Carcioma tiroideo en perros : aspectos moleculares

Esta tesis incluye dos estudios. El primero tuvo por objetivos conocer la casuística del cáncer en perros en general y estudiar factores asociados al mismo; así como también recabar información clínica y análisis colaterales del carcinoma tiroideo en particular. Se realizó un estudio descriptivo retrospectivo tomando como base de datos el Hospital de la Facultad de Veterinaria, Montevideo, Uruguay en el período 2005-2010 para calcular la casuística del cáncer. En los carcinomas localizados en la zona ventral de cuello se registró además signos clínicos, análisis colaterales, resultado histopatológico y presencia de metástasis. Para el año 2010 se registró la edad, sexo y raza de todos los otros casos clínicos atendidos por el Hospital cuyos diagnósticos excluían el cáncer (Grupo control). La edad se categorizó en 3: 0 a 5 años (jóvenes), 6 a 10 años (mediana edad) y 11 a 20 años (gerontes). La casuística de cáncer en perros atendidos en el Hospital de Facultad de Veterinaria en el período 2005-2010 estuvo entre 0.11 y 0.15. El género afectó la frecuencia de aparición de cáncer, el odds ratio (razón de posibilidades) de que una hembra presente cáncer respecto al macho fue de 2.0. El efecto de la edad resultó un factor determinante en la presencia del cáncer: el odds ratio de que un perro de 6 a 10 años presente cáncer respecto de un perro joven (0 a 5 años) es de 8.1; valor que aumenta a 12.5 para animales gerontes (>10 años). La proporción de casos con cáncer respecto de otros casos clínicos no neoplásicos tendió a estar afectada por la raza, siendo la Bóxer la más afectada. El tipo de cáncer presente estuvo afectado por la edad, ya que si bien la mayoría de los canceres fueron más frecuentes en animales de mediana a avanzada edad (> 5 años), los tumores venéreos transmisibles fueron más frecuentes en animales jóvenes, los óseos se distribuyeron equitativamente entre las 3 categorías etarias y los linfáticos fueron más frecuentes en animales jóvenes y gerontes. En el segundo estudio se investigó la expresión de genes específicos de tiroides en la glándula sana, en la glándula carcinomatosa y en el lóbulo tiroideo contralateral (CL). Se determinaron los transcriptos del receptor de tirotropina (TSH-R), tiroglobulina (Tg), tiroperoxidasa (TPO), factor de transcripción tiroideo 1 (TTF-1), gen pareado box 8 (PAX-8), factor de crecimiento similar a insulina 1 (IGF-1) y receptor de estrógeno alfa (ER?) por PCR en tiempo real en carcinomas caninos, en el CL y en glándulas tiroideas sanas. Las concentraciones de mRNA del TSH-R, PAX-8, IGF-1 y ER? no fueron diferentes entre grupos. El grupo carcinoma tuvo un nivel de expresión de mRNA menor para Tg y TPO que el grupo sano y contralateral, no encontrándose diferencias entre estos dos últimos grupos; lo que sugiere que el tejido carcinomatoso tendría alterada la capacidad para sintetizar hormonas tiroideas. La expresión del TTF-1 fue mayor en el grupo CL con respecto a las glándulas tiroideas sanas y carcinomatosas que no difirieron entre sí. Teniendo en cuenta que los resultados de los transcriptos de Tg y TPO en la glándula carcinomatosa reflejan una actividad de síntesis de hormonas tiroideas alterada y que el TTF-1 promueve la proliferación de los tirocitos, se puede sugerir que el aumento de este factor en el CL podría implicar un mecanismo compensatorio para mantener la condición de eutiroideo encontrada en la mayoría de los perros incluidos en el ensayo

Karakterizacija endokrino-metaboličkog profila koji se koristi za procjenu funkcije štitnjače kod pasa pasmina engleski i francuski buldog

