Investigation on the role of apoptosis in aneurysm rupture

Project location: ITALY
Project start date: September 2001 - Project end date: September 2002
Project number: 2001-13
Beneficiary: ATENA Onlus

Aneurysm: 2002 Project Description

The project started in June 2001. Its aim is to identify the out-breaking factors that cause the hemorrhage, that's to say the rupture of the aneurysm. Prof. Giulio Maira hypothesized that apoptosis (programmed cell death) could play a role in aneurysm rupture. Starting from this assumption, the medical staff of Policlinico Gemelli involved in the research financed by the NPF and headed by Prof. Giulio Maira, has already collected some ruptured aneurysms to perform a study on the concrete feasibility of the evaluation of apoptotic features in aneurysm specimens. Such study although performed on few cases, appears clearly encouraging.

Their experience suggests that a minimum period of 1 year appears necessary to obtain reliable specimens (nearly 20 cases) and data. For pure technical reasons, not all specimens obtained at surgery are suitable for a reliable evaluation of apoptosis, nor during all surgical procedures aneurysm specimens can be safely collected. The researchers will be able to obtain reliable preliminary results by the beginning of next year.


Final results 2002

The aim of the research is to identify the out-breaking factors that cause the hemorrhage, that's to say the rupture of the aneurysm. Prof. Maira hypothesized that apoptosis (programmed cell death) could play a role in aneurysm rupture. Starting from this assumption, the medical staff of Policlinico Gemelli involved in the research financed by the NPF and headed by Prof. Giulio Maira, has already collected some ruptured aneurysms to perform a study on the concrete feasibility of the evaluation of apoptotic features in aneurysm specimens. Such study although performed on few cases, appears clearly encouraging. Their experience suggests that a minimum period of 1 year appears necessary to obtain reliable specimens (nearly 20 cases) and data. For pure technical reasons, not all specimens obtained at surgery are suitable for a reliable evaluation of apoptosis, nor during all surgical procedures aneurysm specimens can be safely collected.

Specimens from 27 intracranial aneurysms were collected during surgery, after aneurysm clipping and the verification of its perfect exclusion from brain circulation. Of the 27 specimens, 9 (33%) were obtained from male patients, while 18 (67%), from female patients. 17 (63%) aneurysms were ruptured, 10 were unruptured. The TUNEL (Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling) technique was used to compare by optic microscopy, apoptosis in ruptured and unruptured aneurysms. Such methodology permits the identification of apoptosis, evidentiating its main aspect: DNA fragmentation. Apoptotic levels were also related to: patient's age and sex, aneurismal volume and shape, surgical timing.

Statistically significant differences (p < 0.001), were observed in apoptosis when comparing ruptured and unruptured aneurysms. Higher levels of apoptosis (Class A2-A3), were observed in the majority of rupture aneurysms. Two cases of ruptured aneurysms were classified in class A0-A1. In 9 (90%), unruptured aneurysms apoptosis was not detected. Apoptotic phenomena were evident only in one case of unruptured aneurysm. No statistically significant differences were noted when comparing the levels of apoptosis to patients' age and sex. An irregular aneurysm shape correlated significantly with higher levels of apoptosis. The contact of blood from subarachnoid haemorrhage with aneurismal dome did not influence apoptosis, which appears therefore as a genuine phenomena not depending upon the contact of blood (SAH) with the ruptured aneurysm dome. Studying apoptosis in relation to several variables, brought information on the possible predictability of rupture in unruptured intracranial aneurysm incidentally diagnosed. The evaluation of apoptotic cells by electronic microscopy, permitted to investigate some peculiar ultrastructural carachteristics of apoptosis, evidentiating clearly cells in a "pre-apoptotic" conditions and cells in "mature apoptosis". The features observed clearly suggest for an interpretations of apoptotic phenomena observed, as a cause of aneurysm rupture and not as a consequence of such event.

Conclusions: An innovative approach to the problem of intracranial aneurysms is suggested. The role of apoptosis in aneurysms rupture appeared relevant. Information on rupture predictability of aneurysms was obtained. The development of these studies could influence therapeutic strategies of such lesions. This evaluation of aneurysm in humans, could open the field to future studies, aimed to disclose the exact etiology of intracranial aneurysms enlargement and rupture. We consider our project as a first step toward this target.

Evaluation of Apoptosis

Evaluation Class 0: Absence of apoptosis, or presence of few scattered apoptotic cells in all examined slides
Class A1: Presence of a low number of apoptotic cells in all examined slides
Class A2: Presence of an elevated number of apoptotic cells in all examined slides
Class A3: Presence of a very elevate number of apoptotic cells in all examined slides

Statystical Analyses: test. Values of p < 0.05 were considered as statistically significant.

Compared apoptosis: Ruptured and unruptured aneurysm

Apoptosis A3-A2Apoptosis A1-0
17152
Unruptured A. 1019


test: p < 0.001

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