Immunohistochemical evaluation of cyclin D1 and p63 in odontogenic keratocyst and unicystic ameloblastoma.

INTRODUCTION: Odontogenic keratocyst (OKC) and unicystic ameloblastoma (UA) are lesions of odontogenic origin. Both lesions are morphologically cysts. However, they are classified as developmental cysts and epithelial odontogenic tumours, respectively. Cyclin D1 (CCD1) dysregulation is associated with oncogenic activity and malignancies, while tumour protein p63 (p63) alterations are associated with tumourigenesis. AIM: To evaluate and compare the protein expression of CCD1 and p63 in sporadic OKC (OKC-sp), syndromic OKC (OKC-sy), and UA. MATERIAL AND METHODS: 45 cases from the Anatomical Pathology Department, Faculty of Dentistry, University of Chile were analysed and divided into groups: OKC-sp (n=15), OKC-sy (n=15) and UA (n=15), the latter categorised into intraluminal and/or luminal (n=7) and mural (n=8). Immunohistochemical staining for CCD1 and p63 proteins was performed from paraffin-embedded sections. Statistical analysis included the Shapiro-Wilk test, one-way ANOVA with Tukey's multiple comparisons, and Spearman's correlation coefficient (p<0.05). RESULTS: There was an involvement mainly in women in the mandibular area, and a high frequency of jaw expansion, especially in the mural UA. P63 protein expression was higher than CCD1 in all cystic lesions, particularly in mural UA (p<0.001). No correlation was found between CCD1 and p63 expression. CONCLUSION: P63 may serve as a valuable marker for evaluating cell proliferative activity in odontogenic cystic lesions, providing insights into the aggressive behaviour of mural UA.

p63 expression in granulosa-luteinized cells of infertile patients with peritoneal endometriosis submitted to in vitro fertilization.

OBJECTIVE: Endometriosis is associated with infertility, even without an anatomical abnormality. Furthermore, the peritoneal (mild) phenotype of this disease is the most prevalent and linked to infertility. The present study aimed to investigate the p63 gene and protein expression in granulosa cells from pre-ovulatory follicles in patients with endometriosis and infertility submitted to in vitro fertilization. METHODS: Twenty-eight patients participated in the study and were divided into two groups according to the presence or absence of endometriosis. The p63 gene-expression levels assessment was performed by real-time PCR (qPCR) using the TaqMan assay, and we used immunofluorescence to check the p63 protein expression after IVF. RESULTS: There was no significant difference between the groups regarding age, hormonal levels, oocyte standards, and p63 gene expression. The control group showed an RQ of 1.000 (0.431 to 2.323) and the study group showed an RQ of 0.725 (0.249 to 2.105), p>0.05. Both groups showed a weak expression of the p63 gene (p>0.05). CONCLUSIONS: This study described that endometriosis may not affect the p63 gene expression. Moreover, after follicular recruitment and growth, we found a weak expression of this protein, suggesting it is not part of oocyte maturation and development control.

Can p63 serve as a biomarker for diagnosing giant cell tumor of bone? A systematic review and meta-analysis

ABSTRACT BACKGROUND: Tumor protein p63 (p63) has been reported to be highly expressed in giant cell tumor of bone (GCTB). Whether p63 can be treated as a diagnostic marker for GCTB remains unclear. OBJECTIVE: We conducted a meta-analysis to evaluate the applicability of p63 in diagnosing GCTB. DESIGN AND SETTING: Systematic review and meta-analysis carried out in a public hospital, Hong Kong, China. METHODS: We searched PubMed, EMBASE and the Cochrane Library from inception to April 30, 2019. Literature in English or Chinese about the differential diagnosis of GCTB using p63 were included. ­Animal experiments, reviews, correspondence, case reports, expert opinions and editorials were excluded. Studies were also excluded if they did not provide sufficient information to construct a 2 × 2 contingency table. We calculated individual and pooled sensitivities and specificities. We used I² as an indicator of heterogeneity. RESULTS: Out of 88 records identified, 8 articles on 788 GCTB patients fulfilled the inclusion criteria and were included in the present analysis. Bivariate analyses yielded a pooled mean sensitivity of 0.87 (95% confidence interval, CI, 0.72-0.95) and specificity of 0.71 (95% CI, 0.56-0.82) for using p63 as a biomarker in diagnosing GCTB. The area under the receiver operating characteristic curve was 0.86 (95% CI, 0.82-0.88). CONCLUSION: p63 is a helpful indicator in diagnosing GCTB due to its high sensitivity and specificity. ­Nonetheless, the results need to be carefully interpreted based on other diagnostic methods such as imaging. SYSTEMATIC REVIEW REGISTRATION: 164115 (PROSPERO registration number)