This study investigates whether breed or gender affect serum hormone and metabolite concentrations used to evaluate thyroid function in the Bulldog breed. Sixty-seven healthy adult English Bulldogs (n = 20), French Bulldogs (n = 17), German Shepherds (n = 15) and mongrels (n = 15) of both sexes were selected. Determination of serum total thyroxine (TT4), free thyroxine (FT4), and thyroid stimulating hormone (TSH) was performed via a competitive enzymatic chemiluminescent solid-phase immunoassay. Cholesterol and triglyceride concentrations were analyzed by spectrophotometry. Serum concentrations of TT4, FT4, TSH, cholesterol, and triglycerides for French and English Bulldogs were within the international reference ranges for the canine population. Breed had a significant effect on serum levels of TT4 (P = 0.0012) and FT4 (P<0.0001); English and French Bulldogs had higher serum TT4 and FT4 concentrations than German Shepherds and mongrels. Gender had a significant effect only on serum FT4 levels; females exhibited higher levels (P = 0.0309). Cholesterol, triglycerides, and TSH serum concentrations did not differ with breed or gender. Healthy French and English Bulldogs included in this study had higher serum concentrations of TT4 and FT4 compared with German Shepherds and mongrels, and the concentration of FT4 was also higher in females.Ovim radom istraženo je utječu li pasmina i spol na koncentracije hormona i metabolita u serumu koji služi za procjenu funkcije štitnjače u pasa pasmine buldog. Uključeno je ukupno šezdeset i sedam odraslih zdravih pasa oba spola, među kojima je bilo 20 engleskih buldoga, 17 francuskih buldoga, 15 njemačkih ovčara i 15 križanaca. Ukupni tiroksin (TT4), slobodni tiroksin (FT4) i hormon koji stimulira štitnjaču (TSH) određeni su pomoću kompetitivne enzimske kemiluminoscentne imunoanalize. Koncentracije kolesterola i triglicerida analizirane su spektrofotometrijom. Koncentracije TT4, FT4, TSH, kolesterola i triglicerida u serumu za francuske i engleske buldoge bile su unutar međunarodnih referentnih raspona za populaciju pasa. Pasmina je imala signifikantan učinak na razinu TT4 (p = 0,0012) i FT4 (p< 0,0001) u serumu, pri čemu su engleski i francuski buldozi imali veće koncentracije TT4 i FT4 u serumu nego njemački ovčari i križanci. Spol je imao signifikantan utjecaj samo na razinu FT4 u serumu, pri čemu je ta razina bila viša kod ženki (p = 0,0309). Kolesterol, trigliceridi i koncentracije TSH seruma nisu se razlikovali ovisno o pasmini ili spolu. Zdravi francuski i engleski buldozi uključeni u ovo istraživanje su, u usporedbi s njemačkim ovčarima i križancima, imali veće koncentracije TT4 i FT4 u serumu. Također, koncentracija FT4 u serumu bila je viša kod ženki

