Patologie tiroidee e surrenaliche del cane e del gatto: aspetti epidemiologici, clinici e clinico-patologici

L’ipertiroidismo felino rappresenta oggi la più comune endocrinopatia della specie. I capitoli 2 e 3 costituiscono una revisione della letteratura in merito agli aspetti clinici, diagnostici e terapeutici della patologia. Il capitolo 4 indaga il ruolo della dimetilarginina simmetrica (SDMA) come marker di funzionalità renale nei gatti ipertiroidei prima e dopo terapia medica. La patologia tiroidea più comune nel cane è l’ipotiroidismo. Nello studio riportato al capitolo 5 sono state indagate le performance diagnostiche di freeT3, freeT4, rT3, 3,3-T2 e 3,5-T2, misurati tramite LC-MS/MS, nel differenziare tra cani ipotiroidei, cani con patologie non-tiroidee e cani sani. La presenza di una possibile correlazione tra la gravità della condizione clinica dei pazienti ipotiroidei, le variabili emato-chimiche e le concentrazioni sieriche di cTSH è stata valutata nel capitolo 6. Il capitolo 7 valuta l’andamento dell’SDMA in cani ipotiroidei prima e dopo supplementazione ormonale. A differenza della Sindrome di Cushing dell’uomo, che è considerata una malattia rara, nel cane l’ipercortisolismo spontaneo (HC) è una delle endocrinopatie più comuni. Gli aspetti epidemiologici dell’HC e la ricerca di un metodo di monitoraggio alternativo al test di stimolazione con ACTH nei cani trattati con Trilostano sono stati approfonditi rispettivamente nei capitoli 8 e 9. A differenza dell'HC, l'ipoadrenocorticismo primario (PH) è una patologia rara nel cane. Lo scopo dello studio riportato nel capitolo 10 consiste nel descrivere le frazioni escretorie degli elettroliti urinari nei cani con PH e di indagare se esse possano rappresentare un utile supporto alla diagnosi e al trattamento del PH canino. Il riscontro accidentale di masse surrenaliche rappresenta una criticità diagnostica. Infatti, può essere difficile distinguere morfologicamente tra lesioni corticali e midollari e tra lesioni maligne e benigne. Nel capitolo 11 vengono descritti i rilievi immunoistochimici dell'incidentaloma surrenalico nel cane e viene valutato il ruolo del Ki-67 PI come indicatore di malignità.

Efficacia del Tocilizumab nel trattamaneto del rigetto umorale cronico attivo nel paziente trapiantato di rene

In the last decades significant improvements has been reached in short term graft survival, conversely long-term graft survival in still an open challenge for the scientific community. One of the major causes of long term graft loss is represented by chronic- active antibody mediated rejection (cAMR), a recently identified entity whose diagnosis is based on laboratoristic and histologic elements: the presence of DSA associated to specific morphological lesions as inflammation and microvascular damage associated or not to C4d deposition. Treatment of cAMR is an open field of debate. Tocilizumab, an anti-IL6 monoclonal antibody has been recently proposed as a first line treatment for cAMR, showing encouranging results. We describe our monocentric experience using Tocilizumab as first-line therapy for cAMR. Graft function (eGFR), proteinuria and DSA have been evaluated every 6 month for 24 months; histology have been performed after 12 months of treatment. No adverse events have been observed during study period. 12 patients completed the study with a follow-up of 24 months. Kidney function showed a worsening during follow-up that reaches statistical significance at 12 and 24 months (eGFR from 32.2±13.9 ml/min to 26.9±13 ml/min), but far less than expected for these kind of patients. 4 patients (30%) reached ESRD during follow-up, 3 requiring renal replacement therapy. We did not observed any statically significant variation in proteinuria and in DSA MFI levels. From a histological point of view, we observed a significant improvement in active cAMR lesions (C4d deposition and Acute tissue injury (MTA, g>0/ptc>0, v>0) and no progression among chronic lesions (Transplant glomerulopathy, PTC multilayering and aterial intimal fibrosis) Tocilizumab shown good results, with a stabilization of graft function, a reduction in kidney inflammation and active lesions in kidney biopsy and not allowing progression of chronic lesions.