Short-Term Results Analysis in the Allogenic Transplantation of Limbal Stem Cells Expanded on Amniotic Membrane in Patients with Bilateral Limbal Stem Cell Deficiency.

Purpose: The objective of this study was to describe the short-term results of allogenic transplantation of limbal stem cells expanded on amniotic membrane for the ocular surface reconstruction. Methods: Prospective nonrandomized, nonmasked study in a single ophthalmological center. Ten patients with bilateral total limbal stem cell deficiency (LSCD) were included. Expression and presence of ABCB5 and Δp63α in amniotic membrane-cultured limbal epithelial stem cells were analyzed, in relationship with clinical changes after allogenic transplantation. An objective evaluation was performed to determine corneal transparency and superficial vascularization. Results: In a median follow-up time of 11.6 months, 7 patients (70%) were considered as failure compared with the preoperative status. ABCB5 and Δp63α are expressed in similar amount in the limbal epithelial cells expanded in vitro and transplanted in patients with bilateral LSCD. Conclusions: Transplantation of allogenic epithelial limbal cells expanded in amniotic membrane could be considered in patients with LSCD due to burns or congenital etiologies such as aniridia, but its benefit is limited for patients with immunologic diseases.

A Systematic review and Meta-Analysis of the survival and clinicopathological features of p63 expression in Merkel cell carcinoma.

Merkel cell carcinoma (MCC) is a rare skin tumour of neuroendocrine origin with aggressive behaviour. The aims of this study were to investigate the association of p63 + MCC with clinicopathological features and to estimate survival through a systematic review and meta-analysis. A comprehensive search of PubMed, Embase, Scopus and Virtual Health Library following the PRISMA guidelines was conducted on September 2017. DerSimonian and Lard random-effects models were used to calculate survival-weighted means and their corresponding 95% confidence intervals (CI) among studies. Five studies met our inclusion criteria after screening 77 citations and 36 full-text articles. The included studies enrolled 413 patients with MCC. We observed that p63 + MCC was significantly associated with mortality with OR 2.92 (95% CI [1.66-5.13]). The summary hazard ratio of multivariate analysis was 1.99 (95% CI [1.32-3.01]). The only clinicopathological feature associated with p63 + MCC with statistical significance was the Merkel cell polyomavirus (MCPyV) status. The presence of MCPyV was associated as a protective factor for the expression of p63 (OR 0.25, 95% CI [0.08-0.73]). These results support that p63 + MCC evaluated by immunohistochemistry has a poor outcome. Therefore, we suggest p63 to be performed when staging MCC.

Infrared meibography and molecular assessment of p63 gene mutations in a Mexican patient with EEC syndrome. / Meibografía infrarroja y evaluación molecular de la mutación del gen p63 en un paciente mexicano con síndrome EEC.