Carcinoma trioideo en caninos : mecanismos moleculares y tratamiento

Esta tesis, que se presenta en cuatro artículos, investigó aspectos moleculares del carcinoma tiroideo canino y su tratamiento. En el Artículo I se investigó la expresión de transcriptos determinados por PCR en tiempo real en tiroides sanas, carcinomatosas y en el lóbulo tiroideo contralateral (CL) sano de los perros con cáncer. El grupo carcinoma tuvo una concentración de mRNA menor para Tiroglobulina (Tg) y Tiroperoxidasa (TPO) que el grupo sano y el CL, lo que sugiere que tiene alterada la síntesis de hormonas tiroideas. Las concentraciones de mRNA del receptor de tirotropina (TSH-R), gen pareado box 8 (PAX-8), y receptor de estrógeno alfa (ER?) no fueron diferentes entre grupos. La expresión del TTF-1 y del factor de crecimiento similar a insulina 1 (IGF-I) fue mayor en el grupo CL con respecto a la glándula carcinomatosa, lo que sugiere un mecanismo compensatorio para mantener la condición de eutiroideo en los perros con cáncer. En el Artículo II se determinó la inmunotinción de TSH-R, Tg, TTF-1, antígeno nuclear de proliferación celular (PCNA), factor de crecimiento vascular (VEGF) y de crecimiento fibroblástico 2 (FGF-2) en glándulas tiroideas sanas y carcinomatosas. El número de folículos fue menor y la densidad celular y número de vasos sanguíneos fue mayor en glándulas carcinomatosas respecto las sanas. No se encontraron diferencias en la inmunotinción de TSH-R y Tg entre grupos en células foliculares y fibroblastos. Las células foliculares carcinomatosas presentaron 2 y 3 veces más intensidad de tinción de TTF-1 y PCNA respectivamente respecto las sanas. La tinción de VEGF en células endoteliales fue mayor en los carcinomas. La mayor expresión de proteínas relacionadas a la proliferación y angiogénesis (TTF-1, PCNA y VEGF) encontrada en el carcinoma es consistente con los mecanismos de progresión tumoral. El objetivo del Artículo III fue determinar si marcadores como VEGF, FGF-2, IGF-1 y su receptor (IGF-1R) así como los receptores de ácido retinoico (RAR? y RXR) se asociaban con un patrón histológico diferencial (folicular compacto vs compacto vs sanos). La expresión de IGF-I, IGF-1R y VEGF en fibroblastos y células endoteliales fue mayor en carcinomas compactos que tejido sano y se encontró una mayor expresión de IGF-1R en fibroblastos y FGF-2 en células endoteliales del carcinoma compacto respecto de los foliculares-compactos. La mayor expresión de factores vinculados a la oncogénesis en células endoteliales y fibroblastos encontrada en los carcinomas compactos sostiene la relevancia de las células estromales en la generación del tumor. En el Artículo IV se evaluó el efecto de isotretinoína 9-cis (AR9-cis) sobre la recurrencia tumoral y sobrevida como tratamiento novedoso frente al tradicional (doxorrubicina) en el carcinoma tiroideo post cirugía, incluyendo un grupo control (sin tratamiento). El tipo de carcinoma afectó el tiempo de recurrencia y el de sobrevida, siendo mayor en los carcinomas foliculares respecto de los carcinomas foliculares-compactos, que a su vez fueron mayores que los carcinomas compactos. Existió una interacción entre el tratamiento y el tipo de carcinoma, ya que no se encontró efecto del tratamiento en los carcinomas foliculares, mientras que en los otros tipos de carcinoma, los perros tratados con AR9-cis presentaron mayor tiempo de recurrencia y sobrevida, indicando que este tratamiento utilizado luego de la cirugía es superior al tratamiento tradicional con doxorrubicina.This thesis presented in four articles, investigated the molecular basis of thyroid carcinoma in dogs and its treatment. In Article I, transcript expression was determined by real time PCR in healthy thyroid glands, carcinomas and in the healthy contra lateral lobe (CL) of dogs with cancer. Thyroglobulin (Tg), and thyroperoxidase (TPO) mRNA expression in carcinoma glands was lower than in healthy glands, suggesting an altered capacity to synthesize thyroid hormones. Concentrations of thyrotropin receptor (TSH-R), paired box 8 transcription factor (PAX-8), and estrogen receptor alpha (ERα) mRNA were not different between groups. The expression of thyroid specific transcription factor-1 (TTF-1) and insulin like growth factor (IGF-I) was greater in the CL group than in carcinoma group, suggesting that the CL lobe may function in a compensatory state. In Article II, the average immmunostaining of TSH-R, Tg, TTF-1, vascular endothelial growth factor (VEGF), fibroblastic growth factor-2 (FGF-2) and proliferating cell nuclear antigen (PCNA) were determined in follicular carcinomas and healthy thyroid canine glands. The number of follicles was lower, while cell density and number of blood vessels were greater in thyroid carcinomas than in healthy thyroid glands. No differences were detected in the intensity of staining of TSH-R and Tg in follicular cells and fibroblasts between healthy and carcinomas tissue. Follicular cells of the carcinoma tissue presented 2 to 3-fold greater staining of TTF-1 and PCNA respectively than cells of healthy thyroid tissue. VEGF staining was stronger in endothelial cells of the tumoral tissue when compared with normal thyroid tissue, but no differences were found in the other cell types. The greater protein expression of factors related to proliferation and angiogenesis (TTF-1, PCNA and VEGF) found in the carcinoma glands respect healthy thyroid tissue, is consistent with the mechanisms of tumor progression. The objective of Article III was to determine if markers like VEGF, FGF-2, IGF-1 and its receptor (IGF-1R) and retinoic receptors (RARα and RXR) were associated with the tumor histological type (compact carcinomas vs follicular-compact vs healthy thyroid glands). IGF-I, IGF-1R and VEGF expression was greater in fibroblasts and endothelial cells of compact carcinoma than healthy glands. IGF-1R expression in fibroblasts and FGF-2 expression in endothelial cells of compact carcinoma were greater than follicular-compact carcinoma. The greater expression of factors related to proliferation and angiogenesis in endothelial cells and/or fibroblasts of compact carcinoma when compared to healthy glands supports the relevance of stromal cells on oncogenesis. In Article IV, the effects of isotretinoína 9-cis (AR9-cis) as a novel post surgery treatment of thyroid carcinoma when compared to a traditional treatment (doxorubicin) was investigated and a control untreated group was included. The time to recurrence was significantly shorter in the control group and doxorubicin group vs AR9-cis group, while no differences were found among doxorubicin and control groups. The type of carcinoma affected time of recurrence and survival as follicular carcinomas presented a greater times than follicular-compact carcinomas which in turn presented greater survival time than compact carcinomas. The interaction among treatment and type was significant as recurrence and survival times in follicular carcinomas did not differ between treatments, but AR9-cis dogs had greater times in follicular-compact and compact carcinomas, showing that this treatment is superior to the traditional (doxorubicin)

