Follow‐up of atypia and follicular lesions of undetermined significance in thyroid fine needle aspiration cytology

N. Dincer, S. Balci, A. Yazgan, G. Guney, R. Ersoy, B. Cakir and G. Guler
Follow‐up of atypia and follicular lesions of undetermined significance in thyroid fine needle aspiration cytology Objective: To report our experience of atypia of undetermined significance (AUS)/follicular lesion of undetermined significance (FLUS) rate and outcome. Methods: Among 7658 patients with 19 569 nodules, 524 (2.7%) nodules were diagnosed as AUS/FLUS on fine needle aspiration (FNA). After exclusion of patients with simultaneous nodules that were suspicious for follicular neoplasm or malignancy or that were malignant, 368 (4.8%) patients were diagnosed as AUS/FLUS. The outcome of 146 patients who had undergone surgery or repeated fine needle aspirate at the time of preparation of this study was evaluated. The original FNAs were matched to repeated FNAs and thyroidectomy or diagnostic lobectomy specimens. Results: Seventy‐two (19.6%) of the 368 patients had directly undergone surgery, either a lobectomy or a thyroidectomy: of these, 27 (37.5%) had neoplastic nodules (21 were malignant). Seventy‐four (20.1%) of the 368 patients had repeat FNA. On second FNA, 47 of 74 (63.5%) were benign, three were suspicious for follicular neoplasm, one was malignant and 23 (31.1%) were non‐diagnostic. Four patients had a third FNA: two were AUS/FLUS, one was malignant and one non‐diagnostic. One patient had a fourth FNA, which was diagnosed as AUS/FLUS. Sixteen (21.6%) of 74 patients with repeat FNA had surgery: three of these had neoplastic nodules (two were malignant). Overall, 88 of the 368 (23.9%) patients had a thyroidectomy of which 30 (34.1%) were neoplastic and 23 (26.1%) malignant. The neoplastic rate for patients who were once diagnosed with AUS/FLUS was 8.2% and the malignancy rate 6.3%. The malignancy rate for patients on follow‐up at the time we prepared the study was 15.7% (23/146); 222 remained on follow‐up without surgery or repeat FNA or were managed elsewhere. Conclusions: Although in this category repeat FNA is expected rather than excision, we suggest evaluation of all AUS/FLUS patients in multidisciplinary meetings to decide management and recommend follow‐up of all patients with this diagnosis.     

In vitro inhibition of cytosolic carbonic anhydrases I and II by some new dihydroxycoumarin compounds

A new series of 6, 7-dihydroxy-3-(methylphenyl) chromenones, including three new derivatives, i.e. 6,7-dihydroxy-3-(2-methylphenyl)-2H-chromen-2-one (OPC); 6,7-dihydroxy-3-(3-methylphenyl)-2H-chromen-2-one (MPC); 6,7-dihydroxy-3-(4-methylphenyl)-2H-chromen-2-one (PPC) and one previously described, namely 6,7-dihydroxy-3-phenyl-2H-chromen-2-one (DPC), were synthesized. These compounds were investigated as inhibitors of human carbonic anhydrase I (hCA-I) and human carbonic anhydrase II (hCA-II) which had been purified from human erythrocytes on an affinity gel comprised of L-tyrosine-sulfonamide-Sepharose 4B.     

A molecularly imprinted nanofilm‐based quartz crystal microbalance sensor for the real‐time detection of pirimicarb

This study aimed to prepare a novel quartz crystal microbalance (QCM) sensor for the detection of pirimicarb. Pirimicarb‐imprinted poly (ethylene glycol dimethacrylate‐N‐metacryloyl‐(l )‐tryptophan methyl ester) [p (EGDMA‐MATrp)] nanofilm (MIP) on the gold surface of a QCM chip was synthesized using the molecular imprinting technique. A nonimprinted p (EGDMA‐MATrp) nanofilm (NIP) was also synthesized using the same experimental technique. The MIP and NIP nanofilms were characterized via Fourier transform infrared spectroscopy attenuated total reflectance spectroscopy, contact angle, atomic force microscopy, and an ellipsometer. A competitive adsorption experiment on the sensor was performed to display the selectivity of the nanofilm. An analysis of the QCM sensor showed that the MIP nanofilm exhibited high sensitivity and selectivity for pirimicarb determination. A liquid chromatography‐tandem mass spectrometry method was prepared and validated to determine the accuracy and precision of the QCM sensor. The accuracy and precision of both methods were determined by a comparison of six replicates at three different concentrations to tomato samples extracted by using a Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) method. The limit of detection of the QCM sensor was found to be 0.028 nM. In conclusion, the QCM sensor showed good accuracy, with recovery percentages between 91 and 94%. Also, the pirimicarb‐imprinted QCM sensor exhibited a fast response time, reusability, high selectivity and sensitivity, and a low limit of detection. Therefore, it offers a serious alternative to the traditional analytical methods for pesticide detection in both natural sources and aqueous solutions.     