OBJECTIVE: To report the finding of infrared meibography in a Mexican patient with EEC syndrome (Ectrodactyly-ectodermal dysplasia-cleft syndrome) confirmed by molecular analysis of the p63 gene. CLINICAL CASE: A 31 year-old male patient was seen due to a history of progressive visual loss in both eyes associated with long-term photophobia. The patient was born with cleft lip and palate, ectrodactyly of right hand, and afterwards, displayed nail dysplasia, anodontia and alopecia, with which ectodermal dysplasia was diagnosed. The ophthalmological findings were limited to the adnexa and the ocular surface. In vivo infrared meibography showed total absence of Meibomian glands in the lower eyelids and severe deficiency in the upper eyelids. In addition, it was shown that the patient was a heterozygous carrier of a missense mutation R304W (C → T) in exon 8 of the p63 gene. DISCUSSION: The R304W mutation in the p63 gene region is definitely related to characteristics such as the absence of Meibomian glands.

Lung cancer biopsy: Can diagnosis be changed after immunohistochemistry when the H&E-Based morphology corresponds to a specific tumor subtype?

OBJECTIVES: Advancements in non-small cell lung cancer treatment based on targeted therapies have made the differentiation between adenocarcinoma and squamous cell carcinoma increasingly important. Pathologists are challenged to make the correct diagnosis in small specimens. We studied the accuracy of an immunohistochemical panel in subclassifying non-small cell lung cancer in routine small biopsies and compared the results with the diagnosis from resected lung specimens, autopsy samples or biopsied/resected metastases. METHODS: In total, 340 lung cancer biopsies were investigated for the expression of CK5, TTF1, p63 and surfactant. RESULTS: We characterized 166 adenocarcinomas and 124 squamous cell carcinomas. Overall, 85% of cases displayed binary staining (TTF1 positive/p63 negative, and vice versa). The diagnoses of ten cases with a morphology that indicated a specific tumor subtype were changed after immunohistochemistry (IHC). A second specimen was available for 71 patients, and the first diagnosis at biopsy was confirmed in 95% of these cases. Most non-small cell lung cancer cases present a binary immunohistochemical profile in small biopsies, contributing to good diagnostic accuracy with routine markers. In a small proportion of cases, the diagnosis can be changed after IHC even when the morphological aspects indicate one specific tumor subtype. CONCLUSIONS: We recommend that routine small biopsies of lung cancer without classic morphology should be subjected to a minimum immunohistochemical panel to differentiate adenocarcinoma from squamous cell carcinoma.

A novel c.1037C > G (p.Ala346Gly) mutation in TP63 as cause of the ectrodactyly-ectodermal dysplasia and cleft lip/palate (EEC) syndrome.

Ectrodactyly - ectodermal dysplasia and cleft lip/palate (EEC) syndrome (OMIM 604292) is a rare disorder determined by mutations in the TP63 gene. Most cases of EEC syndrome are associated to mutations in the DNA binding domain (DBD) region of the p63 protein. Here we report on a three-generation Brazilian family with three individuals (mother, son and grandfather) affected by EEC syndrome, determined by a novel mutation c.1037C > G (p.Ala346Gly). The disorder in this family exhibits a broad spectrum of phenotypes: two individuals were personally examined, one presenting the complete constellation of EEC syndrome manifestations and the other presenting an intermediate phenotype; the third affected, a deceased individual not examined personally and referred to by his daughter, exhibited only the split-hand/foot malformation (SHFM). Our findings contribute to elucidate the complex phenotype-genotype correlations in EEC syndrome and other related TP63-mutation syndromes. The possibility of the mutation c.1037C > G being related both to acro-dermato-ungual-lacrimal-tooth (ADULT) syndrome and SHFM is also raised by the findings here reported.

Developmental exposure to bisphenol A alters the differentiation and functional response of the adult rat uterus to estrogen treatment.

We assessed the long-term effect of perinatal exposure to bisphenol A (BPA) on the rat uterus and the uterine response to estrogen (E2) replacement therapy. BPA (0.5 or 50µg/kg/day) was administered in the drinking water from gestational day 9 until weaning. We studied the uterus of female offspring on postnatal day (PND) 90 and 360, and the uterine E2 response on PND460 (PND460-E2). On PND90, BPA-exposed rats showed altered glandular proliferation and α-actin expression. On PND360, BPA exposure increased the incidence of abnormalities in the luminal and glandular epithelium. On PND460-E2, the multiplicity of glands with squamous metaplasia increased in BPA50 while the incidence of glands with daughter glands increased in BPA0.5. The expression of steroid receptors, p63 and IGF-I was modified in BPA-exposed rats on PND460-E2. The long-lasting effects of perinatal exposure to BPA included induction of abnormalities in uterine tissue and altered response to E2 replacement therapy.