Cortisol secretion after adrenocorticotrophin (ACTH) and Dexamethasone tests in healthy female and male dogs

<p>Abstract</p> <p>Background</p> <p>For the conclusive diagnosis of Cushing's Syndrome, a stimulating ACTH test or a low suppressive Dexamethasone test is used. Reports in other species than the dog indicate that plasma cortisol concentration after ACTH administration is affected by gender. We investigated the effect of gender on the cortisol response to ACTH and Dexamethasone tests in dogs.</p> <p>Methods</p> <p>Seven healthy adult Cocker Spaniels (4 females and 3 males) were assigned to a two by two factorial design: 4 dogs (2 females and 2 males) received IV Dexamethasone 0.01 mg/kg, while the other 3 dogs received an IV saline solution (control group). Two weeks later the treatments were reversed. After one month, ACTH was given IV (250 μg/animal) to 4 dogs (2 female and 2 males) while the rest was treated with saline solution (control group). Cortisol concentrations were determined by a direct solid-phase radioimmunoassay and cholesterol and triglycerides by commercial kits.</p> <p>Results and Discussion</p> <p>No effect of treatment was observed in metabolite concentrations, but females presented higher cholesterol concentrations. ACTH-treated dogs showed an increase in cortisol levels in the first hour after sampling until 3 hours post injection. Cortisol concentrations in Dexamethasone-treated dogs decreased one hour post injection and remained low for 3 hours, thereafter cortisol concentrations increased. The increase in cortisol levels from one to two hours post ACTH injection was significantly higher in females than males. In Dexamethasone-treated males cortisol levels decreased one hour post injection up to 3 hours; in females the decrease was more pronounced and prolonged, up to 5 hours post injection.</p> <p>Conclusion</p> <p>We have demonstrated that cortisol response to ACTH and Dexamethasone treatment in dogs differs according to sex.</p

Differential branching fraction and angular analysis of the decay B0→K∗0μ+μ−

The angular distribution and differential branching fraction of the decay B 0→ K ∗0 μ + μ − are studied using a data sample, collected by the LHCb experiment in pp collisions at s√=7 TeV, corresponding to an integrated luminosity of 1.0 fb−1. Several angular observables are measured in bins of the dimuon invariant mass squared, q 2. A first measurement of the zero-crossing point of the forward-backward asymmetry of the dimuon system is also presented. The zero-crossing point is measured to be q20=4.9±0.9GeV2/c4 , where the uncertainty is the sum of statistical and systematic uncertainties. The results are consistent with the Standard Model predictions

