Proteína prion celular em doenças humanas não-priônicas: aspectos moleculares e celulares / Cellular prion protein in non prionic human diseases: molecular and cellular aspects

RESUMO

Encefalopatias espongiformes são doenças priônicas que afetam humanos e animais. O agente infeccioso, PrPsc é uma isoforma anormal de uma proteína celular, PrPc, que é expressa em todas as células e altamente conservada entre as espécies. As funções até então conhecidas da proteína normal PrPc, a relacionam com neuroplasticidade, neuritogenese, neuroproteção, além de participação na proteção ao estresse oxidativo. Este trabalho abordou algumas destas características avaliando a proteína PrPc em algumas doenças humanas não priônicas. Baseado em dados obtidos em camundongos que não expressam a proteína PrPc, onde várias alterações foram descritas como hiperexcitabilidade neuronal, além de alta sensibilidade a agentes convulsivantes, investigamos o envolvimento de PrPccom síndromes epilépticas humanas. Analisamos a seqüência do gene que codifica PrPc (Prnp) em indivíduos com epilepsia refratária ao tratamento farmacológico portadores de epilepsia de lobo temporal mesial associada à esclerose hipocampal (ELTM-EH, n=100) e epilepsias focais associadas a anormalidades do desenvolvimento cortical (ADC, n=68), que foram comparadas com um grupo controle (n=180)... Nossos resultados mostraram que não há variantes alélicas prevalentes nestes pacientes. Investigamos então, a expressão da proteína PrPc por método imunoistoquímico, nos gliomas mais prevalentes, os astrocitomas... Acrescentamos ao estudo a pesquisa da expressão de STI-1 (stress inducible protein-I) ligante de PrPc associado a neuroproteção, n-NOS (neuronal nitric oxide sintase) e I-NOS (inducible nitric oxide sintase), enzimas de síntese de óxido nítrico, que estão relacionadas a função de proteção a estresse oxidativo descrito para a proteína PrPc (AU)

ABSTRACT

Prions are the infectious agents that cause neurodegenerative diseases, called Transmissible Spongiform Encephalopathies, both in humans and animais. In fact, they are a counterpart o f the cellular prion protein (PrPc) which is expressed in most cell types and is conserved among species, suggesting its important role in the cellular physiology. PrPc has a pivotal role in protection against oxidative stress, neuritogenesis, neuroplasticity and neuroprotection. Therefore, the present work aimed to evaluate the PrPc participation human maladies other than prion diseases based on these functions of the normal protein. Mice with the PrPc gene (Prnp) ablated show neuronal hiperexcitability and enhanced sensitivity to seizures, indicating that PrPc might be related to epilepsy. Here~ we evaluated the genetic contribution of Prnp to the human medically untreatable Mesial Temporal Lobe Epilepsy related to Hippocampal Sclerosis MTLE-HS) and in Malformations of Cortical Development (MCD). DNA obtained fron1 peripheral blood cells was used to evaluate Prnp coding sequence from 100 patients with surgically treated MTLE-HS and 68 patients with different forms of (MCD). These data were further compared with a control healthy group with similar demographic characteristics (n=180). A Prnp variant allele at codon 171, Asn171Ser, which was absent in the contro l group, was highly prevalent in patients with MTLEHS (23%) and MCD (13%). This rare polymorphism was also associated with the worst surgical outcome in MTLE-HS. Seizure history has been associated with some brain tumors and our next approach was evaluate Prnp variant alleles in patients with gliomas (n = 49). Some ofthe Prnp polymorphisms were present, however their frequency was similar to that found in the normal population. PrPc has been associated to protection against oxidative stress and its binding to the Stress Inducible protein 1 (STil) mediates neuroprotection. The we investigated PrPc, STI-1 and nvo oxidative enzymes i-NOS and n-NOS, expression in astrocyton1.as (n = 109), using inm1unohistochemistry. Our data shows that astrocitomas grade II and III have lower PrPc, STI-1 and n-NOS leveis when compared with normal brain tissues. However, glioblastomas showed higher PrPc and n-NOS expression than astrocitomas II and III. These results indicate that PrPc expression could be related to tumor malignancy. We further performed the same expression analysis using a tissue microarray constructed with 25 different human tumors neuroblastoma, retinoblastoma, pancreatic and prostatic adenocarcinoma, invasive ductal breast cancer, carcinoid tumor, malignant melanoma and head and neck squamous cell carcinoma. The correspondent normal tissue was also included in this construction. PrPc and i-NOS presented a high expression in invasive ductal breast cancer in contrast with normal breast tissue where no stain was observed in ductal cells. In retinoblasto1nas, STI-1 expression was lower in non differentiated tumors than in differentiated ones. On the other hand, PrPc tends to decrease in invasive and metastatic tumors particularly in neuroblastomas. Finally, our data points that PrPc polymorphisms contribute to a higher predisposition to epilepsies and is also a predictive factor for surgery outcome. Furthermore, we observed an alteration on PrPc and its ligand STil expression pattern in human tumors sue h as astrocytomas, retino blastomas and ductal invasive breast carcinoma, suggesting that these proteins can be implicated in the tumor process. Further investigations will be necessary to delineate the cellular and molecular mechanisms involved with these observations (AU)