Perinatal exposure to diethylstilbestrol alters the functional differentiation of the adult rat uterus.

The exposure to endocrine disrupters and female reproductive tract disorders has not been totally clarified. The present study assessed the long-term effect of perinatal (gestation+lactation) exposure to diethylstilbestrol (DES) on the rat uterus and the effect of estrogen replacement therapy. DES (5µg/kg bw/day) was administered in the drinking water from gestational day 9 until weaning and we studied the uterus of young adult (PND90) and adult (PND360) females. To investigate whether perinatal exposure to DES modified the uterine response to a long-lasting estrogen treatment, 12-month-old rats exposed to DES were ovariectomized and treated with 17ß-estradiol for 3 months (PND460). In young adult rats (PND90), the DES treatment decreased both the proliferation of glandular epithelial cells and the percentage of glandular perimeter occupied by α-smooth muscle actin-positive cells. The other tissue compartments remained unchanged. Cell apoptosis was not altered in DES-exposed females. In control adult rats (PND360), there were some morphologically abnormal uterine glands. In adult rats exposed to DES, the incidence of glands with cellular anomalies increased. In response to estrogens (PND460), the incidence of cystic glands increased in the DES group. We observed glands with daughter glands and conglomerates of glands only on PND460 and in response to estrogen replacement therapy, independently of DES exposure. The p63 isoforms were expressed without changes on PND460. Estrogen receptors α and ß showed no changes, while the progesterone receptor decreased in the subepithelial stroma of DES-exposed animals with estrogen treatment. The long-lasting effects of perinatal exposure to DES included the induction of abnormalities in uterine tissues of aged female rats and an altered response of the adult uterus to estradiol.

Immunohistochemical expression of p63, epidermal growth factor receptor (EGFR) and notch-1 in radicular cysts, dentigerous cysts and keratocystic odontogenic tumors

The aim of this study was to assess the immunohistochemical expression of p63 protein, epidermal growth factor receptor (EGFR) and Notch-1 in the epithelial lining of radicular cysts (RC), dentigerous cysts (DC) and keratocystic odontogenic tumors (KOT). For this study, 35 RC, 22 DC and 17 KOT were used. The clinical and epidemiological data were collected from the patient charts filed in the Oral Pathology Laboratory, University of Ribeirão Preto, Brazil. Immunohistochemical reactions against the p63, EGFR and Notch-1 were performed in 3-µm-thick histological sections. The slides were evaluated according to the following criteria: negative: <5% of positive cells, low expression: 5-50% of positive cells, and high expression: >50% of positive cells. Moreover, the intensity of EGFR and Notch-1 expressions was also evaluated. Fisher's exact test and Spearman's correlation coefficients were used for statistical analysis, considering a significance level of 5%. Almost all cases demonstrated p63, EGFR and Notch-1 expressions. The p63 expression was significantly higher in KOT (p<0.001). Positive correlation between these immunomarkers was observed. These findings suggest the participation of p63, EGFR and Notch-1 in the development, maintenance and integrity of cystic odontogenic epithelial lining, favoring lesion persistence. The high expression of p63 in KOT suggests that it may be related to their more aggressive biological behavior and marked tendency to recurrence.