Insertion site‐based polymorphism markers open new perspectives for genome saturation and marker‐assisted selection in wheat

In wheat, the deployment of marker‐assisted selection has long been hampered by the lack of markers compatible with high‐throughput cost‐effective genotyping techniques. Recently, insertion site‐based polymorphism (ISBP) markers have appeared as very powerful new tools for genomics and genetic studies in hexaploid wheat. To demonstrate their possible use in wheat breeding programmes, we assessed their potential to meet the five main requirements for utilization in MAS: flexible and high‐throughput detection methods, low quantity and quality of DNA required, low cost per assay, tight link to target loci and high level of polymorphism in breeding material. Toward this aim, we developed a programme, IsbpFinder, for the automated design of ISBP markers and adapted three detection methods (melting curve analysis, SNaPshot^® Multiplex System and Illumina BeadArray technology) for high throughput and flexible detection of ISBP or ISBP‐derived SNP markers. We demonstrate that the high level of polymorphism of the ISBPs combined with cost‐effective genotyping methods can be used to efficiently saturate genetic maps, discriminate between elite cultivars, and design tightly linked diagnostic markers for virtually all target loci in the wheat genome. All together, our results suggest that ISBP markers have the potential to lead to a breakthrough in wheat marker‐assisted selection.     

Differential retrotranslocation of mitochondrial Bax and Bak

The Bcl-2 proteins Bax and Bak can permeabilize the outer mitochondrial membrane and commit cells to apoptosis. Pro-survival Bcl-2 proteins control Bax by constant retrotranslocation into the cytosol of healthy cells. The stabilization of cytosolic Bax raises the question whether the functionally redundant but largely mitochondrial Bak shares this level of regulation. Here we report that Bak is retrotranslocated from the mitochondria by pro-survival Bcl-2 proteins. Bak is present in the cytosol of human cells and tissues, but low shuttling rates cause predominant mitochondrial Bak localization. Interchanging the membrane anchors of Bax and Bak reverses their subcellular localization compared to the wild-type proteins. Strikingly, the reduction of Bax shuttling to the level of Bak retrotranslocation results in full Bax toxicity even in absence of apoptosis induction. Thus, fast Bax retrotranslocation is required to protect cells from commitment to programmed death.     

Anti-inflammatory effects of leptin and cholecystokinin on acetic acid-induced colitis in rats: role of capsaicin-sensitive vagal afferent fibers

Leptin and cholecystokinin (CCK) have a synergistic interaction in the suppression of food intake, and afford similar gastroprotective activity. The present study was designed to investigate the putative protective effects of CCK and leptin on acute colonic inflammation. Leptin or CCK-8s was injected to rats intraperitoneally immediately before and 6 h after the induction of colitis with acetic acid. CCK-A receptor antagonist (L-364,718) or CCK-B receptor antagonist (L-365,260) was injected intraperitoneally 15 min before leptin or CCK treatments. In a group of rats, vagal afferent fibers were denervated by topical application of capsaicin on the cervical vagi. Rats were decapitated at 24 h, and the distal 8 cm of the colon were removed for macroscopic scoring, determination of tissue wet weight index (WWI), histologic assessment and tissue myeloperoxidase (MPO) activity. All inflammation parameters were increased by acetic acid-induced colitis compared to control group. Leptin or CCK-8s treatment reduced these parameters in a similar manner, while co-administration of leptin and CCK was found to be more effective in reducing the macroscopic score and WWI. CCK-8s-induced reduction in the score and WWI was prevented by CCK-A, but not by CCK-B receptor antagonist, whereas neither antagonist altered the inhibitory effect of leptin on colitis-induced injury. On the other hand, perivagal capsaicin prevented the protective effects of both CCK-8s and leptin on colitis. Our results indicate that leptin and CCK have anti-inflammatory effects on acetic acid-induced colitis in rats, which appear to be mediated by capsaicin-sensitive vagal afferent fibers involving the reduction in colonic neutrophil infiltration.     

