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A new index of agonal state for neurological disease

A major concern for researchers working on gene expression in post mortem human brain tissue is whether the conditions that occur immediately prior to death, the agonal state, affect RNA integrity within the brain. To study this requires an assessment of the agonal state and the quality of isolated brain RNA. Here, we propose an index of agonal state (IAS) score to quantify the factors most likely to affect RNA isolated from brain tissue. The IAS clearly differentiated death following chronic neurological disease from sudden death, but no correlation was observed between the IAS score and the quality of RNA isolated from the brain. It has been suggested that the agonal state is a major determinant of brain RNA quality [1–9]. Agonal factors that have been postulated to have an impact on RNA quality include hypoxia, dehydration and seizures [3,4,10]. A method for assessing agonal state was proposed by Tomita et al. [11]. This scored as present or absent a list of agonal factors but did not take into account the extent of these factors, disease duration or histopathological features. To include these, we have designed a new measure, the IAS that includes 10 different parameters (Table 1). To test the ability of this score to distinguish between chronic neurological disease and ‘control’ sudden death brains, we determined the IAS for 78 cases sourced from two brain banks, the MRC National Creutzfeldt–Jakob Disease Surveillance Unit (NCJDSU) Brain and Tissue Bank and the MRC Sudden Death Brain and Tissue Bank. The first brain bank contained neuropathologically confirmed cases of prolonged neurological disease (ND), including variant Creutzfeldt–Jakob disease (vCJD) as well as suspected cases of vCJD that on diagnosis were shown to be cases of other neurological diseases (OND). The second brain bank contained control non-neurological disease cases (NND) with effectively no agonal duration. Both banks have ethical approval for the storage and local use of these cases. The scores for each of the parameters in our IAS value were determined from clinical case histories, autopsy reports and histopathological assessments substantiated by discussions with clinicians, nurse practitioners and neuropathologists. Neurological disease duration was defined as the length of time from first onset of symptoms until death. Onset of symptoms for ND ranged from 1 to 72 months with a mean duration of 13 months. For all cases from the NCJDSU brain and tissue bank, an in-depth questionnaire had been completed by next of kin and/or the patient for surveillance purposes, and this provided detailed disease duration. The NND cases were examples of sudden death with no known underlying disease and were scored 0 for disease duration. For vCJD and OND cases, seizures, coma and pyrexia and neurotoxic substances were clearly recorded in clinical notes and scored as absent 0, or present 1. In these groups, if hypoglycaemia or dehydration were mentioned in the clinical notes, these were scored 1 if present or 2 if severe. If there was no mention of these factors, the case was scored 0. For NND cases, if the cause of death was alcohol or drug related, this was taken as evidence of hypoglycaemia and dehydration and given a combined score of 1. It is generally accepted that alcoholor drug-related deaths commonly show this type of biochemical derangement. If the drug was neurotoxic (excluding alcohol), this was also scored 1 for neurotoxic substance. Based on clinical notes and autopsy reports, mode of death was classified into four groups using a modification of the method described by Hardy et al. [3]: (i) violent fast death due, for example, to gun shot (accidental, homicidal or suicidal) or blunt force trauma (typically road traffic accidents); (ii) fast death of natural cause, or sudden unexpected death of an otherwise healthy individual (typically suspension by ligature); (iii) intermediate death, unexpected death of a sick patient, which cannot be classified as fast or slow death (typical cases include myocardial infarctions); and (iv) slow death, death after a long period of illness with a prolonged terminal phase. Multiple organ failure was identified from autopsy reports. The degree of hypoxic and/or ischaemic change was identified from neuropathology reports and scored 1 as present or 2 as severe; if no mention of these features was made, the case was scored 0. To allow for incomplete data from the NND group, two