O objetivo deste estudo foi avaliar a expressão imunoistoquímica da proteína p63, receptor do fator de crescimento epidérmico (EGFR) e Notch-1 no revestimento epitelial de cistos radiculares (CR), cistos dentígeros (CD) e tumores odontogênicos queratocísticos (TOQ). Para este estudo, 35 CR, 22 CD e 17 TOQ foram utilizados. Os dados clínicos e epidemiológicos foram coletados das fichas dos pacientes arquivadas no Laboratório de Patologia Oral, Universidade de Ribeirão Preto, Brasil. Reações imunoistoquímicas contra p63, EGFR e Notch-1 foram realizadas em cortes histológicos de 3 µm. As lâminas foram avaliadas de acordo com os seguintes critérios: negativo <5% das células positivas, baixa expressão - 5%-50% das células positivas e alta expressão >50% das células positivas. Além disso, a intensidade de expressão de EGFR e Notch-1 foi também avaliada. Teste exato de Fisher e coeficiente de correlação de Spearman foram usados para análise estatística, considerando um nível de significância de 5%. Quase todos os casos demonstraram expressão de p63, EGFR e Notch-1. A expressão de p63 foi significativamente maior nos TOQ (p<0.001). Correlação positiva entre os imunomarcadores foi observada. Esses achados sugerem a participação de p63, EGFR e Notch-1 no desenvolvimento, manutenção e integridade do revestimento epitelial cístico, favorecendo a persistência das lesões. A alta expressão de p63 no TOQ sugere que ela pode estar relacionada ao comportamento biológico mais agressivo e marcada tendência a recorrência.

Imunomarcação da p63 em carcinoma espinocelular de boca e orofaringe e comparação com os graus de malignidade / Immunolabelling of p63 in oral and oropharyngeal squamous cell carcinoma and comparison with the degree of differentiation

Introdução: o carcinoma espinocelular (CEC) é a neoplasia maligna mais freqüente da cavidade oral e, apesar dos avanços dos métodos de diagnóstico, a sobrevida ainda é menor que 50%. Marcadores tumorais têm sido pesquisados no intuito de obterem-se informações mais precisas sobre o prognóstico dos pacientes e a proteína p63 é uma das candidatas a desempenhar esse papel. Objetivo: avaliar, por meio de análise imunoistoquímica, a possível associação entre a expressão da p63 e os graus de malignidade de carcinomas de boca e orofaringe. Material e Método: Foram selecionados 14 casos de CEC de boca e orofaringe dos arquivos de dois Serviços de Cirurgia de Cabeça e Pescoço e os cortes obtidos a partir dos blocos de parafina submetidos à análise imunoistoquímica. Os tumores foram classificados em alto e baixo grau de malignidade e os escores comparados com a imunomarcação da p63. Resultados: Seis casos (43%) foram classificados como de baixo grau e, destes, quatro positivos e dois negativos para a p63, enquanto oito casos (57%) classificados como de alto grau, três foram positivos e cinco negativos para p63. Conclusão: houve uma tendência da proteína p63 ser expressa nos tumores com melhor diferenciação, todavia, maior número de casos e estudo em separado das isoformas da p63 são necessários para confirmar esses resultados.

Introduction: Squamous cell carcinoma (SCC) is the most frequent malignant neoplasm of the oral cavity and oropharynx and albeit the advances in diagnosis, the overall survival of the patients is less than 50%. Tumoral markers have been searched in order to obtain more precise information on the prognosis of the patients and p63 protein is one of the candidates do play this role. Objective: to evaluate immunohistochemically the putative association between the expression of p63 protein and the malignity degrees of oral cavity and oropharynx carcinomas. Materials and methods: 14 cases of SCC of oral cavity and oropharynx were selected from the archives of two services of Head and Neck Surgery and the specimens obtained from paraffin blocks underwent imunohistochemical analyzes. The tumors were classified in high or low degree of malignity and the scores compared with the immunolabelling of p63. Results: 6 cases (43%) were classified as low degree and from these, 4 were positive and 3 negative to p63 labelling. The other 8 cases (57%) were classified as high degree and from these, 3 were positive and 5 negative to p63 protein. Conclusion: there was a trend of the p63 protein to be expressed in tumors with higher differentiation, although to confirm these results a higher number of specimens, as well as a study of the different isoforms of the protein are necessary.