A new PCR-based marker on chromosome 4AL for resistance to pre-harvest sprouting in wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

Pre-harvest sprouting (PHS) is a complex trait controlled by multiple genes with strong interaction between environment and genotype that makes it difficult to select breeding materials by phenotypic assessment. One of the most important genes for pre-harvest sprouting resistance is consistently identified on the long arm of chromosome 4A. The 4AL PHS tolerance gene has therefore been targeted by Australian white-grained wheat breeders. A new robust PCR marker for the PHS QTL on wheat chromosome 4AL based on candidate genes search was developed in this study. The new marker was mapped on 4AL deletion bin 13-0.59-0.66 using 4AL deletion lines derived from Chinese Spring. This marker is located on 4AL between molecular markers Xbarc170 and Xwg622 in the doubled-haploid wheat population Cranbrook × Halberd. It was mapped between molecular markers Xbarc170 and Xgwm269 that have been previously shown to be closely linked to grain dormancy in the doubled haploid wheat population SW95-50213 × Cunningham and was co-located with Xgwm269 in population Janz × AUS1408. This marker offers an additional efficient tool for marker-assisted selection of dormancy for white-grained wheat breeding. Comparative analysis indicated that the wheat chromosome 4AL QTL for seed dormancy and PHS resistance is homologous with the barley QTL on chromosome 5HL controlling seed dormancy and PHS resistance. This marker will facilitate identification of the gene associated with the 4A QTL that controls a major component of grain dormancy and PHS resistance.     

L-Cysteine influx and efflux: a possible role for red blood cells in regulation of redox status of the plasma

The objective of this study was to investigate if erythrocytes play a role in the maintenance of redox homeostasis of the plasma. Thus, we studied L-cysteine efflux and influx in vitro in human erythrocytes. In the present study, we exposed the erythrocytes to different concentrations of L-cysteine and then measured the intracellular free -SH concentrations. Erythrocytes treated in the same manner were later utilized for the cysteine efflux studies. The effect of temperature on the influx and the efflux processes were also evaluated. Change in the free -SH content of the buffer was evaluated as a measure for the presence of an efflux process. The effects of free -SH depletion on L-cysteine transport is also investigated. We also determined the rate of L-cysteine efflux in the presence and absence of buthionine sulfoximine (BSO) in erythrocytes that are pretreated with 1-chloro-2,4-dinitro benzene, a glutathione (GSH) depletory. Our L-cysteine influx studies demonstrated that erythrocytes can respond to increases in L-cysteine concentration in the extracellular media and influx L-cysteine in a concentration-dependent manner. Free -SH concentrations in erythrocytes treated with 1 mM L-cysteine reached to 1.64 +/- 0.06 mM in 1 h whereas this concentration reached to 4.30 +/- 0.01 mM in 10 mM L-cysteine treated erythrocytes. The L-cysteine efflux is also determined to be time-and concentration-dependent. Erythrocytes that are pretreated with higher L-cysteine concentrations displayed a higher efflux process. Outside concentration of free -SH in 1 mM L-cysteine pretreated erythrocytes reached to 0.200 +/- 0.005 mM in 1 h whereas this concentration reached to 1.014 +/- 0.002 with 10 mM L-cysteine pretreated erythrocytes. Our results also indicate that the rate of inward and outward transport of L-cysteine is affected by the oxidative status of the erythrocytes. When GSH is depleted and GSH synthesis is blocked, the L-cysteine uptake and the efflux processes are significantly decreased. Depending on our results, it could be concluded that erythrocytes play a role in the regulation of the plasma redox status and intracellular level of GSH determines the rate of the L-cysteine efflux.     

Synthesis of new sulfonamide inhibitors of carbonic anhydrase

Four different derivatives of aromatic sulfonamides have been synthesized: 1,2-bis[(4-sulfonamidobenzamide)ethoxy]ethane (SBAM), 1,2-bis[(4-sulfonamidobenzoate)ethoxy]ethane, 1,2-bis[(2,4-dichloro-5-sulfonamidobenzamide)ethoxy]ethane, and 1,2-bis[(2,4-dichloro-5-sulfonamidobenzoate)ethoxy]ethane. SBAM is a most potent inhibitor on ciliary epithelium carbonic anhydrase and is approximately 13 times more active against carbonic anhydrase isoform II than against isoform I.