Naslov
Chromatography-based isolation of intracranial extracellular vesicles from patients with severe traumatic brain injury

Autori
Krapić, Mia ; Malenica Staver, Mladenka ; Kuharić, Janja ; Kučić, Natalia ; Sotošek Tokmadžić, Vlatka ; Lučin, Pero ; Grabušić, Kristina

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Abstract Book of 8th student congress of neuroscience (NeuRi) / - , 2018, 54-54

Skup
NeuRi 2018 (8th student congress of neuroscience)

Mjesto i datum
Rab, Hrvatska; Rijeka, Hrvatska, 20.04.2018. - 22.04.2018

Vrsta sudjelovanja
Predavanje

Vrsta recenzije
Podatak o recenziji nije dostupan

Ključne riječi
Extracellular vesicles, size-exclusion chromatography, traumatic brain injury, cerebrospinal fluid

Sažetak
Extracellular vesicles (EVs) are nanosized spherical particles enclosed by phospholipid membrane and secreted by cells in physiologic and pathologic conditions. EVs can indicate ongoing processes in tissues and organs since i) they participate in intercellular communication and their content reflects the condition and type of cells they originate from ; and ii) EVs are released into body fluids and thus can be accessed. Recently it was shown that intracranial EVs after severe traumatic brain injury (TBI) change their composition and might indicate neuroregenerative processes in the brain. As such, EVs could be prognostic markers of neuronal recovery after severe TBI in which intracranial damage is caused by external trauma and can have life-threatening consequences due to the impairment of brain functions. However, comprehensive changes in TBI-EV composition are not known and depend on quality of isolated EVs. Here, we show an in- house developed chromatography-based isolation of total EVs from cerebrospinal fluid (CSF) during acute phase after severe TBI. Our study included 6 TBI patients who underwent ventriculostomy placement for intracranial pressure management. The samples of TBI-CSF were collected at days 1, 3, and 7 after injury. Samples of control-CSF were derived from 6 healthy subjects who underwent lumbar puncture as part of the standard diagnostic procedure. Obtained clinical CSF samples were combined into 4 pools (control and TBI-day 1, 3 and 7) and applied to size-exclusion chromatography on sepharose CL-6B. Up to 80 chromatography fractions per analysed CSF-pool were collected by gravity flow and subsequently used for protein characterisation. Soluble proteins as determined by Bradford assay were efficiently separated from EVs which were identified by 2 independent methods: measurement of acetylcholinesterase activity and immunodetection of CD9 tetraspanin. In conclusion, our chromatography protocol provides efficient and cost-effective method to isolate total EVs which can be used for further analyses.

Izvorni jezik
Engleski

Znanstvena područja
Interdisciplinarne prirodne znanosti, Biotehnologija u biomedicini (prirodno područje, biomedicina i zdravstvo, biotehničko područje)

POVEZANOST RADA