92 nodules ATA +; 8/115 ATA-), or between AITD (6/78 AITD+; 7/129 AITD-) was observed. Nodules with FISH+ were observed along the whole cytological spectrum, with prevalence significantly higher in suspect or malignant cytology (13.6% TIR 3-5 vs 3.1% TIR 2 nodules). 7/13 FISH+ nodules were surgically removed and available for I-FISH. FISH+ was confirmed in 3 samples, classified as PTC classic type. Our results suggest no association between RET rearrangement and HT. The percentage of FISH+ nodules increases according to the risk of malignancy. Difference between I-FISH and RT-PCR results was observed, presumibly due to the sensitivity of I-FISH strategy. Since we detected RET-PTC m-RNA transcript in ≥ 6,8% FISH+ samples, the oncogenic potential of nodules below this value remains to be clarified, and FISH+ nodules with benign cytology should be considered for a careful follow-up

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field