High sensitivity analysis of BRAF mutations in neoplastic and non-neoplastic thyroid lesions

The clonal distribution of BRAFV600E in papillary thyroid carcinoma (PTC) has been recently debated. No information is currently available about precursor lesions of PTCs. My first aim was to establish whether the BRAFV600E mutation occurs as a subclonal event in PTCs. My second aim was to screen BRAF mutations in histologically benign tissue of cases with BRAFV600E or BRAFwt PTCs in order to identify putative precursor lesions of PTCs. Highly sensitive semi-quantitative methods were used: Allele Specific LNA quantitative PCR (ASLNAqPCR) and 454 Next-Generation Sequencing (NGS). For the first aim 155 consecutive formalin-fixed and paraffin-embedded (FFPE) specimens of PTCs were analyzed. The percentage of mutated cells obtained was normalized to the estimated number of neoplastic cells. Three groups of tumors were identified: a first had a percentage of BRAF mutated neoplastic cells > 80%; a second group showed a number of BRAF mutated neoplastic cells < 30%; a third group had a distribution of BRAFV600E between 30-80%. The large presence of BRAFV600E mutated neoplastic cell sub-populations suggests that BRAFV600E may be acquired early during tumorigenesis: therefore, BRAFV600E can be heterogeneously distributed in PTC. For the second aim, two groups were studied: one consisted of 20 cases with BRAFV600E mutated PTC, the other of 9 BRAFwt PTCs. Seventy-five and 23 histologically benign FFPE thyroid specimens were analyzed from the BRAFV600E mutated and BRAFwt PTC groups, respectively. The screening of BRAF mutations identified BRAFV600E in “atypical” cell foci from both groups of patients. “Unusual” BRAF substitutions were observed in histologically benign thyroid associated with BRAFV600E PTCs. These mutations were very uncommon in the group with BRAFwt PTCs and in BRAFV600E PTCs. Therefore, lesions carrying BRAF mutations may represent “abortive” attempts at cancer development: only BRAFV600E boosts neoplastic transformation to PTC. BRAFV600E mutated “atypical foci” may represent precursor lesions of BRAFV600E mutated PTCs.

Immunohistochemical and Molecular Prognostic/Predictive Markers in Neoplastic Diseases

Traditional morphological examinations are not anymore sufficient for a complete evaluation of tumoral tissue and the use of neoplastic markers is of utmost importance. Neoplastic markers can be classified in: diagnostic, prognostic and predictive markers. Three markers were analyzed. 1) Insulin-like growth factor binding protein 2 (IGFBP2) was immunohistochemically examined in prostatic tissues: 40 radical prostatectomies from hormonally untreated patients with their preoperative biopsies, 10 radical prostatectomies from patients under complete androgen ablation before surgery and 10 simple prostatectomies from patients with bladder outlet obstruction. Results were compared with α-methylacyl-CoA racemase (AMACR). IGFBP2 was expressed in the cytoplasm of untreated adenocarcinomas and, to a lesser extent, in HG-PIN; the expression was markedly lower in patients after complete androgen ablation. AMACR was similarly expressed in both adenocarcinoma and HG-PIN, the level being similar in both lesions; the expression was slightly lower in patients after complete androgen ablation. IGFBP2 may be used a diagnostic marker of prostatic adenocarcinomas. 2) Heparan surface proteoglycan immunohistochemical expression was examined in 150 oral squamous cell carcinomas. Follow up information was available in 93 patients (range: 6-34 months, mean: 19±7). After surgery, chemotherapy was performed in 8 patients and radiotherapy in 61 patients. Multivariate and univariate overall survival analyses showed that high expression of syndecan-1 (SYN-1) was associated with a poor prognosis. In patients treated with radiotherapy, such association was higher. SYN-1 is a prognostic marker in oral squamous cell carcinomas; it may also represent a predictive factor for responsiveness to radiotherapy. 3) EGFR was studied in 33 pulmonary adenocarcinomas with traditional DNA sequencing methods and with two mutation-specific antibodies. Overall, the two antibodies had 61.1% sensitivity and 100% specificity in detecting EGFR mutations. EGFR mutation-specific antibodies may represent a predictive marker to identify patients candidate to tyrosine kinase inhibitors therapy.