Opposite-side flavour tagging of B mesons at the LHCb experiment

The calibration and performance of the oppositeside flavour tagging algorithms used for the measurements of time-dependent asymmetries at the LHCb experiment are described. The algorithms have been developed using simulated events and optimized and calibrated with B + →J/ψK +, B0 →J/ψK ∗0 and B0 →D ∗− μ + νμ decay modes with 0.37 fb−1 of data collected in pp collisions at √ s = 7 TeV during the 2011 physics run. The oppositeside tagging power is determined in the B + → J/ψK + channel to be (2.10 ± 0.08 ± 0.24) %, where the first uncertainty is statistical and the second is systematic

Search for CP violation in D+→ϕπ+ and D+s→K0Sπ+ decays

A search for CP violation in D + → ϕπ + decays is performed using data collected in 2011 by the LHCb experiment corresponding to an integrated luminosity of 1.0 fb−1 at a centre of mass energy of 7 TeV. The CP -violating asymmetry is measured to be (−0.04 ± 0.14 ± 0.14)% for candidates with K − K + mass within 20 MeV/c 2 of the ϕ meson mass. A search for a CP -violating asymmetry that varies across the ϕ mass region of the D + → K − K + π + Dalitz plot is also performed, and no evidence for CP violation is found. In addition, the CP asymmetry in the D+s→K0Sπ+ decay is measured to be (0.61 ± 0.83 ± 0.14)%

Measurement of the Bs0→J/ψKS0B_s^0\to J/\psi K_S^0 branching fraction

The $B_s^0\to J/\psi K_S^0$ branching fraction is measured in a data sample corresponding to 0.41$fb^{-1}$ of integrated luminosity collected with the LHCb detector at the LHC. This channel is sensitive to the penguin contributions affecting the sin2$\beta$ measurement from $B^0\to J/\psi K_S^0$ The time-integrated branching fraction is measured to be $BF(B_s^0\to J/\psi K_S^0)=(1.83\pm0.28)\times10^{-5}$. This is the most precise measurement to date

Measurement of the CP-violating phase \phi s in Bs->J/\psi\pi+\pi- decays

Measurement of the mixing-induced CP-violating phase phi_s in Bs decays is of prime importance in probing new physics. Here 7421 +/- 105 signal events from the dominantly CP-odd final state J/\psi pi+ pi- are selected in 1/fb of pp collision data collected at sqrt{s} = 7 TeV with the LHCb detector. A time-dependent fit to the data yields a value of phi_s=-0.019^{+0.173+0.004}_{-0.174-0.003} rad, consistent with the Standard Model expectation. No evidence of direct CP violation is found.Comment: 15 pages, 10 figures; minor revisions on May 23, 201

Measurement of the CP-violating phase phi_s in the decay Bs->J/psi phi

We present a measurement of the time-dependent CP-violating asymmetry in B_s -> J/psi phi decays, using data collected with the LHCb detector at the LHC. The decay time distribution of B_s -> J/psi phi is characterized by the decay widths Gamma_H and Gamma_L of the heavy and light mass eigenstates of the B_s-B_s-bar system and by a CP-violating phase phi_s. In a sample of about 8500 B_s -> J/psi phi events isolated from 0.37 fb^-1 of pp collisions at sqrt(s)=7 TeV we measure phi_s = 0.15 +/- 0.18 (stat) +/- 0.06 (syst) rad. We also find an average B_s decay width Gamma_s == (Gamma_L + Gamma_H)/2 = 0.657 +/- 0.009 (stat) +/- 0.008 (syst) ps^-1 and a decay width difference Delta Gamma_s == Gamma_L - Gamma_H} = 0.123 +/- 0.029 (stat) +/- 0.011 (syst) ps^-1. Our measurement is insensitive to the transformation (phi_s,DeltaGamma_s --> pi - phi_s, - Delta Gamma_s.Comment: 9 pages, 3 figure